Changes of Rho kinase activity in peripheral blood T lymphocytes from patients with atopic dermatitis and their significance / 中华皮肤科杂志

ABSTRACT

Objective To evaluate changes of Rho kinase (ROK)activity in peripheral blood T lymphocytes from patients with atopic dermatitis (AD), and to analyze their clinical significance. Methods Eight milliliters of heparin-anticoagulated blood samples were collected from 60 patients with AD and 60 healthy human controls followed by separation of T lymphocytes and sera from these blood samples as well as culture of isolated T lymphocytes with 10% fetal bovine serum for 24 hours. Both patient- and control-derived T lymphocytes were classified into two groups to be cultured with patient- or control-derived sera. In addition, some patient-derived T lymphocytes were classified into 4 groups:Y27632 group treated with the Rho kinase-specific inhibitor Y2763, CD3/CD28 group treated with anti-CD3/anti-CD28 monoclonal antibodies, Y27632 + CD3/CD28 group treated with Y27632 and anti-CD3/anti-CD28 monoclonal antibodies, and control group treated with patient-derived sera. Subsequently, Western-blot analysis was performed to evaluate ROK activity in cells, methyl thiazolyl tetrazolium(MTT)assay to evaluate proliferative activity of T lymphocytes, and ELISA to measure interleukin 6 (IL-6)and IL-10 levels in supernatants of T lymphocytes. Results ROK activity was significantly lower in fresh T lymphocytes from patients than in those from healthy controls (2.47% ± 0.89% vs. 0.65% ± 0.35%, t =2.729, P 0.05). Compared with T lymphocytes cultured with control-derived sera, those cultured with patient-derived sera showed significantly increased ROK activity (F = 8.22, P < 0.001). Concretely speaking, ROK activity was significantly higher in patient-derived T lymphocytes cultured with patient-derived sera than in those cultured with control-derived sera (2.41% ± 0.87% vs. 0.76% ± 0.41%, P < 0.05), and higher in control-derived T lymphocytes cultured with patient-derived sera than in those cultured with control-derived sera(2.17% ± 0.85% vs. 0.64% ± 0.33%, P< 0.05)at 24 hours. Y27632 could significantly inhibit the proliferation of as well as secretion of IL-6 (F = 18.68, 22.95, respectively, both P < 0.001)by patient-derived T lymphocytes, but had insignificant effects on secretion of IL-10. The cellular proliferative activity and IL-6 supernatant level were significantly lower in the Y27632 group than in the control group, and lower in the Y27632 + CD3/CD28 group than in the CD3/CD28 group (all P < 0.05). Conclusion Aberrant activation of ROK exists in T lymphocytes from patients with AD, which may play a certain role in the pathogenesis of AD.