目的 探讨移植脂肪间充质干细胞(adipose tissue derived stem cells,ADSCs)对MRL/lpr小鼠的治疗作用及对MRL/lpr小鼠脾脏Th17/Treg细胞平衡的影响。方法 15只12周龄的雌性MRL/lpr小鼠采用随机数字表法分为ADSCs治疗组(ADSCs组)、对照组(control组)和环磷酰胺治疗组(cyclophosphamide,CTX组),每组5只。ADSCs组经尾静脉注射ADSCs,每次注射的细胞数为1×10⁶/150 μL,每周1次,共注射8次;control组经尾静脉注射等体积磷酸盐缓冲液,每周1次,共注射8次;CTX组经尾静脉注射环磷酰胺,剂量为15 mg/kg(体重),每周1次,连用2次,休息2周后重复,共注射4次。留取治疗前和治疗后尿液检测24 h尿蛋白浓度。治疗8周后,处死MRL/Ipr小鼠,以流式细胞仪分析脾细胞Th17/Treg细胞比例变化。结果 (1)24 h尿蛋白浓度变化:治疗前,3组小鼠的24 h尿蛋白浓度差异无统计学意义( P>0.05);治疗4周后,ADSCs组和CTX组24 h尿蛋白浓度显著低于control组,差异有统计学意义[(5.02±1.61) g/L vs. (7.10±1.63) g/L、(4.90±0.71) g/L vs. (7.10±1.63) g/L, P<0.05],而且治疗时间越长,效果越明显,治疗8周后,ADSCs组和CTX组24 h尿蛋白浓度显著低于control组[(2.24±0.73) g/L vs. (10.36±1.64) g/L、(3.80±1.45) g/L vs. (10.36±1.64) g/L, P<0.01];ADSCs组和CTX组之间24 h尿蛋白浓度差异无统计学意义( P>0.05)。(2)脾脏Treg细胞占CD4⁺T细胞的百分率:ADSCs组和CTX组脾淋巴细胞中Treg细胞占CD4⁺T细胞的百分率高于control组,3组分别为13.62%±1.87%、14.14%±1.29%、0.71%±1.23%,但组间差异无统计学意义( P>0.05)。(3)脾脏Th17细胞占CD4⁺T细胞的百分率:ADSCs组和CTX组脾淋巴细胞中Th17细胞占CD4⁺T细胞的百分率显著低于control组,3组分别为1.43%±0.20%、1.63%±0.65%、6.37%±1.64%,差异有统计学意义( P<0.01)。结论 移植ADSCs能够减少MRL/lpr小鼠尿蛋白浓度,治疗时间越长,效果越显著。ADSCs能够降低MRL/lpr小鼠脾脏Th17细胞水平,提高Treg细胞水平,负向调节Th17/Treg细胞平衡,进而发挥抗炎和免疫调节的作用。

Objective: Preliminary study on therapeutic effects of adipose tissue derived stem cells (ADSCs) on MRL/lpr mice and the effect on imbalance of Th17/Treg.Methods: Fifteen 12-week-old MRL/lpr mice were randomly divided into 3 groups by using random number table, including ADSCs group, control group and cyclophosphamide (CTX) group, with 5 in each group. ADSCs group and control group were injected with 1×10⁶ADSCs or phosphate buffered solution (PBS) via tail vein respectively, once a week, a total of eight times. CTX group was injected CTX at a dose of 15 mg/kg body weight, once a week for 2 weeks, and then repeated after 2 weeks’ rest, a total of four times. The 24-hour proteinuria was measured before and after treatment. All the mice were sacrificed after treatment for 8 weeks. Th17 cells and Treg cells in splenic were examined by flow cytometry.Results: (1) The 24-hour proteinuria in the three groups had no significant difference before treatment ( P>0.05). After therapy for 4 weeks, the 24-hour proteinuria in the ADSCs and CTX groups was much lower than those in control group, and the difference was significant [(5.02±1.61) g/L vs. (7.10±1.63) g/L, (4.90±0.71) g/L vs. (7.10±1.63) g/L, P<0.05], and the longer the duration of treatment (8 weeks), the more obvious effect [(2.24±0.73) g/L vs. (10.36±1.64) g/L, (3.80±1.45) g/L vs. (10.36±1.64) g/L, P<0.01]. There was no significant difference in 24-hour proteinuria between ADSCs group and CTX group ( P>0.05). (2) Percentage of Treg cells /CD4⁺T cells in the spleen lymphocytes: The percentages in ADSCs and CTX groups were higher than that in control group. The levels were 13.62%±1.87%, 14.14%±1.29%, 10.71%±1.23%, respectively, but there was no significant difference ( P>0.05). (3) Percentage of Th17 cells /CD4⁺T cells in the spleen lymphocytes: The percentages in ADSCs and CTX groups were significantly lower than that in control group. The levels were 1.43%±0.20%, 1.63%±0.65%, 6.37%±1.64%, respectively, with statistical significance ( P<0.01).Conclusion: Transplantation of ADSCs can reduce the 24-hour proteinuria in MRL/lpr mice. To prolong the time of treatment, the effect is more significant. Transplantation of ADSCs can up-regulate Treg cells and down-regulate Th17 cells. ADSCs have the ability to regulate the immune balance of Th17/Treg in MRL/lpr mice, suggesting that ADSCs play the role of anti-inflammatory and immune regulation by regulating the Treg and Th17 cells.