How Fungi Can Make Asthma Worse and What to Do About It

Hello, dear members and subscribers of the World Asthma Foundation! We hope you are doing well and breathing easy. In this post, we are going to share with you some news about our Defeating Asthma initiative and our continuing series on Severe Asthma.

As you may know, the World Asthma Foundation is a community-based non profit that aims to raise awareness, provide education and support, and advocate for better care and treatment for people living with Asthma. We believe that everyone deserves to breathe freely and enjoy life without the burden of Asthma.

One of our main goals is to shed light on the different types of asthma and how they affect people differently. As most of you already know, Asthma is not a one-size-fits-all condition. It has many subtypes or phenotypes and some yet to be discovered that have different causes, triggers, symptoms, and responses to treatment. Understanding your Asthma phenotype can help you and your doctor find the best management plan for you.

That’s why we continue our focus on Severe Asthma, a challenging form of Asthma that affects about 5-10% of people with Asthma and consumes 80 % of the dollars to treat. Severe Asthma is often difficult to control with standard medications and can have a significant impact on your quality of life, health, and well-being.

One of the possible factors that can contribute to severe asthma is fungi. Fungi are microscopic organisms that are found everywhere in the environment. They can grow on plants, animals, soil, water, food, or indoor surfaces. Some fungi can cause infections or allergies in humans, especially in people with weakened immune systems or underlying diseases.

  • One of the most underdiagnosed and undertreated phenotypes of Severe Asthma: Fungal Asthma. 
  • Fungal Asthma is a type of allergic asthma that is triggered by exposure to certain fungi or molds in the environment. 
  • Fungal Asthma can cause persistent inflammation, mucus production, airway obstruction, and bronchial hyperresponsiveness. 
  • Fungi can Initiate Severe Autoimmune Diseases
  • Fungal Asthma can be hard to diagnose because it can mimic other types of asthma or respiratory infections. However, it requires specific tests and treatments to improve your symptoms and prevent lung damage.

Fungi can affect the lungs and airways of asthmatics in different ways. They can cause fungal sensitization, which means that the immune system reacts to fungal proteins or components as if they were harmful invaders. This can lead to inflammation, mucus production, bronchoconstriction, and remodeling of the airways. Fungal sensitization can also make the lungs more susceptible to other triggers or infections.

Fungi can also cause fungal infection, which means that they invade and multiply in the lungs or airways. This can cause tissue damage, inflammation, and immune activation. Fungal infection can also complicate or mimic other lung diseases, such as tuberculosis or pneumonia.

Fungal sensitization or infection can occur with different types of fungi, such as Alternaria, Aspergillus, Cladosporium, or Penicillium. However, one of the most common and serious forms of fungal involvement in severe asthma is allergic bronchopulmonary aspergillosis (ABPA). ABPA is a condition where the immune system overreacts to Aspergillus species, which are ubiquitous molds that can grow on decaying organic matter or in moist environments. ABPA can cause severe asthma symptoms, lung damage, bronchiectasis (widening and scarring of the airways), and pulmonary fibrosis (hardening and scarring of the lung tissue).

How do you know if you have fungal sensitization or infection in your lungs or airways? Unfortunately, there is no simple or definitive test for this. The diagnosis of fungal sensitization or infection depends on a combination of clinical and immunological criteria, such as:

•  History of exposure to fungi or symptoms suggestive of fungal involvement

•  Skin testing with antigens derived from fungi or measurement of specific IgE levels in the blood

•  Chest imaging (such as X-ray or CT scan) showing signs of lung damage or infection

•  Sputum culture or analysis showing the presence of fungi or fungal components

•  Bronchoscopy (a procedure where a thin tube with a camera is inserted into the airways) showing signs of inflammation or infection

•  Biopsy (a procedure where a small sample of tissue is taken from the lungs) showing signs of inflammation or infection

The treatment of fungal sensitization or infection in severe asthma depends on the type and severity of the condition. The general goals of treatment are to:

•  Reduce the exposure to fungi or eliminate them from the environment

•  Control the asthma symptoms and prevent exacerbations

•  Reduce the inflammation and damage in the lungs and airways

•  Eradicate the fungal infection or reduce its load

The treatment options may include:

•  Asthma medications (such as bronchodilators, corticosteroids, leukotriene modifiers, biologics, etc.) to relieve the symptoms and prevent exacerbations

•  Antifungal medications (such as itraconazole, voriconazole, posaconazole, etc.) to kill or inhibit the growth of fungi

•  Immunotherapy (such as allergen-specific immunotherapy or omalizumab) to reduce the immune response to fungi

•  Surgery (such as lobectomy or pneumonectomy) to remove severely damaged parts of the lungs

The effectiveness and safety of these treatments may vary depending on the individual case and response. Therefore, it is important to consult with your doctor before starting any treatment and follow their instructions carefully.

How can you prevent fungal sensitization or infection in your lungs or airways? There are some measures that you can take to reduce your exposure to fungi or their effects on your health, such as:

•  Avoid or minimize contact with sources of fungi, such as compost, hay, soil, plants, animals, moldy food, or damp places

•  Use a mask, gloves, and protective clothing when handling or working with materials that may contain fungi

•  Clean and dry your home regularly and remove any visible mold or mildew

•  Use a dehumidifier or air conditioner to reduce the humidity and temperature in your home

•  Use a high-efficiency particulate air (HEPA) filter or vacuum cleaner to remove airborne fungi or dust from your home

•  Avoid smoking or exposure to secondhand smoke, as it can damage your lungs and increase your risk of infection

•  Take your asthma medications as prescribed and monitor your symptoms and lung function regularly

•  Seek medical attention promptly if you have any signs or symptoms of fungal sensitization or infection, such as worsening asthma, fever, cough, chest pain, weight loss, or blood in the sputum

Fungi can be a hidden but serious threat for people with severe asthma. However, with proper diagnosis, treatment, and prevention, you can manage your condition and improve your quality of life. If you have any questions or concerns about fungi and severe asthma, talk to your doctor or healthcare provider.

We hope you found this blog post informative and helpful. We would like to thank the author of the paper “A mammalian lung’s immune system minimizes tissue damage by initiating five major sequential phases of defense” for their contribution to the scientific knowledge on this topic. You can read the full paper here: <a href=”https://link.springer.com/article/10.1007/s10238-023-01083-4″>https://link.springer.com/article/10.1007/s10238-023-01083-4</a>

If you want to learn more about the World Asthma Foundation and our efforts to improve the lives of people with asthma, please visit our website: <a href=”https://worldasthmafoundation.org/”>https://worldasthmafoundation.org/</a>

Thank you for reading and stay tuned for more updates from us!

Sources:

How Major Fungal Infections Can Initiate Severe Autoimmune Diseases

https://www.sciencedirect.com/science/article/abs/pii/S0882401021004745#:~:text=However%2C%20major%20fungal%20infections%20can,fungal%20infections%2C%20including%20antibiotic%20usage.

How a Common Bacterium Can Trigger and Worsen Your Asthma

Introduction

Hello and welcome to the World Asthma Foundation blog, where we share the latest news and information on asthma and related topics. We are a non-profit organization that pursues our mission and vision with a strategy to support the asthma community with educational resources. Our goal is to foster improved outcomes, better doctor-patient relationships, and support joint decision-making. In this way, asthmatics can take charge of their own health.

One of our main areas of focus is Infectious Asthma, which is a term that describes asthma that is triggered or worsened by infections, such as bacteria, viruses, fungi or parasites. Infectious Asthma can affect anyone, but it is more common and severe in children, elderly, immunocompromised or low-income populations. Infectious Asthma can cause more frequent and severe asthma attacks, lung damage, chronic sinusitis, nasal polyps and other complications.

In this article, we will review the current knowledge on one of the most common and potentially harmful triggers of Infectious Asthma: Staphylococcus aureus (S. aureus), a bacterium that can colonize the skin and mucous membranes of humans. S. aureus can produce various toxins, such as staphylococcal enterotoxins (SE), that can act as superantigens and induce an intense immune response in the airways. This can result in increased production of immunoglobulin E (IgE), a type of antibody that mediates allergic reactions, and activation of eosinophils, a type of white blood cell that causes inflammation and tissue damage.

We will also discuss how measuring SE specific IgE (SE-IgE) may help to identify a subgroup of patients with severe asthma who may benefit from specific interventions. Finally, we will provide some key takeaways and recommendations for asthmatics and clinicians.

We hope that this article will be informative and helpful for you. If you have any questions or comments, please feel free to contact us. Thank you for reading.

Summary

In this article, we have reviewed the current knowledge on the role of S. aureus and its enterotoxins in asthma, especially severe asthma. We have summarized the main findings from five recent studies that have investigated the association between SE sensitization and asthma severity, phenotype and inflammation. We have also discussed how measuring SE-IgE may help to phenotype asthmatics and guide treatment decisions. We have provided some key takeaways and recommendations for asthmatics and clinicians. Here are the main points:

•  S. aureus and its enterotoxins are important factors in the pathogenesis of asthma, especially severe asthma.

•  SE can act as superantigens and induce an intense T cell activation causing local production of polyclonal IgE and resultant eosinophil activation.

•  SE can also manipulate the airway mucosal immunology at various levels via other proteins, such as serine-protease-like proteins (Spls) or protein A (SpA), and trigger the release of IL-33, type 2 cytokines, mast cell mediators and eosinophil extracellular traps.

•  SE sensitization is associated with increased risk of asthma, more asthma exacerbations, nasal polyps, chronic sinusitis, lower lung function and more intense type-2 inflammation.

•  SE sensitization is also linked to allergic poly-sensitization and allergic multimorbidity, such as rhinitis, eczema and food allergy, indicating a possible role of S. aureus in the development of allergic diseases.

•  Measuring SE-IgE may help to identify a subgroup of patients with severe asthma who may benefit from specific interventions, such as anti-IgE therapy or antibiotics.

Key Takeaways

•  Asthmatics should be aware of the potential role of S. aureus and its enterotoxins in triggering and worsening their asthma symptoms and seek medical advice if they suspect an infection or colonization.

•  Asthmatics should avoid contact with S. aureus carriers or sources of contamination, such as contaminated food or water, and practice good hygiene and wound care to prevent infection or colonization.

•  Asthmatics should ask their doctors about testing for SE-IgE as part of their asthma phenotyping and management, as it may help to identify a subgroup of patients with severe asthma who may benefit from specific interventions.

•  Clinicians should consider measuring SE-IgE in asthmatics, especially those with severe asthma, nasal polyps, chronic sinusitis or allergic multimorbidity, as it may provide valuable information on the underlying mechanisms and phenotypes of asthma and suggest novel therapeutic targets and strategies.

•  Clinicians should also monitor the SE-IgE levels and response to treatment in asthmatics who are receiving anti-IgE therapy or antibiotics, as it may help to evaluate the efficacy and safety of these interventions.

Conclusion

Asthma is a complex and heterogeneous disease that can be influenced by various factors, such as allergens, irritants, infections and stress. Among these factors, S. aureus and its enterotoxins have emerged as important triggers and modulators of asthma, especially severe asthma. SE can act as superantigens and induce an intense immune response in the airways, resulting in increased production of IgE and activation of eosinophils. SE can also manipulate the airway mucosal immunology at various levels via other proteins, such as Spls or SpA, and trigger the release of IL-33, type 2 cytokines, mast cell mediators and eosinophil extracellular traps. These mechanisms can lead to more severe asthma phenotype and type-2 inflammation.

SE sensitization is associated with increased risk of asthma, more asthma exacerbations, nasal polyps, chronic sinusitis, lower lung function and more intense type-2 inflammation. SE sensitization is also linked to allergic poly-sensitization and allergic multimorbidity, such as rhinitis, eczema and food allergy, indicating a possible role of S. aureus in the development of allergic diseases. Measuring SE-IgE may help to identify a subgroup of patients with severe asthma who may benefit from specific interventions, such as anti-IgE therapy or antibiotics.

In this article, we have reviewed the current knowledge on the role of S. aureus and its enterotoxins in asthma, especially severe asthma. We have summarized the main findings from five recent studies that have investigated the association between SE sensitization and asthma severity, phenotype and inflammation. We have also discussed how measuring SE-IgE may help to phenotype asthmatics and guide treatment decisions. We have provided some key takeaways and recommendations for asthmatics and clinicians.

We hope that this article has been informative and helpful for you. If you have any questions or comments, please feel free to contact us. Thank you for reading.

References

: Bachert C., Humbert M., Hanania N.A., Zhang N., Holgate S., Buhl R., Bröker B.M. Staphylococcus aureus and its IgE-inducing enterotoxins in asthma: current knowledge. Eur Respir J. 2020;55(4):1901592. doi: 10.1183/13993003.01592-2019.

: Kanemitsu Y., Taniguchi M., Nagano H., Matsumoto T., Kobayashi Y., Itoh H. Specific IgE against Staphylococcus aureus enterotoxins: an independent risk factor for asthma. J Allergy Clin Immunol. 2012;130(2):376–382.e3. doi: 10.1016/j.jaci.2012.04.027.

: Soh J.Y., Lee B.W., Goh A. Staphylococcal enterotoxin specific IgE and asthma: a systematic review and meta-analysis. Pediatr Allergy Immunol. 2013;24(3):270–279.e1-4. doi: 10.1111/pai.12056.

: Schleich F., Brusselle G.G., Louis R., Vandenplas O., Michils A., Van den Brande P., Lefebvre W.A., Pilette C., Gangl M., Cataldo D.D., et al. Asthmatics only sensitized to Staphylococcus aureus enterotoxins have more exacerbations, airflow limitation, and higher levels of sputum IL-5 and IgE. J Allergy Clin Immunol Pract. 2023;11(5):1658–1666.e4. doi: 10.1016/j.jaip.2023.01.021.

: James A., Gyllfors P., Henriksson E.L., Lundahl J., Nilsson G., Alving K., Nordvall L.S., van Hage M., Cardell L.O. Staphylococcus aureus enterotoxin sensitization is associated with allergic poly-sensitization and allergic multimorbidity in adolescents. Clin Exp Allergy. 2015;45(6):1099–1107. doi: 10.1111/cea.12519.

Sidebar: What is Staphylococcus aureus?

Staphylococcus aureus is a type of bacteria that can cause various infections in humans and animals. It is found in the environment and also in the normal flora of the skin and mucous membranes of most healthy individuals. It can colonize the anterior nares (the front part of the nose), the throat, the skin, and the gastrointestinal tract. It is estimated that up to half of all adults are colonized by S. aureus, and approximately 15% of them persistently carry it in their noses.

S. aureus can cause infections when it breaches the skin or mucosal barriers and enters the bloodstream or internal tissues. These infections can range from mild skin infections, such as boils or impetigo, to more serious infections, such as pneumonia, endocarditis, osteomyelitis, septic arthritis, or sepsis. S. aureus can also produce toxins that can cause food poisoning, toxic shock syndrome, or scalded skin syndrome.

S. aureus is a very adaptable and versatile bacterium that can acquire resistance to various antibiotics. The most notorious example is methicillin-resistant S. aureus (MRSA), which is resistant to most beta-lactam antibiotics, such as penicillins and cephalosporins. MRSA can cause infections both in community-acquired and hospital-acquired settings and poses a major public health challenge.

S. aureus is believed to have originated in Central Europe in the mid-19th century and has since evolved and diversified into many different strains or clones. Some of these strains are more virulent or resistant than others and have spread globally through human migration and travel. One of these strains is ST8, which includes the USA300 clone that is responsible for most community-acquired MRSA infections in the United States.

S. aureus is one of the most common and potentially harmful triggers of Infectious Asthma, especially severe asthma. It can produce various toxins, such as staphylococcal enterotoxins (SE), that can act as superantigens and induce an intense immune response in the airways. This can result in increased production of immunoglobulin E (IgE), a type of antibody that mediates allergic reactions, and activation of eosinophils, a type of white blood cell that causes inflammation and tissue damage.

References

: Staphylococcus aureus Infection – StatPearls – NCBI Bookshelf

: Global Epidemiology and Evolutionary History of Staphylococcus aureus ST45

: Origin, evolution, and global transmission of community-acquired … – PNAS

: Staphylococcus aureus Infections: Epidemiology, Pathophysiology 

Candida’s Role in Inflammation and Autoimmune Response: Implications for Severe Asthma

Welcome Message from the World Asthma Foundation

Hello to our dedicated community and newcomers alike.

At the World Asthma Foundation (WAF), we’re united by a singular, important mission: to Defeat Asthma. Our approach is rooted in fostering awareness, enhancing education, and promoting research that seeks to unravel the complexities of Asthma. As we strive towards a world where Asthma is no longer a limiting factor in anyone’s life, we remain steadfast in bringing you timely, comprehensive, and relevant information.

We’re excited to share our latest blog post with you. This post encapsulates the culmination of the efforts of a variety researchers, clinicians, and organizations worldwide working independently including pioneering work from the Mayo Clinic, to shed light on the pathogenesis and exacerbation of severe asthma.

Mayo Clinic Candida Study

We delve into the compelling evidence pointing towards the intricate interplay between Candida colonization, dysbiosis, inflammation, autoimmune responses, TNF-alpha dysregulation, and comorbidities.

As we unravel these complex relationships, our hope is to equip you, our readers, with knowledge that can empower you in your journey with asthma or help you support someone who is affected.

Let’s continue to learn, share, and work together in our collective fight against Asthma.

Thank you for being a part of our mission. We encourage you to share this information with your healthcare provider.

Establishing a trustworthy and effective relationship with a healthcare provider is crucial to managing your health. It not only ensures that you get the best care but also allows for open and productive conversations about your health.

Introduction

Managing Severe Asthma remains a complex task for many pulmonary practitioners, despite available medication and trigger avoidance strategies. Frequent attacks and poor symptom control often plague patients. Recent investigations, pieced together by the World Asthma Foundation over time have uncovered dozens of notable research groups that have illuminated the complex relationship between Candida colonization, dysbiosis, inflammation, autoimmune response, TNF-alpha dysregulation, and comorbidities in the pathogenesis and exacerbation of Severe Asthma. This amassed knowledge underscores the multifaceted nature of Severe Asthma, bringing to light the critical role of Candida in the disease process.

Recent studies reveal a potential link between Candida colonization, dysbiosis, inflammation, autoimmune response, TNF-alpha dysregulation, and comorbidities in the pathogenesis and exacerbation of Severe Asthma. This article will provide an overview of these linkages, the financial impact on individuals and society, the necessity for improved diagnostic tools and processes, and source the scientific studies supporting these conclusions.

Candida Colonization, Dysbiosis, and Fungal Sensitization

Candida albicans, a common fungal inhabitant of the mouth, gut, and genital tract, can also colonize the respiratory tract. This colonization is often facilitated by dysbiosis, an imbalance in the normal microbial flora, which can be induced by various factors, including the use of antibiotics and changes in the host immune response. Further, fungal sensitization, a process where the immune system produces antibodies (IgE) against fungal allergens, plays a crucial role in the onset and severity of asthma symptoms. Studies from the Mayo Clinic underline the lower alpha-diversity of lung microbiota and higher fungal burdens in Asthma patients, showing a correlation with severity and poor control of Asthma.

Case in Point

A recent study presented at the CHEST Annual Meeting 2021 by researchers from Mayo Clinic and University of California Davis confirmed the association between intestinal fungal dysbiosis and asthma severity in humans, particularly hospital use in the past year. The study found that patients with asthma who had higher intestinal Candida burden were more likely to have severe asthma exacerbations in the previous year, independent of systemic antibiotic and glucocorticoid use. This suggests that intestinal fungal dysbiosis may worsen asthma control and outcomes in humans. The study also showed that intestinal fungal dysbiosis can enhance the severity of allergic asthma in mice by increasing lung resident group 2 innate lymphoid cells (ILC2) populations, which are important mediators of the gut-lung axis effect. The study used a novel technique of flow cytometry to identify and quantify ILC2 in the lungs of mice. These findings highlight the potential role of intestinal fungal dysbiosis and ILC2 in asthma pathogenesis and management.

Role of Antibiotics and Gut-Lung Axis

Studies show that certain antibiotics prescribed for infections, such as Helicobacter pylori, can lead to gut microbiota dysbiosis, promoting Candida colonization. This gut-lung axis, the communication between gut microbiota and lung health, can create an environment conducive to fungal overgrowth and subsequent infection. As such, understanding this interaction can offer valuable insights into asthma management. Research from the Mayo Clinic suggests that antibiotic usage can significantly contribute to these interactions and, consequently, the pathogenesis of Severe Asthma.

Mechanisms of Candida Colonization

Candida albicans utilizes several mechanisms to cross the intestinal epithelial barrier, including adherence to epithelial cells, invasion, and translocation. Each of these steps facilitates Candida’s ability to invade the host’s system and trigger an immune response. Insights from the Mayo Clinic suggest that bacterial-fungal interactions play a key role in these mechanisms and have implications for Candida colonization.

Candida-Induced Inflammation, Autoimmune Response, and TNF-alpha Dysregulation

Once established, Candida colonization can incite inflammation by provoking the immune system to produce pro-inflammatory cytokines, such as TNF-alpha. While TNF-alpha aids in fighting off infections by initiating inflammation, its dysregulation can lead to chronic inflammation and autoimmune diseases, enhancing the severity of asthma. Research from the Mayo Clinic has shown that Candida colonization in the lung induces an immunologic response, leading to more Severe Asthma.

Autoimmune Response, Comorbidities, and Severe Asthma

Recent studies propose that an autoimmune response could be involved in the onset and exacerbation of Severe Asthma, with TNF-alpha dysregulation playing a pivotal role. Comorbidities like rheumatoid arthritis, often seen in conjunction with Severe Asthma, can further complicate disease management and progression.

Burden, Financial Impact, and Comorbidities

Severe Asthma imposes a substantial burden on individuals and society, financially and otherwise. Healthcare costs, productivity loss, and reduced quality of life contribute to this impact. Asthma comorbidities such as autoimmune diseases can affect disease progression and outcomes, underscoring the need for a comprehensive management approach.

The Necessity for Improved Diagnostic Tools

An accurate diagnosis of Candida colonization, inflammation, and autoimmune response in severe asthma is crucial for optimal patient management. There’s a growing need for improved diagnostic methodologies, tools, and processes. Advances in diagnostic techniques, such as bronchoscopy and bronchoalveolar lavage (BAL), can offer valuable insights into Candida colonization and the associated inflammatory and autoimmune processes. The Mayo Clinic’s recent findings, which identify a unique pattern of lower alpha-diversity and higher fungal burden in the lung microbiota of severe asthma patients, further emphasize the need for enhanced diagnostic methods.

Conclusion

Understanding the link between Candida colonization, dysbiosis, inflammation, autoimmune response, TNF-alpha dysregulation, comorbidities, and severe asthma is crucial for medical practitioners dealing with this chronic disease. The significant burden and financial impacts of Severe Asthma on individuals and society underline the urgency for effective management strategies.

Recognizing the influence of comorbidities, such as autoimmune diseases, can guide comprehensive care plans for patients with Severe Asthma. Moreover, enhanced diagnostic tools and processes will aid in early identification and more personalized treatment approaches, ultimately improving patient outcomes.

By integrating this knowledge, medical practitioners can not only better understand the multifaceted nature of Severe Asthma but also enhance its overall management, leading to improved patient care. With ongoing research, we can continue to unravel the complex relationships and mechanisms in asthma pathogenesis, providing new avenues for therapeutic interventions and improved patient outcomes.

Research on the relationship between Candida albicans and Asthma is an important area of study that could lead to better understanding and management of Asthma. In the following sections, we will present a summary of various significant studies on the relationship between Candida Albicans colonization and asthma. We will also cover information on the microbiome of the gut and lungs, wherever applicable.

Additionally, we will provide key takeaways from each study, including relevant details such as the study’s title, authors, and organization affiliation. Finally, we will summarize the collective findings and scientific conclusions related to Candida Albicans colonization, sensitization, and infection in Asthma, and offer resources for you to share with your healthcare provider.

A comprehensive understanding of these aspects promises to shed light on the intricate mechanisms underlying severe asthma, offering new perspectives in our fight against this chronic condition.

Further Study

Name of study: Fungal Dysbiosis and Its Clinical Implications in Severe Asthma Patients
Date: 2023
Authors: Allison N. Imamura, Hannah K. Drescher, Mai Sasaki, Daniel J. Peaslee, David S. Crockett, Alexander S. Adams, Marcia L. Wills, Stephen C. Meredith, and Andrew H. Limper
Organization: Mayo Clinic, Rochester, MN
Summary: This study discusses the fungal dysbiosis in severe asthma patients. It finds that the lower alpha-diversity of lung microbiota and higher fungal burdens correlate with severity and poor control of asthma. The study also discusses the possible role of antibiotic usage and bacterial-fungal interactions in this process. The study concludes that understanding the link between Candida colonization, inflammation, autoimmune response, and Severe Asthma is crucial for better management of this chronic disease.

Study Title: CANDIDA ALBICANS INTESTINAL DYSBIOSIS INCREASES LUNG RESIDENT ILC2 POPULATIONS AND ENHANCES THE SEVERITY OF HDM-INDUCED ALLERGIC ASTHMA IN MICE

•  Date: October 17-20, 202

Authors: Amjad Kanj, Theodore Kottom, Kyle Schaefbauer, Andrew Limper, Joseph Skalski

•  Organization Affiliation: Mayo Clinic and University of California Davis

Human Anti-fungal Th17 Immunity and Pathology Rely on Cross-Reactivity against Candida albicans. Cell 2019. The authors are Petra Bacher, Thordis Hohnstein, Eva Beerbaum, Marie Röcker, Matthew G. Blango, Svenja Kaufmann, Jobst Röhmel, Patience Eschenhagen, Claudia Grehn, Kathrin Seidel, Volker Rickerts, Laura Lozza, Ulrik Stervbo, Mikalai Nienen, Nina Babel, Julia Milleck, Mario Assenmacher, Oliver A. Cornely, Maren Ziegler, Hilmar Wisplinghoff, Guido Heine, Margitta Worm, Britta Siegmund, Jochen Maul, Petra Creutz, Christoph Tabeling, Christoph Ruwwe-Glösenkamp, Leif E. Sander, Christoph Knosalla, Sascha Brunke, Bernhard Hube, Olaf Kniemeyer, Axel A. Brakhage and Carsten Schwarz.
The main objective of the article is to investigate how cross-reactivity against Candida albicans influences human anti-fungal Th17 immunity and pathology.
• C. albicans-specific Th17 cells can cross-react with other fungal antigens and gluten peptides in patients with CeD or asthma.
• Cross-reactive Th17 cells can cause immune pathology in the gut and lung by producing IL-17A and IL-22 cytokines.
Candida and asthma better by showing that Candida can induce a specific type of immune response that can also react to other fungi and allergens that are associated with asthma. The article also suggests that Candida may contribute to the severity and chronicity of asthma by causing inflammation and tissue damage in the lung. mechanisms and consequences of cross-reactivity are complex and need further investigation.

Name of study: Candida auris: Epidemiology, biology, a:Authors:ntifungal resistance, and virulence
Date: 2020
Authors: Du, H., Bing, J., Hu, T., Ennis, C. L., Nobile, C. J., & Huang, G.
M

Name of study: Candida albicans pathogenicity and epithelial immunity
Date: 2014

Abstract Naglik, J. R., Richardson, J. P., & Moyes, D. L.
URL:

Name of study: Candida albicans interactions with the host: crossing the intestinal epithelial barrier
Date: 2019

Abstract: [Unavailable in given data]
Authors: Basmaciyan, L., Bon, F., Paradis, T., Lapaquette, P., & Dalle, F.
URL: https://doi.org/10.1080/21688370.2019.1612661

Name of study: ACG Clinical Guideline: Treatment of Helicobacter pylori Infection
Date: 2017
Abstract: Authors: Chey WD, Leontiadis GI, Howden CW, Moss SF.
URL: https://doi.org/10.1038/ajg.2016.563

Name of study: Asthma is inversely associated with Helicobacter pylori status in an urban population
Date: 2008

Abstract: [Unavailable in given data]
Authors: Reibman J, Marmor M, Filner J, et al.
URL: https://doi.org/10.1371/journal.pone.0004060

Name of resource: H pylori Probiotics: A Comprehensive Overview for Health Practitioners
Date: 2020
Abstract: Authors: Ruscio M.
URL: https://drruscio.com/h-pylori-probiotics/

Name of resource: Treatment regimens for Helicobacter pylori in adults
Date: 2022

Abstract:
Authors: Lamont JT.

Name of study: Effects of probiotics on the recurrence of bacterial vaginosis: a review
Date: 2014
Abstract:
Authors: Homayouni A, Bastani P, Ziyadi S, et al.

Gut and Lung Connection to Asthma – Rodney Dietert, PhD

In this fifth in a series of interviews with Rodney Dietert PhD, he talks about communication between the gut and lung. Dr. Dietert is Cornell University Professor Emeritus, Health Scientist Head of Translational Science + Education for SEED and the Author of the Human Super-Organism How the Microbiome is Revolutionizing the Pursuit of a Healthy Life we learn about:

* The Gut and lung communication and its relationship to Asthma

World Asthma FoundationDefeating Asthma Series uncovers New Hope for Asthma Managementant

Asthmatics: Our understanding of Asthma and the way we treat it may soon be radically different from what currently exists, due to new research on the human microbiome and how the microbiome affects asthma.

Interview

World Asthma Foundation: Research into the Microbiome and its relationship to health has improved significantly in the last few years. For example, we now know about the relationship between the gut and health. We’ve also learned about communication between the gut and the lung and the impact on Asthma. Dr. Dietert, so there’s some crosstalk, right?

Video interview: Asthma Connection to Gut and Lung Cross Talk – Rodney Dietert, PhD

Dr. Dietert: Tremendous crosstalk, absolutely tremendous. You’re correct that if you’re looking at endpoints, something like risk of asthma or management of asthma, then you really, at a minimum, are going to focus both on the respiratory system microbiome and the gut microbiome. That’s not necessarily the exclusion of others but those two are really important. Just like the gut microbiome can affect the brain, it can affect behavior, mood. You don’t need lots of hardcore meds as an antidepressant when you’ve got the solution sitting right in your gut in terms of the microbiome.

With the respiratory system, you’ve got both the local microbes being extremely important but you have crosstalk, you have chemical interactions that are originating in the gut that are affecting the respiratory system as well.

World Asthma Foundation: Dr. Dietert, we certainly thank you for your time, all that you do for the microbiome and the community. Good afternoon, and thanks again.

Dr. Dietert: Well, and thank you for all you do with the World Asthma foundation, Bill. Pleasure.

To learn more about Dr. Dietert, go here.

Gut and Lung crosstalk interview with Rodney Dietert.

Missing Microbes and Asthma Link Say Multiple Studies – Martin J Blaser MD

Defeating Asthma Series uncovers New Hope for Asthma Management

In this third interview with Martin J Blaser MD, Director of the Center for Advanced Biotechnology and Medicine at Rutgers Biomedical and Health Sciences and the Henry Rutgers Chair of the Human Microbiome and Professor of Medicine and Microbiology at the Rutgers Robert Wood Johnson Medical School in New Jersey and the Author of the “Missing Microbes – How the Overuse of Antibiotics is Fueling Our Modern Plagues.” we learn:

  • About the H. pylori and Asthma connection
  • Additional reserach looking into the connection between H. pylori and Asthma
  • Whether the Microbes can reintroduced
Video: Missing Microbes and Asthma Link Say Multiple Studies – Martin J Blaser MD

Asthma Foundation: Dr. Blaser, we’ve talked about the asthma connection and the H. pylori topic. Can you identify these missing microbes also with tests? 

Dr. Blaser: Yes. The paper with Jakob Stokholm in Nature Communications looked at this– We saw that there was a difference in the microbiome in the kids that were one year old. That was the age at which their microbiome made a difference, whether they’d have a risk of asthma or not. Then we asked, “Okay, what’s the difference in the specific microbes at age one between the positives and the negatives?” We identified about 20 microbes that were significantly different, mostly lost, mostly missing.

What was interesting is that a group from British Columbia, led by Dr. Brett Finlay and colleagues had published about this also. They had found, I think, four or five organisms and we matched on four of the five. Again, two independent studies finding the same relationship makes it stronger

World Asthma Foundation: If I understand correctly, your research is determining whether or not you can repopulate the H. pylori. Is that independent of the intestinal microbes? 

Dr. Blaser: In theory, yes. What’s interesting is that people have been interested in microbes and asthma for quite some time, and most of the concentration was in the large intestine, in the colon. We were interested in the stomach first, but then we got more involved in the colon also. I think that both compartments in the body are important. Both of them are important. They’re both subject to this terrible pressure of the disappearance of microbes because of such things as antibiotics and cesarean sections and the like. They’re both. All of these microbes are potentially replaceable. That’s the hope.

World Asthma Foundation: Fantastic, that’s the hope.

 

Asthma and Indoor Air Pollution:

Key insights for Asthmatics:

  • Makes Asthma Worse
  • Significant Association with Exacerbations
  • Among this panel of relatively moderate to severe asthmatics, the respiratory irritants produced by several domestic combustion sources were associated with increased morbidity.
  • Although there is abundant clinical evidence of asthmatic responses to indoor aeroallergens, the symptomatic impacts of other common indoor air pollutants from gas stoves, fireplaces, and environmental tobacco smoke have been less well characterized. These combustion sources produce a complex mixture of pollutants, many of which are respiratory irritants.
  • Results of an analysis of associations between indoor pollution and several outcomes of respiratory morbidity in a population of adult asthmatics residing in the U.S. Denver, Colorado, metropolitan area. A panel of 164 asthmatics recorded in a daily diary the occurrence of several respiratory symptoms, nocturnal asthma, medication use, and restrictions in activity, as well as the use of gas stoves, wood stoves, or fireplaces, and exposure to environmental tobacco smoke.
  • Multiple logistic regression analysis suggests that the indoor sources of combustion have a statistically significant association with exacerbations of asthma. For example, after correcting for repeated measures and autocorrelation, the reported use of a gas stove was associated with moderate or worse shortness of breath (OR, 1.60; 95% CI, 1.11-2.32), moderate or worse cough (OR, 1.71; 95% CI, 0.97-3.01), nocturnal asthma (OR, 1.01; 95% CI, 0.91-1.13), and restrictions in activity (OR, 1.47; 95% CI, 1.0-2.16
  • The WAF Editorial Board wishes to thank and acknowledge B D Ostro 1 , M J Lipsett, J K Mann, M B Wiener, J Selner
    California Environmental Protection Agency, Berkeley for their contribution to Asthma education and research.

Asthma and COVID-19 Update Study on Risk

Asthma does not appear to increase the risk or influence its severity, according to University study

Whats new

Rutgers researchers say further study is needed but those with the chronic respiratory disease don’t appear to be at a higher risk of getting extremely ill or dying from coronavirus.“Older age and conditions such as heart disease, high blood pressure, chronic obstructive pulmonary disease, diabetes and obesity are reported risk factors for the development and progression of COVID-19,” said Reynold A. Panettieri Jr., a pulmonary critical care physician and director of the Rutgers Institute for Translational Medicine and Science and co-author of a paper published in the Journal of Allergy and Clinical Immunology.

“However, people with asthma — even those with diminished lung function who are being treated to manage asthmatic inflammation — seem to be no worse affected by SARS-CoV-2 than a non-asthmatic person. There is limited data as to why this is the case — if it is physiological or a result of the treatment to manage the inflammation.”

Children and young adults with asthma suffer mainly from allergic inflammation, while older adults who experience the same type of airway inflammation can also suffer from eosinophilic asthma — a more severe form. In these cases, people experience abnormally high levels of a type of white blood cell that helps the body fight infection, which can cause inflammation in the airways, sinuses, nasal passages and lower respiratory tract, potentially making them more at risk for a serious case of COVID-19.

Further Study Needed

Panettieri discusses what we know about asthma and inflammation and the important questions that still need to be answered.

How might awareness of SARS-CoV-2 affect the health of people with asthma?
Since the news has focused our attention on the effects of COVID-19 on people in vulnerable populations, those with asthma may become hyper-vigilant about personal hygiene and social distancing. Social distancing could improve asthma control since people who are self-quarantined are also not as exposed to seasonal triggers that include allergens or respiratory viruses. There is also evidence that people are being more attentive to taking their asthma medication during the pandemic, which can contribute to overall health.

What effect might inhaled steroids have on COVID-19 outcomes?
Inhaled corticosteroids, which are commonly used to protect against asthma attacks, also may reduce the virus’s ability to establish an infection. However, studies have shown that steroids may decrease the body’s immune response and worsen the inflammatory response. Steroids also have been shown to delay the clearing of the SARS and MERS virus — similar to SARS-CoV-2 — from the respiratory tract and thus may worsen COVID-19 outcomes. Future studies should address whether inhaled steroids in patients with asthma or allergies increase or decrease the risks of SARS-CoV-2 infection, and whether these effects are different depending on the steroid type.

In what way does age play a role in how asthma patients react to exposure to the virus?
A person’s susceptibility to and severity of COVID-19 infection increases with age. However, since asthma sufferers tend to be younger than those with reported high-risk conditions, age-adjusted studies could help us better understand if age is a factor in explaining why asthma patients may not be at greater risk for infection.

Children and young adults with asthma suffer mainly from allergic inflammation, while older adults who experience the same type of airway inflammation can also suffer from eosinophilic asthma — a more severe form. In these cases, people experience abnormally high levels of a type of white blood cell that helps the body fight infection, which can cause inflammation in the airways, sinuses, nasal passages and lower respiratory tract, potentially making them more at risk for a serious case of COVID-19.

In addition, an enzyme attached to the cell membranes in the lungs, arteries, heart, kidney and intestines that has been shown to be an entry point for SARS-CoV-2 into cells is increased in response to the virus. This enzyme is also thought to be beneficial in clearing other respiratory viruses, especially in children. How this enzyme affects the ability of SARS-CoV-2 to infect people with asthma is still unclear.

How might conditions in addition to asthma affect a person’s risk of infection?
Asthma tends to be associated with far fewer other conditions than chronic obstructive pulmonary disease or cardiovascular disease. If SARS-CoV-2 is a disease that causes dysfunction in the cells that line blood vessels throughout the body, then diabetes, heart disease, obesity and other diseases associated with this condition may make people more susceptible to the virus than those who are asthmatic.

Important to know

However, older people with asthma who also have high blood pressure, diabetes or heart disease may have similar instances of COVID-19 as non-asthmatics with those conditions.

Gut Health and Asthma

The gut and lungs are anatomically distinct, but potential anatomic communications and complex pathways involving their respective microbiota have reinforced the existence of a gut–lung axis (GLA). Compared to the better-studied gut microbiota, the lung microbiota, only considered in recent years, represents a more discreet part of the whole microbiota associated to human hosts. Gut health is not the only area to think about.

While the majority of studies focused on the bacterial component of the microbiota in healthy and pathological conditions, recent works highlighted the contribution of fungal and viral kingdoms at both digestive and respiratory levels. Moreover, growing evidence indicates the key role of inter-kingdom crosstalks in maintaining host homeostasis and in disease evolution.

In fact, the recently emerged GLA concept involves host–microbe as well as microbe–microbe interactions, based both on localized and long-reaching effects. GLA can shape immune responses and interfere with the course of respiratory diseases. In this review, we aim to analyze how the lung and gut microbiota influence each other and may impact on respiratory diseases.

Due to the limited knowledge on the human virobiota, we focused on gut and lung bacteriobiota and mycobiota, with a specific attention on inter-kingdom microbial crosstalk. These are able to shape local or long-reached host responses within the GLA.

Introduction

Recent advances in microbiota explorations have led to an improved knowledge of the communities of commensal microorganisms within the human body. Human skin and mucosal surfaces are associated with rich and complex ecosystems (microbiota) composed of bacteria (bacteriobiota), fungi (mycobiota), viruses (virobiota), phages, archaea, protists, and helminths (Cho and Blaser, 2012).

The role of the gut bacteriobiota in local health homeostasis and diseases is being increasingly investigated, but its long-distance impacts still need to be clarified (Chiu et al., 2017). Among the relevant inter-organ connections, the gut–lung axis (GLA) remains less studied than the gut–brain axis.

So far, microbiota studies mainly focused on the bacterial component, neglecting other microbial kingdoms. However, the understanding of mycobiota involvement in human health and inter-organ connections should not be overlooked (Nguyen et al., 2015; Enaud et al., 2018).

Viruses are also known to be key players in numerous respiratory diseases and to interact with the human immune system, but technical issues still limit the amount of data regarding virobiota (Mitchell and Glanville, 2018). Therefore, we will focus on bacterial and fungal components of the microbiota and their close interactions that are able to shape local or long-reached host responses within the GLA.

While GLA mycobiota also influences chronic gut diseases such as IBD, we will not address this key role in the present review: we aimed at analyzing how lung and gut bacteriobiota and mycobiota influence each other, how they interact with the human immune system, and their role in respiratory diseases.

Gut Health

Microbial Interactions Within the Gut–Lung Axis

The gut microbiota has been the most extensively investigated in gut health. The majority of genes (99%) amplified in human stools are from bacteria, which are as numerous as human cells and comprise 150 distinct bacterial species, belonging mainly to Firmicutes and Bacteroidetes phyla. Proteobacteria, Actinobacteria, Cyanobacteria, and Fusobacteria are also represented in healthy people (Sekirov et al., 2010; Human Microbiome Project Consortium, 2012).

More recently, fungi have been recognized as an integral part of our commensal flora, and their role in health and diseases is increasingly considered (Huffnagle and Noverr, 2013; Huseyin et al., 2017). Fungi are about 100 times larger than bacteria, so even if fungal sequences are 100 to 1,000 times less frequent than bacterial sequences, fungi must not be neglected in the gastrointestinal ecosystem.

Mycobiota Diversity

In contrast with the bacteriobiota, the diversity of the gut mycobiota in healthy subjects is limited to few genera, with a high prevalence of Saccharomyces cerevisiae, Malassezia restricta, and Candida albicans (Nash et al., 2017).

Note from the WAF editorial board. We wish to acknowledge and thank Raphaël Enaud, Renaud Preve, Eleonora Ciarlo, Fabien Beaufils, Gregoire Wieërs, Benoit Guery and Laurence Delhaes for their support of Asthma education and research. For more information about Asthma or Gut Health, visit the World Asthma Foundation.

Although often dichotomized due to technical and analysis sequencing issues, critical interactions exist between bacteriobiota and mycobiota (Peleg et al., 2010). The most appropriate approach to decipher the role of gut microbiota is therefore considering the gut as an ecosystem in which inter-kingdom interactions occur and have major implications as suggested by the significant correlations between the gut bacteriobiota and mycobiota profiles among healthy subjects (Hoffmann et al., 2013).

Yeasts

Yeasts, e.g., Saccharomyces boulardii and C. albicans, or fungus wall components, e.g., ?-glucans, are able to inhibit the growth of some intestinal pathogens (Zhou et al., 2013; Markey et al., 2018). S. boulardii also produces proteases or phosphatases that inactivate the toxins produced by intestinal bacteria such as Clostridium difficile and Escherichia coli (Castagliuolo et al., 1999; Buts et al., 2006).

In addition, at physiological state and during gut microbiota disturbances (e.g., after a course of antibiotics), fungal species may take over the bacterial functions of immune modulation, preventing mucosal tissue damages (Jiang et al., 2017). Vice versa, bacteria can also modulate fungi: fatty acids locally produced by bacteria impact on the phenotype of C. albicans (Noverr and Huffnagle, 2004; Tso et al., 2018).

Microbiota

Beside the widely studied gut microbiota, microbiotas of other sites, including the lungs, are essential for host homeostasis and disease. The lung microbiota is now recognized as a cornerstone in the physiopathology of numerous respiratory diseases (Soret et al., 2019; Vandenborght et al., 2019).

Inter-Kingdom Crosstalk Within the Lung Microbiota

The lung microbiota represents a significantly lower biomass than the gut microbiota: about 10 to 100 bacteria per 1,000 human cells (Sze et al., 2012). Its composition depends on the microbial colonization from the oropharynx and upper respiratory tract through salivary micro-inhalations, on the host elimination abilities (especially coughing and mucociliary clearance), on interactions with the host immune system, and on local conditions for microbial proliferation, such as pH or oxygen concentration (Gleeson et al., 1997; Wilson and Hamilos, 2014).

The predominant bacterial phyla in lungs are the same as in gut, mainly Firmicutes and Bacteroidetes followed by Proteobacteria and Actinobacteria (Charlson et al., 2011). In healthy subjects, the main identified fungi are usually environmental: Ascomycota (Aspergillus, Cladosporium, Eremothecium, and Vanderwaltozyma) and Microsporidia (Systenostrema) (Nguyen et al., 2015; Vandenborght et al., 2019).

In contrast to the intestinal or oral microbiota, data highlighting the interactions between bacteria and fungi in the human respiratory tract are more scattered (Delhaes et al., 2012; Soret et al., 2019). However, data from both in vitro and in vivo studies suggest relevant inter-kingdom crosstalk (Delhaes et al., 2012; Xu and Dongari-Bagtzoglou, 2015; Lof et al., 2017; Soret et al., 2019).

Several Pathways

This dialogue may involve several pathways as physical interaction, quorum-sensing molecules, production of antimicrobial agents, immune response modulation, and nutrient exchange (Peleg et al., 2010). Synergistic interactions have been documented between Candida and Streptococcus, such as stimulation of Streptococcus growth by Candida, increasing biofilm formation, or enhancement of the Candida pathogenicity by Streptococcus (Diaz et al., 2012; Xu et al., 2014).

In vitro studies exhibited an increased growth of Aspergillus fumigatus in presence of Pseudomonas aeruginosa, due to the mold’s ability in to assimilate P. aeruginosa-derived volatile sulfur compounds (Briard et al., 2019; Scott et al., 2019). However, the lung microbiota modulation is not limited to local inter-kingdom crosstalk and also depends on inter-compartment crosstalk between the gut and lungs.
Microbial Inter-compartment Crosstalk

From birth throughout the entire life span, a close correlation between the composition of the gut and lung microbiota exists, suggesting a host-wide network (Grier et al., 2018). For instance, modification of newborns’ diet influences the composition of their lung microbiota, and fecal transplantation in rats induces changes in the lung microbiota (Madan et al., 2012; Liu et al., 2017).

Gut-Lung Interaction

The host’s health condition can impact this gut–lung interaction too. In cystic fibrosis (CF) newborns, gut colonizations with Roseburia, Dorea, Coprococcus, Blautia, or Escherichia presaged their respiratory appearance, and their gut and lung abundances are highly correlated over time (Madan et al., 2012). Similarly, the lung microbiota is enriched with gut bacteria, such as Bacteroides spp., after sepsis (Dickson et al., 2016).

Conversely, lung microbiota may affect the gut microbiota composition. In a pre-clinical model, influenza infection triggers an increased proportion of Enterobacteriaceae and decreased abundances of Lactobacilli and Lactococci in the gut (Looft and Allen, 2012). Consistently, lipopolysaccharide (LPS) instillation in the lungs of mice is associated with gut microbiota disturbances (Sze et al., 2014).

Although gastroesophageal content inhalations and sputum swallowing partially explain this inter-organ connection, GLA also involves indirect communications such as host immune modulation.

Gut–Lung Axis Interactions With Human Immune System

Gut microbiota effects on the local immune system have been extensively reviewed (Elson and Alexander, 2015). Briefly, the gut microbiota closely interacts with the mucosal immune system using both pro-inflammatory and regulatory signals (Skelly et al., 2019). It also influences neutrophil responses, modulating their ability to extravasate from blood (Karmarkar and Rock, 2013).

Receptor Signaling

Toll-like receptor (TLR) signaling is essential for microbiota-driven myelopoiesis and exerts a neonatal selection shaping the gut microbiota with long-term consequences (Balmer et al., 2014; Fulde et al., 2018). Moreover, the gut microbiota communicates with and influences immune cells expressing TLR or GPR41/43 by means of microbial associated molecular patterns (MAMPs) or short-chain fatty acids (SCFAs) (Le Poul et al., 2003).

Data focused on the gut mycobiota’s impact on the immune system are sparser. Commensal fungi seem to reinforce bacterial protective benefits on both local and systemic immunity, with a specific role for mannans, a highly conserved fungal wall component. Moreover, fungi are able to produce SCFAs (Baltierra-Trejo et al., 2015; Xiros et al., 2019). Therefore, gut mycobiota perturbations could be as deleterious as bacteriobiota ones (Wheeler et al., 2016; Jiang et al., 2017).

Lung Microbiota and Local Immunity

A crucial role of lung microbiota in the maturation and homeostasis of lung immunity has emerged over the last few years (Dickson et al., 2018). Colonization of the respiratory tract provides essential signals for maturing local immune cells with long-term consequences (Gollwitzer et al., 2014).

Pre-clinical studies confirm the causality between airway microbial colonization and the regulation and maturation of the airways’ immune cells. Germ-free mice exhibit increased local Th2-associated cytokine and IgE production, promoting allergic airway inflammation (Herbst et al., 2011).

Consistently, lung exposure to commensal bacteria reduces Th2-associated cytokine production after an allergen challenge and induces regulatory cells early in life (Russell et al., 2012; Gollwitzer et al., 2014). The establishment of resident memory B cells in lungs also requires encountering lung microbiota local antigens, especially regarding immunity against viruses such as influenza (Allie et al., 2019).

Interactions between lung microbiota and immunity are also a two-way process; a major inflammation in the lungs can morbidly transform the lung microbiota composition (Molyneaux et al., 2013).

Gut Health, Long-Reaching Immune Modulation Within Gut–Lung Axis

Beyond the local immune regulation by the site-specific microbiota, the long-reaching immune impact of gut microbiota is now being recognized, especially on the pulmonary immune system (Chiu et al., 2017).

The mesenteric lymphatic system is an essential pathway between the lungs and the intestine, through which intact bacteria, their fragments, or metabolites (e.g., SCFAs) may translocate across the intestinal barrier, reach the systemic circulation, and modulate the lung immune response (Trompette et al., 2014; Bingula et al., 2017; McAleer and Kolls, 2018).

SCFAs, mainly produced by the bacterial dietary fibers’ fermentation especially in case of a high-fiber diet (HFD), act in the lungs as signaling molecules on resident antigen-presenting cells to attenuate the inflammatory and allergic responses (Anand and Mande, 2018; Cait et al., 2018).

SCFA receptor–deficient mice show increased inflammatory responses in experimental models of asthma (Trompette et al., 2014). Fungi, including A. fumigatus, can also produce SCFAs or create a biofilm enhancing the bacterial production of SCFAs, but on the other hand, bacterial SCFAs can dampen fungal growth (Hynes et al., 2008; Baltierra-Trejo et al., 2015; Xiros et al., 2019). The impact of fungal production of SCFAs on the host has not been assessed so far.

Other Elements

Other important players of this long-reaching immune effect are gut segmented filamentous bacteria (SFBs), a commensal bacteria colonizing the ileum of most animals, including humans, and involved in the modulation of the immune system’s development (Yin et al., 2013). SFBs regulate CD4+ T-cell polarization into the Th17 pathway, which is implicated in the response to pulmonary fungal infections and lung autoimmune manifestations (McAleer et al., 2016; Bradley et al., 2017).

Recently, innate lymphoid cells, involved in tissue repair, have been shown to be recruited from the gut to the lungs in response to inflammatory signals upon IL-25 (Huang et al., 2018). Finally, intestinal TLR activation, required for the NF-?B–dependent pathways of innate immunity and inflammation, is associated with an increased influenza-related lung response in mice (Ichinohe et al., 2011).

Mechanisms

Other mechanisms may be involved in modulating the long-reaching immune response related to gut microbiota, exemplified by the increased number of mononuclear leukocytes and an increased phagocytic and lytic activity after treatment with Bifidobacterium lactis HN019 probiotics (Gill et al., 2001). Diet, especially fiber intake, which increases the systemic level of SCFAs, or probiotics influence the pulmonary immune response and thus impact the progression of respiratory disorders (King et al., 2007; Varraso et al., 2015; Anand and Mande, 2018).

The GLA immune dialogue remains a two-way process. For instance, Salmonella nasal inoculation promotes a Salmonella-specific gut immunization which depends on lung dendritic cells (Ruane et al., 2013). Respiratory influenza infection also modulates the composition of the gut microbiota as stated above. These intestinal microbial disruptions seem to be unrelated to an intestinal tropism of influenza virus but mediated by Th17 cells (Wang et al., 2014).

In summary, GLA results from complex interactions between the different microbial components of both the gut and lung microbiotas combined with local and long-reaching immune effects. All these interactions strongly suggest a major role for the GLA in respiratory diseases, as recently documented in a mice model (Skalski et al., 2018).
Gut–Lung Axis in Respiratory Diseases

Acute Infectious Diseases

Regarding influenza infection and the impact of gut and lung microbiota, our knowledge is still fragmentary; human data are not yet available. However, antibiotic treatment causes significantly reduced immune responses against influenza virus in mice (Ichinohe et al., 2011). Conversely, influenza-infected HFD-fed mice exhibit increased survival rates compared to infected controls thanks to an enhanced generation of Ly6c-patrolling monocytes. These monocytes increase the numbers of macrophages that have a limited capacity to produce CXCL1 locally, reducing neutrophil recruitment to the airways and thus tissue damage. In parallel, diet-derived SCFAs boost CD8+ T-cell effector function in HFD-fed mice (Trompette et al., 2018).

Both lung and gut microbiota are essential against bacterial pneumonia. The lung microbiota is able to protect against respiratory infections with Streptococcus pneumoniae and Klebsiella pneumoniae by priming the pulmonary production of granulocyte-macrophage colony-stimulating factor (GM-CSF) via IL-17 and Nod2 stimulation (Brown et al., 2017).

Gut Health and Lung Bacterial Infections

The gut microbiota also plays a crucial role in response to lung bacterial infections. Studies on germ-free mice showed an increased morbidity and mortality during K. pneumoniae, S. pneumoniae, or P. aeruginosa acute lung infection (Fagundes et al., 2012; Fox et al., 2012; Brown et al., 2017). The use of broad-spectrum antibiotic treatments, to disrupt mouse gut microbiota, results in worse outcome in lung infection mouse models (Schuijt et al., 2016; Robak et al., 2018).

Mechanistically, alveolar macrophages from mice deprived of gut microbiota through antibiotic treatment are less responsive to stimulation and show reduced phagocytic capacity (Schuijt et al., 2016). Interestingly, priming of antibiotic-treated animals with TLR agonists restores resistance to pulmonary infections (Fagundes et al., 2012). SFBs appear to be an important gut microbiota component for lung defense against bacterial infection thanks to their capacity to induce the production of the Th17 cytokine, IL-22, and to increase neutrophil counts in the lungs during Staphylococcus aureus pneumonia (Gauguet et al., 2015).

Modulating chronic infectious diseases will similarly depend on gut and lung microbiotas. For instance, Mycobacterium tuberculosis infection severity is correlated with gut microbiota (Namasivayam et al., 2018).

Chronic Respiratory Diseases

Multiple studies have addressed the impact of gut and lung microbiota on chronic respiratory diseases such as chronic obstructive pulmonary disease (COPD), asthma, and CF (Table 1).

Table 1. Gut–lung axis in human chronic respiratory diseases. Gut Health.

Decreased lung microbiota diversity and Proteobacteria expansion are associated with both COPD severity and exacerbations (Garcia-Nuñez et al., 2014; Wang et al., 2016, 2018; Mayhew et al., 2018). The fact that patients with genetic mannose binding lectin deficiency exhibit a more diverse pulmonary microbiota and a lower risk of exacerbation suggests not only association but also causality (Dicker et al., 2018).

Besides the lung flora, the gut microbiota is involved in exacerbations, as suggested by the increased gastrointestinal permeability in patients admitted for COPD exacerbations (Sprooten et al., 2018). Whatever the permeability’s origin (hypoxemia or pro-inflammatory status), the level of circulating gut microbiota–dependent trimethylamine-N-oxide has been associated with mortality in COPD patients (Ottiger et al., 2018). This association being explained by comorbidities and age, its impact per se is not guaranteed. Further studies are warranted to investigate the role of GLA in COPD and to assess causality.

Early Life Perturbation

Early-life perturbations in fungal and bacterial gut colonization, such as low gut microbial diversity, e.g., after neonatal antibiotic use, are critical to induce childhood asthma development (Abrahamsson et al., 2014; Metsälä et al., 2015; Arrieta et al., 2018).

This microbial disruption is associated with modifications of fecal SCFA levels (Arrieta et al., 2018). Causality has been assessed in murine models. Inoculation of the bacteria absent in the microbiota of asthmatic patients decreases airways inflammation (Arrieta et al., 2015).

Fungi

Furthermore, Bacteroides fragilis seems to play a major role in immune homeostasis, balancing the host systemic Th1/Th2 ratio and therefore conferring protection against allergen-induced airway disorders (Mazmanian et al., 2005; Panzer and Lynch, 2015; Arrieta et al., 2018). Nevertheless, it is still not fully deciphered, as some studies conversely found that an early colonization with Bacteroides, including B. fragilis, could be an early indicator of asthma later in life (Vael et al., 2008).

Regarding fungi, gut fungal overgrowth (after antibiotic administration or a gut colonization protocol with Candida or Wallemia mellicola) increases the occurrence of asthma via IL-13 without any fungal expansion in the lungs (Noverr et al., 2005; Wheeler et al., 2016; Skalski et al., 2018). The prostaglandin E2 produced in the gut by Candida can reach the lungs and promotes lung M2 macrophage polarization and allergic airway inflammation (Kim et al., 2014).

Mouse & Human Gut Health

In mice, a gut overrepresentation of W. mellicola associated with several intestinal microbiome disturbances appears to have long-reaching effects on the pulmonary immune response and severity of asthma, by involving the Th2 pathways, especially IL-13 and to a lesser degree IL-17, goblet cell differentiation, fibroblasts activation, and IgE production by B cells (Skalski et al., 2018).

These results indicate that the GLA, mainly through the gut microbiota, is likely to play a major role in asthma.

Cystic Fibrosis and Gut Health

In CF patients, gut and lung microbiota are distinct from those of healthy subjects, and disease progression is associated with microbiota alterations. (Madan et al., 2012; Stokell et al., 2015; Nielsen et al., 2016). Moreover, the bacterial abundances at both sites are highly correlated and have similar trends over time (Madan et al., 2012). This is especially true regarding Streptococcus, which is found in higher proportion in CF stools, gastric contents, and sputa. (Al-Momani et al., 2016; Nielsen et al., 2016).

Moreover, CF patients with a documented intestinal inflammation exhibit a higher Streptococcus abundance in the gut (Enaud et al., 2019). That suggests the GLA’s involvement in intestinal inflammation. Of note, gut but not lung microbiota alteration is associated with early-life exacerbations. Some gut microbiota perturbations, such as a decrease of Parabacteroides, are predictive of airway colonization with P. aeruginosa (Hoen et al., 2015).

Furthermore, oral administration of probiotics to CF patients leads to a decreased number of exacerbations (Anderson et al., 2016). While the mycobiota has been recently studied in CF (Nguyen et al., 2015; Soret et al., 2019), no data on the role of the fungal component of the GLA are currently available in CF. This deserves to be more widely studied.

Improving Health in the Gut

The role of inter-compartment and inter-kingdom interactions within the GLA in those pulmonary diseases now has to be further confirmed and causality assessed. Diet, probiotics, or more specific modulations could be, in the near future, novel essential tools in therapeutic management of these respiratory diseases.

Conclusion

The gut–lung axis or GLA has emerged as a specific axis with intensive dialogues between the gut and lungs, involving each compartment in a two-way manner, with both microbial and immune interactions (Figure 1). Each kingdom and compartment plays a crucial role in this dialogue, and consequently in host health and diseases. The roles of fungal and viral kingdoms within the GLA still remain to be further investigated. Their manipulation, as for the bacterial component, could pave the way for new approaches in the management of several respiratory diseases such as acute infections, COPD, asthma, and/or CF.

WAF: Gut health is an important area of research for the foundation.

Gut Health and asthma, an interview with Rodney Dietert, PhD.

See also Dr. Dietert’s interview about the Gut and Lung connection.

Non-Eosinophilic Asthma (NEA)

Although non-eosinophilic asthma (NEA) is not the best known and most prevalent asthma phenotype, its importance cannot be underestimated. NEA is characterized by airway inflammation with the absence of eosinophils, subsequent to activation of non-predominant type 2 immunologic pathways. This phenotype, which possibly includes several not well-defined subphenotypes, is defined by an eosinophil count <2% in sputum. NEA has been associated with environmental and/or host factors, such as smoking cigarettes, pollution, work-related agents, infections, and obesity. These risk factors, alone or in conjunction, can activate specific cellular and molecular pathways leading to non-type 2 inflammation.

Note from the WAF: We wish to acknowledge and thank Darío Antolín-Amérigo, Javier Domínguez-Ortega,3,4 and Santiago Quirce Department of Allergy, Hospital General de Villalba, Madrid, Spain, for their contribution to Asthma education and research.

The most relevant clinical trait of NEA is its poor response to standard asthma treatments, especially to inhaled corticosteroids, leading to a higher severity of disease and to difficult-to-control asthma. Indeed, NEA constitutes about 50% of severe asthma cases. Since most current and forthcoming biologic therapies specifically target type 2 asthma phenotypes, such as uncontrolled severe eosinophilic or allergic asthma, there is a dramatic lack of effective treatments for uncontrolled non-type 2 asthma. Research efforts are now focusing on elucidating the phenotypes underlying the non-type 2 asthma, and several studies are being conducted with new drugs and biologics aiming to develop effective strategies for this type of asthma, and various immunologic pathways are being scrutinized to optimize efficacy and to abolish possible adverse effects.

Why Cell Biology of Asthma Matters

Cell types responsible for the major pathology in asthma:
1. Epithelial cells – initiate airway inflammation mucus, and
2. Smooth muscle cells – contract excessively to cause airway narrowing.

The clinical manifestations of asthma are caused by obstruction of the conducting airways of the lung. Two airway cell types are critical for asthma pathogenesis: epithelial cells and smooth muscle cells. Airway epithelial cells, which are the first line of defense against inhaled pathogens and particles, initiate airway inflammation and produce mucus, an important contributor to airway obstruction. The other main cause of airway obstruction is contraction of airway smooth muscle. Complementary experimental approaches involving cultured cells, animal models, and human clinical studies have provided many insights into diverse mechanisms that contribute to airway epithelial and smooth muscle cell pathology in this complex disease. Continued attention to the study of the cell biology of asthma will be crucial for generating new ideas for asthma prevention and treatment based on normalizing epithelial and smooth muscle function.

Note from the WAF editorial board: We wish to acknowledge and thank David J. Erle and Dean Sheppard, Lung Biology Center and Department of Medicine, University of California, San Francisco for their support for Asthma research and education.

Asthma is a common disease that affects up to 8% of children in the United States (Moorman et al., 2007) and is a major cause of morbidity worldwide. The principal clinical manifestations of asthma are repeated episodes of shortness of breath and wheezing that are at least partially reversible, recurrent cough, and excess airway mucus production. Because asthma involves an integrated response in the conducting airways of the lung to known or unknown triggers, it is a multicellular disease, involving abnormal responses of many different cell types in the lung (Locksley, 2010). Here we focus on the two cell types that are ultimately responsible for the major symptomatic pathology in asthma—epithelial cells that initiate airway inflammation in asthma and are the source of excess airway mucus, and smooth muscle cells that contract excessively to cause symptomatic airway narrowing. The current thinking about cell–cell communications that drive asthma (Fig. 1) is that known and unknown inhaled stimuli (i.e., proteases and other constituents of inhaled allergens, respiratory viruses, and air pollutants) stimulate airway epithelial cells to secrete the cytokines TSLP, interleukin (IL)-25, and IL-33, which act on subepithelial dendritic cells, mast cells, and innate lymphoid cells (iLCs) to recruit both innate and adaptive hematopoietic cells and initiate the release of T helper 2 (Th2) cytokines (principally IL-5 and IL-13; Locksley, 2010; Scanlon and McKenzie, 2012; Bando et al., 2013; Barlow et al., 2013; Nussbaum et al., 2013). Environmental stimuli also activate afferent nerves in the airway epithelium that can themselves release biologically active peptide mediators and also trigger reflex release of acetylcholine from efferent fibers in the vagus nerve. This initial response is amplified by the recruitment and differentiation of subsets of T cells that sustain secretion of these cytokines and in some cases secrete another cytokine, IL-17, at specific strategic sites in the airway wall. The released cytokines act on epithelial cells and smooth muscle cells and drive the pathological responses of these cells that contribute to symptomatic disease. The cell biology underlying the responses of the relevant hematopoietic lineages is not specific to asthma and has been discussed elsewhere (Locksley, 2010; Scanlon and McKenzie, 2012). We focus our discussion on the contributions of epithelial cells and airway smooth muscle cells.
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Figure 1.

Cell–cell communication in the airway wall in asthma. Environmental triggers concurrently act on airway afferent nerves (which both release their own peptide mediators and stimulate reflex release of the bronchoconstrictor acetylcholine) and airway epithelial cells to initiate responses in multiple cell types that contribute to the mucous metaplasia and airway smooth muscle contraction that characterize asthma. Epithelial cells release TSLP and IL-33, which act on airway dendritic cells, and IL-25, which together with IL-33 acts on mast cells, basophils, and innate type 2 lymphocytes (iLC2). These secreted products stimulate dendritic cell maturation that facilitates the generation of effector T cells and triggers the release of both direct bronchoconstrictors and Th2 cytokines from innate immune cells, which feed back on both the epithelium and airway smooth muscle and further facilitate amplification of airway inflammation through subsequent adaptive T cell responses.

Cell biology of airway epithelium

The airway is covered with a continuous sheet of epithelial cells (Crystal et al., 2008; Ganesan et al., 2013). Two major airway cell types, ciliated and secretory cells, establish and maintain the mucociliary apparatus, which is critical for preserving airway patency and defending against inhaled pathogens and allergens. The apparatus consists of a mucus gel layer and an underlying periciliary layer. Ciliated cells each project ?300 motile cilia into the periciliary layer that are critical for propelling the mucus layer up the airway. In addition, cilia are coated with membrane-spanning mucins and tethered mucopolysaccharides that exclude mucus from the periciliary space and promote formation of a distinct mucus layer (Button et al., 2012). Secretory cells produce a different class of mucins, the polymeric gel-forming mucins. The two major airway gel-forming mucins are MUC5AC and MUC5B. Some secretory cells, known as mucous or goblet cells, produce mucins and store them within easily visualized collections of mucin granules, whereas other cells produce and secrete mucins (especially MUC5B) but lack prominent granules. Gel-forming mucins are secreted into the airway lumen and are responsible for the characteristic viscoelastic properties of the mucus gel layer.
Airway epithelial injury and remodeling in asthma

A variety of structural changes in the epithelium and other portions of the airway, termed “airway remodeling,” is frequently seen in individuals with asthma (Elias et al., 1999). These changes include airway wall thickening, epithelial hypertrophy and mucous metaplasia, subepithelial fibrosis, myofibroblast hyperplasia, and smooth muscle cell hyperplasia and hypertrophy. Airway remodeling is thought to represent a response to ongoing tissue injury caused by infectious agents, allergens, or inhaled particulates and by the host responses to these stimuli. Signs of frank epithelial injury, including loss of epithelial integrity, disruption of tight junctions, impairment of barrier function, and cell death, have been identified in some studies and may correlate with asthma severity (Laitinen et al., 1985; Jeffery et al., 1989; Barbato et al., 2006; Holgate, 2007). However, in many individuals asthma symptoms and features of airway remodeling, including mucous metaplasia and subepithelial fibrosis, are seen in the absence of signs of active airway infection or overt tissue injury (Ordoñez et al., 2000), suggesting that other processes account for the persistence of asthma in these individuals. Substantial evidence suggests that the persistence of asthma is driven by ongoing host immune responses that generate mediators driving airway remodeling and airway dysfunction. The epithelium is both a site of production of these mediators and a source of cells that respond to mediators produced by immune cells and other cells within the airway. How airway epithelial cells recognize and respond to viruses, allergens, and other stimuli has been comprehensively reviewed elsewhere (Lambrecht and Hammad, 2012). Here we will focus on the contribution of the epithelium to production of and responses to Th2 cytokines.
Airway epithelial contributions to Th2 responses.

Th2 cytokines, especially IL-13, play critical roles in asthma. Multiple cytokines, including TSLP, GM-CSF, IL-1, IL-25, and IL-33, are produced by the epithelium and promote production of Th2 cytokines by immune cells (Cates et al., 2004; Hammad et al., 2009; Locksley, 2010; Nagarkar et al., 2012). Genome-wide association studies implicate multiple Th2-related genes, including IL13, IL33, and TSLP, in asthma (Moffatt et al., 2010; Torgerson et al., 2011). IL-13 is produced by innate lymphoid cells (Neill et al., 2010; Price et al., 2010; Saenz et al., 2010; Hasnain et al., 2011) and Th2 cells (Grünig et al., 1998; Wills-Karp et al., 1998) during allergic inflammation and by macrophages in a mouse model of virus-induced airway disease (Kim et al., 2008). IL-13 induces characteristic changes in airway epithelial mRNA (Kuperman et al., 2005b; Woodruff et al., 2007; Zhen et al., 2007) and miRNA (Solberg et al., 2012) expression patterns in airway epithelial cells. The IL-13 transcriptional “signature” can be used to identify individuals with “Th2 high” and “Th2 low” asthma (Woodruff et al., 2009). The IL-13–induced protein periostin is secreted basally from airway epithelial cells and can be used as a biomarker for Th2 high asthma (Jia et al., 2012; Parulekar et al., 2014). Roughly half of individuals with asthma are Th2 high, and these individuals have better responses to treatment with inhaled corticosteroids (Woodruff et al., 2009) or anti–IL-13 antibody (Corren et al., 2011). The key drivers of Th2 low asthma remain poorly understood, although Th17 family cytokines may be important (Newcomb and Peebles, 2013).

Mucous metaplasia.

Although mucus is critical for host defense, pathological mucus production is an important contributor to asthma morbidity and mortality. In fatal asthma, airways are often plugged with tenacious mucus plugs that obstruct movement of gas (Kuyper et al., 2003). This catastrophic phenomenon likely reflects increased mucin production and secretion as well as changes in mucin cross-linking, mucus gel hydration, and mucus clearance. Abnormalities in mucus are not limited to severe asthma exacerbations because an increase in intracellular mucin stores (mucous metaplasia) is seen even in individuals with stable, mild to moderate asthma (Ordoñez et al., 2001). In mouse allergic airway disease models of asthma, mucous metaplasia results from increased production and storage of mucins (especially MUC5AC) in preexisting secretory cells, including club cells (Evans et al., 2004), rather than transdifferentiation of ciliated cells (Pardo-Saganta et al., 2013). However, in virus-driven models of asthma mucous cells might arise from transdifferentiation of ciliated cells (Tyner et al., 2006). A variety of stimuli and signaling pathways have been shown to regulate mucin production and secretion in airway epithelial cells.
IL-13 stimulates mucin production in Th2 high asthma.

Direct effects of IL-13 on airway epithelial cells induce mucous metaplasia in human airway epithelial cells in culture (Laoukili et al., 2001; Zhen et al., 2007) and in mouse airway epithelial cells in vivo (Kuperman et al., 2002). IL-13 is necessary for mucous metaplasia in many mouse asthma models (Grünig et al., 1998; Wills-Karp et al., 1998; Tyner et al., 2006). Individuals with Th2 high asthma have elevated levels of bronchial epithelial cell MUC5AC mRNA compared with healthy controls or individuals with Th2 low asthma (Woodruff et al., 2009). Recent transgenic mouse studies demonstrate roles for MUC5AC in clearance of enteric nematode infections (Hasnain et al., 2011) and protection against influenza infection (Ehre et al., 2012). Increased MUC5AC expression is therefore part of an integrated immune response that contributes to host defense against pathogens or inhaled particulates. A less well-recognized feature of Th2-high asthma is the substantial decrease in expression of MUC5B (Woodruff et al., 2009). The recent discovery that MUC5B is required for normal mucociliary clearance and defense against airway infection (Roy et al., 2014) suggests further attention should be directed to the possibility that a reduction in MUC5B may be an important contributor to airway dysfunction in asthma.

IL-13 is recognized by cell surface receptors expressed on almost all cell types, including airway epithelial cells (Fig. 2). The airway epithelial cell IL-13 receptor that is critical for mucous metaplasia is a heterodimer composed of IL-13R?1 and IL-4R?. Removal of this receptor in airway epithelial secretory cells (driven by the CCSP promoter) prevented mucous metaplasia in an allergic asthma model (Kuperman et al., 2005a). IL-13 binding leads to activation of Jak kinases associated with the receptor cytoplasmic domain and subsequent phosphorylation of signal transducer and activator of transcription 6 (STAT6). STAT6 activation is required for IL-13–induced mucous metaplasia (Kuperman et al., 2002).
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Figure 2.

Mechanisms of IL-13–induced mucous metaplasia. IL-13 binds to its receptor on the surface of mucous cell progenitors (e.g., club cells) leading to phosphorylation of STAT6 and translocation of STAT6 heterodimers to the nucleus, where they bind to promoters of STAT6-responsive genes. STAT6-dependent processes that contribute to mucous metaplasia include a CLCA1-dependent pathway, a Serpin-dependent pathway, and a 15-lipoxygenase-1–dependent pathway. The transcription factor SPDEF is a master regulator of mucous cell differentiation. It inhibits FOXA2, which represses mucous cell differentiation, and activates transcription of other genes that are expressed in mucous cells.

The series of events that link STAT6 activation to mucous metaplasia are only partly understood. STAT6 does not appear to directly regulate MUC5AC transcription (Young et al., 2007) and the critical direct targets of STAT6 have not been determined. One pathway that depends upon STAT6 activation involves the protein calcium-activated chloride channel 1 (CLCA1). CLCA1 is among the most highly induced genes in airway epithelial cells from individuals with asthma (Hoshino et al., 2002; Toda et al., 2002). Despite its name, CLCA1 does not appear to function as an ion channel but instead undergoes extracellular secretion and cleavage. Extracellular CLCA1 can induce MUC5AC expression via activation of the MAP kinase MAPK13 (p38?-MAPK; Alevy et al., 2012), although the presumed CLCA1 receptor and the relevant MAPK13 targets have not yet been identified. A second pathway involves the protease inhibitor Serpin3a, the mouse orthologue of human SERPINB3 and SERPINB4. These serpins are induced by IL-13 in a STAT6-dependent fashion (Ray et al., 2005). After allergen challenge, Serpin3a?/? mice had less mucous metaplasia than wild-type mice (Sivaprasad et al., 2011), despite an intact inflammatory response. These results suggest that serpins inhibit proteases that normally degrade one or more proteins required for mucous metaplasia, although the relevant proteases and their protein substrates are not yet known. Another IL-13–induced pathway involves the enzyme 15-lipoxygenase-1 (15-LO-1; Zhao et al., 2009). 15-LO-1 converts arachidonic acid to 15-hydroxyeicosatetraenoic acid, which was shown to enhance MUC5AC expression in human airway epithelial cells.

IL-13– and STAT6-mediated mucous metaplasia depends upon changes in the activity of a network of transcription factors. Allergen-induced IL-13–mediated STAT6 activation leads to increased expression of the SAM-pointed domain–containing Ets-like factor (SPDEF; Park et al., 2007; Chen et al., 2009). The induction of SPDEF depends at least in part on FOXM1, a member of the Forkhead box (FOX) family of transcription factors (Ren et al., 2013). The SPDEF program is also important for mucous metaplasia triggered by other stimuli, including rhinoviruses (Korfhagen et al., 2012). Although SPDEF does not appear to directly regulate mucin gene transcription, SPDEF initiates a transcriptional program that is necessary and sufficient to induce mucous metaplasia. One of the effects of SPDEF is inhibition of the expression of another FOX family gene, FOXA2. In mice, deletion of Foxa2 in mucous cell precursors is sufficient to induce mucous metaplasia, and overexpression of FOXA2 inhibits allergen-induced mucous metaplasia (Zhen et al., 2007; G. Chen et al., 2010). The relationship between IL-13 and FOXA2 is complex. IL-13 inhibits expression of FOXA2, which contributes to mucous metaplasia. However, deletion of Foxa2 in airway epithelial cells during fetal development resulted in Th2 inflammation and production of IL-13 in the airway (G. Chen et al., 2010). The direct targets that are responsible for these effects of FOXA2 are not yet known.
The EGFR pathway induces mucin gene expression and mucous metaplasia.

Epidermal growth factor receptor (EGFR) binds multiple ligands including EGF, TGF-?, heparin-binding EGF, amphiregulin, ?-cellulin, and epiregulin. Ligand binding activates the EGFR kinase domain, initiating signaling cascades that are central to many fundamental biological processes, including cell proliferation, differentiation, survival, and migration. EGFR ligands induce expression of MUC5AC in human airway epithelial cell lines and a tyrosine kinase inhibitor that inhibits EGFR kinase prevents mucous metaplasia induced either by an EGFR ligand or by allergen challenge (Takeyama et al., 1999). Subsequent studies showed that bronchial epithelial EGFR levels are increased in asthma and correlate with disease severity (Takeyama et al., 2001a), and that epithelial EGFR signaling contributes to mucous metaplasia in a chronic asthma model (Le Cras et al., 2011).

Various stimuli, including bacterial products (Kohri et al., 2002; Lemjabbar and Basbaum, 2002;Koff et al., 2008), viruses (Tyner et al., 2006; Zhu et al., 2009; Barbier et al., 2012), cigarette smoke (Takeyama et al., 2001b; Basbaum et al., 2002), and inflammatory cell products (Burgel et al., 2001) can activate the EGFR pathway in airway epithelial cells. Some stimuli have been shown to initiate the EGFR signaling cascade by activating the PKC isoforms PKC ? and PKC ?, leading to recruitment of the NADPH oxidase subunits p47phox and p67phox to membrane-associated dual oxidase-1 and the generation of reactive oxygen species (ROS) at the cell surface (Shao and Nadel, 2005). ROS in turn activate latent TGF-?–converting enzyme resulting in cleavage of surface EGFR pro-ligands (Shao et al., 2003). EGFR ligand binding leads to activation of the Ras–Raf–MEK1/2–ERK1/2 pathway and MUC5AC transcriptional induction, which depends upon the Sp1 transcription factor and Sp1-binding sites within the MUC5AC promoter (Takeyama et al., 2000; Perrais et al., 2002). The IL-13 and EGFR pathways make critical but distinct contributions to gene regulation in airway epithelial cells (Zhen et al., 2007). Both pathways inhibit expression of FOXA2, suggesting that this transcription factor may represent a final common pathway for IL-13– and EGFR-induced mucous metaplasia.

Notch signaling regulates mucous cell differentiation.

Notch signaling is also important for mucous metaplasia (Tsao et al., 2011). Notch is a transmembrane receptor that binds to cell-surface ligands in the Delta-like and Jagged families. Ligand binding activates ?-secretase–mediated proteolytic cleavage and liberates the Notch intracellular domain, which enters the nucleus, associates with transcription factors, and drives expression of downstream Notch genes. Genetic manipulation of Notch signaling in mice has different effects depending on the developmental stage. In explanted embryonic lungs, addition of Notch ligand or expression of a constitutively active form of Notch increased MUC5AC-containing mucous cells, whereas a ?-secretase inhibitor reduced mucous cells (Guseh et al., 2009). Notch-induced mucous metaplasia did not require STAT6 activation, suggesting that the Notch and STAT6 pathways may operate in parallel. In contrast, in postnatal mouse lung, disruptions of Notch signaling induced mucous metaplasia (Tsao et al., 2011), a process that principally depends on the Notch ligand Jagged1 (Zhang et al., 2013). The Notch target Hes1 appears to be critical for inhibition of mucous metaplasia and MUC5AC transcription, although inactivation of Hes1 was not sufficient to induce mucous metaplasia (Ou-Yang et al., 2013). The observation that a ?-secretase inhibitor reduced IL-13–induced mucous metaplasia in cultured human airway epithelial cells (Guseh et al., 2009) suggests that further attention to the role of epithelial Notch signaling in asthma is warranted.

The secretory pathway in mucous cells

Mucin monomers are large (?5,000 amino acid residue) proteins that require extensive processing in the ER and Golgi. Each mucin monomer contains ?200 cysteine residues that can potentially participate in intra- and intermolecular disulfide bonds. The ER of mucous cells contains specialized molecules that are not widely expressed in other cell types and are required for efficient processing of mucins. One of these is anterior gradient 2 (AGR2) homologue, a member of the protein disulfide isomerase family. An active site cysteine residue in AGR2 forms mixed disulfide bonds with mucins in the ER and mice deficient in AGR2 have profound defects in intestinal mucin production (Park et al., 2009). In a mouse model of allergic asthma, AGR2-deficient mice had reduced mucus production compared with allergen-challenged wild-type mice (Schroeder et al., 2012). The reduction in mucus production was associated with activation of the unfolded protein response, a characteristic response to ER stress (Walter and Ron, 2011). AGR2 may therefore either have a direct role in mucin folding or another function necessary for maintaining normal function of the mucous cell ER. Another molecule found in the mucous cell ER is inositol-requiring enzyme 1? (IRE1?), a transmembrane ER stress sensor. IRE1? is found in mucus-producing cells in the intestine and the airways, but not in other cells. IRE1? regulates AGR2 transcription, and mice deficient in IRE1? had reduced AGR2 expression and impaired airway mucin production in an allergic asthma model (Martino et al., 2013). AGR2 and IRE1? have apparently evolved to meet the unusual demands posed by the need to produce large amounts of mucins.

ORMDL3, a member of the Orm family of transmembrane ER proteins, has also been implicated in asthma. Genetic polymorphisms at loci close to ORMDL3 were strongly associated with asthma in multiple genome-wide association studies (Moffatt et al., 2007; Galanter et al., 2008). Allergen challenge induced ORMDL3 expression in airway epithelial cells in a STAT6-dependent fashion, although ORMDL3 does not appear to be a direct target of STAT6 (Miller et al., 2012). Studies involving overexpression or knockdown of ORDML3 in HEK293 cells indicate that ORMDL3 is involved in regulating ER stress responses and ER-mediated calcium signaling (Cantero-Recasens et al., 2010). In addition, Orm proteins form complexes with serine palmitoyl-CoA transferase (SPT), the first and rate-limiting enzyme in sphingolipid production, and may thereby help coordinate lipid metabolism in the secretory pathway (Breslow et al., 2010). Genetic and pharmacologic reductions in SPT activity induced airway hyperresponsiveness in the absence of inflammation or mucous metaplasia (Worgall et al., 2013). Further studies are required to determine whether ORMDL3’s role in modulating sphingolipid production, ER stress, calcium signaling, or other ER functions in airway epithelial cells or other cells is important in asthma.

Mucins travel from the ER to the Golgi and then are packaged into large granules for secretion. In the Golgi, mucins are extensively O-glycosylated and undergo further multimerization before being released from the cell by regulated exocytosis. Throughout the airways of normal mice and in distal (smaller) airways of humans, basal secretion accounts for most mucin release, and mucin-producing cells retain too little mucin to detect using histological stains. However, mucous cells found in larger airways of humans and allergen-challenged mice contain readily detectable accumulations of mucin-containing granules that can be released by various stimuli, including the P2Y2 receptor ligands ATP and UTP and proteases that cleave protease-activated receptors. Mice lacking the exocytic priming protein Munc13-2 accumulate mucin in secretory cells that normally have minimal intracellular mucin (club cells) but can secrete mucin in response to stimulation (Zhu et al., 2008). In contrast, allergen-challenged mice lacking the low affinity calcium sensor synaptotagmin-2 have a severe defect in acute agonist-stimulated airway mucin secretion, but have preserved basal secretion and do not accumulate mucins in club cells (Tuvim et al., 2009). Agonist-stimulated secretion also depends upon the IL-13–inducible calcium-activated chloride channel TMEM16A, which is increased in mucous cells from individuals with asthma (Huang et al., 2012). Because increased production of MUC5AC via transgenic overexpression was not in itself sufficient to cause airway obstruction (Ehre et al., 2012), it seems likely that qualitative defects in mucin processing, secretion, or hydration that affect the physicochemical properties of mucus contribute to airway obstruction in asthma. Epithelial transport of water and ions, including H+ and bicarbonate, is important in maintaining the normal properties of mucus (E. Chen et al., 2010; Paisley et al., 2010; Garland et al., 2013). Rapid secretion of stored mucin, which is not fully hydrated, may result in the formation of concentrated, rubbery mucus that cannot be cleared normally by cilia or by coughing (Fahy and Dickey, 2010). Hence, IL-13 (Danahay et al., 2002; Nakagami et al., 2008) and other asthma mediators that affect airway epithelial cell water and ion transport could contribute to airway obstruction by altering the physicochemical properties of mucus.
Ciliated cell structure and function in asthma

In comparison with the extensive asthma literature regarding mucous cells, relatively few reports have focused on ciliated cells. One study of epithelial cell strips obtained by endobronchial brushing found decreased ciliary beat frequency and increases in abnormal ciliary beating patterns and ciliary ultrastructural defects in individuals with asthma compared with healthy controls (Thomas et al., 2010). These abnormalities were more pronounced in severe asthma. Ciliary abnormalities were accompanied by increases in the numbers of dead cells and evidence of loss of epithelial structural integrity, which suggests that ciliary dysfunction may be a consequence of a generalized epithelial injury. In any case, these results suggest that ciliary dysfunction might be an important contributor to impaired mucociliary clearance in asthma.
Cell biology of airway smooth muscle in asthma

The excessive airway narrowing that can lead to severe shortness of breath, respiratory failure, and death from asthma is largely due to contraction of the bands of smooth muscle present in the walls of large- and medium-sized conducting airways in the lung. In the large central airways of humans, these bands of muscle are present in the posterior portion of the airways and attach to the anterior airway cartilage rings, but in more peripheral airways smooth muscle is present circumferentially around the airways. In both locations, contraction of smooth muscle, which can be physiologically induced by release of acetylcholine from efferent parasympathetic nerves or by release of histamine and cysteinyl leukotrienes from mast cells and basophils, causes airway narrowing, with the most extensive narrowing in medium-sized airways. In healthy mammals, including humans, physiological responses to release of acetylcholine from efferent nerves or release of histamine and leukotrienes from mast cells and basophils causes only mild and generally asymptomatic airway narrowing. Normal mammals are also generally resistant to marked airway narrowing in response to pharmacologic administration of high concentrations of these contractile agonists directly into the airways. However, people with asthma have a marked increase in sensitivity to all of these agonists that can readily be demonstrated by dramatic increases in airway resistance and associated drops in maximal expiratory airflow rates during forced expiratory maneuvers (Boushey et al., 1980). Recent comparisons between responses to inhaled allergens in allergic asthmatic subjects and other subjects with similarly severe cutaneous immune responses to allergens makes it clear that all allergic humans release largely similar amounts of bronchoconstrictors into the airways (i.e., histamine and leukotrienes), but only asthmatics develop exaggerated airway narrowing in response to these mediators (Becky Kelly et al., 2003).
Mechanisms regulating generation of force by airway smooth muscle actin–myosin coupling

Force generation by airway smooth muscle is mediated by interactions between actin and myosin that depend on phosphorylation of the myosin light chain by the serine–threonine kinase, myosin light chain kinase (Fig. 3). This process is negatively regulated by myosin phosphatase. Increases in intracellular calcium concentration in smooth muscle cells induce contraction by two parallel pathways. When bound to calcium, the serine–threonine kinase calmodulin directly phosphorylates, and thereby activates, myosin light chain kinase. Increased calcium also increases GTP loading of the GTPase, RhoA, which increases the activity of its downstream effector kinases Rho-associated coiled-coil–containing protein kinases 1 and 2 (ROCK 1 and 2). ROCKs directly phosphorylate myosin light chain phosphatase, an effect that inactivates the phosphatase, further enhancing myosin phosphorylation. RhoA can also be activated independently of increases in intracellular calcium.

Core signaling pathways responsible for airway smooth muscle contraction. Airway smooth muscle contractile force is generated by cyclic cross-bridging of actin and smooth muscle myosin, which depends on myosin phosphorylation. Myosin phosphorylation is regulated by cyclic increases in cytosolic calcium (Ca2+) that activate calmodulin (CaM) to phosphorylate myosin light chain kinase (MLCK), which directly phosphorylates myosin. In parallel, the small GTPase, RhoA, is activated by both calcium-dependent and -independent pathways. Rho directly activates Rho-associated coiled-coil protein kinase (ROCK) which, in turn, phosphorylates and thereby inactivates myosin light chain phosphatase (MLCP), which normally dephosphorylates myosin. The most important physiological pathway for increasing cytosolic calcium in airway smooth muscle involves activation of G?q by G protein–coupled receptors that respond to extracellular contractile agonists, such as methacholine (Mch), serotonin (5-HT), and histamine. G?q activates phospholipase C ? (PLC?), which generates IP3 to bind to IP3 receptors on the sarcoplasmic reticulum and release sequestered Ca2+.

There are multiple upstream paths to increased i[Ca] in airway smooth muscle. Acetylcholine, released from post-ganglionic parasympathetic efferent nerves that innervate the muscle, activates G protein–coupled M2 muscarinic receptors, which are coupled to G?q. GTP-loaded G?q activates its downstream effector, PLC?, which phosphorylates PIP2 to generate IP3. IP3, in turn, binds to IP3 receptors on the sarcoplasmic reticulum to trigger translocation of calcium into the cytosol. Other contractile agonists, including histamine, bradykinin, and serotonin (5-HT; the specific agonists and receptors vary across mammalian species) bind to different G protein–coupled receptors to trigger the same pathway. Agonist-induced airway smooth muscle contraction is usually associated with cyclic oscillations in i[Ca], thought to be induced by local changes in cytosolic calcium triggering reuptake of calcium by the sarcoplasmic reticulum, and the magnitude of contractile force induced is most closely associated with the frequency of these calcium oscillations rather than their amplitude (Bergner and Sanderson, 2002).

Increases in cytosolic calcium concentration can also be induced by an influx of calcium from the extracellular space, generally due to the opening of voltage-gated calcium channels in the plasma membrane. These channels can be opened experimentally by increasing the extracellular concentration of potassium ions, which also induces airway smooth muscle contraction. Increased extracellular potassium concentrations also increase release of acetylcholine from post-ganglionic efferent nerves, so proper interpretation of the effects of KCl requires simultaneous addition of a muscarinic antagonist such as atropine.
Regulation of airway smooth muscle force generation by integrin-containing adhesion complexes

For smooth muscle cell contraction to be translated into the force required for airway narrowing, the contracting smooth muscle cell must be firmly tethered to the underlying ECM. Linkage to the ECM is accomplished through the organization of multi-protein complexes nucleated by integrins. The short cytoplasmic domains of integrins can organize surprisingly large multi-protein machines that modulate multiple signaling pathways and link integrins (and thus their ECM ligands) to the actin–myosin cytoskeleton (Yamada and Geiger, 1997; Zaidel-Bar et al., 2007). Many of the contractile agonists that stimulate myosin phosphorylation and actin–myosin interaction simultaneously enhance the formation of integrin signaling complexes, induce actin polymerization at sites of adhesion, and strengthen coupling between the actin–myosin cytoskeleton and the ECM (Mehta and Gunst, 1999; Tang et al., 1999, 2003; Gunst and Fredberg, 2003; Gunst et al., 2003; Opazo Saez et al., 2004). These events appear to also be quite important for generation of maximal contractile force because interventions that inhibit the formation or activity of adhesion complexes can inhibit the strength of contraction without affecting myosin phosphorylation (Mehta and Gunst, 1999; Tang et al., 2003; Opazo Saez et al., 2004).
Lessons from abnormal behavior of airway smooth muscle in animal models
Mice lacking ?9?1 integrin in airway smooth muscle.

Although there are large differences between the organization of airways in mice and humans, in vivo abnormalities in airway narrowing seen in mouse models do provide some insight into pathways that potentially contribute to abnormal airway smooth muscle contraction in asthma. For the purposes of this review, we will cite three illustrative examples. The integrin ?9?1 is highly expressed in airway smooth muscle (Palmer et al., 1993). Conditional knockout of the integrin ?9 subunit (uniquely found in the ?9?1 integrin) results in a spontaneous increase in in vivo airway responsiveness (as measured by increases in pulmonary resistance in response to intravenous acetylcholine), and to increased contractile responses to cholinergic agonists of both airways in lung slices and tracheal rings studied in an organ bath (Chen et al., 2012). Interestingly, although tracheal rings from these mice also have increased contractile responses to other G protein–coupled receptor agonists (e.g., serotonin), they have normal contractile responses to depolarization with KCl. These findings suggest that loss of ?9?1 increases airway responsiveness at some step upstream of calcium release from the sarcoplasmic reticulum (Fig. 4 A). In this case, increased airway responsiveness appears to be due to loss of co-localization of the polyamine-catabolizing enzyme spermidine/spermine N1-acetyltransferase (SSAT), which binds directly to the ?9 cytoplasmic domain (Chen et al., 2004), and the lipid kinase, PIP5K1?, which binds directly to talin, an integrin ?1 subunit binding partner. Spermine and spermidine are critical cofactors for PIP5K1?, so its juxtaposition with SSAT effectively reduces enzymatic activity. PIP5K1? converts PI4P to PIP2 and is responsible for most of the PIP2 produced in airway smooth muscle cells (Chen et al., 1998). PIP2 is the substrate for IP3 generation by PLC?, so when ?9?1 is present and ligated, contractile agonists that activate receptors coupled to G?q induce less IP3 generation (Chen et al., 2012) and thus less Ca2+ release through IP3 receptors in the sarcoplasmic reticulum. The importance of this pathway was confirmed by the observations that the frequency of Ca2+ oscillations induced by cholinergic agonists was reduced in lung slices from mice lacking ?9?1, and that all of the abnormalities in smooth muscle from these animals could be rescued by addition of a cell-permeable form of PIP2 (Chen et al., 2012).

Pathways that negatively regulate airway smooth muscle contraction. (A) The integrin ?9?1 negatively regulates airway smooth muscle contraction by colocalizing the polyamine-catabolizing enzyme, spermine spermidine acetyltransferase (SSAT), which directly binds to the ?9 subunit with the lipid kinase, PIP5K1?, the major source of PIP2 in airway smooth muscle, which binds to talin, a direct interactor with the ?1 subunit. PIP5K1? depends on spermine and spermidine for maximal activity, so the local breakdown of spermine and spermidine reduces PIP5K1? activity, thereby decreasing PIP2 concentrations and the amount of IP3 that is generated by activation of contractile G protein–coupled receptors (such as those activated by acetylcholine or serotonin [5-HT]). (B) The secreted scaffold protein, milk fat globule-EGF factor 8 (MFGE8), inhibits the smooth muscle hypercontractility induced by IL-13, IL-17, and tumor necrosis factor ? (TNF) by inhibiting the induction and activation of the small GTPase, RhoA. Active RhoA contributes to smooth muscle contraction by directly activating Rho-associated coiled-coil protein kinase (ROCK) which, in turn, phosphorylates and thereby inactivates myosin light chain phosphatase (MLCP), which normally dephosphorylates myosin.
Effects of T cell cytokines on airway smooth muscle contractility.

Several studies conducted over the past 15 years have suggested that cytokines released from T cells can contribute to airway hyperresponsiveness in allergic asthma (Locksley, 2010). The Th2 cytokine IL-13 has been most extensively studied, and can induce both mucous metaplasia and airway hyperresponsiveness when administered directly into the airways of mice (Grünig et al., 1998; Wills-Karp et al., 1998). In vitro, incubation of tracheal rings or lung slices increases narrowing of airways in lung slices and increases force generation by mouse tracheal rings, at least in part by inducing a dramatic increase in expression of the small GTPase, RhoA (Chiba et al., 2009), which is a critical effector of airway smooth muscle contraction (Fig. 4 B). Chronic allergen challenge or direct administration of IL-13 into the airways of mice also increased RhoA expression, in association with induction of airway hyperresponsiveness. A recent study suggested that IL-17 can also increase airway smooth muscle contractility and airway narrowing by induction of RhoA in airway smooth muscle cells (Kudo et al., 2012). In that study, mice lacking the ?v?8 integrin specifically on antigen-presenting dendritic cells were protected from allergen-induced airway hyperresponsiveness. These mice had the same degree of general airway inflammation and mucous metaplasia in response to allergen as wild-type control mice, but had a very specific defect in the generation of antigen-specific Th17 cells, an important source of IL-17 in lungs (Kudo et al., 2012). In vitro, IL-17 was shown to directly increase the contractility of mouse tracheal rings and to increase the levels of RhoA protein and its downstream effector, ROCK2, and to increase phosphorylation of the direct ROCK target, myosin phosphatase. Phosphorylation of myosin phosphatase inhibits its function, and IL-17 was also shown to consequently increase phosphorylation of myosin light chain kinase. Importantly, all of these biochemical effects were dramatically induced in vivo in airway smooth muscle of control mice in response to allergen sensitization and challenge, but all were markedly reduced in mice lacking ?v?8 on dendritic cells. Furthermore, tracheal rings removed from these knockout mice after allergen challenge had decreased in vitro contractility compared with rings from allergen challenged control mice, but this difference in contractility was eliminated by exogenous addition of IL-17. These findings strongly suggest that both IL-13 and IL-17 can contribute to airway hyperresponsiveness by directly inducing RhoA expression in airway smooth muscle (Fig. 4 B). Tumor necrosis factor ?, also implicated in asthma pathogenesis, has been shown to increase airway smooth muscle contractility by a similar mechanism (Goto et al., 2009).
Enhanced cytokine-mediated airway smooth muscle contraction in MFGE8-deficient mice.

Milk fat globule EGF factor 8 (MFGE8) is a secreted protein composed of two EGF repeats and two discoidin domains. MFGE8 was originally described to facilitate uptake of apoptotic cells by phagocytes (Hanayama et al., 2004). Mice lacking MFGE8 have normal baseline lung morphology and function, but have exaggerated airway responsiveness after allergen sensitization and challenge (Kudo et al., 2013). However, this abnormality did not appear to be related to any effects on reuptake of apoptotic cells. Immunostaining demonstrated that secreted MFGE8 was concentrated adjacent to airway smooth muscle. Tracheal rings removed from MFGE8 knockout mice had normal contractile responses at baseline, but had markedly enhanced contractile responses after overnight incubation with IL-13, and this increase in contractility could be rescued by addition of recombinant MFGE8 to the muscle bath. Importantly, rescue required the presence of at least one of the discoidin domains and of the integrin-binding RGD motif of the second EGF repeat. In mouse tracheal rings and cultured airway smooth muscle, loss of MFGE8 greatly enhanced the IL-13–induced increase in RhoA protein. These findings suggest that ligation of one or more RGD-binding integrins on airway smooth muscle by extracellular MFGE8 normally serves as a brake on cytokine-mediated RhoA induction and thereby limits maximal cytokine-induced airway hyperresponsiveness (Fig. 4 B). The specific integrin(s) involved in this response, the molecular mechanisms linking integrin ligation to inhibition of RhoA, and the role and binding partner(s) of the MFGE8 discoidin domains that are required for RhoA inhibition all remain to be determined.

Conclusions

Rapid progress has been made toward identifying epithelial and smooth muscle cell molecules and pathways that can produce many of the abnormalities found in individuals with asthma. Because these discoveries were made in diverse experimental systems, we still face major challenges in understanding how these molecules and pathways interact in vivo and in identifying the pathways that are most relevant in people with asthma. Asthma is a heterogeneous disease, and recent progress toward identifying subtypes with distinct pathophysiologic mechanisms promises to focus attention on certain pathways in epithelial and smooth muscle cells (Lötvall et al., 2011). It will be especially important to understand mechanisms underlying severe asthma. Approximately 5–10% of individuals with asthma have severe disease, with symptoms that persist despite standard therapy with bronchodilators and inhaled corticosteroids (Brightling et al., 2012). These individuals have high rates of asthma exacerbations leading to hospitalization and are at relatively high risk for fatal asthma attacks. Continued attention to the study of the cell biology of asthma will be crucial for generating new ideas for asthma prevention and treatment based on normalizing epithelial and smooth muscle function.