Characterization of VHL promoter variants in patients suspected of Von Hippel-Lindau disease

Von Hippel-Lindau (VHL) disease is a hereditary tumor syndrome in which carriers are at an increased risk of developing a variety of tumors in multiple organ systems. A clinical diagnosis of VHL is determined by the presence of specific clinical manifestations while a molecular genetic diagnosis results from a pathogenic variant in the VHL gene. The majority of mutations occur in VHL coding exons and DNA analysis of these regions has a reported sensitivity of nearly 100%. However, rare variants in the VHL gene promoter may be detected in some cases of suspected VHL disease. We report two cases where VHL promoter variants were detected and describe the role of multi-step mRNA and protein analysis in the diagnostic evaluation of these cases.     

U6/H1双启动子siRNA表达框架构建及其对肿瘤细胞端粒酶活性的干扰作用

目的：构建针对端粒酶hTERT基因的U6/H1双启动子siRNA表达框架（SEC）,并观察其转录产物对HeLa细胞端粒酶活性的干扰作用。 方法：利用融合PCR技术,针对人端粒酶hTERT基因开放阅读区构建3条U6/H1双启动子SEC以及针对其3''端非翻译区构建1条U6/H1双启动子SEC,对各SEC鉴定后,分别转染人宫颈癌HeLa细胞,用端粒重复序列扩增法（TRAP）检测HeLa细胞的端粒酶活性。 结果：4种针对端粒酶hTERT基因的U6/H1双启动子SEC均成功构建,转染HeLa细胞后,对端粒酶活性的抑制率分别为36.8%、57.39%、80.47%、70.31%。 结论：针对端粒酶hTERT基因的U6/H1双启动子SEC的成功构建,为开展肿瘤端粒酶基因干扰的实验研究提供了新的有效手段。     

TNFα基因启动子多态性与小儿哮喘的相关性

目的：探讨肿瘤坏死因于(tumor necrosis factor alpha,TNFα)基因多态性、等位基因在儿童哮喘发病机制中的意义。方法：采用顺序特异引物聚合酶链反应(polymerase chain reaction with sequence spe-cifie primer,PCR—SSP)方法对30例小儿哮喘病儿(哮喘组)及26例非哮喘呼吸道疾病患儿(非哮喘组)作TNFα基因多态性分析。结果：哮喘组的TNFα基因启动于—308位点基因型中，G／G9例，G／A16例，A／A5例3非哮喘组相应为G／G14例，G／A10例，A／A2例，两组TNFα基因启动于—308位点基因型差异无统计学意义。哮喘组的TNFα基因启动于—308位点等位基因G和A的频率为0.57和0.43，非哮喘组TNFα基因启动于—308位点等位基因G和A的频率为0.73和0．27，哮喘组TNFα基因启动于—308位点等位基因A的频率比非哮喘组高，相对危险度0.482，95％可信区间0.217～1.070，但两组TNFα基因启动因于—308位点等位基因频率差异无统计学意义。结论：本研究未能证实小儿哮喘易感性与TNFα基因启动于—308位点上多态性相关联。     

肝脂酶启动子250 G/A多态性与血脂的关系

目的探讨各项血脂水平与肝脂酶(HL)启动子250 G/A基因多态性的关系。方法运用聚合酶链反应-限制性内切酶片段长度多态性技术(PCR-RFLP),对112例三酰甘油(TG)正常者和103例高TG者的HL启动子250 G/A基因多态性进行研究。结果发现HL启动子250 AA等位基因频率在高TG组为0.447 7明显高于正常TG组(0.263 4,P<0.05)。在总研究组发现AA型的TG、载脂蛋白B(ApoB)浓度明显高于GG型和GA型,而载脂蛋白A l(ApoA l)则明显降低(P<0.05)。按性别分层后发现AA型女性和男性的TG、低密度脂蛋白胆固醇(LDL-C)的浓度明显高于非AA型同性(P<0.05),而高密度脂蛋白胆固醇(HDL-C)、ApoA l浓度分别低于非AA型的同性(P<0.05)。经Logistic回归分析显示年龄、LDL-C、ApoB、AA型是危险因子,而HDL-C是保护因子。结论HL启动子250 G/A多态性与高TG血症的发生相关,并可影响血浆脂类的代谢,可能是高TG所致疾病的危险因素。     

High Hb A2 β-Thalassemia Due to a 468 bp Deletion in a Patient with Hb S/β-Thalassemia

We describe a case of Hb S/β-thalassemia (thal) involving a 468 bp deletion that removes the β-globin gene promoter but leaves the coding regions intact. This is the second report of this deletion, and our family study establishes that this deletion causes β⁰-thal with unusually high levels of Hb A₂ and Hb F. As with other genotypes involving deletions of the 5′ region of the β-globin gene, our patient had a mild form of Hb S/β-thal.     

Rap1GAP regulates renal cell carcinoma invasion

Although patients with localized and regional kidney tumors have a high survival rate, incidence of mortality significantly increases for patients with metastatic disease. It is imperative to decipher the molecular mechanisms of kidney tumor migration and invasion in order to develop effective therapies for patients with advanced cancer. Rap1, a small GTPase protein, has been implicated in cancer cell growth and invasion. Here, we profile migratory and invasive properties of commonly used renal cell carcinoma (RCC) cell lines and correlate that with expression and function of the Rap inactivator Rap1GAP. We report that levels of Rap1GAP inversely correlate with invasion but not migration. We also report that forced over-expression of Rap1GAP decreases invasion of RCC cells but does not impact their rate of proliferation. Low expression levels of Rap1GAP in RCC cells are due, at least in part, to promoter hypermethylation. Rescued expression of Rap1GAP with a demethylating drug, decitabine (5-azadC), decreases the RCC SN12C cell invasion of collagen, fibronectin, and Matrigel matrices. RCC cell lines express distinct levels of cell adhesion proteins and the forced over-expression of Rap1GAP attenuated levels of both cadherins and integrins that are known to regulate the cancer cells invasion. These results demonstrate that targeted restoration of Rap1GAP expression may serve as a potential therapeutic approach to reduce metastasis of kidney cancers.     

乙型肝炎病毒基因型与YMDD基因区序列突变及BCP突变关系探讨

目的 对湘潭地区HBV感染者的基因型、YMDD基因区序列突变及BCP区突变情况进行研究,并对三者之间的关系进行探讨.方法 针对湘潭地区952例不同类型的HBV感染者的样本同时进行基因型、YMDD基因区序列突变及BCP区突变检测,并对检测结果进行统计分析.结果 湘潭地区HBV各种基因型分布比例为:B型698例、占73.32%,C型115例、占12.08%,B、C型混合感染139例、占14.60%.YMDD基因区序列突变结果显示:YMDD野生型844例、占88.66%,其余为YMDD突变型,其中YVDD 54例、占5.67%,YIDD 53例、占5.57%,1例为YMDD与YVDD混合感染.BCP区突变结果显示:1762A/1764G(野生型)672例、占70.59%,1762T/1764A(突变型)188例、占19.75%,其余92例为1762T/1764A和1762A/1764G混合型、占9.66%.基因型、YMDD基因区序列突变及BCP区突变三者相关性分析显示:HBV B型和C型YMDD基因区序列突变率分别为10.04%、10.43%,差异无统计学意义(χ2=0.017,P＞0.05),HBV B型和C型YMDD基因区序列突变类别差异有统计学意义(χ2=4.836,P＜0.05),HBV C型YVDD突变率(9.57%)要高于B型(5.88%).HBV B型和C型BCP区突变率分别为27.36%、46.09%,差异有统计学意义(χ2=16.478,P＜0.01),C型BCP区突变率要高于B型.HBV YMDD野生型和突变型的BCP区突变率分别为2 8.67%、35.51%,差异无统计学意义(χ2=2.139,P＞0.05),但YVDD BCP区突变率(61.11%)要高于其他类别.结论 (1)湘潭地区流行的HBV基因型主要为B型和C型,其中B型为优势基因型,具有南方地区的特点.(2)拉米夫定治疗前通过HBV基因型检测来预测抗病毒应答可能并无实际意义.(3)HBV基因分型、YMDD基因区序列突变、BCP区突变检测的应用,将有助于临床上对乙肝患者的预后和转归进行正确评价,为及时合理的实施干预措施提供了重要依据.

Abstract：

Objective To investigate the relationship between HBV genotypes and YMDD motif mutations or BCP mutations in Xiangtan of Hunan Province. Methods HBV genotypes, YMDD motif mutations and BCP mutations were analyzed in 952 HBV infected patients. Results HBV genotyping showed that 698 HBV type B patients and 115 HBV type C patients accounted for 73.32% and 12.08% respectively of all the participants. The rest 139( 14.60% )were genotype B and C mixed infection( B + C ). The analysis of YMDD motif mutations showed that 844 YMDD wild-type which accounted for 88.66% of all the subjects and the remainder were YMDD mutation types, of which 54( 5.67% ) carried YVDD, 53( 5.57% ) YIDD,and 1 YVDD and YIDD mixed infection. Basic Core Promoter mutations showed that 1762A/1764G ( wild type )accounted for 70.59% and 1762T/1764A( mutant ) accounted for 19.75%. The rest 92 patients were 1762T/1764A and 1762A/1764G mixed infection. This study showed no significant difference in the rate of YMDD mutation( 10.04% vs 10.43% ,χ2 =0.017,P＞0.05 ) ,but a significant difference in the types of YMDD mutation(χ2 = 4.836, P ＜ 0.05 )between HBV types B and C. The YVDD mutation was more commonly seen in genotype C( 9.57% ) than in genotype B( 5.88% ). The BCP mutation rate showed a significant difference( 27.36% vs 46.09%, χ2 = 16.478, P ＜ 0.01 ). Genotype C was more frequent than genotype B. The BCP mutation rate showed no significant difference between YMDD Wild-type and YMDD mutation types( 28.67% vs 35.51%, χ2 = 2.139, P ＞ 0.05 ), but most of BCP mutations happened in YVDD mutant type( 61.11% ). Conclusions ( 1 ) The predominant HBV genotypes in Xiangtan were genotype B and genotype C, the major genotype was type B, which display the characteristics of epidemiology in Southern China. ( 2 ) Determination of HBV genotypes before lamivudine therapy was probably not an important pretreatment investigation to predict antiviral responses. ( 3 ) Detection of HBV genotypes, YMDD motif mutations and BCP mutations will contribute to the correct evaluation of prognosis and timely proper management of HBV patients.