朱苗蕊　權(quán)卓　楊麗霞　程濤　張定華　高漢媛　孫文

摘要：目的 觀察新加糖腎康對(duì)高糖環(huán)境培養(yǎng)下人腎小管上皮細(xì)胞（HK-2）α-平滑肌肌動(dòng)蛋白（α-SMA）和E-鈣黏蛋白（E-cadherin）的影響，探討其防治糖尿病腎間質(zhì)纖維化的作用機(jī)制。方法 含10%胎牛血清1640培養(yǎng)基體外培養(yǎng)HK-2細(xì)胞。實(shí)驗(yàn)分為空白對(duì)照組、高糖組、空白血清組和新加糖腎康低、中、高劑量組。藥物干預(yù)后，MTT法檢測(cè)細(xì)胞增殖，ELISA檢測(cè)α-SMA和E-cadherin的含量。結(jié)果 與空白對(duì)照組比較，HK-2細(xì)胞經(jīng)高糖培養(yǎng)后細(xì)胞數(shù)量和α-SMA含量顯著增加、E-cadherin含量明顯降低（P<0.05）；與高糖組比較，新加糖腎康組α-SMA含量降低、E-cadherin含量增加、細(xì)胞增殖受到抑制。結(jié)論 新加糖腎康能夠抑制腎小管上皮細(xì)胞表型轉(zhuǎn)化及細(xì)胞增殖，具有防治糖尿病腎間質(zhì)纖維化的作用。

關(guān)鍵詞：新加糖腎康；高糖；人腎小管上皮細(xì)胞；表型轉(zhuǎn)化；細(xì)胞增殖

DOI：10.3969/j.issn.1005-5304.2016.03.015

中圖分類(lèi)號(hào)：R285.5 文獻(xiàn)標(biāo)識(shí)碼：A 文章編號(hào)：1005-5304（2016）03-0054-04

Abstract： Objective To observe he effects of new Tangshenkang on α-SMA and E-cadherin of human renal tubular epithelial cell HK-2 in high concentrations of glucose； To explore the mechanism of new Tangshenkang on the prevention and treatment of diabetic renal fibrosis. Methods The HK-2 cells were cultured and divided into control group， high glucose group， animal serum control group， new Tangshenkang low-， medium-， and high-dosage group. After medicine intervention， cell proliferation was tested by MTT assay， and contents of α-SMA and E-cadherin were observed by ELISA assay. Results Compared with control group， α-SMA of HK-2 cultured with high glucose was much notable， but the content of E-cadherin significantly decreased， with statistical significance （P<0.05）. The content of α-SMA of HK-2 cultured with new Tangshenkang decreased， and the content of E-cadherin increased； cell proliferation was markedly inhibited in culture medium supernatant of HK-2 cells cultured with high glucose plus new Tangshenkang compared with only high glucose， with statistical significance. Conclusion New Tangshenkang can inhibit cell proliferation and epithelial-myofibroblast transdifferentiation of HK-2 cell induced by high glucose， and prevent the development of diabetic renal fibrosis to a certain extent.

Key words： new Tangshenkang； high glucose； human renal tubular epithelial cell HK-2； epithelial-myofibroblast transdifferentiation； cell proliferation

糖尿病腎病（DN）是糖尿病常見(jiàn)的慢性微血管并發(fā)癥之一，其發(fā)病率高、危害大，在糖尿病所有并發(fā)癥中占主要地位。但其發(fā)病機(jī)制尚未完全闡明。既往研究認(rèn)為，早期DN病變?cè)谀I小球部位，而近年來(lái)研究發(fā)現(xiàn)，腎小管上皮細(xì)胞表型轉(zhuǎn)化引起的腎間質(zhì)纖維化與其關(guān)系更為密切[1]。中藥復(fù)方新加糖腎康是在甘肅省中醫(yī)院劉國(guó)安和張定華研究的基礎(chǔ)上，結(jié)合北京中醫(yī)藥大學(xué)劉銅華教授治療DN的學(xué)術(shù)思想，經(jīng)組方優(yōu)化，最終確立的臨床經(jīng)驗(yàn)方，具有益氣養(yǎng)陰、清熱祛濕、活血化瘀的功效。本課題組前期研究發(fā)現(xiàn)，新加糖腎康能夠減少高糖環(huán)境培養(yǎng)下人腎小管上皮細(xì)胞細(xì)胞外基質(zhì)成分Ⅰ型膠原（ColⅠ）、Ⅲ型膠原（Col Ⅲ）和纖連蛋白（FN）的分泌與沉積，并能調(diào)控纖維化因子基質(zhì)金屬蛋白酶-9（MMP-9）及其抑制劑-1（TIMP-1）的分泌與釋放[2-3]。本實(shí)驗(yàn)在此基礎(chǔ)上，觀察其對(duì)高糖環(huán)境下培養(yǎng)人腎小管上皮細(xì)胞標(biāo)志蛋白α-平滑肌肌動(dòng)蛋白（α-SMA）和E-鈣黏蛋白（E-cadherin）的影響，進(jìn)一步探討其對(duì)DN腎間質(zhì)纖維化的作用機(jī)制。

1 實(shí)驗(yàn)材料

1.1 動(dòng)物與細(xì)胞株

SPF級(jí)雄性Wistar大鼠20只，甘肅中醫(yī)藥大學(xué)動(dòng)物中心，許可證號(hào)SCXK（甘）2004-0006，飼養(yǎng)于甘肅省中醫(yī)藥研究院中藥研究所動(dòng)物室。人腎小管上皮細(xì)胞（HK-2），上海斯信生物科技有限公司，甘肅省中醫(yī)藥研究院中心實(shí)驗(yàn)室細(xì)胞培養(yǎng)室培養(yǎng)。