| BackgroundSystemic lupus erythematosus (SLE) is a chronic, multi-system-involved autoimmunedisease, and greatly threatens the health of human beings. There are about1million sufferersin our country, but its pathogenesis is not fully understood to date. The major treatment forSLE now is using glucocorticoids combined with a variety of drugs such ascyclophosphamide, methotrexate, leflunomide, and so on. Since hydroxychloroquine (HCQ)has widely been applied in treating SLE and showed a certain effect, it is receiving more andmore attention in its role in rheumatism treatment. However, the exact mechanism has notbeen known. In recent years, HCQ was used to treat SLE in our department, and found thatthe number of leukocytes in patients' peripheral blood, especially that of lymphocytes wasdecreased to some degree. This indicates that HCQ might improve this condition throughaffecting lymphocytes.In this study, it is aimed to investigate the effect of HCQ on the apoptosis of lymphocytesand T helper cell type17(Th17) and the level of its secreting cytokine IL-17, in order toexplore the possible mechanism of HCQ in treating SLE.Objective1. To observe the effect of HCQ on peripheral blood mononuclear cells (PBMC) of SLEpatients.2. To investigate the effect of HCQ on the apoptosis of PBMC of SLE patients.3. To investigate the effect of HCQ on Th17and the expression level of IL-17, and toexplore the possible mechanism of HCQ in treating SLE.Methods1. In vitro studies:40cases of incipient SLE patients still in a stable period wererandomly divided into Cyclophosphamide (CTX) treated group and HCQ+CTX treated group,effects on lymphocytes were compared between these two groups. Indexes observed includethe number of White Blood Cells (WBCs) and lymphocytes (LY), systemic lupus erythematosus disease activity index (SLEDAI), Erythrocyte Sedimentation Rate (ESR) andcomplement C3. Follow up time points include0w (baseline),6w, and12w.2êIn vivo studies:(1) Peripheral blood of20cases of incipient SLE patients in activestage was drawn, and PBMC were separated for cell culture. Using HCQ and ChlormethineHydrochloride (NH2), respectively, with different concentrations, as intervention, and aftercultured for24h and48h, using Flow cytometry (FCM) to test the apoptosis of PBMC andthe level of Th17; supernatant of the cultured cells was collected to test the level of cytokineIL-17in it by ELISA.Results1. Evaluation of clinical indexes:At the baseline, comparing CTX treated group and HCQ+CTX treated group, there wereno significant difference in WBCs, LY, SLEDAI, ESR, and Complement C3(P>0.05).At6w, as compared with those at the baseline: SLEDAI scores of both groups weredecreased significantly (P<0.05), but there were no significant differences in the degrees ofchange and between the two groups (P>0.05); while there were no significant differences inthe indexes as WBCs, LY, ESR, and Complement C3, and their degrees of change (P>0.05).At12w, as compared with those at the baseline: SLEDAI scores and ESR of both groupswere decreased significantly (P<0.05), and Complement C3was increased significantly(P<0.05); as for WBC and LY in HCQ+CTX treated group compared with those in CTXtreated group, they were decreased to some degree, but more significant (P<0.05); and ifcompared with those at6w, there was no significant difference for each index (P>0.05); from6w to12w, the decrease of SLEDAI score in HCQ+CTX treated group was more significantthan that in CTX treated group (P<0.05); but there was no significant difference in the degreeof change of ESR and Complement C3between the two groups (P>0.05).2. PBMC from SLE patients were extracted for cell culture, adding HCQ and NH2withdifferent concentrations: HCQ lower concentration group (final concentration was5g/ml),HCQ higher concentration group (final concentration was25g/ml), NH2lower concentrationgroup (final concentration was0.1g/ml), and NH2higher concentration group (finalconcentration was1g/ml). The apoptosis rate at24h was tested by FCM, and the results ofHCQ lower concentration group and NH2lower concentration group indicated(10.325±0.713)%and (8.490±1.065)%(P<0.05); and the results of HCQ higherconcentration group and NH2higher concentration group were (18.863±4.101)%and(17.654±3.543)%(P<0.05), and there were significant differences between them.3. The percentage of Th17was tested by FCM, with results of HCQ lower concentration group and NH2lower concentration group of SLE patients at24h as (0.81±0.13)%and(0.76±0.26)%, as compared with the control, both had statistical difference (P<0.05).4. Levels of IL-17in the supernatant of HCQ group and NH2group of SLE patients weretested by ELISA, and there were significant differences (P<0.05).Conclusions1. HCQ could significantly improve the condition of SLE patients, but also induced thedecrease of WBCs, especially lymphocytes, to some degree.2. Effect of inducing PBMC apoptosis and decreasing Th17might be the possiblemechanism of HCQ in treating SLE.3. Secretion of cytokine IL-17in peripheral blood was decreased in HCQ treating SLE. |
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