自噬基因在系统性红斑狼疮患者T细胞中表达的检测

Detection of autophagy gene expression in T cells from patients with systemic lupus erythematosus

目的通过检测SLE患者T细胞中自噬基因微管相关蛋白1轻链3（LC3）、Atg5、Beelin-1、Atg7表达及T细胞中自噬发生率，探讨其与SLE发病的关系。方法选择67例SLE患者（SLE组）和31名健康体检者（健康对照组），应用实时定量PCR检测2组受试者T细胞中自噬基因LC3、Atg5、Beclin-1、Atg7表达水平，同时采用流式细胞术检测2组受试者T细胞中自噬发生情况，进一步分析自噬基因表达水平与自噬发生、SLE患者临床特点相关性。2组计量资料比较，采用t检验，计数资料比较采用矿检验。所有数据采用SPSS17．0软件分析。结果LC3mRNA和Atg7 mRNA在SLE患者T细胞中的表达低于健康对照组（t=2．623，3．162；P=0．010，0．002），而Beclin1 mRNA和Atg5mRNA表达在2组间差异则无统计学意义（P均〉0．05）。LC3-II蛋白表达水平（LC3—II／GAPDH比值）在SLE外周血T细胞中为（0．21±0．08），明低于健康对照组的（0．34±0．11）（t=1．846，P=0．047）。。T细胞自噬发生率在SLE患者为（3．7±1．9）％，低于健康对照组的（6．6±1．4）％，（t=4．132，P=0．000），而且发现自噬发生率与Atg7mRNA表达呈显著性相关（r=0．492，P=0．008）。T细胞中Atg7mRNA表达水平低（△CT值〉9．86）的SLE患者更容易出现肾脏受累（P=0．008）及抗dsDNA抗体阳性（P=0．018），且疾病活动度越高（P=0．035），而关节炎、血液系统及中枢神经系统损害发生率虽有增高趋势，但差异无统计学意义。结论SLE患者T细胞中LC3和Atg7 mRNA表达下凋，从而导致T细胞自噬下调，进而自噬形成障碍，致使不能有效清除衰老、变性的生物分子，提示自噬在SLE发病及病情进展中发挥重要作用。

Objective In order to explore the role of autophagy in the development of systemic lupus erythematosus （SLE）, we measmed the expression of autophagy related gene microtubule-associated protein 1 light chain 3 （LC3）, Atg5, Beclin1, Atg7 and the incidence of autophagy in T cells from patients with SLE. Methods The mRNA levels of LC3, Atg5, Beclinl, Atg7 in T cells from 67 SLE patients and 3l healthy individuals were detected by real-time quantitative polymerase chain reacton （qPCR） technique. Autophagy in T cells from 17 SLE patients and 11 healthy controls was also determined by flow cytometry （FACs）. The correlation of Atg7 mRNA expression with clincal features was then analyzed. The differences between the two groups were tested by t-test andx2 test, all data were analyzed by statistical and service solutions （SPSS） 17.0 software. Results The mRNA levels of LC3 and Atg7 （△CT value） in SLE patients were obviously down- regulated as compared to healthy populations （P=0.010, P=0.002）, paralleled with the decreased autophagy rate detected by flow cytometry in T cells of SLE patients [（3.7±1.9）% vs （6.6±1.4）%, t=4.132, P=0.000]. Also, the protein expression levels of LC3-II in T cells of SLE patients （LC3-II/GAPDH） was significantly lower than those in healthy controls （0.21±0.08 vs 0.34±0.11, t=1.846, P=0.047）. Moreover, Atg7 mRNA expression levels were found to be negatively correlated to autophagy rate （r=-0.492, P=0.008）. However, when comparing the clinical features of 24 SLE patients with decreased Atg7 mRNA expression （ACT value〉9.86） to 43 SLEpatients with normal or high Atg7 mRNA expression （ACT value〈9.86）, increasing trend of incidence of arthritis, blood involvement and CNS was noted in patients with decreased Atg7 mRNA expression. However, there was a significant difference between the two groups in the incidence of renal involvement and anti- dsDNA antibody and SLEDAI （P=0.008, P=0.018, P=0.035）. Conclusion The impaired autophagy resahed from down-regulated LC3 and Atg7 mRNA levels in T cells from SLE patients indicates that autophagy plays a role in mediating the occurrence and development of SLE, which might be through unable to clean harmful molecules effectively.