贲门癌中染色体8p21-p23杂合性丢失的研究

目的 研究贲门癌中染色体8p21-p23杂合性丢失的情况。方法 采用激光捕获显微切割技术获得均质的肿瘤细胞及正常的胃粘膜细胞,多重置换扩增技术扩增捕获细胞的基因组DNA。PCR结合硝酸银染色方法分析19例贲门癌染色体8p21-p23的杂合性丢失。结果 在贲门癌中染色体8p21-p23的缺失频率非常高（63.2%）,我们确定了一个最小丢失区域. 结论 进一步明确此最小丢失区域内的抑癌基因将有助于贲门癌发生机制的阐明。     

8p杂合性缺失与膀胱肿瘤的关系

肿瘤遗传学和分子生物学研究表明,癌症的发生是由癌基因的激活及抑癌基因失活的结果,而抑癌基因的失活可能在其中起着更为关键的作用。抑癌基因（tumor suppressor gene）是能够抑制细胞的恶性转化,对正常细胞的增殖起负性调节的基因。抑癌基因的失活常常表现为一个等位基因丢失（allelic loss）和另一个存留等位基因（retained allele）突变。其中,等位基因丢失就是由肿瘤中常见的染色体区域或节段缺失所致,它同时还会伴有抑癌基因座位相邻区域的杂合性缺失（loss of heterozygosity,LOH）。IDH是指肿瘤基因中特定染色体上某种DNA多态标记的等位基因片段由同一患正常组织基因组的两种变成一种,即等位基因型由杂合子变成纯合子。IDH是肿瘤细胞中染色体缺失的表现。     

脑胶质瘤中P53基因突变及其与染色体17p杂合性丢失的比较

为进一步阐明P53基因突变和染色体17p杂合性丢失(LOH)的关系及两者与脑胶质瘤发生发展的相关性。应用PCR－SSCP、DNA序列测定及RFLP分析方法对55例脑胶质瘤中的P53基因突变及染色体17p的杂合性丢失进行了检测。发现P53基因在高级别星型细胞肿瘤(III－IV级)、低级别星型细胞肿瘤(I－II级)和非星型细胞肿瘤中的突变频率分别为：53％(9/17)、7％(1/15)和9％(2/23)。而55例肿瘤组织对应的淋巴细胞中未见P53突变。22％的胶质瘤丢失1个17p等位基因,这部分肿瘤中P53基因的突变频率为50％(6/12),而在43例持有2个17p等位基因的肿瘤中,P53基因的突变频率降为14％,两组相比差异显著(P<0?025)。结果提示P53基因突变是星型细胞肿瘤演变过程中的一个常见遗传事件,可能标志着肿瘤的恶性进展;散发性脑胶质瘤中的P53突变是体细胞型的突变;另外,丢失1个17p等位基因的肿瘤中有50％缺少P53基因突变,提示染色体17p上可能存在另一个参与了部分肿瘤恶性演变的肿瘤抑制基因。 Abstract：To further illustrate the roles of P53 gene and loss of heterozygosity (LOH) on chromosome 17p in the development of malignant gliomas,mutations in the P53 gene were analyzed in 55 gliomas of various malignant grades and histological types by the polymerase chain reaction-single-strand conformation polymorphism (PCR-SSCP) technique and were confirmed by sequencing.Loss of heterozygosity (LOH) for chromosome 17p was also assessed using restriction fragment length polymorphism (RFLP) analysis in same tumors.The mutations did not follow a random distribution among various different subtypes,but occurred in 9 of 17 high-grade astrocytomas (53％),in 1 of 15 low-grade astrocytomas (7％) and in 2of 23 (9％) non-astrocytic tumors.Most of the mutations were missense ones and 42％ (5/12) occurred at the sites of CpG dinucleotide.P53 mutations were not detected in any of the 55 leukocyte DNA samples from patients with brain tumors.These studies demonstrated that P53 inactivation is a common genetic event in astrocytoma progression that may signal the transition from benign to malignant tumor stages.P53 gene mutations in sporadic human brain tumors are somatic in origin (i.e.,nonprenatally determined).The majority of glomas (43/55) analyzed here retained both 17p alleles.The frequency of P53 mutations was 14％ in this group of tumors and increased to 50％ (6/12) in tumors with one 17p allele (P<0.025).Allelic loss for chromosome 17p occurred in 6 of 12 gliomas independently of mutations in the P53 gene.Absence of P53 mutations in 50％ of the tumors with one allele suggests that a tumor suppressor gene other than P53 may be located on chromosome 17p and involved in progression to malignancy of some gliomas.     

人肝癌中的抑癌基因

杂合性丢失分析资料显示,在肝癌的发生发生和演讲过程中涉及多种抑癌基因失活。国内外研究表明,肝癌中存在抑癌基因ＴＰ５３突变失活,ＴＰ５３基因突变表现特异性的和病因相关性。作者通过检查１５个高度多态的微卫星标记,进行染色体缺失精神定位,将 染色体１３ｑ缺失的最小叠区域限定于１３ｑ１４,强烈提示ＲＢ１基因为染色体缺失的靶基因;进一步应用ＰＣＲ－ＳＳＣＰ分析和ＤＮＡ测序发现２例发生染色体１３ｑ臂内缺失的肝     

SEMA3B基因在鼻咽癌组织中的表达、杂合性丢失和甲基化分析

SEMA3B基因定位于鼻咽癌高频缺失区域3p21.3上,最近被证明具有抑瘤基因的功能.分析了鼻咽癌组织中SEMA3B基因的表达、杂合性丢失(LOH)和甲基化情况.首先应用逆转录-聚合酶链式反应(RT-PCR)方法检测了33例鼻咽癌组织和15例慢性鼻咽炎组织中SEMA3B基因的表达,结果显示75.8%(25/33)鼻咽癌组织中SEMA3B基因表达缺失或下调,显著低于慢性鼻咽炎组织中的表达(P=0.001).进一步选取3个微卫星位点D3S1568、D3S1621和D3S4597分析了20例鼻咽癌组织中SEMA3B基因LOH的情况,结果表明3个位点的丢失率分别为10%、20%和15%,总的丢失率为45%,统计分析发现LOH与基因表达之间存在明显相关(P=0.023).最后,采用甲基化特异性PCR方法分析了SEMA3B基因启动子区甲基化,结果发现在100%的鼻咽癌组织和73.3%的慢性鼻咽炎组织中检测到SEMA3B基因启动子区高甲基化.由此得出结论,SEMA3B基因在鼻咽癌组织中表达缺失或下调,LOH是引起其表达异常的原因之一.     

原发性肝癌染色体8p、16q 遗传变异的研究

目的:分析人肝癌(HCC)组织中染色体8p、16q部分基因及染色体片段的遗传变异及与临床病理关系,初步筛选HCC相关的抑癌基因。方法:应用聚合酶链反应-变性聚丙烯酰胺凝胶-银染法分析45例HCC组织标本中染色体8p和16q的杂合性丢失(LOH)及微卫星不稳定性(MSI)。结果:发生LOH的总频率为68.89%(31/45),其中D16S511位点的发生LOH率最高为53.33%(24/45),其次是D8S261(39.02%,16/41)和D8S499(34.88%,15/43)。MSI出现的总频率为11.11%(5/45),出现在三个微卫星位点(D8S261、D8S499及D16S511)上。结论:染色体16q23、8p22-21.3及8p12区域的LOH发生频率高,其可能存在与HCC发生发展相关的新的抑癌基因,特定位点的遗传变异可能与HBV感染、临床病理恶性程度等预后因素相关。     

原发性肝癌染色体8p、16q遗传变异的研究

目的：分析人肝癌（HCC）组织中染色体8p、16q部分基因及染色体片段的遗传变异及与临床病理关系,初步筛选HCC相关的抑癌基因。方法：应用聚合酶链反应-变性聚丙烯酰胺凝胶-银染法分析45例HCC组织标本中染色体8p和16q的杂合性丢失（LOH）及微卫星不稳定性（MSI）。结果：发生LOH的总频率为68.89%（31/45）,其中D16S511位点的发生LOH率最高为53.33%（24/45）,其次是D8S261（39.02%,16/41）和D8S499（34.88%,15/43）。MSI出现的总频率为11.11%（5/45）,出现在三个微卫星位点（D8S261、D8S499及D16S511）上。结论：染色体16q23、8p22-21.3及8p12区域的LOH发生频率高,其可能存在与HCC发生发展相关的新的抑癌基因,特定位点的遗传变异可能与HBV感染、临床病理恶性程度等预后因素相关。     

非小细胞肺癌6号染色体长臂的杂合性缺失的微卫星扫描分析

为探讨6号染色体长臂上与非小细胞肺癌发生相关的微卫星位点,应用多重PCR对41例非小细胞肺癌中6号染色体长臂上的36个微卫星位点进行扩增。PCR产物应用聚丙烯酰胶凝胶电泳分离,电泳结果用GeneScan^TM、Genotyper^TM软件进行分析。结果发现各个位点有明显不同的LOH频率,除D6S1579在41例非小细胞肺癌中未发现杂合性缺失外,其余35个位点均有至少一个位点发生突变,LOH频率从3.57%到75%不等,总LOH频率为78%（32/41）,其中D6S302位点的LOH频率最高（75%）。LOH频率超过20%的位点共有14个,主要分布在2个区域。其中有6个位点：D6S458（21.43%)、D6S1694（26.92%）、D6S1717（35.71%）、D6S1565（40%）、D6S302（75%）、D6S1706（36.36%）分布在6q21附近,具体区域是6q16.3-q21;有5个位点：D6S1550（38.4%）、D6S264（20%）、DS1585（25%）、D6S446（33.3%）、D6S281（30.77%）分布在6q27附近,具体区域是6q26-q27。因此6q21、q27附近可能存在与非小细胞肺癌发生相关的巳知或未知肿瘤抑制基因。     

Cloning and characterization of a novel gene（C17orf25）from the deletion region on chromosome 17p13.3 in hepatocelular carcinoma

INTRODUCTIONLoss of heterozygosity (LOH) at chromosoma1loci associated with tumor suppressor genes has beenimplicated in the genesis of many types of humanmalignancies. On the basis of frequent LOH in tu-mors, coupled with linkage analysis in some heredi-tary cancer syndromes, a number of tumor suppres-sor genes, such as RB[l], DCC[2], NF2[3], VHLI4],MTh1[5], DML/OM1[6], and PrsN/rmC1[7l have been successively isolated.It has beell reported that LOH occurred at l7p invarious ty…     

Genetic markers for idiopathic scoliosis on chromosome 19p 13.3 among Saudi Arabian girls: A pilot study

BACKGROUND AND OBJECTIVE:

Genetic locus linked to chromosome 19p for Adolescent idiopathic scoliosis (AIS) has been described. This study was carried out with the aim to find any significant linkage or association between three microsatellite markers (D19S216, D19S894, and DS1034) of chromosome 19p13.3 in Saudi Arabian girls with AIS.

MATERIALS AND METHODS:

In eleven unrelated Saudi Arabian girls who were treated for AIS with Cobb angle of ≥30 degrees and in 10 unrelated healthy individuals, linkage analysis was performed using parametric and nonparametric methods by use of GENEHUNTER version 2.1. Multipoint linkage analysis was used in specifying an autosomal dominant trait with a gene frequency of 0.01 and an estimated penetrance of 80% at the genotype and the allele level. Fisher''s exact test was used in the analysis of contingency tables for the D19S216, D19S894, and DS1034 markers.

RESULTS:

The analysis between the patient group and healthy girls showed that at genotypic level there was no significant association of the markers and scoliosis D19S216 (P = 0.21), D19S894 (P = 0.37), and DS1034 (P = 0.25). Whereas, at the allele level, there was statistically significant association between the marker DS1034 (P = 0.008) and no significant association with the other two markers D19S216 (P = 0.25) and D19S894 (P = 0.17).

CONCLUSIONS:

Our study shows that at genotypic level none of the markers reported earlier were associated with scoliosis but at allele level, marker DS1034 was significantly associated with patients with AIS. This allele marker on chromosome 19p appears important in the etiology of AIS.     

Loss of heterozygosity at chromosome 6 as a marker of early genetic alterations in cervical intraepithelial neoplasias and microinvasive carcinomas

Oncogenic human papillomaviruses (mostly HPV types 16 and 18) are the major cause of cervical intraepithelial neoplasia (CIN), which progresses into cervical cancer (CC). To reveal early genetic alterations of chromosome 6 that are important for CC progression, we analyzed the loss of heterozygosity (LOH) in DNAs from 45 CIN cases, 47 microcarcinomas, and 19 invasive squamous cell carcinomas stage IB. LOH analysis of DNA samples prepared with microdissection from all CIN foci, as well as from CC lesions and synchronous CIN, permitted investigation of CIN and CC heterogeneity. Out of all CC stage I cases, 79% showed LOH with six microsatellite markers at chromosome 6. LOH with the microsatellite markers D6S276 (6p22) and TNFa (6p21.3) was found in 50% of the CC cases. LOH frequency in CIN lesions synchronous with CC was higher then in CIN cases without cancer; the statistical significance (P = 0.004) was shown for D6S291 (6p21.2). The finding suggests that the high frequency of LOH in CIN lesions is a marker of unfavorable prognosis for CIN. Progression from microcarcinoma to invasive CC of stage IB was associated with a higher LOH frequency at D6S344 (6p25) and TNFa (6p21.3). Early genetic alterations were found in CIN with microsatellites D6S273 and TNFa located at 6p21.3. Moreover, LOH frequency at D6S273 remained the same in both CIN and CC cases. Based on HPV typing, LOH analysis, and X-chromosome inactivation, the polyclonality of CC lesions, as well as CIN, was observed in a few patients.     

Loss of heterozygosity of the p53 gene and deregulated expression of its mRNA and protein in human brain tumors

Tumor-specific alterations at the p53 gene locus were analyzed in 40 human brain tumor samples. Gliomas were more prevalent in young males and meningiomas in old females. Structural changes at the intron 1 region of the p53 gene were analyzed in these tumors by Southern blotting. Among the 40 tumors, 33 were informative and 21 of these (63.6%) informative cases showed loss of heterozygosity (LOH). This is the first report showing LOH at the intron 1 region of p53 gene in human brain tumors. The level of p53 mRNA, p53 protein and Ser 392 phosphorylated p53 protein were also analyzed in all tumor samples. Normal sized p53 mRNA and protein were present in all the tumor samples; however, their levels were 1.5- to 4-fold higher compared to the control suggesting deregulated p53 pathway in these tumors. No correlation was found between LOH status and the levels of p53 mRNA and protein. In all high-grade glioblastomas majority of the p53 protein existed as Ser 392 phosphorylated form as compared to low-grade gliomas. In addition, the percentage of Ser 392 phosphorylated form of p53 protein was lower in meningiomas and other brain tumor types irrespective of tumor grade. These results suggest involvement of Ser 392 phosphorylated form of p53 protein during the later stages of glioma development. These results also indicate that deregulation of p53 gene could occur at various steps in p53 pathway and suggest an overall deregulation of p53 gene in most brain tumor types.     

Loss of heterozygosity and microsatellite instability in tumor-associated stromal cells and tumor epithelium of prostate cancer

In the past decade, intense studies of the tumor microenvironment yielded ample data testifying to the critical role of stroma in carcinogenesis. Genetic lesions accumulate not only in the tumor epithelium, but also in tumor-associated fibroblasts. The epithelial and stromal components of prostate cancer (PC) and prostatic intraepithelial neoplasia (PIN) were isolated by laser capture microdissection and subjected to microsatellite analysis of chromosome regions 8p22, 13q14, and 16q23. The frequency of allele alterations in the epithelium was 48% for 8p22, 72% for 13q14, and 37% for 16q23. Slightly higher frequencies of the loss of heterozygosity (LOH) and microsatellite instability in these loci were observed in tumor-associated stroma. Molecular alterations were also found in both epithelial (16–27%) and stromal (8–22%) components in PIN. LOH on chromosomes 16 and 13 in the epithelium was significantly associated with the Gleason score, PC stage, and metastasis into regional lymph nodes. Thus, multiple genetic aberrations occur in the stromal component of PC as frequently as in the tumor epithelium.     

人类第五染色体5q13.3带区一新基因的分子克隆初报

人类第五染色体５ｑ１３．３带区是毛发样白血病、ｒｅｆａｃｔｏｒｙ白血病、骨髓增生不良综合征、卵巢癌、肺癌等恶性肿瘤的共同畸变区域,表明该区域可能存在一个或多个同恶性肿瘤发生有关的基因（癌基因或抑癌基因）．克隆这些有关基因并进一步鉴定,对于阐明恶性肿瘤的发病机理,进而应用于基因治疗具有重要的理论和实践意义．本文报道位于该区域的一个新基因的ｃＤＮＡ克隆及在人体不同组织表达的初步结果．