Aim： To detect the expression of VASA in human ejaculated spermatozoa, and to compare the expression of VASA between normozoospermic men and patients with oligozoospermia. Methods： Ejaculated spermatozoa were collec...Aim： To detect the expression of VASA in human ejaculated spermatozoa, and to compare the expression of VASA between normozoospermic men and patients with oligozoospermia. Methods： Ejaculated spermatozoa were collected from normozoospermic men and patients with oligozoospermia by masturbation, and subsequently segregated through a discontinuous gradient of Percoll to obtain the spermatozoa. Reverse transcription polymerase chain reaction （RT- PCR）, quantitative RT-PCR （QRT-PCR）, immunoflurescence and Western blotting were used to detect the expression of VASA in mRNA and protein levels. Results： VASA mRNA was expressed in the ejaculated spermatozoa. QRT-PCR analysis showed that VASA mRNA level was approximately 5-fold higher in normozoospermic men than that in oligozoospermic men. Immunofluorescence and Western blotting analysis showed that VASA protein was located on the cytoplasmic membrane of heads and tails of spermatozoa, and its expression was significantly decreased in oligozoospermic men, which is similar to the result of QRT-PCR. Conclusion： The expression of VASA mRNA and protein was significantly decreased in the sperm of oligozoospermic men, which suggested the lower expression of the VASA gene might be associated with pathogenesis in some subtypes of male infertility and VASA could be used as a molecular marker for the diagnosis of male infertility.展开更多
Androgen and androgen receptor （AR） play important roles in male spermatogenesis and fertility, yet detailed androgen/AR signals in Sertoli cells remain unclear. To identify AR target genes in Sertoli cells, we anal...Androgen and androgen receptor （AR） play important roles in male spermatogenesis and fertility, yet detailed androgen/AR signals in Sertoli cells remain unclear. To identify AR target genes in Sertoli cells, we analyzed the gene expression profiles of testis between mice lacking AR in Sertoli cells （S-AR-/y） and their littermate wild-type （WT） mice. Digital gene expression analysis identified 2276 genes downregulated and 2865 genes upregulated in the S-AR-/y mice testis compared to WT ones. To further nail down the difference within Sertoli cells, we first constructed Sertoli cell line TM4 with stably transfected AR （named as TM4/AR） and found androgens failed to transactivate AR in Sertoli TM4 and TM4/AR cells. Interestingly, additional transient transfection of AR-cDNA resulted in significant androgen responsiveness with TM4/AR cells showing 10 times more androgen sensitivity than TM4 cells. In the condition where maximal androgen response was demonstrated, we then analyzed gene expression and found the expression levels of 2313 genes were changed more than twofold by transient transfection of AR-cDNA in the presence of testosterone. Among these genes, 603 androgen-/ AR-regulated genes, including 164 upregulated and 439 downregulated, were found in both S-AR-/y mice testis and TM4/AR cells. Using informatics analysis, the gene ontology was applied to analyze these androgen-/AR-regulated genes to predict the potential roles of androgen/AR in the process of spermatogenesis. Together, using gene analysis in both S-AR-/y mice testis and TM4/AR cells may help us to better understand the androeen/AR signals in Sertoli cells and their influences in spermatogenesis.展开更多
This study assesses the long-term outcomes in Han Chinese patients with clinical stage I non-seminomatous germ cell testicular cancer （CSI NSGCT） treated with surveillance, retroperitoneal lymph node dissection （RP...This study assesses the long-term outcomes in Han Chinese patients with clinical stage I non-seminomatous germ cell testicular cancer （CSI NSGCT） treated with surveillance, retroperitoneal lymph node dissection （RPLND） and adjuvant chemotherapy. We retrospectively evaluated 89 patients with a mean age of 26.5 years. After orchiectomy, 37 patients were treated with surveillance, 34 underwent RPLND and 18 were managed with chemotherapy. The overall survival rate, the recurrence-free survival rate and the risk factors were evaluated. The median follow-up length was 92 months （range： 6-149 months）. Thirteen of the 89 patients （14.6%） had relapses, and one died by the evaluation date. The overall survival rate was 98.9%. The cumulative 4-year recurrence-free rates were 80.2%, 92.0% and 100% for the surveillance, RPLND and chemotherapy groups, respectively. The disease-free period tended to be briefer in patients with a history of cryptorchidism and those with stage Is. Therefore, surveillance, RPLND and adjuvant chemotherapy might be reliable strategies in compliant patients with CSI NSGCT. Surveillance should be recommended for patients with the lowest recurrence rate, especially those without lymphovascular invasion. This study might aid the establishment of a standard therapy for CSI NSGCT in China.展开更多
Objective To identify genes that involved in spermatogenesis. Methods In order to screen the testis-specific genes, testes cDNA samples from BALB/c mice of different postnatal days （days 4, 9, 18, 35, 54 and 6 months...Objective To identify genes that involved in spermatogenesis. Methods In order to screen the testis-specific genes, testes cDNA samples from BALB/c mice of different postnatal days （days 4, 9, 18, 35, 54 and 6 months） were performed with mouse whole genome Affymetrix chip. The characteristics of the selected gene were analyzed by various bioinformatic tools. The expression profile of the selected gene was identified by RT-PCR. Results By analyzing the hybridization signals, a gene with a differential expression in the developmental stages of testis was identified. This gene was designated as TSF22. The full length cDNA of 1 597 bp contained an open reading frame of 570 bp which encoded a putative protein of 190 amino acids and a molecular weight of 22.106 kD. RT-PCR analysis revealed that TSF22 mRNA was exclusively expressed in mice testis. Conclusions TSF22, functions as a testis-specific transcription factor, may play important roles during spermatogenesis.展开更多
Objective To analyze the expression of SPACA4 in human and mice. Methods Testes cRNA samples from Balb/c mice of different postnatal days were performed with mouse affymetrix chip to screen the expression of SPACA4 in...Objective To analyze the expression of SPACA4 in human and mice. Methods Testes cRNA samples from Balb/c mice of different postnatal days were performed with mouse affymetrix chip to screen the expression of SPACA4 in mice. Sub-quantitative RT-PCR and bioinformatic tools were used here to describe the expression profile of SPACA4 in mice and human. Results The results of gene chip analysis indicated that the expression of mSPACA4 began after d 35 of postnatal testis in mice. Sub-quantitative RT-PCR assay showed that SPACA4 gene expressed exclusively in mouse and human testis, and mouse mSPACA4 gene expressed after d 35 of postnatal testis that was consistency with the results of gene chip analysis. By bioinformatics analysis, mSPACA4 is located in cell membrane （34.8%） or plasma membrane （34.8%）, the signal peptide cleavage site between position 19 and 20 amino acids, transmembrane region between 2-20 and 101-126 amino acids, respectively, on mSPACA4 protein. Conclusion mSPACA4 and hSPACA4 were testis-specific genes, and the expression of mSPACA4 begins after d 35 of postnatal testis in mice. SPACA4 is a candidate for targeting in a sperm-based contraceptive vaccine.展开更多
Aim:To investigate the stepwise development and germ cell gene expression in allografted neonatal mouse testes and the differentiation of immature human testicular cells in xenografted human testes.Methods:Immunodefic...Aim:To investigate the stepwise development and germ cell gene expression in allografted neonatal mouse testes and the differentiation of immature human testicular cells in xenografted human testes.Methods:Immunodeficient nude mice were used as hosts for allografting of neonatal mouse testes and xenografting of human fetal testicular tissues. Stepwise development and stage-specific gene expression of germ cells in allografts were systematically evaluated and parallel compared with those in intact mice by periodically monitoring the graft status with measurement of graft weight,histological analysis and determination of five stage-specific genes.Human testicular tissues from 20 and 26 weeks fetuses were used for the xenografting study.Histological analysis of xenografts was performed 116 and 135 d after the grafting procedure.Results:In the allografting study,progressive increase in tissue volume and weight as well as in tubule diameter in grafts was observed;the appearance time of various germ cells in seminiferous tubules,including spermatogonia,spermatocytes,round and elongate spermatids and sperm,was comparable with that in intact donors;the initiation of gene transcription in grafts showed a similar trend as in normal mice.Graft weight ceased to increase after 7-8 weeks and degradation of grafts was observed after 5 weeks with progressive damage to seminiferous epithelium.In the xenografting study using immature human testicular tissues,graft survival and development was indicated by increasing graft weight,Sertoli cells differentiation into advanced stage,germ cells migration and location to the basal lamina and formation of a niche-like structure.Conclusion:The developmental course and gene expression pattern of germ cells in allografts were similar to those in intact mice.The best time point for retrieval of mouse sperm from grafts was 5-7 weeks after grafting procedure.An accelerated development of immature human testicular cells could be achieved by ectopic xenografting of human testes.(Asian J Androl 2006 Jul;8:393-403)展开更多
基金We would like to thank Mr Jian-Rong Zhang, Mr Li-Bing Zhang and Dr Zhen-Dong Yu for technical assistance. This work was supported by grants from the National Natural Science Foundation of China （No. 30500543）, Ministry of Education ＂985 project＂ （No. 985-2-054-29）, and Shenzhen Foundation of Science ＆ Technology （JH200505270413B）.
文摘Aim： To detect the expression of VASA in human ejaculated spermatozoa, and to compare the expression of VASA between normozoospermic men and patients with oligozoospermia. Methods： Ejaculated spermatozoa were collected from normozoospermic men and patients with oligozoospermia by masturbation, and subsequently segregated through a discontinuous gradient of Percoll to obtain the spermatozoa. Reverse transcription polymerase chain reaction （RT- PCR）, quantitative RT-PCR （QRT-PCR）, immunoflurescence and Western blotting were used to detect the expression of VASA in mRNA and protein levels. Results： VASA mRNA was expressed in the ejaculated spermatozoa. QRT-PCR analysis showed that VASA mRNA level was approximately 5-fold higher in normozoospermic men than that in oligozoospermic men. Immunofluorescence and Western blotting analysis showed that VASA protein was located on the cytoplasmic membrane of heads and tails of spermatozoa, and its expression was significantly decreased in oligozoospermic men, which is similar to the result of QRT-PCR. Conclusion： The expression of VASA mRNA and protein was significantly decreased in the sperm of oligozoospermic men, which suggested the lower expression of the VASA gene might be associated with pathogenesis in some subtypes of male infertility and VASA could be used as a molecular marker for the diagnosis of male infertility.
基金This work was supported by the National Natural Science Foundation of China （no. 30971636）, and the George H. Whipple Professorship Endowment, and National Science Council, Talwan, China （96-2314-B-182A-023-MY2 and 97- 2314-B-182A-077-MY3）. Supplementary Information accompanies the paper on Asian lournal of Andrology website （http：Hwww.nature.com/aja）.
文摘Androgen and androgen receptor （AR） play important roles in male spermatogenesis and fertility, yet detailed androgen/AR signals in Sertoli cells remain unclear. To identify AR target genes in Sertoli cells, we analyzed the gene expression profiles of testis between mice lacking AR in Sertoli cells （S-AR-/y） and their littermate wild-type （WT） mice. Digital gene expression analysis identified 2276 genes downregulated and 2865 genes upregulated in the S-AR-/y mice testis compared to WT ones. To further nail down the difference within Sertoli cells, we first constructed Sertoli cell line TM4 with stably transfected AR （named as TM4/AR） and found androgens failed to transactivate AR in Sertoli TM4 and TM4/AR cells. Interestingly, additional transient transfection of AR-cDNA resulted in significant androgen responsiveness with TM4/AR cells showing 10 times more androgen sensitivity than TM4 cells. In the condition where maximal androgen response was demonstrated, we then analyzed gene expression and found the expression levels of 2313 genes were changed more than twofold by transient transfection of AR-cDNA in the presence of testosterone. Among these genes, 603 androgen-/ AR-regulated genes, including 164 upregulated and 439 downregulated, were found in both S-AR-/y mice testis and TM4/AR cells. Using informatics analysis, the gene ontology was applied to analyze these androgen-/AR-regulated genes to predict the potential roles of androgen/AR in the process of spermatogenesis. Together, using gene analysis in both S-AR-/y mice testis and TM4/AR cells may help us to better understand the androeen/AR signals in Sertoli cells and their influences in spermatogenesis.
文摘This study assesses the long-term outcomes in Han Chinese patients with clinical stage I non-seminomatous germ cell testicular cancer （CSI NSGCT） treated with surveillance, retroperitoneal lymph node dissection （RPLND） and adjuvant chemotherapy. We retrospectively evaluated 89 patients with a mean age of 26.5 years. After orchiectomy, 37 patients were treated with surveillance, 34 underwent RPLND and 18 were managed with chemotherapy. The overall survival rate, the recurrence-free survival rate and the risk factors were evaluated. The median follow-up length was 92 months （range： 6-149 months）. Thirteen of the 89 patients （14.6%） had relapses, and one died by the evaluation date. The overall survival rate was 98.9%. The cumulative 4-year recurrence-free rates were 80.2%, 92.0% and 100% for the surveillance, RPLND and chemotherapy groups, respectively. The disease-free period tended to be briefer in patients with a history of cryptorchidism and those with stage Is. Therefore, surveillance, RPLND and adjuvant chemotherapy might be reliable strategies in compliant patients with CSI NSGCT. Surveillance should be recommended for patients with the lowest recurrence rate, especially those without lymphovascular invasion. This study might aid the establishment of a standard therapy for CSI NSGCT in China.
基金This study was supported by grants from Chinese Natural Science Funds(No.30471728,No.30500543)Natural Science Funds of Guangdong Province(No.04007303)
文摘Objective To identify genes that involved in spermatogenesis. Methods In order to screen the testis-specific genes, testes cDNA samples from BALB/c mice of different postnatal days （days 4, 9, 18, 35, 54 and 6 months） were performed with mouse whole genome Affymetrix chip. The characteristics of the selected gene were analyzed by various bioinformatic tools. The expression profile of the selected gene was identified by RT-PCR. Results By analyzing the hybridization signals, a gene with a differential expression in the developmental stages of testis was identified. This gene was designated as TSF22. The full length cDNA of 1 597 bp contained an open reading frame of 570 bp which encoded a putative protein of 190 amino acids and a molecular weight of 22.106 kD. RT-PCR analysis revealed that TSF22 mRNA was exclusively expressed in mice testis. Conclusions TSF22, functions as a testis-specific transcription factor, may play important roles during spermatogenesis.
基金This study was supported by grants from National Natural Science Foundation of China (No.30700824, No.30770810)973 Program(2008CB517412)Guangdong Natural Science Foundation (No.7008952)
文摘Objective To analyze the expression of SPACA4 in human and mice. Methods Testes cRNA samples from Balb/c mice of different postnatal days were performed with mouse affymetrix chip to screen the expression of SPACA4 in mice. Sub-quantitative RT-PCR and bioinformatic tools were used here to describe the expression profile of SPACA4 in mice and human. Results The results of gene chip analysis indicated that the expression of mSPACA4 began after d 35 of postnatal testis in mice. Sub-quantitative RT-PCR assay showed that SPACA4 gene expressed exclusively in mouse and human testis, and mouse mSPACA4 gene expressed after d 35 of postnatal testis that was consistency with the results of gene chip analysis. By bioinformatics analysis, mSPACA4 is located in cell membrane （34.8%） or plasma membrane （34.8%）, the signal peptide cleavage site between position 19 and 20 amino acids, transmembrane region between 2-20 and 101-126 amino acids, respectively, on mSPACA4 protein. Conclusion mSPACA4 and hSPACA4 were testis-specific genes, and the expression of mSPACA4 begins after d 35 of postnatal testis in mice. SPACA4 is a candidate for targeting in a sperm-based contraceptive vaccine.
文摘Aim:To investigate the stepwise development and germ cell gene expression in allografted neonatal mouse testes and the differentiation of immature human testicular cells in xenografted human testes.Methods:Immunodeficient nude mice were used as hosts for allografting of neonatal mouse testes and xenografting of human fetal testicular tissues. Stepwise development and stage-specific gene expression of germ cells in allografts were systematically evaluated and parallel compared with those in intact mice by periodically monitoring the graft status with measurement of graft weight,histological analysis and determination of five stage-specific genes.Human testicular tissues from 20 and 26 weeks fetuses were used for the xenografting study.Histological analysis of xenografts was performed 116 and 135 d after the grafting procedure.Results:In the allografting study,progressive increase in tissue volume and weight as well as in tubule diameter in grafts was observed;the appearance time of various germ cells in seminiferous tubules,including spermatogonia,spermatocytes,round and elongate spermatids and sperm,was comparable with that in intact donors;the initiation of gene transcription in grafts showed a similar trend as in normal mice.Graft weight ceased to increase after 7-8 weeks and degradation of grafts was observed after 5 weeks with progressive damage to seminiferous epithelium.In the xenografting study using immature human testicular tissues,graft survival and development was indicated by increasing graft weight,Sertoli cells differentiation into advanced stage,germ cells migration and location to the basal lamina and formation of a niche-like structure.Conclusion:The developmental course and gene expression pattern of germ cells in allografts were similar to those in intact mice.The best time point for retrieval of mouse sperm from grafts was 5-7 weeks after grafting procedure.An accelerated development of immature human testicular cells could be achieved by ectopic xenografting of human testes.(Asian J Androl 2006 Jul;8:393-403)
