Human papillomavirus type 45 E7 is a transforming protein inducing retinoblastoma protein degradation and anchorage-independent cell cycle progression

High-risk human papillomaviruses (HPV) cause cervical cancer. The biological properties of HPV-45, the third most prevalent high-risk HPV-genotype, are unknown. We demonstrate here that the HPV-45 E7 protein transforms immortalized NIH3T3 fibroblasts, while mutations in either the conserved LXCXE sequence (C28G) or the carboxyl-terminus (Δ87LQQLF91) significantly abolish this activity. To address the mechanisms underlying cell transformation by HPV-45 E7, we investigated its impact on the cell cycle. We show that HPV-45 E7 associates with the hypophosphorylated form of the retinoblastoma protein (pRb) and induces a significant reduction in the pRb half-life which can be blocked by epoxomicin. Moreover, HPV-45 E7 induces anchorage-independent cell cycle progression of NIH3T3 cells and extends the lifespan of primary human keratinocytes. HPV-45 E7C28G did not bind pRb and could neither induce pRb-proteolysis nor promote cell cycle progression. HPV-45 E7Δ87LQQLF91 had intermediate pRb-binding affinity and retained a residual activity to induce the degradation of pRb but lost the capability to promote cell cycle progression in suspension. Another carboxyl-terminal mutant, HPV-45 E7Δ81AEDL84, showed a trend to reduced transforming activity, had reduced pRb-binding activity and lost the capability to induce pRb-degradation; however, this mutant could induce anchorage-independent cell cycle progression with the same efficiency as HPV-45 E7 wild type. In summary, these data suggest that HPV-45 E7 is a transforming protein and that abrogation of cell cycle control contributes to its oncogenic potential.     

A comparison of TRECs and flow cytometry for naive T cell quantification

Assessment of thymic output by measurement of naive T cells is carried out routinely in clinical diagnostic laboratories, predominantly using flow cytometry with a suitable panel of antibodies. Naive T cell measurements can also be made using molecular analyses to quantify T cell receptor excision circle (TRECs) levels in sorted cells from peripheral blood. In this study we have compared TRECs levels retrospectively with CD45RA⁺CD27⁺ T cells and also with CD45RA⁺CD31⁺ T cells in 134 patient samples at diagnosis or during follow‐up. Both panels provide naive T cell measurements that have a strongly positive correlation with TRECs numbers and are suitable for use with enumerating naive T cell levels in a clinical laboratory.     

敲低膜联蛋白A5对人胃癌MGC-803和MKN-45细胞周期的影响

目的 探讨敲低膜联蛋白A5（ANXA5）对人胃癌细胞系MGC-803、MKN-45细胞周期相关蛋白表达的影响。 方法 将细胞分为干扰组、阴性对照组及空白对照组,采用脂质体转染法将靶向膜联蛋白A5的siRNA以及阴性siRNA分别转染干扰组和阴性对照组MGC-803、MKN-45细胞,空白对照组不加任何试剂。转染后48 h采用Real-time PCR和Western bloting分别从mRNA和蛋白水平检测ANXA5的表达,确定ANXA5被敲低之后,Real-time PCR和Western bloting检测各组p21^cip1 mRNA和P21^cip1的表达,Western bloting检测各组细胞周期蛋白D1（cyclinD1）蛋白的表达,流式细胞术检测各组细胞周期的变化情况。 结果 敲低ANXA5后,与阴性对照组和空白对照组相比,两种细胞的p21^cip1mRNA和P21^cip1蛋白均显著降低（P<0.05）,cyclinD1（P<0.05）也显著降低。 结论 敲低ANXA5可下调MGC-803、MKN-45细胞中p21^cip1mRNA和P21^cip1蛋白以及cyclinD1的表达,推测ANXA5可能通过作用于细胞周期相关蛋白而引发细胞周期阻滞从而影响着胃癌的发展。     

Monotypic epithelioid angiomyolipoma of the adrenal gland: an unusual site for a rare extrarenal tumor

Epithelioid angiomyolipoma (AML) is an uncommon renal mesenchymal tumor with malignant potential and is frequently associated with tuberous sclerosis. Extrarenal AMLs are rare, and to the best of our knowledge, this is the first reported case of a primary monotypic epithelioid AML of adrenal gland in a patient without evidence of tuberous sclerosis. The patient is a 42-year old man who presented with retroperitoneal hemorrhage resulting from spontaneous rupture of adrenal mass. Histologically, the tumor showed a prominent component of epithelioid smooth muscle cells with slightly pleomorphic nuclei, sometimes with prominent nucleoli and eosinophilic cytoplasm resembling oncocytic tumors. Epithelioid cells were positive for melanoma (HMB45 and positive MelanA) and smooth muscle markers (α-smooth muscle–specific actin), but not for epithelial markers (cytokeratin, EMA). Differential diagnosis from renal cell carcinoma, adrenal gland carcinoma, and metastatic carcinoma is often challenging because of its epithelioid morphology. Because primary and secondary malignant tumors are much more common and aggressive neoplasms, establishing the correct diagnosis has important therapeutic and prognostic implications.     

热变性对血细胞流式细胞术分析结果的影响

目的 初步探讨热变性对外周血细胞在流式细胞术分析结果中的影响,从而提高流式-荧光原位杂交（Flow-FISH）技术的应用.方法 收集5例非骨髓造血干细胞疾病患者肝素抗凝外周血标本,以CD45-Alexa Fluor（R）647标记细胞表面抗原,高温变性后,采用流式细胞术分析热变性前后外周血有核细胞的散射光信号和荧光信号变化.结果 热变性后,外周血粒细胞的侧向散射光明显缩小;单核细胞不易通过散射光被区分和设门.所有细胞CD45表达强度均减弱,以淋巴细胞为著;通过侧向散射光和CD45设门虽能大致区分各群细胞,但不及未热变性细胞清晰.结论 热变性后,外周血细胞在流式细胞术中的散射光信号和荧光信号均发生了变化,按照常规FSC/SSC和CD45/SSC方法设门进行细胞亚群分析的结果不精确,利用系列特异性荧光抗体标记是一种有前景的方法.     

Apoptosis of glomerular mesangial cells induced by sublytic C5b‐9 complexes in rats with Thy‐1 nephritis is dependent on Gadd45γ upregulation

The complement C5b‐9 complexes can result in cell apoptosis, but the mechanism of sublytic C5b‐9‐mediated glomerular mesangial cell (GMC) apoptosis in Thy‐1 nephritis (Thy‐1N) remains largely unclear. The Gadd45 gene is involved in the cellular response to DNA damage and can promote cell apoptosis. In this study, both Gadd45γ expression patterns and pathologic changes of renal tissue were examined in rat Thy‐1N. Both Gadd45γ expression and GMC apoptosis were significantly decreased in Thy‐1N rats upon the depletion of complement with cobra venom factor. Our in vitro studies showed that Gadd45γ over‐expression increased sublytic C5b‐9‐induced GMC apoptosis, while Gadd45γ gene knockdown by siRNA greatly reduced GMC apoptosis. Moreover, Gadd45γ gene silencing in vivo markedly inhibited the pathologic changes in the renal tissue of Thy‐1N rats. These data suggest that Gadd45γ gene expression is involved in regulating GMC apoptosis mediated by sublytic C5b‐9 in Thy‐1N.     

脂联素基因45T/G多态性对高糖低脂膳食诱导的健康青年血脂比值变化的影响

目的 探讨脂联素基因(APM1)45T/G多态性与健康中国汉族青年血脂比值的关系及其对高糖低脂(high carbohydrate and low fat,HC/LF)膳食诱导的血脂比值变化的影响.方法 56名健康青年志愿者[(22.89±1.80)岁],给予7 d平衡膳食和6 d HC/LF膳食.在第1 d、第8 d、第14 d清晨取空腹12 h静脉血,测定血脂,计算甘油三酯(TG)/高密度脂蛋白胆固醇(HDL-C)、log(TG/HDL-C)、低密度脂蛋白胆固醇(LDL-C)/HDLC、总胆固醇(TC)/HDL-C比值,提取血基因组DNA,聚合酶链反应-限制性酶切法分析APM145T/G多态性.结果 无论是整体还是按性别分组,TT基因型受试者与G等位基因携带者之间血脂比值基础值均无明显差异.男性G等位基因携带者HC/LF膳食后TC/HDL-C高于TT基因型受试者(P<0.05);与膳食前相比,TT基因型受试者HC/LF膳食后LDL-C/HDL-C和TC/HDL-C均降低(P<0.05),G等位基因携带者仅TC/HDL-C降低(P<0.01).在女性,无论HC/LF膳食前还是HC/LF膳食后,G等位基因携带者TG/HDL-C和log(TG/HDL-C)均低于TT基因型个体(P<0.05);与膳食前相比,TT基因型受试者HC/LF膳食后TG/HDL-C和log(TG/HDL-C)升高(P<0.05)、TC/HDL-C降低(P<0.001),G等位基因携带者LDL-C/HDL-C(P<0.05)和TC/HDL-C(P<0.01)降低,而TG/HDL-C和log(TG/HDL-C)无显著变化.结论 APM145T/G G等位基因能抑制HC/LF膳食诱导的健康青年女性TG/HDL-C和log(TG/HDL-C)升高并使LDL-C/HDL-C降低,但可能使男性的LDL-C/HDL-C降低消失、TC/HDL-C升高.     

010 CD45—淋巴细胞活化的重要调节分子

CD45是一种蛋白酪氨酸磷酸酶，它可以通过其蛋白酪氨酸磷酸活性使蛋白酪氨酸激酶抑制位点去磷酸化从而激活激酶，在淋巴细胞的活化中发挥重要作用。本文就CD45在淋巴细胞活化中的作用作一简单综述。