缓激肽对肾系膜细胞增殖与细胞外基质分泌的作用及有关机制的探讨

Effects of bradykinin on proliferation and extracellular matrix secretion in renal mesangial cells and associated mechanism

目的研究缓激肽（BK）对肾系膜细胞增殖与细胞外基质分泌的作用及探讨与血小板源生长因子BB（PDGF-BB）有关的ERK信号途径机制。方法BK单独或与PDGF-BB共同孵育系膜细胞后,以MTF法测细胞增殖情况;ELISA法测胶原（C01）Ⅰ、ColⅣ分泌;Western杂交测ERK蛋白表达。用BK受体特异阻断剂HOE-140、酪氨酸磷酸酶抑制剂（OV）和ERK途径阻断剂U0126预孵育,以BK和PDGF-BB共同刺激系膜细胞,Western杂交测ERK蛋白表达。结果BK（10-1000μg/L）可单独刺激系膜细胞增殖、ColⅠ和ColⅣ分泌及诱导ERK1/2磷酸化。20μg/L PDGF-BB预孵育也有类似作用,但可被BK呈剂量依赖性抑制。1μmol/L HOE-140和0.5 mmol/L OV能分别阻断BK对PDGF-BB-ERK1/2途径磷酸化的抑制作用,而U0126抑制了HOE-140和OV的作用。结论BK刺激系膜细胞增殖的作用主要是通过ERK途径介导的,与PDGF-BB共同作用时则呈拮抗作用,并且与ERK途径的受抑有关。缓激肽B2受体和酪氨酸磷酸酶参与了BK的双向调节作用。

Objective To investigate the effects of bradykinin （BK） on proliferation and collagen secretion in cultured renal mesangial cells （MCs） and to elucidate the mechanism associated with PDGF-BB and ERK signaling pathway. Methods Cultured MCs in vitro were stimulated by BK alone or following incubation of PDGF-BB. Proliferation of MCs was measured by MTI＇, secretion of collagen by ELISA and protein expression by Western blotting. After incubation with HOE-140 （BK receptor specific inhibitor）, OV （tyrosine phosphatase inhibitor） and U0126 （ERK pathway inhibitor）, MCs were co-stimulated by BK and PDGF-BB, then protein expression of ERK was examined by Western blotting. Results BK （10-1000 μg/L） alone increased the proliferation, secretion of Col Ⅰ , Col Ⅳ and induced phosphorylation of ERK1/2 in MCs. Similar results were found in MCs stimulated by PDGF-BB, but such effects were inhibited by BK in a dose-dependent manner. Meanwhile above inhibition of BK against PDGF-BB was blocked by 1 μmol/L HOE-140 and 0.5 mmol/L OV. In other hand, the effects of HOE-140 and OV were inhibited by U0126. Conclusions Stimulation of BK on MCs is mainly mediated through ERK signaling pathway. BK can inhibit the similar stimulation of PDGF-BB and possesses a double effect in which B2 receptor and tyrosine phosphatase may be involved.