HHV6感染激活卡波济肉瘤相关疱疹病毒(KSHV)的溶解性周期复制

运用细胞融合、细胞混合培养、条件培养基培养和病毒直接刺激等方法,研究人类疱疹病毒6型(HHV6)对卡波济肉瘤相关疱疹病毒(KSHV)溶解性周期复制的影响。①将HHV6感染的JJhan细胞(T淋巴细胞系)与BCBL-1细胞(原发性渗出性淋巴瘤,PEL)进行细胞融合形成异核体细胞。②将HHV6感染的JJhan细胞与BcBL-1细胞进行混合培养。③收集HHV6感染的JJhan细胞培养上清液作为条件培养基进行灭活处理,以灭活前后的条件培养基培养BcBL-1细胞。进一步离心纯化HHV6病毒颗粒,并感染BCBL-1细胞,分别设紫外线和热灭活的HHV6病毒颗粒感染BCBL-1细胞为对照。提取上述的实验细胞总RNA,RT-PCR和／或实时定量(Real-time)PCR检测卡波济肉瘤相关疱疹病毒(KSHV)次要衣壳蛋白编码基因ORF26 mRNA转录。结果显示：①细胞融合后15h开始出现明显细胞病变,RT-PCR检测不同时间的实验组ORF26 mRNA转录水平均明显高于对照组;Real-time PCR检测各时间ORF26 mRNA转录水平是对照组的2．3倍以上;②细胞混合培养72h时,实验组ORF26 mRNA转录水平是对照组的1．8倍;混合培养5天时,实验组KSHV裂解周期蛋白K8．1表达水平是对照组的2．46倍;③灭活前后的HHV6感染细胞培养上清液培养BCBL-1细胞96h时,ORF26 mRNA转录水平分别是对照组的2．73倍和2．22倍;④灭活前后的HHV6均可增强BCBL-1细胞中KStHV ORF26 mRNA转录水平。提示：KHV6感染可激活KSHV的溶解性周期复制。     

Propranolol Decreases Proliferation of Endothelial Cells Transformed by Kaposi's Sarcoma-Associated Herpesvirus and Induces Lytic Viral Gene Expression

Kaposi''s sarcoma (KS) is common in Africa, but economic constraints hinder successful treatment in most patients. Propranolol, a generic β-adrenergic antagonist, decreased proliferation of KS-associated herpesvirus (KSHV)-infected cells. Downregulation of cyclin A2 and cyclin-dependent kinase 1 (CDK1) recapitulated this phenotype. Additionally, propranolol induced lytic gene expression in association with downregulation of CDK6. Thus, propranolol has diverse effects on KSHV-infected cells, and this generic drug has potential as a therapeutic agent for KS.     

The Kaposi's Sarcoma-Associated Herpesvirus (KSHV)-Induced 5-Lipoxygenase-Leukotriene B4 Cascade Plays Key Roles in KSHV Latency,Monocyte Recruitment,and Lipogenesis

Kaposi''s sarcoma-associated herpesvirus (KSHV) is etiologically associated with Kaposi''s sarcoma (KS) and primary effusion lymphoma (PEL). KS lesions are characterized by endothelial cells with multiple copies of the latent KSHV episomal genome, lytic replication in a low percentage of infiltrating monocytes, and inflammatory cytokines plus growth factors. We demonstrated that KSHV utilizes inflammatory cyclooxygenase 2/prostaglandin E₂ to establish and maintain latency (Sharma-Walia, N., A. G. Paul, V. Bottero, S. Sadagopan, M. V. Veettil, N. Kerur, and B. Chandran, PLoS Pathog 6:e1000777, 2010 [doi:10.1371/journal.ppat.1000777]). Here, we evaluated the role of 5-lipoxygenase (5LO) and its chemotactic metabolite leukotriene B4 (LTB4) in KSHV biology. Abundant staining of 5LO was detected in human KS tissue sections. We observed elevated levels of 5LO and high levels of secretion of LTB4 during primary KSHV infection of endothelial cells and in PEL B cells (BCBL-1 and BC-3 cells). Blocking the 5LO/LTB4 cascade inhibited viral latent ORF73, immunomodulatory K5, viral macrophage inflammatory protein 1 (MIP-1), and viral MIP-2 gene expression, without much effect on lytic switch ORF50, immediate early lytic K8, and viral interferon-regulatory factor 2 gene expression. 5LO inhibition significantly downregulated latent viral Cyclin and latency-associated nuclear antigen 2 levels in PEL cells. 5LO/LTB4 inhibition downregulated TH2-related cytokine secretion, elevated TH1-related cytokine secretion, and reduced human monocyte recruitment, adhesion, and transendothelial migration. 5LO/LTB4 inhibition reduced fatty acid synthase (FASN) promoter activity and its expression. Since FASN, a key enzyme required in lipogenesis, is important in KSHV latency, these findings collectively suggest that 5LO/LTB4 play important roles in KSHV biology and that effective inhibition of the 5LO/LTB4 pathway could potentially be used in treatment to control KS/PEL.