高糖条件下基质细胞趋化因子1α抑制人肾小管上皮细胞Ⅳ型胶原表达

Stromal cell-derived factor-1α suppressese the expression of type Ⅳ collagen in human renal proximal tubular epithelial cells in high glucose

目的 本文探讨基质细胞趋化因子1α （stromal cell-derived factor-1,SDF-1α）对人肾小管上皮细胞（HK细胞）细胞外基质（Ⅳ型胶原）表达的影响.方法 不同浓度的葡萄糖培养HK细胞,分为正常葡萄糖组（NG组：含5.5 mmol/L葡萄糖）和高糖组（HG组：含25.0 mmol/L葡萄糖）,分别用转化生长因子β1 （TGF-β1）和SDF-1α干预HK细胞12h,再分为空白对照组（NC组）、阴性对照组（DMSO组）、T组（TGF-β1组）、S组（SDF-1α组）、T＋S组（TGF-β1和SDF-1α共同干预组）,采用实时定量聚合酶链反应（RT-PCR）和Western blotting分别检测Ⅳ型胶原mRNA和蛋白的表达.两组间比较采用t检验.结果 （1）HK细胞能持续表达SDF-1α和其受体CXCR4 mRNA.高糖条件下SDF-1α和CXCR4mRNA表达呈时间梯度增加,24 h两者表达量达到高峰.24 h CXCR4 mRNA表达量是12h表达量的1.2倍（t=6.08,P＜0.05）,SDF-1αmRNA表达量是12h的1.58倍（t=7.52,P＜0.05）.（2）高糖条件下SDF-1α抑制HK细胞Ⅳ型胶原mRNA和蛋白的表达,并能降低由TGF-β1诱导的Ⅳ型胶原表达.S＋T组Ⅳ型胶原mRNA表达量是T组的0.33倍（t=12.57,P＜0.05）,蛋白表达量是T组的0.47倍（t=14.23,P＜0.05）.但在正常葡萄糖条件下,SDF-1α对由TGF-β1诱导的Ⅳ型胶原表达没有影响.（3）TGF-β1促进HK细胞Ⅳ型胶原mRNA和蛋白表达,高糖条件下其促进作用更为显著.T组Ⅳ型胶原mRNA是正常对照组的6.63倍（t=19.21,P＜0.05）,蛋白表达量是其2.59倍（t=15.71,P＜0.05）.结论 高糖条件下SDF-1α能减轻TGF-β1诱导的HK细胞Ⅳ型胶原表达,可能由此改善肾脏纤维化.TGF-β1促进HK细胞Ⅳ型胶原的表达,促进肾脏纤维化的发生发展.

Objective To explore the effect of stromal cell-derived factor-1α （SDF-1α） on extracellular matrix （type Ⅳ collagen） in human renal proximal tubular epithelial cells （HKC） with the different glucose conditions.Methods HK cell were cultured exposed to SDF-1α and transform growth factor-β1（TGF-β1） of the different concentration with normal （NG group：5.5 mmol/L glocuse） or high （HG group：25.0 mmol/L glucose） glucose.With the stimulation of TGF-β1 and SDF-1α for 12 hours later,the HKC were divided to the following groups：control group（NC group）,S group （SDF-1α alone）,T group （TGF-β 1 alone）,S＋T group（SDF-1α and TGF-β1 together）.Real-time PCR and Western blotting were used to detect the m RNA and protein expression of type Ⅳ collagen in HKC.Comparison between two groups was using t test.Results （1）The m RNA expression of SDF-1α and CXCR4 mRNA was detected continuously in HKC.High glucose could increase the expression of SDF-1α and CXCR4 mRNA in HKC with a time-dependent manner,which reached the highest level at 24 h.The mRNA level of CXCR4 at 24 h was 1.2 times of those at 12 h（t-6.08,P＜0.05）,meanwhile the mRNA level of SDF-1α at 24 h was 1.58 times of those at 12 h（t=7.52,P＜0.05）.（2）With high glucose SDF-1α suppressed the mRNA and protein expression of type Ⅳ collagen induced by TGF-β1 in HKC.The mRNA and protein expression level of type Ⅳ collagen in S＋T group were 0.33 times （t=12.57,P＜0.05）,and 0.47 times （t=14.23,P＜0.05） of those in T group,respectively.Under the normal glucose condition,SDF-1α had no effect on the expression of type Ⅳ collagen induced by TGF-β1.（3）TGF-β1 stimulated mRNA and protein expression of type Ⅳ collagen in HKC,especially in high glucose.The mRNA and protein expression level of type Ⅳ collagen in T group were 6.63 times （t=19.21,P＜0.05）,and 2.59 times （t=15.71,P＜0.05） of those in control group respectively.Conclusions （1）SDF-1α maybe attenuate the renal fibrosis by suppressing the expression of type Ⅳ collagen induced by TGF-β1 in high glucose.（2）TGF-β1 promotes the development of DN by increasing the expression of type Ⅳ collagen in HKC.