Paradoxically decreased intracellular collagenase activity in macrophages from bronchoalveolar lavage fluid in current smokers and patients with obstructive ventilatory disorder.

The roles of matrix metalloproteinases (MMPs) in chronic obstructive pulmonary disease (COPD) have been studied. Macrophages are considered to release MMPs in the lung tissue. We measured intracellular collagenase activity in intact CD14(+)CD45++ cells from the bronchoalveolar lavage fluid (BALF) of patients with obstructive ventilatory impairment and other respiratory diseases. Collagenase activity in current smokers was significantly lower than those in non-smokers (3.25+/-0.54 versus 5.48+/-0.55; P=0.006), and also lower than those in ex-smokers (versus 6.54+/-1.54; P=0.019). We found a lower activity of collagenase in patients with FEV1/FVC below 70% than those with 70% or higher (2.68+/-0.59 versus 4.51+/-0.44; P=0.034). Mean value of FEV1/FVC in patients with collagenase activity of 4 or higher was significantly elevated as compared to those with the activity lower than 4 (83.3+/-3.3 versus 71.8+/-4.9; P=0.021). The discrepancy between increased release of MMPs by previous reports and decreased intracellular activity in our results, may be explained by the production of inactive form of MMPs is relatively increased in COPD. Our study may provide the future direction in investigating the mechanism of COPD. In clinics, this measurement in patients with smoking habits may be helpful to advise them to stop smoking, and to avoid progression to the irreversible obstructive disease.

Direct demonstration of the productive capability of cytokines at the single cell level in lung sarcoidosis using multicolor cytometry.

BACKGROUND: Sarcoidosis is a chronic systemic disorder of unknown etiology characterized by noncaseating epithelioid cell granulomatous lesions, around which an increasing number of CD4+ T cells infiltrate. These CD4+ T cells may release interferon-gamma (IFN-gamma) and interleukin-2 (IL-2). These cytokines are considered to play an important role in pathogenesis of sarcoidosis. METHODS: We employed a modification of Jung's method using multicolor flow cytometry to assess the capability of single cells obtained from bronchoalveolar lavage (BAL) fluid to produce various cytokines. BAL CD4+ T cell production of IFN-gamma, IL-2 and IL-4 after phorbol ester and ionomycin stimulation were studied. RESULTS: The percentage of IFN-gamma- and IL-2-producing CD4+ T cells was significantly higher in patients with sarcoidosis compared to healthy volunteers [84.7 +/- 7.5 vs. 51.2 +/- 14.8% (p < 0.005), and 75.3 +/- 8.7 vs. 39.8 +/-11.0% (p < 0.001), respectively]. No significant difference in the percentage of IL-4-producing CD4+ T cells was noted (1.2 +/- 0.6 vs. 3.5 +/- 2.6%; not significant), whereas the absolute number of IL-4-producing CD4+ T cells was significantly higher in patients with sarcoidosis compared to healthy volunteers (563.6 +/- 330.2 vs. 50.9 +/- 66.9/ml; p < 0.005). In the IL-4-producing CD4+ T cells, about 80% of cells concomitantly produced IFN-gamma and more than 60% of cells also produced IL-2. CONCLUSION: We demonstrate that Th1-like-producing cells are predominant in the CD4+ as well as in the CD8+ T cell subset of patients with sarcoidosis. We for the first time demonstrated concomitant capabilities of BAL CD4+ T cells to produce Th1 and Th2 cytokines at the single cell level by multicolor flow cytometry.