PRS-CTGF-siRNA attenuates matrix production and the expression of vascular endothelial growth factors induced by transforming growth factor β1 in cultured human peritoneal mesothelial cells / 中华肾脏病杂志

ABSTRACT

Objective To investigate the effects of connective tissue growth factor (CTGF) siRNA delivered by pRetro-Super (PRS) retrovirus vector on extracellular matrix and VEGF expression in human peritoneal mesothelial cells (HPMC). Methods Four pairs of oligonucleotides including 64 bp DNA were designed and synthesized in vitro according to siRNA target sequence and PRS retrovirus desire.PRS-CTGF-siRNA1-4 recombinant retrovirus vectors were constructed.The recombinant retrovirus vectors containing CTGF-siRNA were transferred into PT67 packaging cell lines with lipefectamine 2000,then infected HPMC.mRNA expression was determined by semi-quantitative RT-PCR and protein expression was determined by Western blot.Results Both mRNA and protein expressions of CTGF,FN,Col I,laminin (LN) and VEGF were significantly increased in HPMC with 5 μg/L TGF-β1 stimulation (P＜0.01,respectively).CTGF,FN,Col I,LN mRNA and protein and VEGF mRNA expression stimulated by TGF-β1 were significantly decreased in HPMC infected with PRS-CTGF-siRNA1～4 retrovirus vectors (P＜0.01,respectively).The inhibitory rates on CTGF were 69.3%,22.2%,27.4% and 38.8%,respectively (P＜0.01).At the same time,there was also a significant reduction of VEGF protein expression in HPMC infected with PRS-CTGF-siRNA1 vector (P＜0.01).There was no significant difference in HPMC infected with PRS void vector. Conclusion CTGF siRNA delivered by PRS retrovirus vector can effectively inhibit the enhancement of extracellular matrix and VEGF expression stimulated by TGF-β1 in HPMC.