Kaposi sarcoma-associated herpesvirus g protein-coupled receptor enhances endothelial cell survival in part by upregulation of bcl-2.
Kaposi sarcoma-associated herpesvirus g protein-coupled receptor enhances endothelial cell survival in part by upregulation of bcl-2.
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发表时间：
2013-03
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通讯作者：
Elizabeth R Abboud;B. Shelby;Magdalena Angelova;Anne B Nelson;Marybeth Ferris;Harris McFerrin;C. Morris;D. Sullivan
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作者：
Elizabeth R Abboud;B. Shelby;Magdalena Angelova;Anne B Nelson;Marybeth Ferris;Harris McFerrin;C. Morris;D. Sullivan
卡波西肉瘤相关疱疹病毒（KSHV）编码的G蛋白偶联受体（vGPCR）是一种组成型活性裂解相蛋白，与人白细胞介素-8受体具有显著同源性。vGPCR是诱导血管生成以及卡波西肉瘤（KS）病变特征性的梭形细胞增殖所必需且足够的。我们以前证明，Bcl-2，抗凋亡蛋白，上调KS病变。本研究的目的是确定vGPCR是否通过上调Bcl-2表达来增强内皮细胞存活，并阐明所涉及的信号通路。方法用表达vGPCR的逆转录病毒转导原代人脐静脉内皮细胞，然后进行血清饥饿。通过荧光激活细胞分选分析细胞活力和凋亡。实时定量逆转录聚合酶链反应和免疫印迹法测定Bcl-2的表达。磷脂酰肌醇3-激酶（PI 3 K）/Akt和雷帕霉素（mTOR）的哺乳动物靶标的特异性药理学抑制剂被用来阐明所涉及的信号通路。Bcl-2表达用小干扰RNA（siRNA）敲低。结果表达vGPCR的内皮细胞在血清饥饿后存活率增加，Bcl-2 mRNA和蛋白表达上调。vGPCR诱导的Bcl-2 mRNA和蛋白水平的增加依赖于PI 3 K信号传导，而不是mTOR。此外，Bcl-2的siRNA抑制导致所观察到的vGPCR介导的细胞存活优势的显著消除。结论：综上所述，结果表明Bcl-2是vGPCR诱导的内皮细胞存活的介质，并且是Akt在此过程中的下游效应物。
BACKGROUND Kaposi sarcoma-associated herpesvirus (KSHV) encoded G protein-coupled receptor (vGPCR) is a constitutively active lytic phase protein with significant homology to the human interleukin-8 receptor. vGPCR is necessary and sufficient to induce angiogenesis as well as the spindle cell proliferation characteristic of Kaposi sarcoma (KS) lesions. We previously demonstrated that Bcl-2, an antiapoptotic protein, is upregulated in KS lesions. The aim of this study was to determine if vGPCR enhances endothelial cell survival through upregulation of Bcl-2 expression and to elucidate the signaling pathways involved. METHODS Primary human umbilical vein endothelial cells were transduced with a recombinant retrovirus expressing vGPCR and then subjected to serum starvation. Cell viability and apoptosis were analyzed by fluorescence-activated cell sorting. Bcl-2 expression was determined by real-time quantitative reverse transcription polymerase chain reaction and immunoblotting. Specific pharmacological inhibitors of phosphatidylinositol 3-kinase (PI3K)/Akt and the mammalian target of rapamycin (mTOR) were employed to elucidate the signaling pathways involved. Bcl-2 expression was knocked down using small interfering RNA (siRNA). RESULTS Endothelial cells expressing vGPCR showed increased survival after serum starvation and upregulation of Bcl-2 messenger RNA (mRNA) and protein. The vGPCR-induced increases in both Bcl-2 mRNA and protein levels were dependent on PI3K signaling but not on mTOR. Moreover, siRNA inhibition of Bcl-2 resulted in significant abrogation of the observed vGPCR-mediated cell survival advantage. CONCLUSIONS Taken together, the results demonstrate that Bcl-2 is a mediator of vGPCR-induced endothelial cell survival and is a downstream effector of Akt in this process.