不同肾功能进展IgA肾病患者血清IgA1对人肾小球系膜细胞增殖及TGF-β1表达的影响

Effect of serum IgA1 on human mesangial cells and expression of TGF-β1 in patients of IgA nephropathy with different renal function progress

目的·探讨肾功能快速进展为终末期肾病(end stage renal disease,ESRD)的IgA肾病患者与肾功能长期稳定的IgA肾病患者血清IgA1对体外培养的正常人肾小球系膜细胞(human mesangial cells,HMCs)增殖的刺激作用及对转化生长因子β1(transforming growth factor-β1,TGF-β1)表达水平的影响。方法·9例经肾活检确诊为原发性IgA肾病的患者,根据病情进展情况分为肾功能快速进展组(n=6)和肾功能长期稳定组(n=3)。采用亲和层析及分子筛提取血清IgA1;用不同质量浓度(10、50、250、1 000μg/m L)的IgA1分别干预HMC 12、24 h。CCK8法测定细胞增殖率;real-time PCR测定TGF-β1 m RNA表达;ELISA法测定TGF-β1蛋白表达。结果·肾功能快速进展组及肾功能长期稳定组患者血清IgA1对HMC有明显的促增殖作用,且表现为时间依赖性和浓度依赖性,作用高峰质量浓度分别为250μg/m L和1 000μg/m L。2组患者血清IgA1均能显著上调TGF-β1的m RNA和蛋白的表达。在肾功能快速进展组,IgA1质量浓度为10μg/m L时TGF-β1的m RNA及蛋白水平达到高峰,其后随刺激浓度升高而递减;在肾功能长期稳定组,TGF-β1水平则与血清IgA1呈现平行的浓度依赖性。结论·肾功能快速进展及长期稳定的患者血清IgA1在一定浓度和时间范围内均具有刺激HMC增殖的作用。在较低质量浓度(10～50μg/m L)范围内,促增殖作用没有明显差异,但肾功能快速进展的患者血清IgA1上调TGF-β1表达的作用更强,提示血清IgA1具有一定的预后评估作用。

Objective·To extract serum IgA1 from patients with IgA nephropathy(IgAN)(end stage renal disease vs long-term stable renal function)to explore the effect on proliferation rate of human mesangial cells(HMCs)and the level of transforming growth factor-β1(TGF-β1).Methods·Nine patients with primary IgAN were divided into rapidly progressive group(n=6)and long-term stable group(n=3).Jacalin affinity chromatography and sephacryl S-200HR molecular sieves were used to distract serum IgA1.HMCs were cultured and co-cultivated with different IgA1 concentration(10,50,250 and 1 000μg/mL)at point of 12 h and 24 h respectively.The proliferation rate was measured by cell counting kit-8(CCK8).The expression of TGF-β1 mRNA was measured via quantitative real-time PCR(qRT-PCR).Enzyme-linked immunosorbent assay(ELISA)was used to detect TGF-β1 protein.Results·Serum IgA1 from IgAN patients with different renal functions(end stage renal disease vs long-term stable renal function)activated proliferation of HMC significantly,presenting with time-dependence and concentration-dependence.The highest value showed at 250μg/mL and 1 000μg/mL respectively.Serum IgA1 in two groups of patients statistically increased the expression of TGF-β1 mRNA and protein.In group with end stage renal disease,the summit stood at 10μg/mL and started to decrease by degrees afterwards;while in group with long-term stable renal function,the level of TGF-β1 increased in a parallel manner with the serum IgA1.Conclusion·Serum IgA1 from IgAN patients with different renal functions(end stage renal disease vs long-term stable renal function)can both promote the proliferation of HMC.There is no dramatical difference observed with in10-50μg/mL,but the IgA1 from group with end stage renal disease reveals a stronger effect on TGF-β1,in accordance with the pathological type of the patients(IgA sclerosis),suggesting the prognostic value of serum IgA.