microRNA－21反义寡核苷酸对肝星状细胞胶原合成的影响

Effect of microRNA-21 antisense oligonucleotide on collagen synthesis in the rat hepatic stellate cells

目的：探讨microRNA－21（ miR－21）反义寡核苷酸对肝星状细胞胶原合成的影响。方法肝星状细胞分离和培养，利用脂质体2000将miR－21反义寡核苷酸转染活化的肝星状细胞中，转染48 h后，收集肝星状细胞，采用实时RT－PCR法和Western blotting分别检测细胞miR－21表达和Ⅰ、Ⅲ型胶原蛋白表达；MTT法检测转染后细胞增殖活性。结果与无关反义寡核苷酸组相比，miR－21反义寡核苷酸转染后miR－21表达较无关对照组下调约76％（ P ＜0．01）；细胞增殖活性降低（26±3）％（ P ＜0．01）；Ⅰ、Ⅲ型胶原蛋白分别降低（61±7）％和（48±6）％（均P ＜0．01）。结论 miR－21反义寡核苷酸能显著下调miR－21的表达，抑制肝星状细胞增殖及胶原的合成。

Objective To investigate the effect of microRNA-21 （miR-21） antisense oligonucleoti-de on collagen synthesis in the rat hepatic satellite cells （ HSCs） .Methods Rat hepatic stellate cells were isolated and cultured; the miR-21 antisense oligonucleotide was transfected into HSCs with lipofectamine 2000;after incubation 48 h, the HSCs were collected.The expression of miR-21 was detected with reverse transcription polymerase chain reaction （ RT-PCR） , andα-smooth muscle actin （α-SMA） protein in HSCs with Western blot.The cell proliferation was assayed with methyl thiazolyl tetrazolium （ MTT） method.Re-sults Compared to scrambled control group, the expression of miR-21 was reduced by 76%（ P 〈0.01）, the proliferation activity of HSCs was reduced by（26 ±3）%（ P 〈0.01）, the expressions of type I and III collagen proteins were reduced by（61 ±7）%and （48 ±6）%（ P 〈0.01）.Conclusions The miR-21 an-tisense oligonucleotide could reduce significantly the expression of miR-21, and inhibit HSC proliferation and extracellular matrix production.