吸烟与男性生殖细胞的基因损伤

辅助生殖技术的应用 ,使检测人类配子的临床结局、减数分裂的成熟过程和基因完整性的异常成为可能。吸烟可引起生殖细胞的基因损伤 ,但此损伤在减数分裂期间可以修复 ;而在已射出的精子中 ,修复基因损伤的能力极大地下降。吸烟能改变精母细胞减数分裂的纺锤体 ,导致染色体错误 ,从而影响生殖结局。     

1例45,X不育症男性的分子遗传学特征

目的从1例45,X不育症男性的分子遗传学特征探讨无精子症或少弱精子症患者的遗传缺陷与精子生成障碍的关系,分析部分患者不育原因是否可能与染色体异常有关。方法采用常规染色体、FISH及分子检测技术,对该患者进行遗传学分析。结果常规染色体检测显示该患者核型为45,X,dic(Y;13)(q12;pl1)。且易位的染色体中只有一个着丝粒具有活性,其中72%的细胞Y染色体着丝粒失活,13号染色体着丝粒收缩,具有活性,其余28%的细胞13号染色体着丝粒失活。FISH与分子遗传学分析证实患者13号短臂增加部分来源于Y染色体,Y断裂位点位于q12异染色质区。近端包括AZF和SRY在内的所有Y常染色体部分与13号染色体短臂融合,形成一条双着丝粒染色体。其13号染色体与Y染色体功能基因区未发生任何缺失。核型描述为45,X,dic(Y;13)(q12;p11).ish(SRY+,DYZ3+,GLP13+)。结论本例遗传学分析结果提示部分男性不育症可能是由Y/常染色体易位造成减数分裂异常,导致生精受阻,精子生成大量减少而引发不育。     

吸烟与男性生殖细胞的基因损伤

辅助生殖技术的应用，使检测人类配子的临床结局、减数分裂的成熟过程和基因完整性的异常成为可能。吸烟可引起生殖细胞的基因损伤，但此损伤在减数分裂期间可以修复；而在已射出的精子中，修复基因损伤的能力极大地下降。吸烟能改变精母细胞减数分裂的纺锤体，导致染色体错误，从而影响生殖结局。     

1例环状22号染色体综合征致无精子症并文献复习

目的:探讨1例患有环状22号染色体综合征的无精子症患者的临床表型和遗传学特征。方法:收集1例环状22号染色体综合征患者的临床信息,结合文献加以分析。结果:患者身材矮小,体检双侧睾丸小,质地软,精液检查示无精子症。染色体核型为46,XY,r(22)(p11,q25),性激素示睾酮低下,睾丸大体病理示组织脆、易拉断,病理镜检示生精小管内支持细胞及生殖细胞数量均减少,细胞层次变薄,生殖细胞均为精原细胞,未见精母细胞和精子细胞,完全无精子发生。部分生精小管的管壁可见间质轻度纤维化。结论:环状22号染色体综合征的临床表型基本正常,但这种遗传异常使患者的睾丸组织严重受损,生精过程被阻滞而导致了无精子症。     

比较基因组杂交研究非综合征性唇腭裂遗传变异

目的 了解非综合征性唇腭裂(nonsyndromic cleft lip with cleft palate,NSCLP)遗传学改变的特征.方法 应用比较基因组杂交技术研究7例NSCLP患者基因组的不平衡即遗传物质的丢失或扩增情况.结果 6、7、10、13、14、16、20、22号染色体出现DNA拷贝数缺失.5、15、18、19号染色体发生DNA拷贝数扩增.出现高频率的DNA拷贝数缺失的染色体是13q(71.4%).结论 NSCLP患者的染色体均出现DNA拷贝数的变化,以DNA拷贝数的缺失为主,提示NSCLP的发生主要是其抑制基因丢失所造成的,13q极可能存在相关的NSCLP抑制基因.     

男性生精障碍相关基因的研究进展

精子发生经历精原干细胞的增殖分化、精母细胞的减数分裂和精子形成等3个主要阶段，最终形成精子。在这个漫长的过程中受到诸多有序表达的基因的调控；如果染色体核型异常，与精子生成相关的基因缺失、突变或表达异常都可能导致精子生成障碍，形成少精子症或无精子症，导致男性不育。引起男性不育的基因很多，包括影响精子发生和精子成熟等基因，     

比较基因组杂交研究瘢痕疙瘩遗传变异

目的 了解瘢痕疙瘩遗传学改变的特征。方法 应用比较基因组杂交技术研究6例瘢痕疙瘩基因组的不平衡即遗传物质的丢失或扩增情况。结果 瘢痕疙瘩出现高频率的DNA拷贝数缺失的染色体是1p(66．7％)、16号染色体(83．3％)、20号染色体(83．3％)、22号染色体(83．3％)，其最小重叠区分别为1pter-32．2、16p13．2-p11．1、20q11．1-q13．2、16p13．2-p11．1，未发现特异区域的DNA拷贝数的高频率扩增。结论 1p、16号染色体、20号染色体、22号染色体极可能存在相关的瘢痕疙瘩抑制基因，这些基因的丢失可能参与了瘢痕疙瘩的发生、发展。     

用双色荧光原位杂交技术检测罗伯逊易位携带者的精子染色体

目的 :探讨用双色荧光原位杂交技术 (D FISH)检测罗伯逊易位携带者的精子染色体的实验方法和应用价值。　方法 :采用荧光原位杂交 (FISH)技术 ,以Biotin标记的 13q 14 .3特异性探针和以Digoxigenin标记的 14 q11.1特异性探针对 2例罗伯逊易位携带者精子标本进行原位杂交 ,并统计精子间期核 13、14号染色体的杂交信号颗粒数量。　结果 :在显微镜下可见精子头部有以Biotin标记的 13q 14 .3特异性探针显示 1个绿色杂交信号 ,以Digoxi genin标记的 14 q11.1特异性探针显示 1个红色杂交信号 ,间期核背景经DAPI复染显示蓝色 ;共统计 30 0 0个精子间期核 ,显示 1个绿色 1个红色杂交信号的精子为 13q/ 14q ,占39.33% ;显示 1个绿色 2个红色杂交信号为 13q/ 14q ,14q ,占 11.5 7% ;仅显示 1个绿色杂交信号为 13q/ ,占 9.2 7% ;显示 2个绿色 1个红色杂交信号为 13q ,13q/ 14 q ,占12 .87% ;仅显示 1个红色杂交信号为 / 14 q ,占9.87% ;显示 2个绿色 2个红色杂交信号为 13q ,13q/ 14q ,14 q ,占12 .2 6 %。　结论 :用双色荧光原位杂交技术检测染色体结构异常患者的精子 ,可以分析其减数分裂过程中染色体分离规律 ,在人类生殖如显微授精和植入前胚胎遗传学诊断等方面具有非常广泛的应用价值     

罗氏易位伴少弱精子症来源的胚胎干细胞向生殖细胞诱导分化的研究

目的:建立罗氏易位伴少弱精子症来源的人胚胎干细胞(ESC),利用体外诱导ESC向生殖细胞分化作为模型,评估辅助生殖技术子代的潜在风险。方法:利用罗氏易位伴少弱精子症患者夫妇捐赠的胚胎,通过分离囊胚内细胞团,培养、传代、扩增,建立ESC CCRM16,其核型为46,XY,+14,rob(13;14)(q10;q10);添加2 mol/L维甲酸体外诱导分化,分析其向生殖细胞分化过程及其相关基因的表达,并与遗传背景完全正常的ESC CCRM23相比较。结果:CCRM16表达OCT4,TRA-1-81,NANOG及SSEA4多能性标记基因,体外能形成拟胚体(EB),体内、体外都能向3个胚层分化。添加维甲酸可直接诱导ESC向生殖细胞分化。原始生殖细胞标志基因DAZL和减数分裂标记基因SCP3的表达水平在CCRM16中较正常夫妇捐赠胚胎建立的ESC CCRM23明显减少。结论:核型为46,XY,+14,rob(13;14)(q10;q10)CCRM16具有ESC典型特征,但其向早期精子分化的过程异常。体外诱导CCRM16向精子分化作为研究模型,可应用于评估少弱精子症患者辅助生殖出生子代的健康风险。     

1例与Y染色体畸变相关的无性腺症、矮身材和蹠骨畸形

目的:报告1例与Y染色体畸变相关的无性腺症、矮身材和蹠骨畸形的罕见病例。方法:对患者进行常规的染色体核型分析;选用X/Y着丝粒探针和Y染色体上性别决定基因(SRY)探针对患者进行原位荧光杂交(FISH);对SRY基因的HMG进行序列分析;应用多重PCR检测Y染色体微缺失;进行矮身材同源框基因(SHOXY)微卫星多态性分析;进行人绒毛膜促性腺激素(HCG)兴奋试验。结果:染色体核型分析显示患者核型为46,X del(Y)(q11.23);患者FISH显示只有1个X染色体和Y染色体杂交信号,但SRY有2个信号,分别位于Y染色体的短臂和长臂上。结合染色体核型和FISH结果,患者核型疑为46,X del(Y)(q11.23)(pter→p11.3::q11.23→p11.3:).ish del(Y)(SRY++,DYZ3+);Y染色体微缺失检测发现有AZFc的缺失;患者的SRY的HMG序列分析结果未显示有突变;SHOXY微卫星多态性分析未发现父亲来源的基因有缺失;兴奋试验显示HCG刺激后,雄激素、雌激素无升高反应,提示为无性腺症。结论:患者的无性腺症、矮身材和蹠骨畸形与Y染色体结构畸变有关,推测与SRY基因,SHOXY基因相关。     

Meiotic pairing error in an infertile male bearing reciprocal deletion of chromosome 13

A series of complex processes takes place during the first meiotic division, including pairing, synapsis, recombination, and segregation of homologous chromosomes. When any of these processes is altered, cellular checkpoints arrest the progression of meiosis and induce cell loss, causing a severe reduction in fertility, or even sterility. In this study, we report on a 29-year-old, healthy, but severe oligozoospermic male with a supernumerary, ring-neocentric 13q12.3 → 13q22 chromosome and a reciprocal deletion, which interfere with the meiotic pairing of chromosomes 13, causing spermatogenesis failure.     

广西地区乙肝病毒/黄曲霉毒素双暴露因素下肝细胞癌染色体遗传学改变的初步研究

目的 探讨广西地区人群乙肝病毒/黄曲霉毒素(HBV/AFB1)双暴露相关性肝细胞癌(HCC)染色体遗传学改变的特点.方法 32例HCC的癌组织,运用微阵列比较基因组杂交技术(Array CGH)检测其22对染色体的变化.结果 (1)32例HCC中,大部分的染色体拷贝数都有不同程度的变化.发生扩增的区段有1 q、7q、8q,其中1q、8q为高频扩增区段.发生缺失区段有1 p、4q、8p、9p、13q、14q、16p、16q、17p、18q、19p、Y,其中1p、4q、8p、16q、17p、19p为高频缺失区段;(2)同时,还存在着若干DNA拷贝数扩增或缺失的小区段.缺失显著的小区段有:2p25.1-p25.2、3q22.3-q23、7p14.1-p14.3.扩增显著的小区段有:9p13.2-9p21;(3)聚类分析发现:13q缺失发生率在HBsAg(+)/AFB1(+)、HBsAg(+)/AFB1(-)、HBsAg(-)/AFB1(+)、HBsAg(-)/AFB1(-)4个亚组中呈依次递减(x2=6.452,P＜0.05).4p在HBsAg(+)/AFB1(-)组中以扩增为主,而在HBsAg(-)/AFB1(+)组与HBsAg(-)/AFB1(-)组则以缺失为主.19q在HBsAg(+)/AFB1(+)组中以扩增为主,在HBsAg(-)/AFB1(+)组与HBsAg(-)/AFB1(-)组中以缺失为主.结论 广西地区肝癌染色体遗传学改变具有多样性,其中19p、2p25.1-25.2、3q22.3-q23的缺失及9p13.2-9p21的扩增可能为该地区肝癌特有的遗传学特征之一.13q的缺失可能与该地区乙肝病毒/黄曲霉毒素双因素的协同作用有关.     

Long arm deletion of chromosome 10 in a boy with monorchidism

We report on a boy with long-arm deletion of chromosome 10 and compare this case to 10 previously reported patients. He had right cryptorchidism and absence of the left testis, but the size of his penis was normal. Cytogenetic analysis of the case showed the deficiency of 10q26.1-26.3 and the presence of 10qter. Four of 10 previously reported patients had an intersex phenotype, and all others had combinations of cryptorchidism, micropenis and hypospadias. These facts indicate that the terminal of chromosome 10q is strongly associated with abnormal male development.     

Detection of chromosome 15 deletion in Prader-Willi syndrome using fluorescence in situ hybridization

Deletion of chromosome 15 was investigated in 6 patients with Prader-Willi syndrome (PWS) using chromosome and fluorescence in situ hybridization (FISH) analysis. Although chromosome analysis using G-banding methods revealed the deletion of 15q11-q13 in only 3 cases, staining by FISH using D15S11 and/or small nuclear ribonucleoprotein polypeptide N (SNRPN) probes detected chromosome 15 deletion in all cases. It would appear that FISH analysis is an effective diagnostic test for the detection of chromosome 15 deletion in patients with PWS.     

Chromosomal Rearrangements in Three Infertile Men

Summary:  Three chromosomal rearrangements: a balanced reciprocal translocation, t(14;10) (q22;q13), a Y-autosome translocation, t(Y;16) (q11;p13) and a deleted Y chromosome, Yq- were detected among 100 infertile men. The autosomal translocation, associated with oligozoospermia was found to be familial with various effects on the female carriers and the proband's father. The patients with the chromosome Y abberations were found to be azoospermic and might have lost the genes necessary for normal sperma-togenesis.
Zusammenfassung:  Unter 100 infertilen Männern wurden drei Chromosomenneuan-ordnungen entdeckt: eine balancierte reziproke Translokation, t(14;10) (q22;q13), eine Y-autosome Translokation, t(Y;16) (q11;p13) und eine Deletion des Y-Chromosoms, Yq-. Die autosomale Translokation bei Oligozoospermie zeigte sich familiär mit verschiedenen Auswirkungen bei den weiblichen Überträgern und dem Vater des Probanden. Die Patien-ten mit chromosomalen Y-Aberrational wiesen eine Azoospermie auf und scheinen die zur normalen Spermatogenese notwendigen Gene verloren zu haben.     

KLF5 is frequently deleted and down-regulated but rarely mutated in prostate cancer

BACKGROUNDS: Previous studies mapped a region at the q21 band of chromosome 13 (13q21), which is frequently deleted in various human cancers including prostate cancer, suggesting the existence of a tumor suppressor gene at 13q21. The target gene of deletion in prostate cancer, however, has not been identified at present. METHODS: We examined four non-neoplastic and 18 neoplastic prostatic cell lines or xenografts. Homozygous/hemizygous deletion was detected by assays of duplex PCR and real-time PCR. Expression levels of genes were determined by the methods of RT-PCR, real time PCR, and northern blot analysis. Mutations of KLF5 were detected by the approaches of single strand conformational polymorphism (SSCP) and direct sequencing. For the detection of promoter methylation, Southern blotting of genomic DNA and restriction digestion or SSCP analysis of methylation specific PCR products were used. Finally, an expression plasmid of KLF5 was introduced into prostate cancer cell lines with reduced KLF5 expression to investigate colony formation for cell growth. RESULTS: A 2-Mb region of homozygous deletion at 13q21 was detected in the LUCaP70 xenograft of prostate cancer. This region of deletion was further narrowed to 142 Kb by a hemizygous deletion in the NCI-H660 cell line. KLF5 was identified as the only complete gene in the smallest region of deletion. Quantitative deletion of KLF5 genome occurred in six of the 18 (33%) prostate cancer xenografts/cell lines. Each of the six samples with deletion also showed loss of expression for KLF5, suggesting that hemizygous deletion is one mechanism for loss of KLF5 expression. In total, 16 of the 18 cases (89%) showed loss of KLF5 expression at different degrees. In contrast, mutations and promoter methylations were not detected in any of the samples. Functionally, restoration of KLF5 in DU 145 and 22Rv1 cell lines significantly inhibited their growth in vitro. CONCLUSIONS: Frequent genomic deletion and loss of expression as well as cell growth suppression indicate that KLF5 is a reasonable candidate for the tumor suppressor gene at 13q21 in prostate cancer. Mutation and promoter methylation are not common mechanisms for the inactivation of KLF5 in prostate cancer.     

Mapping of metastasis suppressor genes for prostate cancer by microcell-mediated chromosome transfer

Aim: To identify the metastasis suppressor genes for prostate cancer. Methods: A copy of human chromosomes was introduced into the highly metastatic Dunning R-3327 rat prostate cancer cells by the use of microcell-mediated chromosome transfer. Relationships between the size of human chromosomes introduced into microcell hybrid clones and the number of lung metastases produced by the clones were analyzed to determine which part of human chromosomes contained the metastasis suppressor gene (s) for prostate cancer. To determine portions of human chromosomes introduced, G-banding chromosomal analysis, fluorescence in situ hybridization analysis, and polymerase chain reaction analysis were performed. Results: Each of microcell hybrid clones containing human chromosomes 7, 8, 10, 11, 12, or 17 showed decreased ability to metastasize to the lung without any loss of ttmaorigenicity. This demonstrates that these human chromosomes contain metastasis suppressor genes for prostate cancer. Spontaneous deletion of portions of human chromosomes was observed in the human chromosome 7, 10, 11, 12, and 17 studies. In the human chromosome 8 study, irradiated microcell-mediated chromosome transfer was performed to enrich chromosomal ann deletions of human chromosome 8. Molecular and cytogenetic analyses of microcell hybrid clones demonstrated that metastasis suppressor genes on human chromosomes were located on 7q21-22, 7q31.2-32, 8p21-12, 10q11-22, 11p13-11.2, 12p11-q13, 12q24-ter, and 17pter-q23. KAI1 and MKK4/SEKI were identified as metastasis suppressor genes from 11p11.2 and 17p12, respectively. Conclusion: This assay system is useful to identify metastasis suppressor gene (s) for prostate cancer.     

Genome-wide scan of brothers: replication and fine mapping of prostate cancer susceptibility and aggressiveness loci

BACKGROUND: Substantial evidence suggests that genetic factors play an important role in both the risk of prostate cancer and its biologic aggressiveness. Here we investigate prostate cancer susceptibility and aggressiveness with genome-wide linkage analyses of affected brothers. METHODS: We first undertook a new genome-wide linkage study of 259 brothers with prostate cancer. Our analyses tested whether the proportion of marker alleles shared by brothers was correlated with disease status or Gleason score. To further clarify 11 linkage regions observed here or previously, we genotyped and analyzed an additional 101 finely spaced markers in the 259 men, and in 594 previously studied brothers, allowing for a pooled genome-wide analysis of 853 affected brothers. RESULTS: In the new study, we detected linkage to prostate cancer on chromosome 16q23 (P = 0.009), replicating previous results, and to chromosome 11q24 (P = 0.001). In the pooled analysis, the 16q23 linkage was strengthened (P = 0.0005), as was our previous linkage to chromosome 16p (P = 0.0001), and we detected linkage to chromosome 2q32 (P = 0.009). When evaluating Gleason score, our new study detected linkage to chromosome 7q32 (P = 0.0009), again replicating previous results, and to chromosomes 5p15 (P = 0.003), 9q34 (P = 0.009), 10q26 (P = 0.03), and 18p11 (P = 0.02). In the pooled analysis of Gleason score, we observed stronger linkage to chromosome 7q32 (P = 0.0002), but slightly weaker linkage to chromosomes 5q33 (P = 0.005) and 19q13 (P = 0.009) than previously reported. In addition, the new linkages to chromosomes 10q26 and 18p11 were strengthened (P = 0.0002 and P = 0.002, respectively). CONCLUSIONS: Our results provide compelling evidence for loci harboring prostate cancer susceptibility and tumor aggressiveness genes, especially on chromosomes 16q23 and 7q32.     

Assignment of the gene for human intra-acrosomal protein SP-10 to the p12----q13 region of chromosome 11.

The human sperm antigen SP-10 is a testis-specific, intra-acrosomal protein associated with the membranes of the acrosomal vesicle. The molecule has been designated a "primary vaccine candidate" by a World Health Organization (WHO) Taskforce on Contraceptive Vaccines. cDNA cloning and sequencing have indicated that SP-10 is encoded by a 795-base-pair (bp) reading frame that predicts a 265-amino acid protein of 28.3 kd. In this study, we used a 634-bp fragment (bp 68 through 700, amino acids 3 through 222) of the SP-10 sequence to probe, by Southern blotting, EcoRI-digested DNA from 33 mouse/human somatic cell hybrids involving 16 unrelated human cell lines and 4 mouse cell lines. The hybrids were characterized by karyotypic analysis and by mapped enzyme markers. The presence or absence of positive human bands was scored on the blots and the percent of concordance and discordance with a specific human chromosome was determined. The DNA probe for SP-10 showed a concordance of 31 and a discordancy of 0 for human chromosome 11, mapping SP-10 unequivocally to this chromosome. The hybrid XER-7 with the 11/X translocation: 11p12 or 11p11----11qter:: Xq11----Xqter and the hybrid EXR-5CSAZ with the X/11 translocation: Xpter----Xq22::11q13----11qter localized the SP-10 gene to the p12----q13 region. The SP-10 locus has been assigned the gene symbol ACRV1 (acrosomal vesicle protein-1).     

Analysis of loss of heterozygosity on chromosomes 10q, 11, and 16 in medulloblastomas

OBJECT: The loss of genetic material from specific chromosome loci is a common feature in the oncogenesis of tumors and is often indicative of the presence of important tumor suppressor genes at these loci. Recent molecular genetic analyses have demonstrated frequent loss of chromosomes 10q, 11, and 16 in medulloblastomas. The aim of this study was to localize the targeted deletion regions on the three aforementioned chromosomes in medulloblastomas. METHODS: Loss of heterozygosity (LOH) was examined on chromosomes 10q, 11, and 16 in a series of 22 primary and two recurrent medulloblastomas by using polymerase chain reaction-based microsatellite analysis. The DNA extracted from the tumors and corresponding normal blood samples were amplified independently in the presence of radioactively labeled microsatellite primers, resolved by denaturing gel electrophoresis and processed for autoradiography. The DNA obtained from control blood samples that displayed allelic heterozygosity at a given microsatellite locus were considered informative. Loss of heterozygosity was inferred when the allelic signal intensity of the tumor sample was reduced by at least 40%, relative to that of the constitutional control. The LOH analysis demonstrated that deletions of chromosomes 10q, 11p, and 16q are recurrent genetic events in the development of medulloblastomas. Three subchromosomal regions of loss have been identified and are localized to the deleted in malignant brain tumors 1 [DMBT1] gene site on chromosomes 10q25, 11p13-11p15.1, and 16q24.1-24.3. CONCLUSIONS: These results indicate that DMBT1 is closely associated with the oncogenesis of medulloblastomas and highlight regions of loss on chromosomes 11p and 16q for further fine mapping and cloning of candidate tumor suppressor genes that are important for the genesis of medulloblastoma.