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.


CONCLUSIONS
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.
Kunisawa
Akira
A
First Department of Internal Medicine, Tokyo Medical University, Tokyo 160-0023, Japan. a-kuni@mwd.biglobe.ne.jp
Kawanishi
Yoshikazu
Y
Tago
Hidekazu
H
Nagate
Akira
A
Kasuga
Ikuma
I
Ohyashiki
Kazuma
K
D000328:Adult; D001992:Bronchoalveolar Lavage Fluid; D015704:CD4 Antigens / Q000378:metabolism; D016827:CD8 Antigens / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism*; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D015847:Interleukin-4 / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D017565:Sarcoidosis, Pulmonary / Q000378:metabolism*; D016176:T-Lymphocyte Subsets / Q000378:metabolism*
D015704:CD4 Antigens; D016827:CD8 Antigens; D016207:Cytokines; D015847:Interleukin-4