Interleukin-17 and airway inflammation: a longitudinal airway biopsy study after lung transplantation.

BACKGROUND: Interleukin-17 (IL-17) is a pro-inflammatory cytokine produced from CD4+ T cells and is associated with neutrophilia in infection, ischemia-reperfusion injury, and possibly acute and chronic rejection (bronchiolitis obliterans syndrome, or BOS) after lung transplantation (LTx). Everolimus (ERL) decreases acute rejection, possibly via decreasing airway CD4+ cells and neutrophils. This prospective study aims to assess: (1) the possible role of IL-17 as a link between LTx clinical outcomes (such as infection, acute rejection and BOS) and airway immunopathologic measures from endobronchial biopsy (EBB) and bronchoalveolar lavage (BAL); and (2) any differences in IL-17 production between ERL and azathioprine (AZA)-based immunosuppression. METHODS: This sub-study, from a larger, prospective clinical ERL vs AZA randomized, controlled trial, examines EBB IL-17 expression, relating this to clinical outcomes, BAL and EBB cell counts. EBB IL-17 staining was measured by immunohistologic techniques and expressed as cells per square millimeter of lamina propria. RESULTS: Thirty-four LTx patients were randomized in a double-blind study (ERL = 19, AZA = 15) and underwent a total of 113 bronchoscopies over a 3-year follow-up period. Twenty-six EBBs were taken from LTx recipients with BOS of at least Grade 0p (10 patients). Univariate associations correlated IL-17 positively with EBB CD8+ cells (R2 = 0.010, p = 0.001) and negatively with days post-LTx (R2 = 0.07, p = 0.002). In a multivariate model, IL-17 variability was explained by: days post-LTx (6.2%, p = 0.02); EBB CD8+ (5.9%, p = 0.02); cytomegalovirus mismatch (6.1%, p = 0.02); BAL lymphocyte percentage (4.2%, p = 0.05); and clinical infection (3.7%, p = 0.06). CONCLUSIONS: IL-17 is associated with the early post-LTx time period and airway CD8+ cells. Unexpectedly, rejection grade, BOS, BAL IL-8 and neutrophil counts are not associated. ERL appears not to directly affect IL-17, despite its effects on CD4 cells.

Airway vascular changes after lung transplant: potential contribution to the pathophysiology of bronchiolitis obliterans syndrome.

BACKGROUND: Bronchiolitis obliterans syndrome (BOS) remains the primary factor limiting successful lung transplantation. In asthma and lung transplantation BOS-increased sub-mucosal vascularity has been shown to contribute to airflow limitation. Vascularity has 2 components: sprouting angiogenesis (more vessels) and microvascular enlargement (larger vessels). We hypothesized that the lack of a reanastomosed bronchial arterial blood supply at the time of transplant might stimulate angiogenesis and be a risk factor for subsequent BOS. METHODS: Twenty-seven initially stable lung transplant recipients (BOS 0) were recruited at 148 +/- 80 days post-transplant and underwent clinical and bronchoscopic longitudinal follow-up for at least 3 years. Eight remained stable and BOS developed in 19. Nine normal controls were also recruited. Airway vasculature was examined immunohistochemically in endobronchial biopsy (EBB) specimens with collagen IV antibody, quantified by computer image analysis, and expressed as average vessel size, vessel number, and overall vascularity. The effects of demographic, clinical, bronchoalveolar lavage (BAL), and EBB variables on airway vasculature were analyzed in a multivariate model. RESULTS: No significant differences in airway vascularity were found between stable and BOS lung transplant recipients cross-sectionally or longitudinally. However, both lung transplant groups at baseline showed significantly greater airway vascularity compared with normal controls (p < .05). Multivariate analysis suggested that the percentage of BAL CD3+ cells and acute rejection are the most influential variables on airway vasculature. CONCLUSIONS: This study suggests early and persistent airway vasculature changes occur in lung transplant recipients, mainly manifested as microvascular enlargement. Potentially this baseline change contributes to airway obstruction and also puts all lung transplant recipients at risk for further exponential loss of airway caliber with any subsequent airway inflammatory insult.

Early changes in basement membrane thickness in airway walls post-lung transplantation.

BACKGROUND: Identification of early histopathologic markers of future bronchiolitis obliterans syndrome (BOS) may enable preemptive targeted intervention, delaying and perhaps preventing the onset of BOS. This study aimed to determine if early changes in airway epithelial basement membrane thickness predisposes transplant recipients to the subsequent development of BOS. METHODS: Basement membrane thickness was measured in serial endobronchial biopsies taken from 29 initially stable lung transplant recipients (sLTR) recruited 148 +/- 80 days post-transplant and followed for 3 years. A further 2 years of clinical follow-up was undertaken without biopsies to follow lung function and define ultimate BOS status. Nine healthy subjects (non-atopic, non-asthmatic) were recruited as controls. Sections of paraffinized endobronchial biopsies were stained for collagen type I immunohistochemically, and basement membrane thickness was assessed by computer image analysis. RESULTS: BOS developed in 21 of 29 patients in the 5 years of follow-up, 16 of which had endobronchial biopsies available for analysis before BOS developed (ever-BOS). The first endobronchial biopsies showed increased BMT in the combined sLTR and ever-BOS patients compared with the controls. This initial increase in basement membrane thickness resolved to normal levels within 300 days post-transplant, with a strong negative correlation (r2 = 0.424, p < 0.0001) of basement membrane thickness vs time. Paradoxically, the sLTR tended to have the greatest basement membrane thickness at baseline. CONCLUSION: An initial increase in basement membrane thickness is seen in the airway walls of all lung transplant recipients. This is transient and does not appear to be a risk factor for the subsequent development of BOS in lung allograft recipients.

Longitudinal comparisons of lymphocytes and subtypes between airway wall and bronchoalveolar lavage after human lung transplantation.

BACKGROUND: T lymphocytes are crucial in lung allorejection. The contribution of lymphocyte subtypes to the pathogenesis of chronic rejection (bronchiolitis obliterans syndrome [BOS]) remains unclear. METHODS: Twenty-nine initially healthy lung transplant recipients underwent 136 bronchoscopic assessments, including bronchoalveolar lavage (BAL) (with flow cytometry) and endobronchial biopsies (EBB) (with immunohistochemistry) over 3 years of follow-up. RESULTS: Of the 29 patients studied over 3 years, 23 developed BOS category 0 p and 17 went on to BOS 1. Compared with controls, the BAL percentage of CD4 cells was lower and the percentage of CD8 cells was increased significantly early posttransplant. Subsequent BAL lymphocyte subtype changes with time, or with the development of BOS, were minimal. By contrast, the early posttransplant EBB lymphocyte numbers were normal (P>0.05 vs. controls); subsequently, CD3 and CD8 (but not CD4) cells were increased with time in patients who did not develop BOS (P<0.05) and, more strikingly, in patients who eventually developed BOS (P<0.01). Multivariate analyses suggested an association between BAL lymphocytes (percentage) and azathioprine dose, female gender, rejection grade A on transbronchial biopsies, and pre-BOS status, whereas EBB CD8 cell counts were associated with time posttransplant, pretransplant diagnosis, and rejection grade B on TBB. CONCLUSIONS: There is an early, persistent low percentage of BAL CD4 T cells, high BAL CD8 T cells, and progressively increasing airway wall CD3 and CD8 T cells with time posttransplant in healthy patients (but more predominantly in BOS patients) after transplantation. These immunopathologic changes may suggest that CD8 T cells could escape current immunosuppression and participate in chronic lung rejection.

Everolimus alters the bronchoalveolar lavage and endobronchial biopsy immunologic profile post-human lung transplantation.

Everolimus has recently shown promise in terms of short- and long-term clinical lung transplant outcomes. This study aims to determine the altered lung allograft cellular and cytokine mileau when everolimus is substituted for azathioprine (AZA). Twenty-three stable lung transplantation (LTx) recipients were randomized in a double-blinded study to receive everolimus (13) or AZA (10) plus standard cyclosporine/prednisolone. Bronchoalveolar lavage (BAL) and endobronchial biopsies (EBB) were performed on three occasions (T(0)-T(2)) to elucidate cellular and cytokine profiles via immunocytochemistry, immunohistology and enzyme-linked immunosorbent assay (ELISA) techniques. There were no group differences for demographics or clinical events throughout the study nor baseline cellular/cytokine differences. BAL lymphocyte percentage fell in the AZA group by T(2) (p = 0.05). BAL and EBB CD4 measures significantly declined in the everolimus group by T(2) (p < 0.05). EBB neutrophils rose significantly in the AZA group, with a fall in the everolimus group resulting in a significant difference at T(2) (p = 0.01). In conclusion, everolimus has contributed to potentially important differences in BAL and EBB cellular profiles.

Addition of inhaled corticosteroids to systemic immunosuppression after lung transplantation: a double-blind, placebo-controlled trial.

BACKGROUND: It is postulated that bronchiolitis obliterans syndrome (BOS) is preceded by airway inflammation that has been described even in stable lung transplant recipients. Airway inflammation is known to be suppressed by inhaled steroids in other chronic inflammatory lung diseases, e.g., asthma and chronic obstructive pulmonary disease. BOS is the major cause of morbidity and mortality after lung transplantation. OBJECTIVE: To examine the effect of inhaled corticosteroids on the development of BOS in lung transplant recipients. METHODS: Thirty patients were recruited and randomized in a double-blind fashion to receive either 750 microg of fluticasone propionate (FP) or an identical-appearing placebo twice daily for 3 months; 20 of this group continued until 2 years posttransplantation. Detailed spirometry was performed regularly throughout the study. RESULTS: In the short-term study no differences were found in any examined parameters. In the long-term component of the study no differences were found in the development of neither BOS nor survival. There were minor differences in bronchoalveolar lavage (BAL) lymphocyte percentages. CONCLUSIONS: FP is ineffective for the prevention of BOS after lung transplantation despite the airway inflammation that characterizes this condition. Inadequate local delivery, timing of the therapy relative to transplantation and inherent steroid resistance of this condition may explain the negative finding of this study.

Clinical relevance of airway 11beta-hydroxysteroid dehydrogenase type II enzyme in asthma.

11beta-hydroxysteroid dehydrogenases (11beta-HSD) are responsible for the conversion of bioactive glucocorticoids to and from inactive metabolites. 11beta-HSD2 is generally considered a high-affinity inactivator of natural glucocorticoids, although its activity with synthetic compounds in vivo is unknown. Inhaled corticosteroids (ICS) remain the primary antiinflammatory agents for treating asthma, but little is known about their metabolism in the lung. The aims of this study were to determine whether the 11beta-HSD2 enzyme can be localized to human airway tissue and whether differential expression of this enzyme relates to asthma severity and ICS needs. We studied airway biopsy specimens from 22 asthmatic subjects, in two groups: (1) a group not treated with ICS (n = 7); and (2) a group treated with ICS (range: 200 to 1,500 microg/d; n = 15). A control population consisted of nine nonasthmatic subjects. Immunostaining was done with an immunopurified antibody to human 11beta-HSD2. Immunoreactivity was generally localized to the endothelium of vessels in the lamina propria and to airway epithelium both in asthmatic patients and nonasthmatic controls. There was a statistically significant inverse relationship between the ICS dose required for effective treatment and the extent of epithelial 11beta-HSD2 staining (r = -0.44; p = 0.04). This is consistent with 11beta-HSD2 acting as an oxidoreductase that regenerates rather than inactivates ICS. This study suggests that glucocorticoid sensitivity in the lung is not determined by ICS breakdown, but may be related to 11beta-HSD2 sustaining the activation of synthetic glucocorticoids.