趋化因子受体在肝癌细胞中的表达及其意义

目的 观察趋化因子受体(CCR1 CCR7 CXCB3 CXCR4)在人肝癌细胞系(Hep3BHepG2 HLE和HLF)的表达,并探讨其作用机制.方法 采用逆转录-聚合酶链反应(RT-PCR)、细胞免疫组织化学、流式细胞仪测定趋化因子受体在肝癌细胞表面的表达,通过肝癌细胞Hep3B体外侵袭实验即重组大鼠巨噬细胞激动蛋白(CCL3)-1对Hep3B细胞穿透人工重组基底膜能力的影响来检测趋化因子受体在肝癌侵袭转移时所起的作用,并通过激光共聚焦显微镜观察趋化因子受体在肝癌细胞侵袭转移过程中的作用机制.结果 通过RT-PCR,流式细胞术可检测到CCR1 mR-NA及蛋白在肝癌细胞中表达,免疫组织化学显示CCR1在肝癌细胞包膜和胞质内都有表达.细胞侵袭实验提示Hep3B在CCL3的刺激下,实验组Hep-3B细胞通过基底膜的数量(213±12)多于对照组细胞通过基底膜的数量(102±11),差异有统计学意义(P<0.01).运用激光共聚焦检测到实验组胞内钙离子的浓度增加到.结论 肝癌细胞系(Hep3B HepG2 HLE和HLF)的表面的表达CCRl,且CCR1在肝癌细胞侵袭转移中发挥重要的作用,其作用机制与细胞内钙离子浓度有很关.     

Yellow fever virus strains Asibi and 17D-204 infect human umbilical cord endothelial cells and induce novel changes in gene expression

Yellow fever (YF) is a zoonotic infection with more than 200,000 cases reported annually. Relatively little is known about YF pathogenesis in humans. In this study, we demonstrate that human vascular endothelial cells are susceptible to infection with wild-type and vaccine strains of the YFV and that these infections lead to a differential cellular response to infection. The infection of endothelial cells with either virus resulted in a significant induction of interferon-inducible genes p 78 and Cig 5 while wild-type virus induced a much more pronounced IL 6 and Bc l2 response than did the vaccine strain. Both viruses induced RANTES gene expression, but only the wild-type virus had corresponding increases in RANTES protein expression. The results demonstrate that the wild-type and vaccine strains of YFV elicit significantly different responses to infection in endothelial cells, despite being nearly identical genetically. These differences may account for the attenuated phenotype of the YFV vaccine strain, though the mechanism remains unclear. These data also point to a role for vascular endothelial cells in YF hemorrhagic fever and also suggest that IL 6 may play a role in increased viral pathogenesis, perhaps by influencing coagulation via release of coagulation co-factors such as fibrin or fibrinogen.     

HIV-1协同受体CXCR4、CCR5及相关趋化因子SDF-1在胎盘的表达

目的：研究HIV-1协同受体CXCR4、CCR5及CXCR4的特异性配体SDF-1在人胎盘组织的表达，探索HIV-1子宫内垂直传播的分子机制。方法：半定量RT-PCR检测早、中、晚孕期胎盘及早孕滋养细胞CXCR4、CCR5 mRNA水平；免疫组化和免疫细胞化学检测早孕胎盘及原代培养滋养细胞CXCR4、CCR5蛋白表达；原位杂交及免疫组化分析SDF-1在早孕胎盘的表达；ELISA测定滋养细胞SDF-1的动态分泌水平。结果：各孕期胎盘表达CXCR4及CCR5 mRNA；CXCR4蛋白定位于滋养细胞，而CCR5蛋白定位于绒毛基质中。滋养细胞可转录并翻译SDF-1，且能分泌可溶性SDF-1。结论：滋养细胞同时表达CXCR4及SDF-1，SDF-1可能通过降调CXCR4而拮抗X4-HIV-1感染胎儿细胞；R5-HIV-1或许能通过滋养层裂隙感染CCR5^#基质细胞和/或Hotbauer细胞，从而发生子宫内垂直传播。     

Outer membrane protein A renders dendritic cells and macrophages responsive to CCL21 and triggers dendritic cell migration to secondary lymphoid organs

Outer membrane protein A (OmpA) is a class of bacterial cell wall protein that is immunogenic without adjuvant. As specific immune responses are initiated in the lymph nodes (LN, we analyzed the effect of the OmpA from Klebsiella pneumoniae (KpOmpA) onchemokine/ chemokine receptor expression by APC and on cell migration to the LN. Upon contact with KpOmpA, human immature DC and macrophages acquire CCR7 expression and responsiveness to CCL21. In parallel, CCR1 and CCR5 expression is down-regulated and CXCL8, CCL2, CCL3 and CCL5 production is up-regulated. Mice injected subcutaneously with KpOmpA present a transient inflammatory reaction at the site of injection accompanied by an enlargement of the draining LN with a higher proportion of DC and macrophages. Lastly, when exposed to KpOmpA prior injection, DC but not macrophages migrate to the draining LN. In conclusion, KpOmpA confers a migratory phenotype to DC and triggers their migration to the regional LN. This property contributes to explain how innate cells initiate adaptive immune response upon recognition of conserved bacterial components and also why OmpA is immunogenic in the absence of adjuvant.     

趋化性细胞因子受体在Tc1和Tc2亚群的差异表达

目的研究趋化性细胞因子受体在体外长期培养的人Te1和Tc2细胞系中的表达。方法将新鲜分离的PBMC在特定细胞因子及细胞因子抗体存在条件下，定向诱导为Ⅰ型和Ⅱ型T细胞系，用免疫荧光染色结合流式细胞术分析鉴定，并以膜表面及胞内三色流式细胞术检测趋化性细胞因子受体在不同T细胞亚群的表达。结果①在Ⅰ型和Ⅱ型T细胞整体水平上，CXCR4的表达水平接近，CXCR3和CCR5的表达在Ⅰ型T细胞明显高于Ⅱ型T细胞，CCR3在2种T细胞系的表达水平均较低，但在Ⅱ型T细胞的表达略高于Ⅰ型T细胞；②趋化性细胞因子受体在Tc1和Tc2亚群细胞的表达与上述结果类似，即CXCR的表达水平接近，CXCR3和CCR5的表达在Tc1高于Tc2，CCR3的表达水平较低，但在Tc2则略高于Tc1。结论趋化性细胞因子受体在Tc1和Tc2亚群的表达具有差异性。     

Antitumor immune response by CX3CL1 fractalkine gene transfer depends on both NK and T cells

The CX3C chemokine fractalkine (CX3CL1) exists as both a membrane-bound form promoting firm cell-cell adhesion and a soluble form chemoattracting leukocytes expressing its receptor CX3CR1. When adenoviral vector expressing mouse fractalkine (AdFKN) was transduced to the tumor cells, fractalkine was expressed as both membrane-bound form on the tumor cells and soluble form in the supernatant in vitro. Intratumoral injection of AdFKN (1 x 10(9)PFU/tumor) into C26 and B16F10 tumors resulted in marked reduction of tumor growth compared to control (C26: 86.5%, p<0.001; B16F10: 85.5%, p<0.001). Histological examination of tumor tissues revealed abundant infiltration of NK cells, dendritic cells, and CD8(+) T lymphocytes 3 and/or 6 days after treatment with AdFKN. Splenocytes from mice treated by AdFKN developed tumor-specific cytotoxic T cells, and thereby protected from rechallenging with parental tumor cells. Antitumor effects by AdFKN were completely abrogated in both NK cell-depleted mice and CD8(-/-) mice, and partially blocked in CD4(-/-) mice. These data indicated that fractalkine mediates antitumor effects by both NK cell-dependent and T cell-dependent mechanisms. This study suggests that fractalkine can be a suitable candidate for immunogene therapy of cancer because fractalkine induces both innate and adaptive immunity.     

Heat shock protein 60 from Chlamydia pneumoniae elicits an unusual set of inflammatory responses via Toll-like receptor 2 and 4 in vivo

Heat shock protein 60 (HSP60) from Chlamydia pneumoniae was described to trigger in vitro inflammatory and cytokine responses including TNF and IL-12p40. Although it can be found in atherosclerotic plaques of patients, the stimulatory potential of chlamydial and other HSP60 in vivo is unclear. We now report that chlamydial HSP60 fails to induce TNF expression in vivo, and significant serum levels of IL-12p40 are only found upon intraperitoneal injection of high doses of HSP60 or after intravenous application. Upon purification of chlamydial HSP60 with polymyxin B-agarose columns, its ability to induce TNF secretion in vitro is much reduced. However, purified chlamydial HSP60 causes increased serum levels of the CXC chemokines KC and MIP2 in vivo, as well as a strong accumulation of polymorphonuclear neutrophils (PMN) in the peritoneal cavity upon intraperitoneal challenge. With respect to PMN accumulation, chlamydial HSP60 is more potent than endotoxin or the CpG oligonucleotide 1668. The responses observed are completely abolished in Toll-like receptor (TLR)2/4-double-deficient mice, while single-deficient mice respond almost normally. Furthermore, KC induction and PMN accumulation are largely dependent on MyD88. In conclusion, HSP60 from C. pneumoniae triggers inflammatory responses in vivo that differ from responses induced by endotoxin or CpG oligonucleotides and are dependent on TLR2 and 4.