糖耐康对转化生长因子-β1诱导的HK-2细胞分泌细胞外基质成分的影响

摘要：目的 观察糖耐康对转化生长因子-β1(TGF-β1)诱导的人肾小管上皮细胞(HK-2)对细胞外基质成分表达的影响,探讨糖耐康治疗肾间质纤维化的作用机制。方法 将HK-2 细胞用含10%胎牛血清的DMEM/F12(1∶1)培养基培养,并分为6组：空白对照组、TGF-β1诱导组(TGF-β110 ng/mL)、空白血清对照组(TGF-β110 ng/mL＋10%空白血清)、干预1组(TGF-β110 ng/mL＋5%糖耐康药物血清)、干预2组(TGF-β110 ng/mL＋10%糖耐康药物血清)、干预3组(TGF-β110 ng/mL＋20%糖耐康药物血清)。药物干预24 h,荧光定量PCR检测Ⅰ型胶原(ColⅠ)、Ⅲ型胶原(ColⅢ)和纤连蛋白(FN)的mRNA表达。结果 HK-2细胞经TGF-β1诱导后,ColⅠ、ColⅢ和FN的mRNA表达显著上升,与空白对照组比较,差异有统计学意义(P＜0.05)。经糖耐康药物血清干预后,其表达逐步下降,与TGF-β1诱导组比较,差异有统计学意义(P＜0.05)。而空白血清无此作用。结论 糖耐康在一定程度上能够抑制TGF-β1诱导的人肾小管上皮细胞ColⅠ、ColⅢ和FN的mRNA表达,减少细胞外基质成分的分泌,具有防治肾间质纤维化的作用。

关键词：糖耐康；转化生长因子-β1；HK-2细胞；Ⅰ型胶原；Ⅲ型胶原；纤连蛋白

DOI：10.3969/j.issn.1005-5304.2013.08.014

中图分类号：R285.5

文献标识码：A

文章编号：1005-5304(2013)08-0039-03Effect of Tangnaikang on Extracellular Matrix Components of Human Renal Tubular Epithelial Cell HK-2 Induced by TGF-β1 LI Qin1, YANG Li-xia2, ZHANG Bang-neng2, LIU Tong-hua3, WU Li-li3, SUN Wen3, Margetts Peter Joseph4 (1.Gansu School of TCM, Lanzhou 730050, China；2.Gansu Province Hospital of TCM, Gansu Province Academy of Chinese Medicine, Lanzhou 730050, China；3. Beijing University of Chinese Medicine, Beijing 100029, China；4.McMaster University, Hamilton Ontario T3309, Canada)

Abstract：Objective To explore the effect of Tangnaikang (TNK) on extracellular matrix expression of human tubular epithelial cell HK-2 induced by TGF-β1 and explore its mechanism on the renal fibrosis. Methods The HK-2 cells were cultured by DMEM/F12 (1∶1) with 10% fetal bovine serum and divided into control group, TGF-β1 group (TGF-β110 ng/mL), rat serum control group (TGF-β110 ng/mL +10% rat serum), TNK-containing rat serum therapy groups (TGF-β110 ng/mL+5% Tangnaikang, or +10% Tangnaikang, or +20% TNK). After 24 h of administration, the expression of ColⅠ, Col Ⅲ and FN mRNA were tested by fluorescence quantitative PCR assay. Results The expression of ColⅠ, Col Ⅲ and FN mRNA of HK-2 cultured with TGF-β1 were much higher than the control, and significantly decreased in HK-2 cultured with TGF-β1 plus Tangnaikang compared with only TGF-β1 (P＜0.05), but rat serum control had no effect. Conclusion TNK could inhibit the expression of ColⅠ, Col Ⅲ and FN mRNA of HK-2 cell induced by TGF-β1, and prevent the development of renal fibrosis to some extent.

Key words：Tangnaikang；TGF-β1；human renal tubular epithelial cell；ColⅠ；Col Ⅲ；FN基金项目：国家自然科学基金面上项目(30973909)；北京中医药大学创新团队项目(2011-CXTD-19)通讯作者：刘铜华,E-mail：thliu@tom.com

糖尿病肾病(DN)是糖尿病常见的慢性微血管并发症之一,其发病机制尚未明确。既往认为DN的早期病变部位在肾小球,近年来研究发现肾小管病变及肾间质纤维化与其关系更为密切[1]。肾间质纤维化是所有肾脏疾病发展到终末期的共同病理 in Alzheimercs disease[J]. Exp Geronto,2005,40(10)：774-783.[12] Pouzet B, Zhang WN, Feldon J, et al. Hippocampal lesioned rats are able to learn a spatial position using non-spatial strategies[J]. Behav Brain Res,2002,133(2)：279-291.[13] 高琳娜,贺晓丽,唐千淇.淫羊藿苷对快速老化小鼠SAMP10脑组织单胺类及氨基酸类神经递质的影响[J].中国临床药理学与治疗学,2012, 17(10)：1081-1085.(收稿日期：2013-03-18,编辑：华强)基础,其特征是细胞外基质成分的过度沉积。因此,抑制细胞外基质成分过表达是抗肾间质纤维化治疗DN的主要途径之一。本课题所选糖耐康是北京中医药大学刘铜华教授治疗糖尿病的临床经验方,既往研究表明,糖耐康的主要功效为调节糖尿病小鼠糖代谢、提高胰岛素敏感指数、减轻胰岛素抵抗、促进胰岛β细胞的修复和再生、调节脂代谢异常等[2]。我们研究发现,糖耐康能够抑制高糖诱导大鼠肾小球系膜细胞的增殖,并能降低糖尿病GK大鼠尿蛋白量,表明糖耐康对DN的发生和发展具有一定的治疗作用[3-4]。本实验通过观察糖耐康干预转化生长因子-β1(TGF-β1)诱导的人肾小管上皮细胞对细胞外基质成分表达的变化,进一步探讨其对肾间质纤维化的作用,为其防治DN肾间质纤维化提供依据。