人组织激肽释放酶基因转移对血管平滑肌细胞迁移的影响

Effects of human tissue kallikrein gene transfer on the migration of vascular smooth muscule cells

目的探讨重组腺病毒介导的人激肽释放酶(hKLK1)基因转移对血小板源性生长因子-BB(PDGF-BB)诱导下的自发性高血压大鼠(SHR)的血管平滑肌细胞(VSMCSHR)迁移的影响。方法自行构建携带目的基因hKLK1和强绿色荧光蛋白(EGFP)基因的双顺反子重组腺病毒载体。采用组织贴块法培养SHR胸主动脉血管平滑肌细胞,接种第3～6代VSMCSHR于6孔板,分别加入含hKLK1重组腺病毒和对照病毒干预,3d后加入PDGF-BB诱导24h,采用改良Boyden微孔膜双槽法测定VSMCSHR迁移,并加入缓激肽受体B2特异性阻断剂Hoe140。RT-PCR法测定缓激肽B1受体、B2受体的mRNA表达。结果hKLK1基因转移可明显抑制PDGF-BB诱导的VSMCSHR细胞迁移,抑制率为34.6%,且Hoe140干预不产生影响。PDGF-BB诱导的VSMCSHR缓激肽B2受体mRNA表达明显增加(P<0.001),而Hoe140可明显降低其表达(P<0.01)。结论重组腺病毒介导的hKLK1基因转移可抑制PDGF-BB诱导的VSMCSHR迁移,这一效应可能不通过B2受体途径介导。

Objective To investigate the effects of adenovirus-mediated human tissue kallikrein （Ad-hKLK1） gene transfer on platelet-derived growth factor-BB （PDGF-BB）-induced migration of vascular smooth muscle cells from spontaneously hypertensive rats （VSMCSHR）.Methods A bicistronic recombinant adenovirus vector （Ad-hKLK1） carrying the target hKLK1 gene and the reporter gene EGFP was constructed.VSMCs isolated from the thoracic aorta of male SHR were passaged,and the quiescent VSMCSHR in passages 3-6 seeded in 6-well plates were treated with Ad-hKLK1 and control virus.Human PDGF-BB or icatibant Hoe140,a BK B2 antagonistat,was used as the chemoattractant and placed in the bottom chamber of the Boyden chamber.The mRNA expressions of bradykinin B1 receptor and B2 receptor were detected by RT-PCR in VSMCSHR.Results hKLK1 gene transfer significantly inhibited PDGF-BB-induced migration of VSMCSHR,with the peak inhibition rate of 34.6% （P0.001）.PDGF-BB significantly increased the mRNA expression of B2 receptor but not B1 receptor in VSMCSHR.Conclusion hKLK1 gene transfer can inhibit the migration of VSMCSHR induced by PDGF-BB,and the inhibitory effects may be not mediated by bradykinin B2 receptor.