慢病毒介导的过表达NR4A1单核细胞缓解狼疮小鼠狼疮性肾炎的研究

Lentivirus-mediated overexpressed NR4A1 monocytes in the treatment of lupus nephritis in lupus mice

摘要:

目的： 探索过表达NR4A1是否能够缓解狼疮性肾炎（lupus nephritis，LN）的肾炎症状。方法： 通过腹腔注射0.5 mL pristane构建狼疮小鼠模型，分为生理盐水对照（SC）组、pristane对照（PC）组、空载体对照（VC）组及过表达NR4A1治疗（NR4A1）组。第20周尾静脉注射过表达NR4A1的脾脏单核细胞进行治疗。第28周测量小鼠踝关节左右直径。HE染色及半定量评分评估肾脏病理改变，ELISA检测血清肿瘤坏死因子-α（tumor necrosis factor α，TNF-α）及抗ds-DNA浓度。荧光定量qPCR检测脾脏NR4A1、Bcl-2基因mRNA表达。结果： 与SC组相比，PC组LN相关活动指标明显升高，踝关节直径[4.69（4.26，5.14）mm，P=0.000]、抗ds-DNA抗体浓度[（173.11±25.49）ng/mL，P=0.000]、肾脏病理评分[2.78（2.68，2.81），P=0.000]和NR4A1表达（0.40±0.09，P=0.008）明显降低。与PC组、VC组相比，NR4A1组LN活动指标明显改善，踝关节直径[3.80（3.44，4.20）mm，P_PC=0.002，P_VC=0.033]、抗ds-DNA抗体浓度[（51.92±10.91）ng/mL，P_PC=0.008，P_VC=0.025]、肾脏病理评分[1.39（1.30，1.49），P_PC=0.002，P_VC=0.018]。ELISA结果显示NR4A1组TNF-α浓度[（542.86±108.28）pg/mL]明显低于PC组[（1 779.00±358.40）pg/mL，P=0.008]、VC组[（1 759.71±280.27）pg/mL，P=0.010]。Real-time PCR结果显示NR4A1组Bcl-2基因表达（1.92±0.58）明显高于PC组（0.48±0.20，P=0.000）。结论： 过表达NR4A1缓解LN小鼠LN症状，降低肾炎活动指标，可能涉及NR4A1、TNF-α及Bcl-2之间的相互作用。

Abstract:

Objective: To explore whether overexpression of NR4A1 can alleviate lupus nephritis (LN). Methods: A mouse model of LN was constructed by intraperitoneal injection of 0.5 mL pristane, divided into saline control group (SC), pristane control group (PC), vector control group (VC) and overexpressed NR4A1 group. At the end of 20^thweek, the monocytes overexpressed NR4A1 were injected into the tail vein. The diameter of ankles was measured at the end of 28^thweek. Renal pathological sections were stained with HE staining and semi-quantitative scores were used to assess renal pathological damage. Serum TNF-α and anti-ds-DNA concentrations were measured by ELISA. The expression of Bcl-2 and NR4A1 in the spleen was detected by real-time PCR. Results: The results showed that compared with the SC group, LN-related activity indicators in the PC group were significantly increased, but the ankle joint diameter[4.69 (4.26, 5.13) mm, P=0.000], the anti-ds-DNA antibody concentration [(173.11±25.49) ng/mL, P=0.000], renal pathology score[2.78 (2.68, 2.81), P=0.000], and the expression of NR4A1 (0.40±0.09, P=0.008) was significantly reduced. Compared with the PC and VC groups, the NR4A1 group LN activity indicators were significantly improved, the ankle joint diameter[3.80 (3.44, 4.20) mm, P_PC=0.002, P_VC=0.033], the concentration of anti-ds-DNA antibody [(51.92±10.91) ng/mL, P_PC=0.008, P_VC=0.025], renal pathology score[1.39 (1.30, 1.49), P_PC=0.002, P_VC=0.018].ELISA results showed that the concentration of TNF-α in the NR4A1 group [(542.86±108.28) pg/mL]was significantly lower than that of the PC group [(1 779.00±358.40) pg/mL, P=0.008]and the VC group [(1 759.71±280.27) pg/mL, P=0.010].Real-time PCR results showed that the expression of Bcl-2 gene in the NR4A1 group (1.92±0.58) was significantly higher than that of the PC group (0.48±0.20, P=0.000). Conclusion: Overexpression of NR4A1 alleviates the symptoms of LN and reduces LN activity indicators in LN mice, which may involve the interaction among NR4A1, TNF-α and Bcl-2.