The aim of the present study was to determine the effect of somatostatin (SRIF) on mitogen-induced regeneration of rat aortic vascular smooth muscle cells (VSMC) and for comparison Chinese hamster ovary (CHO)-K1 cells expressing human recombinant sst5 receptors (CHOsst5), following partial denudation of a confluent cell monolayer. Regeneration was assessed by measuring areas of recovery into the denuded area and by counting total cell numbers. 2. In VSMC, SRIF (0.1 nM - 1 microM) had no effect on the basal levels of regeneration but caused a concentration-dependent inhibition (pIC50 8.0-8.6) of the stimulated regeneration induced by submaximal concentrations of basic fibroblast growth factor (bFGF, 10 ng ml[-1]), platelet-derived growth factor-BB (PDGF, 5 ng ml[-1]) or endothelin-1 (ET-1, 100 nM). SRIF (pIC50 8.8) also inhibited bFGF-induced regeneration of CHOsst5 cells. 3. In VSMC, the inhibitory action of SRIF on the regeneration induced by bFGF (10 ng ml[-1]) was due to an anti-proliferative effect, rather than an effect on cell migration, as SRIF (0.1 nM - 1 microM) abolished bFGF-induced increases in total cell numbers. The bFGF-induced increase in cell numbers was also abolished by actinomycin D (0.1 microg ml[-1]). 4. The sst5 receptor-selective agonist, L-362,855 (pIC50 10.5), was about 100 times more potent than SRIF at inhibiting bFGF-induced regeneration of both VSMC and CHOsst5 cells whilst the sst2 receptor-selective agonist, BIM-23027 (pIC50 6.8), was approximately 20 times weaker than SRIF. 5. The sst5 receptor antagonist, BIM-23056 (100 nM), antagonized SRIF-induced inhibition of bFGF-induced regeneration in both VSMC and CHOsst5 cells (estimated pKB values 8.8 and 8.3, respectively). 6. SRIF-induced inhibition of bFGF-induced regeneration of VSMC and CHOsst5 cells was abolished by pretreating cells with pertussis toxin (100 ng ml[-1]) for 20 h. 7. These findings suggest that SRIF-induced inhibition of the proliferation of rat aortic VSMC is mediated via activation of receptors which are similar to human sst5 receptors. Furthermore this inhibitory effect is transduced via pertussis toxin-sensitive Gi/Go proteins.
摘要
本研究的目的是确定生长抑素（SRIF）对丝裂原诱导的大鼠主动脉血管平滑肌细胞（VSMC）再生的影响，并与表达人重组sst5受体的中国仓鼠卵巢（CHO）-K1细胞（CHOsst5）进行比较，实验采用汇合细胞单层部分剥脱的方法。通过测量裸露区域的恢复面积和计算总细胞数来评估再生情况。2. 在VSMC中，SRIF（0.1 nM - 1 microM）对基础再生水平无影响，但对亚最大浓度的碱性成纤维细胞生长因子（bFGF，10 ng ml[-1]）、血小板衍生生长因子-BB（PDGF，5 ng ml[-1]）或内皮素-1（ET-1，100 nM）诱导的刺激再生产生浓度依赖性抑制（pIC50 8.0 - 8.6）。SRIF（pIC50 8.8）也抑制bFGF诱导的CHOsst5细胞再生。3. 在VSMC中，SRIF对bFGF（10 ng ml[-1]）诱导的再生的抑制作用是由于抗增殖作用，而非对细胞迁移的影响，因为SRIF（0.1 nM - 1 microM）消除了bFGF诱导的总细胞数增加。放线菌素D（0.1 microg ml[-1]）也消除了bFGF诱导的细胞数增加。4. sst5受体选择性激动剂L-362,855（pIC50 10.5）在抑制bFGF诱导的VSMC和CHOsst5细胞再生方面比SRIF强约100倍，而sst2受体选择性激动剂BIM-23027（pIC50 6.8）比SRIF弱约20倍。5. sst5受体拮抗剂BIM-23056（100 nM）拮抗SRIF诱导的对bFGF诱导的VSMC和CHOsst5细胞再生的抑制作用（估计pKB值分别为8.8和8.3）。6. 用百日咳毒素（100 ng ml[-1]）预处理细胞20小时后可消除SRIF诱导的对bFGF诱导的VSMC和CHOsst5细胞再生的抑制作用。7. 这些发现表明，SRIF诱导的大鼠主动脉VSMC增殖抑制是通过与人sst5受体相似的受体激活介导的。此外，这种抑制作用是通过百日咳毒素敏感的Gi/Go蛋白转导的。
