Relationships between endothelial nitric oxide synthase gene polymorphisms and osteoporosis in postmenopausal women

Objective: To investigate the relationships between endothelial nitric oxide synthases (eNOS) G894T and 27 bp-variable number tandem repeat (VNTR) gene polymorphisms and osteoporosis in the postmenopausal women of Chinese Han nationality. Methods: In the present study, 281 postmenopausal women from Xi’an urban area in West China were recruited, and divided into osteoporosis, osteopenia, and normal groups according to the diagnostic criteria of osteoporosis proposed by World Health Organization (WHO). The bone mineral density (BMD) values of lumbar vertebrae and left hips were determined by QDR-2000 dual energy X-ray absorptiometry. Blood samples were tested for plasma biochemical indicators including testosterone, estradiol, calcitonin, osteocalcin, and procollagen type I amino-terminal propeptide by enzyme-linked immunosorbent assay (ELISA), tartrate-resistant acid phosphatase by spectrophotometric method, and the content of nitric oxide by Griess method. Genome DNA was extracted from whole blood, and G894T polymorphism of eNOS gene was analyzed by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method and 27 bp-VNTR polymorphism of eNOS gene was genotyped by PCR method. Then the relationships between genotypes and biochemical indicators, genotypes and osteoporosis, and haplotypes and osteoporosis were analyzed. Results: The average BMD values of the femoral neck, ward’s triangle and lumbar vertebrae 1~4 (L1~L4) in the subjects with T/T genotype in eNOS G894T locus were significantly higher than those in the subjects with G/T and G/G genotypes (P<0.05). The average BMD of the femoral neck in the subjects with a/a genotype of eNOS 27 bp-VNTR locus was evidently higher than that in the subjects with b/b genotype (P<0.05). The plasma testosterone and osteocalcin concentrations in the subjects of eNOS G894T G/T genotype were evidently higher than those in the subjects of other genotypes (P<0.05); the plasma estradiol concentration in the subjects of eNOS 27 bp-VNTR a/a genotype was obviously higher than that in the subjects of b/b genotype (P<0.01). eNOS G/G homozygous frequencies in osteoporosis women, osteopenia women, and normal women were 85.37%, 76.38%, and 83.87%, respectively (P>0.05). 0% osteoporosis woman, 0.79% osteopenia women, and 3.23% normal women were eNOS a/a homozygous (P<0.05). The frequencies of eNOS 27 bp-VNTR a allele were 5.33% in the osteoporosis group, 10.24% in the osteopenia group, and 16.13% in the normal group (P<0.05, odds ratio (OR)=0.29, 95% confidence interval (CI)=0.11~0.77), suggesting that a/a genotype and a allele might have protective effects on osteoporosis. The haplotype analysis showed that G-b was 87.7% (214/244) in the osteoporosis group (P<0.05, OR=2.48, 95% CI=1.18~5.18). G-a was 5.3% (13/244) in the osteoporosis group (P<0.05, OR=0.29, 95% CI=0.11~0.77). G-b was a risk factor for osteoporosis, and G-a a protective factor. Conclusion: eNOS G894T G/T genotype influenced the plasma testosterone and osteocalcin concentrations, and T/T genotype influenced BMD. eNOS 27 bp-VNTR a/a genotype increased plasma estradiol concentration to have a protective effect on osteoporosis.     

p53蛋白在胃癌和结肠癌中的表达

目的　研究 p5 3蛋白在胃癌和结肠癌中的表达 ,分析其与临床及病理因素的关系 .方法　采用免疫组织化学S P法 ,对外科手术切除的胃癌 2 0例、结肠癌 39例的癌体标本组织中 p5 3蛋白的表达进行检测 .结果　p5 3蛋白在胃癌和结肠癌标本组中阳性的表达在细胞核 ,其阳性表达率在胃癌和结肠癌标本中分别为 80 %和 4 8.7% .结论　p5 3蛋白的阳性表达与肿瘤的浸润深度无关 (P >0 .0 5 ) ,而与肿瘤的分化程度及淋巴结转移呈相关关系 (P <0 .0 5 ) .     

Mutations in the p53 gene occur in diverse human tumour types

The p53 gene has been a constant source of fascination since its discovery nearly a decade ago. Originally considered to be an oncogene, several convergent lines of research have indicated that the wild-type gene product actually functions as a tumour suppressor gene. For example, expression of the neoplastic phenotype is inhibited, rather than promoted, when rat cells are transfected with the murine wild-type p53 gene together with mutant p53 genes and/or other oncogenes. Moreover, in human tumours, the short arm of chromosome 17 is often deleted. In colorectal cancers, the smallest common region of deletion is centred at 17p13.1; this region harbours the p53 gene, and in two tumours examined in detail, the remaining (non-deleted) p53 alleles were found to contain mutations. This result was provocative because allelic deletion coupled with mutation of the remaining allele is a theoretical hallmark of tumour-suppressor genes. In the present report, we have attempted to determine the generality of this observation; that is, whether tumours with allelic deletions of chromosome 17p contain mutant p53 genes in the allele that is retained. Our results suggest that (1) most tumours with such allelic deletions contain p53 point mutations resulting in amino-acid substitutions, (2) such mutations are not confined to tumours with allelic deletion, but also occur in at least some tumours that have retained both parental 17p alleles, and (3) p53 gene mutations are clustered in four 'hot-spots' which exactly coincide with the four most highly conserved regions of the gene. These results suggest that p53 mutations play a role in the development of many common human malignancies.     

AgNOR和p53在大肠癌中的表达

目的研究AgNOR和p53在大肠癌中的表达及其与大肠癌发生、发展及预后的关系.方法选用大肠癌组织标本80例及相应的癌旁组织标本40例,采用AgNOR染色技术和免疫组织化学法检测大肠癌组织及癌旁组织中AgNOR计数和p53的表达.结果大肠癌中AgNOR计数显著升高,细胞核内AgNOR计数、颗粒分布及形态特点与大肠癌的分化程度、淋巴结转移有相关性(P<0.05);p53在大肠癌组的阳性表达率为66.3%,在癌旁组的阳性表达率为20.8%.两者之间存在显著性差异(P<0.01);大肠癌组织中p53的阳性表达与大肠癌的分化程度、淋巴结转移有相关性(P<0.05),而与大肠癌的发病年龄、性别无关(P>0.05).结论 p53的突变在大肠癌的形成和发展中起了促进作用,AgNOR测定对大肠癌的病理学分级有一定的价值,同时说明AgNOR和p53有可能作为大肠癌发生、发展及预后的肿瘤标志.     

中国汉族人群MC1R基因多态与皮肤颜色等表型特征关联研究

皮肤癌是一种比较常见的肿瘤,在西方人群中研究表明,通过影响皮肤颜色等色素表型特征,黑色素生成通路中相关基因多态性(MC1R等)可增加皮肤癌发生风险.本研究首次在中国汉族人群中选择MC1R基因编码区的多态位点为研究对象,分析其与中国汉族人群的皮肤颜色、对阳光的敏感性、雀斑情况及头发颜色的关联性.在MC1R基因编码区发现7个SNP位点:c.200G>A,c.274G>A,c.359T>C,c.421G>A,c.488G>A,c.497C>G和c.942A>G.其中c.421G>A为新发现的SNP位点,c.497C>G则首次在中国汉族人群中报道.c.942A>G与皮肤颜色相关,其突变纯合基因型GG与皮肤颜色浅显著相关(P=0.044,OR=0.16,95%CI:0.03～0.95).c.359T>C杂合基因型TC与雀斑数目多显著相关(P=0.040,OR=5.76,95%CI:1.05～31.53),表明MC1R基因多态与中国汉族人群的皮肤颜色等色素表型特征具有一定相关性,为相关临床研究提供了参考.     

头颈部鳞癌中p53-mdm2基因异常的研究

目的:检测头颈部鳞癌患者p53基因突变、mdm2基因扩增的状况,了解其与头颈部鳞癌患者的性别、鳞癌分级、淋巴结转移等的相关性及两异常基因的相关性。方法:收集50例行手术切除的头颈部鳞癌患者的新鲜肿瘤组织及其相应的癌旁正常组织,提取标本DNA;用PCR-SS-CP-银染法检测p53基因第5~8外显子的突变状况;用dPCR法检测mdm2基因扩增情况;采用SPSS 10.0统计软件包行2检验分析实验结果。结果:在50例头颈部鳞癌患者标本中,检测出17例存在p53基因突变,突变率为34%,所有的癌旁正常组织未发现p53基因突变;在50例鳞癌标本中检测出6例标本存在mdm2基因扩增,扩增率为12%,其中有一例同时存在p53基因突变;p53基因突变、mdm2基因扩增与患者的鳞癌分级、淋巴结转移、性别等相关性分析结果均无统计学意义(P>0.05)。结论:p53基因突变与mdm2基因扩增在头颈部鳞癌中较常见,可能是头颈部鳞癌发生发展的主要分子机制。     

ACE基因多态性与异常体液型乳腺癌的易感性

 为探讨血管紧张素转换酶(ACE)基因插入/缺失(I/D)多态性与新疆汉族人群乳腺癌的相关性, 按维吾尔医将乳腺癌患者分为4 种体液型, 采用聚合酶链式反应(PCR)技术对新疆汉族139 例乳腺癌患者和72 例正常对照组ACE 基因I/D 多态性进行检测, 比较各组间等位基因和基因型频率分布的差异。结果显示, 异常黏质乳腺癌患者组II 基因型频率(P=0.018)和I 等位基因频率(P=0.004)都显著高于正常对照组;异常黏液质乳腺癌患者组I 等位基因频率显著高于异常黑胆质乳腺癌患者组(P=0.012)。由此得出, ACE 基因I 等位基因和II 基因型可能增加新疆汉族维吾尔医异常黏液质型乳腺癌的发病风险。     

A ribonucleotide reductase gene involved in a p53-dependent cell-cycle checkpoint for DNA damage

The p53 gene is frequently inactivated in human cancers. Here we have isolated a p53-inducible gene, p53R2, by using differential display to examine messenger RNAs in a cancer-derived human cell line carrying a highly regulated wild-type p53 expression system. p53R2 contains a p53-binding sequence in intron 1 and encodes a 351-amino-acid peptide with striking similarity to the ribonucleotide reductase small subunit (R2), which is important in DNA synthesis during cell division. Expression of p53R2, but not R2, was induced by ultraviolet and gamma-irradiation and adriamycin treatment in a wild-type p53-dependent manner. Induction of p53R2 in p53-deficient cells caused G2/M arrest and prevented cells from death in response to adriamycin. Inhibition of endogenous p53R2 expression in cells that have an intact p53-dependent DNA damage checkpoint reduced ribonucleotide reductase activity, DNA repair and cell survival after exposure to various genotoxins. Our results indicate that p53R2 encodes a ribonucleotide reductase that is directly involved in the p53 checkpoint for repair of damaged DNA. The discovery of p53R2 clarifies a relationship between a ribonucleotide reductase activity involved in repair of damaged DNA and tumour suppression by p53.     

Polymorphisms in the genes for coagulation factor Ⅱ,Ⅴ,Ⅶ in patients undergoing coronary angiography

OBJECTIVE: To determine whether polymorphisms in the genes for coagulation factor II, V, VII could predispose an individual to increase risk for coronary artery disease (CAD) and/or myocardial infarction (MI) in Chinese. METHODS: We screened coagulation factor II(G20210A),V(G1691A),VII (R353Q and HVR4) genotype in 374 patients undergoing coronary angiography by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) assay. RESULTS: The R353Q and HVR4 genotype of the factor VII distribution was in accordance with Hardy-Weinberg equilibrium. The frequencies of FVII genotype or allele did not show statistically significant differences between CAD group and controls or between male and female. The frequencies of the Q allele and (RQ + QQ) genotype were significantly higher among the CAD patients without myocardial infarction (MI) history than among those with MI history (P < 0.05). However, HVR4 polymorphism was not significantly different within groups. We only find one normal control of factor II (G20210A) mutation. No coagulation factor V(G1691A) mutation was found in the CAD patients and controls. CONCLUSION: The factor II(G20210A) ,V(G1691A) mutation is absent and may not be a major genetic factor for CAD and/or MI; the Q allele of the R353Q polymorphism of the factor VII gene may be a protective genetic factor against myocardial infarction in Chinese.     

Fbxw7/Cdc4 is a p53-dependent, haploinsufficient tumour suppressor gene

The FBXW7/hCDC4 gene encodes a ubiquitin ligase implicated in the control of chromosome stability. Here we identify the mouse Fbxw7 gene as a p53-dependent tumour suppressor gene by using a mammalian genetic screen for p53-dependent genes involved in tumorigenesis. Radiation-induced lymphomas from p53+/- mice, but not those from p53-/- mice, show frequent loss of heterozygosity and a 10% mutation rate of the Fbxw7 gene. Fbxw7+/- mice have greater susceptibility to radiation-induced tumorigenesis, but most tumours retain and express the wild-type allele, indicating that Fbxw7 is a haploinsufficient tumour suppressor gene. Loss of Fbxw7 alters the spectrum of tumours that develop in p53 deficient mice to include a range of tumours in epithelial tissues such as the lung, liver and ovary. Mouse embryo fibroblasts from Fbxw7-deficient mice, or wild-type mouse cells expressing Fbxw7 small interfering RNA, have higher levels of Aurora-A kinase, c-Jun and Notch4, but not of cyclin E. We propose that p53-dependent loss of Fbxw7 leads to genetic instability by mechanisms that might involve the activation of Aurora-A, providing a rationale for the early occurrence of these mutations in human cancers.     

P53和MDM2蛋白表达对肝细胞肝癌预后判断的意义

目的 :研究肝细胞肝癌中突变型P5 3和MDM2蛋白的表达对临床预后判断的意义。方法 :应用免疫组织化学方法 ,检测 72例原发性肝细胞肝癌手术切除标本突变型P5 3、MDM2蛋白的表达 ;与临床病理学指标和术后生存期进行分析比较。结果 :突变型P5 3蛋白阳性 2 8例(38 89% ) ,MDM2蛋白阳性 2 3例 (31 94 % ) ,二者阳性表达有相关性 (r =0 2 4 8,P <0 0 5 )。突变型P5 3、MDM2蛋白阳性表达病例生存率明显低于突变型P5 3、MDM2蛋白阴性表达病例 (P <0 0 1)。突变型P5 3和MDM2蛋白表达均阳性病例 13例 (18 0 6 % ) ,中位生存期的生存率最低。单因素及多因素分析显示 ,突变型P5 3蛋白表达、MDM2蛋白表达、肿瘤大小与中位生存期的生存率有关 ,MDM2是统计学上最有意义的独立预后指标 (P <0 0 0 0 1)。结论 :应用免疫组织化学方法检测突变型P5 3和MDM2蛋白的表达可作为原发性肝细胞肝癌预后判断的指标。     

脑肿瘤中p53基因突变的研究

应用ＰＣＲＳＳＣＰ技术及ＤＮＡ 测序技术对３７ 例原发性脑肿瘤及相应外周血淋巴细胞中ｐ５３ 基因５ ～８ 外显子的突变情况进行了检测,结果表明,ｐ５３ 基因在原发性脑肿瘤中的突变频率为１９ ％ （７／３７） ．并且突变频率在不同病理类别脑肿瘤中的分布是非随机的,其中星形细胞肿瘤中的突变频率最高,为３６ ％ （５／１４） ．所有突变均为错义点突变,５７ ％ （４／７） 的突变位于ＣｐＧ位点．突变仅发现于脑组织中,外周血淋巴细胞中未检出突变,这些结果提示,ｐ５３ 基因突变在脑肿瘤的发生发展过程中起一定的作用,ｐ５３ 基因在散发性脑肿瘤中的突变为体细胞型的突变     

GRTH基因SNPrs551373与严重少精症的相关性

目的：研究GRTH基因的单核苷酸多态位点（SNP）rs551373（G〉T）的多态性与严重少精症的关系。方法：用PCR—RFLP技术,在119例严重少精症患者和252个正常生育男性中,对SNPrs551373的基因频率和基因型频率的分布进行调查。结果：严重少精症患者中基因型GG的频率明显低于正常男性（72．3％vs．83．3％,降0．013,OR=0．503,95％CI0．299～0．848）;而等位基因T（13．7％vs．8．7％,P=0．021,OR=1．742,95％CI1．082～2．807）和基因型GT（26．9％vs．15．9％,P=0．012,OR=1．949,95％CI1．150～3．304）的频率则显著高于正常男性。结论：GRTH基因的SNPrS551373的多态性与严重少精症的易感性相关。     

胃癌中p53基因失活及其蛋白产物的免疫组化分析

应用ＰＣＲ－ＳＳＣＰ银染，Ｓｏｕｔｈｅｒｎ杂我及免疫组化等方法同步研究２５例胃癌标本的ｐ５３基因突变，杂合性丢失及蛋白持表达，在２２例胃癌中同时获得有关ｐ５３基因突变和杂合性丢失的检测结果，被检出ｐ５３基因突变的５例胃癌中，２例伴有杂合性丢失，３例仅有ｐ５３基因突变。     

Comprehensive diagnostic value of P53, p21WAF1 and proliferating cell nuclear antigen for lung cancer

The expression of P53, p21WAF1 and proliferating cell nuclear antigen (PCNA) was detected in 114 samples of lung cancer patients (with 89 cases benign lung tissue as control) and the diagnostic value of these markers was evaluated. The results show the following: ➀ The positive expression rates of P53, p21WAF1 and PCNA in samples of lung cancer were 47.37%, 75.44% and 80.70%, respectively, which were significantly higher than that in the samples of benign lung diseases ( p < 0.001). The odds ratios were 39.15, 5.75, and 6.76, respectively. This indicates that the expression of P53, p21WAF1 and PCNA was helpful for the diagnosis of lung cancer. ➁ For the diagnosis of lung cancer, the positive likelihood ratio of P53 was 21.08, which were significantly higher than that of p21WAF1 (2.16), PCNA (2.11) and of all the combined tests. This shows that P53 expression was the most valuable for diagnosis of lung cancer. ➂ For the diagnosis of lung cancer, the negative likelihood ratio of P53/p21WAF1/PCNA parallel test was 0.057 1, which was lower than that of other single and combined tests. This indicates that P53/p21WAF1/PCNA parallel test has high diagnostic value for exclusion of lung cancer.     

Cytokinesis failure generating tetraploids promotes tumorigenesis in p53-null cells

A long-standing hypothesis on tumorigenesis is that cell division failure, generating genetically unstable tetraploid cells, facilitates the development of aneuploid malignancies. Here we test this idea by transiently blocking cytokinesis in p53-null (p53-/-) mouse mammary epithelial cells (MMECs), enabling the isolation of diploid and tetraploid cultures. The tetraploid cells had an increase in the frequency of whole-chromosome mis-segregation and chromosomal rearrangements. Only the tetraploid cells were transformed in vitro after exposure to a carcinogen. Furthermore, in the absence of carcinogen, only the tetraploid cells gave rise to malignant mammary epithelial cancers when transplanted subcutaneously into nude mice. These tumours all contained numerous non-reciprocal translocations and an 8-30-fold amplification of a chromosomal region containing a cluster of matrix metalloproteinase (MMP) genes. MMP overexpression is linked to mammary tumours in humans and animal models. Thus, tetraploidy enhances the frequency of chromosomal alterations and promotes tumour development in p53-/- MMECs.     

应用PCR－SSCP银染技术检测消化系统肿瘤的P53基因突变

应用ＰＣＲ－ＳＳＣＰ银染技术，初步研究了肝细胞癌、胃癌和大肠癌中Ｐ５３基因的第６和第７外显子的分子结构改变。对来自癌组织ＤＮＡ和正常组织ＤＮＡ的ＰＣＲ－ＳＳＣＰ电泳带迁移作对比分析，发现３０例肝癌病人中，６例肝癌样品电泳带迁移异常；２６例胃癌病人中，４例胃癌样品电泳带迁移异常；２９例大肠癌病人中，６例大肠癌样品电泳带迁移异常。依据ＤＮＡ单链构象与分子电泳迁移的关系，研究结果表明：该三组病人中，Ｐ５３基因第６、７外显子的突变率分别为２０．０％，１５．４％和２５．０％。同时，也间接提示了Ｐ５３基因突变可能是肝细胞癌、胃癌和大肠癌中一种较多见的分子结构改变。     

CYP1A1基因Ile-Val和Msp1位点多态性与结直肠癌易感性研究

目的探讨细胞色素P450(CYP1A1)基因异亮氨酸(Ile)-缬氨酸(Val)位点和Msp1位点多态性和结直肠癌的相关关系.方法以病例对照的研究方法,采用PCR-RFLP和AS-PCR技术检测79例结直肠癌和110例对照者的CYP1A1基因Ile-val位点和Msp1位点多态性.结果 Ile-Va三种多态基因型在结直肠癌组和对照组分布差异有显著性(P<0.05),Ile/Val,Val/Val基因型在结直肠癌组的分布频率明显高于对照组;Ile/Val,Val/Val基因型患结直肠癌的危险分别是Ile/Ile基因型的2.113倍和4.203倍;当按吸烟分层后(将Ile/Val,Val/Val基因型合并分析),吸烟组中Ile/Val、Val/Val合并基因型患结直肠癌的危险是Ile/Ile基因型的2.98倍(P<0.05);Msp1位点多态性在结直肠癌和对照组差异无统计学意义.结论 CYP1A1第7外显子的Ile/Val,Val/Val基因型与结直肠癌的易感性有关,突变基因型增加了结直肠癌的患病风险;尚不能认为Msp1多态性与结直肠癌的易感性有关.     

Deleted in colorectal carcinoma suppresses metastasis in p53-deficient mammary tumours

Since its discovery in the early 1990s the deleted in colorectal cancer (DCC) gene, located on chromosome 18q21, has been proposed as a tumour suppressor gene as its loss is implicated in the majority of advanced colorectal and many other cancers. DCC belongs to the family of netrin 1 receptors, which function as dependence receptors as they control survival or apoptosis depending on ligand binding. However, the role of DCC as a tumour suppressor remains controversial because of the rarity of DCC-specific mutations and the presence of other tumour suppressor genes in the same chromosomal region. Here we show that in a mouse model of mammary carcinoma based on somatic inactivation of p53, additional loss of DCC promotes metastasis formation without affecting the primary tumour phenotype. Furthermore, we demonstrate that in cell cultures derived from p53-deficient mouse mammary tumours DCC expression controls netrin-1-dependent cell survival, providing a mechanistic basis for the enhanced metastatic capacity of tumour cells lacking DCC. Consistent with this idea, in vivo tumour-cell survival is enhanced by DCC loss. Together, our data support the function of DCC as a context-dependent tumour suppressor that limits survival of disseminated tumour cells.