Natural killer T cells in the lungs of patients with asthma
2009; Elsevier BV; Volume: 123; Issue: 5 Linguagem: Inglês
10.1016/j.jaci.2009.02.013
ISSN1097-6825
AutoresPonpan Matangkasombut, Gautham Marigowda, Aaron Ervine, Luaie Idris, Muriel Pichavant, Hye Young Kim, Takahiro Yasumi, S. Brian Wilson, Rosemarie H. DeKruyff, John L. Faul, Elliot Israel, Omid Akbari, Dale T. Umetsu,
Tópico(s)IL-33, ST2, and ILC Pathways
ResumoTo the Editor:On the basis of studies in mice, an important role for invariant natural killer T (iNKT) cells in the pathogenesis of asthma has been proposed.1Akbari O. Stock P. Meyer E. Kronenberg M. Sidobre S. Nakayama T. et al.Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity.Nat Med. 2003; 9: 582-588Crossref PubMed Scopus (600) Google Scholar, 2Lisbonne M. Diem S. de Castro Keller A. Lefort J. Araujo L.M. Hachem P. et al.Cutting edge: invariant V alpha 14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model.J Immunol. 2003; 171: 1637-1641PubMed Google Scholar, 3Pichavant M. Goya S. Meyer E.H. Johnston R.A. Kim H.Y. Matangkasombut P. et al.Ozone exposure in a mouse model induces airway hyperreactivity that requires the presence of natural killer T cells and IL-17.J Exp Med. 2008; 205: 385-393Crossref PubMed Scopus (256) Google Scholar, 4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar However, the role of iNKT cells in human asthma is controversial because there is disagreement on whether iNKT cells are present in the lungs of patients with asthma.5Akbari O. Faul J.L. Hoyte E.G. Berry G.J. Wahlstrom J. Kronenberg M. et al.CD4+ invariant T-cell-receptor+ natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 1117-1129Crossref PubMed Scopus (360) Google Scholar, 6Vijayanand P. Seumois G. Pickard C. Powell R.M. Angco G. Sammut D. et al.Invariant natural killer T cells in asthma and chronic obstructive pulmonary disease.N Engl J Med. 2007; 356: 1410-1422Crossref PubMed Scopus (175) Google Scholar To address this controversy, we examined bronchoalveolar lavage fluid (BALF) from asthmatic subjects with a broad range of severity and symptom control.The study was approved by the Institutional Review Boards at Children's Hospital Boston and Brigham and Women's and Connolly Hospitals. All 18 subjects provided written informed consent and underwent fiberoptic bronchoscopy. Five had severe asthma (based on the National Heart, Lung, and Blood Institute Severe Asthma Research Program criteria),7Moore W.C. Bleecker E.R. Curran-Everett D. Erzurum S.C. Ameredes B.T. Bacharier L. et al.Characterization of the severe asthma phenotype by the National Heart, Lung, and Blood Institute's Severe Asthma Research Program.J Allergy Clin Immunol. 2007; 119: 405-413Abstract Full Text Full Text PDF PubMed Scopus (757) Google Scholar 8 had well-controlled asthma (based on the National Heart, Lung, and Blood Institute Expert Panel Report 3 Guidelines), and 5 had no history of asthma (Table I). BALF samples were analyzed for the presence of iNKT cells by means of flow cytometry (8-Color BD FACSCanto, FlowJo Software; TreeStar, Inc, Ashland, Ore) by investigators who were blinded to clinical data at the time of the analysis. BALF cells were filtered through 70-μm cell strainers to remove mucous and examined for viability with trypan blue (mean viability, 95%) before treatment with human IgG to block Fc receptors. Cells were then stained with PBS57 (analog of α-GalCer)–loaded CD1d tetramer–phycoerythrin (or unloaded CD1d tetramer–phycoerythrin control; National Institutes of Health tetramer facility) and anti-CD3–Alexa750 (BD Biosciences, San Jose, Calif). We gated on the CD3+ low side-scatter population to exclude alveolar macrophages, which have high side scatter and high autofluorescence (Fig 1, A). The percentage of iNKT cells of total T cells was then analyzed (PBS57-loaded CD1d tetramer–positive CD3+ cells/total CD3+ cells). Background staining (unloaded CD1d tetramer–positive CD3+/CD3+ cells) was then subtracted (Fig 1) to calculate the adjusted percentage of iNKT cells of total T cells for statistical analysis (Fig 2).Table ISubjects' demographic and clinical informationGroupSubject IDSexAge (y)FEV1 (%)∗Percent predicted FEV1 at the time of study entry.Exacerbation†Number of acute asthma exacerbation requiring oral corticosteroid, an emergency department visit, or an urgent care visit in the past 12 months.Duration (y)‡Duration that the subject has received a diagnosis of asthma.ICS dose§Inhaled corticosteroid dose (fluticasone or fluticasone equivalent in micrograms per day). Medications were not changed for at least 4 weeks before bronchoscopy.Other medication‖Other medications used for asthma control (not including rescuer medication).Allergies¶Positive response on allergen skin prick testing.Concomitant diseaseSevere asthmaP1Female50781501000Salmeterol, MontelukastCockroachHypertensionP2Male31562251000Salmeterol, Tiotropium, MontelukastNoneNoneP3Male50412501000Salmeterol, TheophyllineCockroachDiabetes, HypertensionP4Male21581151000SalmeterolCockroach, Cat, MiteNoneP5Male49731401880Salmeterol MontelukastAlternaria, Tree, Dog, CatDiabetes HypertensionMean40611.4361176Well-controlled asthmaW1Female3587025500SalmeterolNoneNoneW2Male41109020500SalmeterolNoneNoneW3Female229003500SalmeterolCatNoneW4Male33103030500SalmeterolCockroachNoneW5Female4786040440NoneCockroach, Rat, MiteNoneW6Female568313800MontelukastNoneNoneW7Female598211.5264NoneNoneHypertension, Ischemic heart diseaseW8Female4799020NoneNDPeptic ulcer, Dilated cardiomyopathyMean43920.2516438No asthmaH1Male43ND000NoneNDSarcoidosisH2Male39ND000NoneNDBronchiectasisH3Male4999000NoneNDPeptic ulcerH4Female31114000NoneNDSarcoidosisH5Female5089000NoneNoneNoneMean41107000Six other BALF samples were evaluated by using a different methodology. These samples could not be directly compared with the final 18 samples because the sensitivity for detecting natural killer T cells was not comparable. The trend of results with the 6 other samples was similar to what is reported for the 18 samples.ND, Not done.∗ Percent predicted FEV1 at the time of study entry.† Number of acute asthma exacerbation requiring oral corticosteroid, an emergency department visit, or an urgent care visit in the past 12 months.‡ Duration that the subject has received a diagnosis of asthma.§ Inhaled corticosteroid dose (fluticasone or fluticasone equivalent in micrograms per day). Medications were not changed for at least 4 weeks before bronchoscopy.‖ Other medications used for asthma control (not including rescuer medication).¶ Positive response on allergen skin prick testing. Open table in a new tab Fig 2Adjusted percentages of iNKT cells/T cells were calculated by subtracting background from the percentage of iNKT/T cells. Statistical comparison of adjusted percentages of iNKT/T cells in BALF from patients with poorly controlled or well-controlled asthma and nonasthmatic control subjects was performed by using the Mann-Whitney U test.View Large Image Figure ViewerDownload Hi-res image Download (PPT)We found that patients with severe asthma had a significant increase in the number of BALF iNKT cells when compared with the number seen in nonasthmatic control subjects (P = .004, Mann-Whitney U test; Fig 2). Some, but not all, patients with well-controlled asthma had an increase in the number of BALF iNKT cells compared with the number seen in nonasthmatic control subjects (P = .02). We conclude that iNKT cells are present in the BALF of some, but not all, patients with asthma. The specific number of iNKT cells present in the BALF was quite variable, but patients with severe asthma appeared to more consistently have an increase in BALF iNKT cells than patients with well-controlled asthma, the majority of whom did not have an increase in the number of BALF iNKT cells over that seen in nonasthmatic individuals.These results suggest that asthma severity and control might be loosely related to the number of pulmonary iNKT cells. However, it is also likely that many other factors are involved in determining the number of pulmonary iNKT cells. The number of iNKT cells in the BALF appears to be extremely dynamic and fluctuates rapidly (in mice iNKT cels in BAL fluid range from 1% of lung lymphocytes in naive mice to 14% after allergen challenge). We do not yet fully understand the factors that regulate the number of iNKT cells in the lung. In our previous study5Akbari O. Faul J.L. Hoyte E.G. Berry G.J. Wahlstrom J. Kronenberg M. et al.CD4+ invariant T-cell-receptor+ natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 1117-1129Crossref PubMed Scopus (360) Google Scholar of subjects with poorly controlled asthma, all of the asthmatic patients had higher numbers of pulmonary iNKT cells compared with numbers seen in patients with sarcoidosis or healthy individuals, but the specific number of iNKT cells present was much higher than in the current study. This difference in the number of pulmonary iNKT cells could be due to different patient selection criteria and possibly to improved methods for analyzing iNKT cells.Importantly, our current results help to explain the results observed in the previous 10 studies of iNKT cells in human asthma. In 7 of the studies, a significant increase in the number of iNKT cells in the lungs of patients with asthma compared with that seen in the healthy individuals was observed (see the References in this article's Online Repository at www.jacionline.org).4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar, 5Akbari O. Faul J.L. Hoyte E.G. Berry G.J. Wahlstrom J. Kronenberg M. et al.CD4+ invariant T-cell-receptor+ natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 1117-1129Crossref PubMed Scopus (360) Google Scholar In an eighth study there was a trend toward an increase in the number of iNKT cells in asthma above that seen in control subjects, but this did not reach statistical significance.8Mutalithas K. Croudace J. Guillen C. Siddiqui S. Thickett D. Wardlaw A. et al.Bronchoalveolar lavage invariant natural killer T cells are not increased in asthma.J Allergy Clin Immunol. 2007; 119: 1274-1276Abstract Full Text Full Text PDF PubMed Scopus (40) Google Scholar In 2 additional studies6Vijayanand P. Seumois G. Pickard C. Powell R.M. Angco G. Sammut D. et al.Invariant natural killer T cells in asthma and chronic obstructive pulmonary disease.N Engl J Med. 2007; 356: 1410-1422Crossref PubMed Scopus (175) Google Scholar, 9Thomas S.Y. Lilly C.M. Luster A.D. Invariant natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 2613-2616Crossref PubMed Scopus (48) Google Scholar several asthmatic patients were observed to have an increase in the number of iNKT cells in BALF (up to 2.1% to 2.7% of BALF lymphocytes). However, most of the patients had much fewer pulmonary iNKT cells, such that the authors concluded that iNKT cells were not important in the pathogenesis of asthma, although BALF from healthy individuals was not examined.6Vijayanand P. Seumois G. Pickard C. Powell R.M. Angco G. Sammut D. et al.Invariant natural killer T cells in asthma and chronic obstructive pulmonary disease.N Engl J Med. 2007; 356: 1410-1422Crossref PubMed Scopus (175) Google Scholar, 9Thomas S.Y. Lilly C.M. Luster A.D. Invariant natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 2613-2616Crossref PubMed Scopus (48) Google Scholar In our current study of patients with a very broad range of asthma severity and control, we observed an increase in the number of pulmonary iNKT cells in all 5 patients with severe asthma and in some, but not all, of the patients with well-controlled asthma, which is consistent with all 10 previous studies taken together.Although addressing the major and controversial question of whether iNKT cells are present in the lungs of asthmatic subjects, our study did not focus on other important issues regarding iNKT cells, including whether the number of iNKT cells in the lung varies over time or with exacerbations, analysis of the phenotype, and the subsets of iNKT cells present in the lungs and whether distinct subsets of iNKT cells are present in different forms of asthma, as suggested by murine studies.1Akbari O. Stock P. Meyer E. Kronenberg M. Sidobre S. Nakayama T. et al.Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity.Nat Med. 2003; 9: 582-588Crossref PubMed Scopus (600) Google Scholar, 2Lisbonne M. Diem S. de Castro Keller A. Lefort J. Araujo L.M. Hachem P. et al.Cutting edge: invariant V alpha 14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model.J Immunol. 2003; 171: 1637-1641PubMed Google Scholar, 3Pichavant M. Goya S. Meyer E.H. Johnston R.A. Kim H.Y. Matangkasombut P. et al.Ozone exposure in a mouse model induces airway hyperreactivity that requires the presence of natural killer T cells and IL-17.J Exp Med. 2008; 205: 385-393Crossref PubMed Scopus (256) Google Scholar, 4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar The limitations of our study were due in part to the small number of patients included in our report and the fact that repeat bronchoscopy of patients with severe asthma cannot be easily accomplished. However, our results strongly suggest that additional studies with a larger number of subjects are warranted to examine these questions.The observation that iNKT cells are present in some patients with asthma suggests that iNKT cells might contribute to the pathogenesis of asthma. This question is of much greater importance but must be addressed by means of examination of iNKT cell function in asthma. Such functional studies are difficult to perform in human subjects for ethical and safety reasons, but studies in mice and nonhuman primates provide a very compelling argument regarding the importance of iNKT cells in the development of airway hyperreactivity (AHR). Thus in 3 distinct models of asthma in mice induced with allergen1Akbari O. Stock P. Meyer E. Kronenberg M. Sidobre S. Nakayama T. et al.Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity.Nat Med. 2003; 9: 582-588Crossref PubMed Scopus (600) Google Scholar, 2Lisbonne M. Diem S. de Castro Keller A. Lefort J. Araujo L.M. Hachem P. et al.Cutting edge: invariant V alpha 14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model.J Immunol. 2003; 171: 1637-1641PubMed Google Scholar or virus4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar or by means of exposure to ozone (a component of air pollution),3Pichavant M. Goya S. Meyer E.H. Johnston R.A. Kim H.Y. Matangkasombut P. et al.Ozone exposure in a mouse model induces airway hyperreactivity that requires the presence of natural killer T cells and IL-17.J Exp Med. 2008; 205: 385-393Crossref PubMed Scopus (256) Google Scholar AHR did not develop in the absence of iNKT cells. In each of these models, a different subset of iNKT cells was required for AHR. Moreover, functional studies of iNKT cells, performed by using direct and specific activation of iNKT cells in naive mice10Meyer E.H. Goya S. Akbari O. Berry G.J. Savage P.B. Kronenberg M. et al.Glycolipid activation of invariant T cell receptor+ NK T cells is sufficient to induce airway hyperreactivity independent of conventional CD4+ T cells.Proc Natl Acad Sci U S A. 2006; 103: 2782-2787Crossref PubMed Scopus (181) Google Scholar or in cynomolgus monkeys,11Matangkasombut P. Pichavant M. Yasumi T. Hendricks C. Savage P.B. Dekruyff R.H. et al.Direct activation of natural killer T cells induces airway hyperreactivity in nonhuman primates.J Allergy Clin Immunol. 2008; 121: 1287-1289Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar resulted in the development of AHR, suggesting that even the few iNKT cells present in the normal lung, when directly activated, can result in AHR. These functional studies of iNKT cells together suggest that iNKT cells might represent an important common element that links many different forms of asthma, including allergic and nonallergic asthma. These results also suggest that the absolute number of iNKT cells present in the lungs might not be as important as their functional capacity when activated to contribute to the development of AHR. To the Editor: On the basis of studies in mice, an important role for invariant natural killer T (iNKT) cells in the pathogenesis of asthma has been proposed.1Akbari O. Stock P. Meyer E. Kronenberg M. Sidobre S. Nakayama T. et al.Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity.Nat Med. 2003; 9: 582-588Crossref PubMed Scopus (600) Google Scholar, 2Lisbonne M. Diem S. de Castro Keller A. Lefort J. Araujo L.M. Hachem P. et al.Cutting edge: invariant V alpha 14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model.J Immunol. 2003; 171: 1637-1641PubMed Google Scholar, 3Pichavant M. Goya S. Meyer E.H. Johnston R.A. Kim H.Y. Matangkasombut P. et al.Ozone exposure in a mouse model induces airway hyperreactivity that requires the presence of natural killer T cells and IL-17.J Exp Med. 2008; 205: 385-393Crossref PubMed Scopus (256) Google Scholar, 4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar However, the role of iNKT cells in human asthma is controversial because there is disagreement on whether iNKT cells are present in the lungs of patients with asthma.5Akbari O. Faul J.L. Hoyte E.G. Berry G.J. Wahlstrom J. Kronenberg M. et al.CD4+ invariant T-cell-receptor+ natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 1117-1129Crossref PubMed Scopus (360) Google Scholar, 6Vijayanand P. Seumois G. Pickard C. Powell R.M. Angco G. Sammut D. et al.Invariant natural killer T cells in asthma and chronic obstructive pulmonary disease.N Engl J Med. 2007; 356: 1410-1422Crossref PubMed Scopus (175) Google Scholar To address this controversy, we examined bronchoalveolar lavage fluid (BALF) from asthmatic subjects with a broad range of severity and symptom control. The study was approved by the Institutional Review Boards at Children's Hospital Boston and Brigham and Women's and Connolly Hospitals. All 18 subjects provided written informed consent and underwent fiberoptic bronchoscopy. Five had severe asthma (based on the National Heart, Lung, and Blood Institute Severe Asthma Research Program criteria),7Moore W.C. Bleecker E.R. Curran-Everett D. Erzurum S.C. Ameredes B.T. Bacharier L. et al.Characterization of the severe asthma phenotype by the National Heart, Lung, and Blood Institute's Severe Asthma Research Program.J Allergy Clin Immunol. 2007; 119: 405-413Abstract Full Text Full Text PDF PubMed Scopus (757) Google Scholar 8 had well-controlled asthma (based on the National Heart, Lung, and Blood Institute Expert Panel Report 3 Guidelines), and 5 had no history of asthma (Table I). BALF samples were analyzed for the presence of iNKT cells by means of flow cytometry (8-Color BD FACSCanto, FlowJo Software; TreeStar, Inc, Ashland, Ore) by investigators who were blinded to clinical data at the time of the analysis. BALF cells were filtered through 70-μm cell strainers to remove mucous and examined for viability with trypan blue (mean viability, 95%) before treatment with human IgG to block Fc receptors. Cells were then stained with PBS57 (analog of α-GalCer)–loaded CD1d tetramer–phycoerythrin (or unloaded CD1d tetramer–phycoerythrin control; National Institutes of Health tetramer facility) and anti-CD3–Alexa750 (BD Biosciences, San Jose, Calif). We gated on the CD3+ low side-scatter population to exclude alveolar macrophages, which have high side scatter and high autofluorescence (Fig 1, A). The percentage of iNKT cells of total T cells was then analyzed (PBS57-loaded CD1d tetramer–positive CD3+ cells/total CD3+ cells). Background staining (unloaded CD1d tetramer–positive CD3+/CD3+ cells) was then subtracted (Fig 1) to calculate the adjusted percentage of iNKT cells of total T cells for statistical analysis (Fig 2). Six other BALF samples were evaluated by using a different methodology. These samples could not be directly compared with the final 18 samples because the sensitivity for detecting natural killer T cells was not comparable. The trend of results with the 6 other samples was similar to what is reported for the 18 samples. ND, Not done. We found that patients with severe asthma had a significant increase in the number of BALF iNKT cells when compared with the number seen in nonasthmatic control subjects (P = .004, Mann-Whitney U test; Fig 2). Some, but not all, patients with well-controlled asthma had an increase in the number of BALF iNKT cells compared with the number seen in nonasthmatic control subjects (P = .02). We conclude that iNKT cells are present in the BALF of some, but not all, patients with asthma. The specific number of iNKT cells present in the BALF was quite variable, but patients with severe asthma appeared to more consistently have an increase in BALF iNKT cells than patients with well-controlled asthma, the majority of whom did not have an increase in the number of BALF iNKT cells over that seen in nonasthmatic individuals. These results suggest that asthma severity and control might be loosely related to the number of pulmonary iNKT cells. However, it is also likely that many other factors are involved in determining the number of pulmonary iNKT cells. The number of iNKT cells in the BALF appears to be extremely dynamic and fluctuates rapidly (in mice iNKT cels in BAL fluid range from 1% of lung lymphocytes in naive mice to 14% after allergen challenge). We do not yet fully understand the factors that regulate the number of iNKT cells in the lung. In our previous study5Akbari O. Faul J.L. Hoyte E.G. Berry G.J. Wahlstrom J. Kronenberg M. et al.CD4+ invariant T-cell-receptor+ natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 1117-1129Crossref PubMed Scopus (360) Google Scholar of subjects with poorly controlled asthma, all of the asthmatic patients had higher numbers of pulmonary iNKT cells compared with numbers seen in patients with sarcoidosis or healthy individuals, but the specific number of iNKT cells present was much higher than in the current study. This difference in the number of pulmonary iNKT cells could be due to different patient selection criteria and possibly to improved methods for analyzing iNKT cells. Importantly, our current results help to explain the results observed in the previous 10 studies of iNKT cells in human asthma. In 7 of the studies, a significant increase in the number of iNKT cells in the lungs of patients with asthma compared with that seen in the healthy individuals was observed (see the References in this article's Online Repository at www.jacionline.org).4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar, 5Akbari O. Faul J.L. Hoyte E.G. Berry G.J. Wahlstrom J. Kronenberg M. et al.CD4+ invariant T-cell-receptor+ natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 1117-1129Crossref PubMed Scopus (360) Google Scholar In an eighth study there was a trend toward an increase in the number of iNKT cells in asthma above that seen in control subjects, but this did not reach statistical significance.8Mutalithas K. Croudace J. Guillen C. Siddiqui S. Thickett D. Wardlaw A. et al.Bronchoalveolar lavage invariant natural killer T cells are not increased in asthma.J Allergy Clin Immunol. 2007; 119: 1274-1276Abstract Full Text Full Text PDF PubMed Scopus (40) Google Scholar In 2 additional studies6Vijayanand P. Seumois G. Pickard C. Powell R.M. Angco G. Sammut D. et al.Invariant natural killer T cells in asthma and chronic obstructive pulmonary disease.N Engl J Med. 2007; 356: 1410-1422Crossref PubMed Scopus (175) Google Scholar, 9Thomas S.Y. Lilly C.M. Luster A.D. Invariant natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 2613-2616Crossref PubMed Scopus (48) Google Scholar several asthmatic patients were observed to have an increase in the number of iNKT cells in BALF (up to 2.1% to 2.7% of BALF lymphocytes). However, most of the patients had much fewer pulmonary iNKT cells, such that the authors concluded that iNKT cells were not important in the pathogenesis of asthma, although BALF from healthy individuals was not examined.6Vijayanand P. Seumois G. Pickard C. Powell R.M. Angco G. Sammut D. et al.Invariant natural killer T cells in asthma and chronic obstructive pulmonary disease.N Engl J Med. 2007; 356: 1410-1422Crossref PubMed Scopus (175) Google Scholar, 9Thomas S.Y. Lilly C.M. Luster A.D. Invariant natural killer T cells in bronchial asthma.N Engl J Med. 2006; 354: 2613-2616Crossref PubMed Scopus (48) Google Scholar In our current study of patients with a very broad range of asthma severity and control, we observed an increase in the number of pulmonary iNKT cells in all 5 patients with severe asthma and in some, but not all, of the patients with well-controlled asthma, which is consistent with all 10 previous studies taken together. Although addressing the major and controversial question of whether iNKT cells are present in the lungs of asthmatic subjects, our study did not focus on other important issues regarding iNKT cells, including whether the number of iNKT cells in the lung varies over time or with exacerbations, analysis of the phenotype, and the subsets of iNKT cells present in the lungs and whether distinct subsets of iNKT cells are present in different forms of asthma, as suggested by murine studies.1Akbari O. Stock P. Meyer E. Kronenberg M. Sidobre S. Nakayama T. et al.Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity.Nat Med. 2003; 9: 582-588Crossref PubMed Scopus (600) Google Scholar, 2Lisbonne M. Diem S. de Castro Keller A. Lefort J. Araujo L.M. Hachem P. et al.Cutting edge: invariant V alpha 14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model.J Immunol. 2003; 171: 1637-1641PubMed Google Scholar, 3Pichavant M. Goya S. Meyer E.H. Johnston R.A. Kim H.Y. Matangkasombut P. et al.Ozone exposure in a mouse model induces airway hyperreactivity that requires the presence of natural killer T cells and IL-17.J Exp Med. 2008; 205: 385-393Crossref PubMed Scopus (256) Google Scholar, 4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar The limitations of our study were due in part to the small number of patients included in our report and the fact that repeat bronchoscopy of patients with severe asthma cannot be easily accomplished. However, our results strongly suggest that additional studies with a larger number of subjects are warranted to examine these questions. The observation that iNKT cells are present in some patients with asthma suggests that iNKT cells might contribute to the pathogenesis of asthma. This question is of much greater importance but must be addressed by means of examination of iNKT cell function in asthma. Such functional studies are difficult to perform in human subjects for ethical and safety reasons, but studies in mice and nonhuman primates provide a very compelling argument regarding the importance of iNKT cells in the development of airway hyperreactivity (AHR). Thus in 3 distinct models of asthma in mice induced with allergen1Akbari O. Stock P. Meyer E. Kronenberg M. Sidobre S. Nakayama T. et al.Essential role of NKT cells producing IL-4 and IL-13 in the development of allergen-induced airway hyperreactivity.Nat Med. 2003; 9: 582-588Crossref PubMed Scopus (600) Google Scholar, 2Lisbonne M. Diem S. de Castro Keller A. Lefort J. Araujo L.M. Hachem P. et al.Cutting edge: invariant V alpha 14 NKT cells are required for allergen-induced airway inflammation and hyperreactivity in an experimental asthma model.J Immunol. 2003; 171: 1637-1641PubMed Google Scholar or virus4Kim E.Y. Battaile J.T. Patel A.C. You Y. Agapov E. Grayson M.H. et al.Persistent activation of an innate immune response translates respiratory viral infection into chronic lung disease.Nat Med. 2008; 14: 633-640Crossref PubMed Scopus (421) Google Scholar or by means of exposure to ozone (a component of air pollution),3Pichavant M. Goya S. Meyer E.H. Johnston R.A. Kim H.Y. Matangkasombut P. et al.Ozone exposure in a mouse model induces airway hyperreactivity that requires the presence of natural killer T cells and IL-17.J Exp Med. 2008; 205: 385-393Crossref PubMed Scopus (256) Google Scholar AHR did not develop in the absence of iNKT cells. In each of these models, a different subset of iNKT cells was required for AHR. Moreover, functional studies of iNKT cells, performed by using direct and specific activation of iNKT cells in naive mice10Meyer E.H. Goya S. Akbari O. Berry G.J. Savage P.B. Kronenberg M. et al.Glycolipid activation of invariant T cell receptor+ NK T cells is sufficient to induce airway hyperreactivity independent of conventional CD4+ T cells.Proc Natl Acad Sci U S A. 2006; 103: 2782-2787Crossref PubMed Scopus (181) Google Scholar or in cynomolgus monkeys,11Matangkasombut P. Pichavant M. Yasumi T. Hendricks C. Savage P.B. Dekruyff R.H. et al.Direct activation of natural killer T cells induces airway hyperreactivity in nonhuman primates.J Allergy Clin Immunol. 2008; 121: 1287-1289Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar resulted in the development of AHR, suggesting that even the few iNKT cells present in the normal lung, when directly activated, can result in AHR. These functional studies of iNKT cells together suggest that iNKT cells might represent an important common element that links many different forms of asthma, including allergic and nonallergic asthma. These results also suggest that the absolute number of iNKT cells present in the lungs might not be as important as their functional capacity when activated to contribute to the development of AHR. We thank the National Institutes of Health tetramer facility for providing CD1d tetramers and Dr Leslie Kalish, Children's Hospital Boston, for help with the statistical analysis. Natural killer T cells in bronchial biopsies from human allergen challenge model of allergic asthmaJournal of Allergy and Clinical ImmunologyVol. 124Issue 4PreviewTo the Editor: Full-Text PDF
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