Testicular malignant Leydig cell tumor: A case report / 中华男科学杂志

<p><b>OBJECTIVE</b>To investigate the clinicopathological features of testicular malignant Leydig cell tumor (TMLCT) and improve the non-invasive diagnosis of the disease.</p><p><b>METHODS</b>We retrospectively analyzed the clinicopathological data on a case of TMLCT, detected the circulating tumor cells (CTC) in the peripheral venous blood, and reviewed the related literature.</p><p><b>RESULTS</b>The patient, a 47-year-old male, underwent radical orchidoepididymectomy under general anesthesia. Postoperative pathology confirmed the lesion to be TMLCT, which was mainly composed of Leydig cells and suspected with vessel carcinoma embolus. Immunohistochemistry showed the tumor cells to be positive for α-inhibin, Ki67, CD30, vimentin, EMA, and PLAP, but negative for CK, CK7, S100, CD10, SMA, Des, AFP, hCG, CEA, CK19, CD117, Oct-4, LCA, CD20, Pax-5, CD3, and CD43. Two CTCs were detected in the peripheral venous blood. The patient received 3 courses of chemotherapy for retroperitoneal multiple lymph nodes metastasis post-operatively. Subsequent CT imaging manifested no obvious reduction of the retroperitoneal lymph nodes and consequently the patient again underwent retroperitoneal lymphadenectomy and cryoablation. At 8 months after treatment, CT examination revealed notably enlarged retroperitoneal lymph nodes with the right adrenal gland evidently invaded.</p><p><b>CONCLUSION</b>TMLCT is an extremely rare sex-gonad stromal tumor with high malignancy and poor prognosis, and CTCs may be used for its early diagnosis and prognostic prediction.</p>

Updated genomics of testicular germ cell tumor / 中华男科学杂志

Testicular germ cell tumor (TGCT) is a most common testicular malignancy with an increasing incidence, and its pathogenesis and mechanisms are not yet clear. The next generation sequencing has become the main tool to uncover the underlying mechanisms of TGCT. The differential gene expressions, gene mutation, predisposing gene-dominated signaling pathways, and changes of the relevant genes in the sex chromosome are largely involved in the occurrence and development of TGCT. Studies on the genomics of TGCT contribute a lot to identifying the pivotal pathogenic genes and paving a theoretical ground for the early screening and targeted therapy of TGCT. This paper summarizes the advances in the studies of the genomics of TGCT so as to reveal thetmechanisms of the disease at the genetic level.

UBE2B gene and male infertility: an update / 中华男科学杂志

Male infertility is a worldwide problem, and about 15% of the cases are associated with spermatogenesis-related gene mutation. The mammalian gene UBE2B is the homolog of the RAD6 gene of yeast, belonging to the ubiquitin proteasome system and playing an important role in spermatogenesis. Mice lacking the UBE2B gene are infertile, with reduced sperm motility, increased morphologically abnormal sperm, and inhibited meiosis of spermatogonia. Accumulated evidence shows that UBE2B gene mutants and single nucleotide polymorphisms are associated with male infertility. This article reviews the relation between the UBE2B gene and male infertility, offering some theoretical evidence for the diagnosis and treatment of male infertility.

Mouse-induced pluripotent stem cells have the potential to differentiate into induced primordial germ cells / 中华男科学杂志

<p><b>OBJECTIVE</b>To investigate whether mouse-induced pluripotent stem (iPS) cell line IP14D-1 has the potential to differentiate into induced primordial germ cells (iPGCs), and to explore the changes in the expression of iPGCs-differentiation associated genes and their possible mechanisms.</p><p><b>METHODS</b>Undifferentiated IP14D-1 was cultured to proliferate and then differentiated to form 4-, 7- and 9-day-old induced embryoid bodies (iEBs) in vitro, respectively. RT-PCR and immunofluorescence were used to detect the expressions of Lin28, Blimpl, Stra8 and Mvh, as well as the localization of the corresponding protein in iEBs.</p><p><b>RESULTS</b>The expression of Blimpl was higher than that of Lin28 in the undifferentiated IP14D-1 and mouse embryonic stem cells (mESCs). Mvh and Stra8 as well as mESCs and EBs were also expressed in IP14D-1 and iEBs, but with no significant differences. The expression of Lin28 was gradually increased in the IP14D-1-derived iEBs from 4 to 7 days, but decreased at 9 days, and the expression of Blimp1 was gradually reduced with the prolonged growing time of iEBs.</p><p><b>CONCLUSION</b>A stable system was established for the culture and differentiation of IP14D-1 and IP14D-1-derived iEBs. The expressions of Lin28, Blimp1, Mvh and Stra8 were not significantly different between the undifferentiated IP14D-1 and mESCs, nor were the expressions of Mvh and Stra8 between iEBs and EBs. IP14D-1 and iEBs had the potential to differentiate into iPGCs, which increased in number in the 7-day-old iEBs, and the expression of iPGC-differentiation associated Lin28 became lower in the older iEBs.</p>

Y chromosome microdeletion and male infertility: past, present and future / 中华男科学杂志

The spermatogenesis failure with a genetic defect is one of the major causes of male infertility. The Y chromosome is considered a lack of important functional genes. It was the discovery of the sex determining region Y that rekindled scientists'attention to the Y chromosome. The successful sequencing of the Y chromosome uncovered its actual structure and the molecular base of its microdeletion. Of the 220 Y chromosome genes (104 coding genes, 111 pseudogenes, and 5 other uncategorized genes), 16 coding genes have been found in the azoospermia factor region (AZF) and related with male fertility. To date, more than 12 Y chromosome microdeletions have been discovered in the AZF region. The amplicons regions in the Y chromosome are the genetic base of microdeletion occurrence. The Y chromosome microdeletions in the AZF region have been identified as a relatively common cause of male infertility and diagnosed by multiplex PCR in the clinical laboratory. Genomics has brought many revolutionized tools beneficial for better understanding the genetics of mal infertility and defining the role of the Y chromosome gene in spermatogenesis.

Expression of the voltage-dependent anion channel gene in human ejaculated spermatozoa / 中华男科学杂志

<p><b>OBJECTIVE</b>To identify the genes involved in sperm motility.</p><p><b>METHODS</b>We hybridized asthenospermia and normal motile sperm cDNA samples with the human whole genome Affymetrix chip to screen differentially expressed genes. Then we detected the mRNA expressions of the voltage-dependent anion channel genes (VDACs) in human organs and spermatozoa by RT-PCR and compared their expressions in the poor and normal motility spermatozoa.</p><p><b>RESULTS</b>Differentially expressed genes VDACs were identified by analysis of the hybridization signals, including the 3 subtypes VDAC1, VDAC2 and VDAC3. The expression of VDAC2 mRNA was significantly decreased in the poor motility sperm (0.568 +/- 0.036), as compared with the healthy men (0.803 +/- 0.043, P < 0.01).</p><p><b>CONCLUSION</b>The decreased expression of VDAC2 in the ejaculated spermatozoa is possibly associated with the reduction of sperm motility.</p>

Advances in the researches of spermatogenic protein, Ropporin / 中华男科学杂志

Ropporin has been identified as a spermatogenic cell-specific protein and may be involved in sperm maturation, motility, capacitation, hyperactivation and acrosome reaction. However, latest studies have shown that Ropporin is expressed weakly in normal non-testis tissues and highly in hematologic malignancies. Its highly conservative expression in mammalians demonstrates its importance to life. This paper updates the characterization, expression and its distribution, and biological function of Ropporin, and the advances in the clinical researches of the protein.

Differential expression of ODF1 in human ejaculated spermatozoa and its clinical significance / 中华男科学杂志

<p><b>OBJECTIVE</b>To compare the expressions of ODF1 (outer dense fiber of the sperm tail 1) in ejaculated spermatozoa from normozoospermic and asthenozoospermic men with low sperm motility.</p><p><b>METHODS</b>Semen analyses were performed on the semen samples obtained from normozoospermic (n=20) and asthenozoospermic (n=20) volunteers according to the WHO criteria. To rule out the contamination of germ cells and leucocytes, the human ejaculated spermatozoa were purified by a discontinuous Percoll density gradient centrifugation. RT-PCR and Western blot were used to detect the expressions of ODF1 in the spermatozoa from the two groups.</p><p><b>RESULTS</b>RT-PCR showed that the expression of ODF1 mRNA was significantly lower in the spermatozoa from the asthenozoospermic patients than in those from the normozoospermic men (1.35 +/- 0.25 vs. 2.79 +/- 0.28, P < 0.05). Western blot confirmed the results from RT-PCR and revealed an obviously decreased expression of ODF1 in the spermatozoa of the asthenozoospermic patients, with statistically significant difference from the normozoospermic group (1.44 +/- 0.26 vs. 3.64 +/- 0.34, P < 0.05).</p><p><b>CONCLUSION</b>The expression of ODF1 was significantly decreased in the ejaculated spermatozoa of asthenozoospermic men, which might be responsible for low sperm motility.</p>

Partial deletions in the AZFc region of the Y chromosome are associated with male infertility / 中华男科学杂志

Microdeletion of the azoospermia factor in the Yq of the Y chromosome is one of the important causes of male infertility. Complete deletion of the AZFc region (b2/b4 deletion) is the most common type of AZF deletion. Recent studies have shown a variety of deletions of the AZFc region, including partial deletions, such as gr/gr deletion, b1/b3 deletion and b2/b3 deletion, which may also be associated with male infertility.

Extracellular matrix regulates expressions of germ cell differentiation associated genes in mouse embryonic stem cells / 中华男科学杂志

<p><b>OBJECTIVE</b>Interactions of cells with the extracellular matrix (ECM) are essential for cell differentiation. The authors sought to determine the roles of different ECMs in the expressions of germ cell differentiation associated genes after mouse embryonic stem cells (mESCs) differentiated into embryoid bodies (EBs).</p><p><b>METHODS</b>EBs derived from mESCs were maintained in suspension for 3 days and then cultured on the plates coated with various ECMs, including fibronectin (F), laminin (L), matrigel (M), collagen (C) and nonadhensive agarose (A), respectively, for 1, 2, 3 or 4 days, followed by evaluation of the expressions of the genes associated with germ cell differentiation by RT-PCR.</p><p><b>RESULTS</b>The EBs of the F and L groups exhibited facilitated adherent differentiation. The expressions of the Blimp-1, Stella, Mvh and Stra8 genes were increased gradually in the F and L but not obviously in the M and C groups. The overall gene expressions were low in the A group, but high and then gradually decreased in the blank control group. Endogenous fibronectin, laminin and integrin beta1 were obviously expressed in the L and control groups.</p><p><b>CONCLUSION</b>Laminin /integrin beta1 signaling may play a role in regulating the differentiation of mESCs into primordial germ cells (PGCs). Exogenous laminin can facilitate the differentiation of mESC-derived EBs into PGCs by acting on the integrin beta1 subunit, while exogenous fibronectin may be involved in the regulation of the differentiation through other integrin subunit. Endogenous laminin and fibronectin secreted by EBs may also facilitate cell differentiation in the absence of exogenous ECMs.</p>

Differential expression of VASA gene in ejaculated spermatozoa from normozoospermic men and patients with oligozoospermia / 亚洲男科学杂志(英文版)

<p><b>AIM</b>To detect the expression of VASA in human ejaculated spermatozoa, and to compare the expression of VASA between normozoospermic men and patients with oligozoospermia.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>

Functional expression of adenylyl cyclase and phosphodiesterase in ejaculated human spermatozoa / 中华男科学杂志

<p><b>OBJECTIVE</b>To compare the differences of expressions of adenylyl cyclase (AC) and phosphodiesterase (PDE) in ejaculated spermatozoa between healthy volunteers and the patients with asthenospermia.</p><p><b>METHODS</b>Ejaculated spermatozoa were collected from healthy volunteers and the patients with asthenospermia. Reverse transcription polymerase chain reaction (RT-PCR) was used to detect mRNA expression of AC and PDE subtypes in human spermatozoa. The concentrations of cAMP and cGMP in the samples were detected by enzyme-linked immunosorbent assay (ELISA).</p><p><b>RESULTS</b>Compared with healthy volunteers, expression of sAC mRNA and concentration of cAMP were significantly decreased in the patients with asthenospermia (P < 0.01) , while the expression of PDE4C mRNA was significantly increased at the same time (P <0.01). There were no marked differences in the expression of ACIII mRNA and concentration of cGMP between the two groups.</p><p><b>CONCLUSION</b>The sAC down-regulation and PDE4C up-regulation are possible reasons for asthenospermia.</p>

Predictive value of serum inhibin B levels as an indicator of the presence of testicular spermatozoa in nonobstructive azoospermia / 中华男科学杂志

<p><b>OBJECTIVE</b>To evaluate the predictive value of serum inhibin B (INH B) levels as an indicator of the presence of testicular spermatozoa in nonobstructive azoospermia.</p><p><b>METHODS</b>Forty patients with nonobstructive azoospermia (NOA), 20 patients with obstructive azoospermia (OA), and 10 fertile volunteers were involved in this study. A chemoluminescence method was used to measure the levels of FSH; Inhibin B was analysed by using sandwich enzyme-linked immuno-sorbent assay.</p><p><b>RESULTS</b>Patients with nonobstructive azoospermia has significantly higher levels of serum FSH [(21.34 +/- 12.15) IU/L] and significantly lower levels of inhibin B [(53.15 +/- 58.74) ng/L] than patients with obstructive azoospermia [FSH: (3.94 +/- 1.52) IU/L, INH B: (162.49 +/- 78.38) ng/L, P < 0.01] and fertile volunteers [FSH: (4.27 +/- 2.84) IU/L, INH B: (228.49 +/- 110.68) ng/L, P < 0.01]. Mean serum inhibin B were significantly higher in patients with nonobstructive azoospermia who had spermatozoa on TESE than in those in whom no spermatozoa was found on TESE [INHB: (90.31 +/- 72.18) ng/L vs (19.54 +/- 20.38) ng/L, r = 0.528, P < 0.01], but mean FSH levels did not have similar predictive power (P > 0.05).</p><p><b>CONCLUSION</b>Serum INH B level seems to be more accurate than serum FSH in the prediction of presence of testicular spermatozoa in patients with nonobstructive azoospermia. Serum inhibin B determination may be substitute of TESE as a diagnostic index.</p>

In-vitro differentiation of human testicular round spermatids to elongating spermatids / 中华男科学杂志

<p><b>OBJECTIVE</b>To investigate the differentiation of human testicular spermatogenic cells during in vitro culture.</p><p><b>METHODS</b>Testicular cells of obstructive azoospermic patients' testis biopsies were dispersed employing mechanic methods. Then, (1) mixed testicular cells were applied to in vitro culture, and changes of the ratio of elongating spermatids and all round cells were analyzed during mixed cell culture; (2) round spermatids were picked up from the mixed cells employing micromanipulator, followed by differentiation of the isolated round spermatids during microdrop culture.</p><p><b>RESULTS</b>The ratio of the elongating spermatids increased significantly (P < 0.05) after 24 hours of mixed cell culture in HTF medium supplemented with FSH and testosterone. During single round spermatid culture, transformation of the round spermatid to elongating spermatid with newly formed flagellum was observed, and the transformation ratio within 48 hours of microdrop culture was 3.54%. The differentiation of human testicular spermatogenic cells cultured in Vero cell conditioned medium was similar to that cultured in HTF medium.</p><p><b>CONCLUSION</b>Human testicular round spermatids can differentiate to elongating spermatids during in vitro culture. Vero cell conditioned medium does not promote the differentiation of human testicular round spermatids to elongating spermatids.</p>

Research progress in ectopic grafting of testicular tissues / 中华男科学杂志

Since Nature published the first report in 2002 on using immunodeficient mice as recipients and allogeneous or heterogeneous testes as donor tissues to study the ectopic development of spermatogenic cells, the technique has been widely applied in various species (including human). In comparison with other in vitro maturation methods for male germ cells, testicular allografting or xenografting technique has such advantages as similar environment for the development of germ cells in physiological conditions, and better reproducibility. Up to now, sperm has been successfully produced by this technique from the testicular tisues of the immature mouse, hamster, cat, rabbit, pig, goat, bovine and rhesus monkey, and their offspring have even been generated by ICSI technique using the mouse and rabbit sperm derived from testis grafts. This article comprehensively reviews the development of the technique by discussing the influencing factors on the germ cell development in grafts including the variety and age of donors, the sex, integrity and immunity of recipients, the graft location and grafting time. And the applications of the technique and the existing problems are discussed as well.

Development of neonatal mouse and fetal human testicular tissue as ectopic grafts in immunodeficient mice / 亚洲男科学杂志(英文版)

<p><b>AIM</b>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.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>

DNA microarray and its application in andrological research / 中华男科学杂志

This article briefly retrospect the development of microarray, introduces the basic working procedures and the current challenges of DNA microarray, and reviews its application to andrological research, as on the testis, spermatogenic cells, epididymis and sperm. We hope it could play a directive role in the studies of male infertility.

A genetic study on microdeletion of azoospermia factor region on Y chromosome of azoospermia and oligozoospermia patients / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate the genetic causes of azoospermia and severe oligozoospermia.</p><p><b>METHODS</b>Cytogenetic analysis and multiplex polymerase chain reaction(PCR) analysis were done on the 148 patients with azoospermia and serious oligozoospermia.</p><p><b>RESULTS</b>Eleven of the 148(7.4%) cases showed microdeletion of at least one STS. In fifteen STS of AZFa, AZFb,AZFd, AZFc, thirteen STS, eleven STS,two STS and one STS microdeletion were found in each case respectively, including two with 12 STS, five with 5 STS microdeletion.Seven cases had chromosomal morphologic changes(4.7%),four had deletion and one had deletion with translocation of long arm on Y chromosome. One had enlarged region one band two(q12) on long arm of Y chromosome and one had reciprocal translocation of autosomes.</p><p><b>CONCLUSION</b>The findings indicated that AZF microdeletion and chromosomal abnormality should be important causes of male infertility.</p>

Using multiplex PCR to analyze the breakpoint of a severe Y-chromosome deletion / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To elucidate the relationship between azoospermia factor(AZF) microdeletion of Y chromosome and azoospermia, the exact breakpoint of a severe Y-chromosome deletion was determined according to the physical map of AZF.</p><p><b>METHODS</b>Multiplex polymerase chain reaction was used to amplify fifteen sequence tagged sites (STS), namely sY82, sY84, sY86 in AZFa, sY124, sY127, sY128, sY133, sY134, sY143 in AZFb, sY239, sY242 sY254, sY255 in AZFc, and sY145, sY152 in AZFd; sex-determining region Y(SRY) was taken as an internal control. And then sY82,sY86,sY85,sY84 were further analyzed using the sample of the patient who had Y-chromosome deletion by G band analysis to map the breakpoint at molecular level.</p><p><b>RESULTS</b>All 15 STS and sY85 were amplified in positive control while only sY82, sY86 were amplified in the clinical sample, thus the breakpoint was found to be between sY86 and sY85.</p><p><b>CONCLUSION</b>This study on the patient provided the direct biomolecular evidence of the exact breakpoint of the severe Y-chromosome deletion and established the deletion map of acrocentric chromosome. It also proved that the patient's azoospermia was due to the deletion of AZF.</p>

Applied anatomic investigation of phalloplasty / 中华整形外科杂志

<p><b>OBJECTIVE</b>To investigate the anatomy of penis and its adjacent organ for phalloplasty.</p><p><b>METHODS</b>Anatomic dissection of penis and perineum was performed on 30 adult male cadavers (60 sides). Observation and measurement were focused on the penile length of different parts, the morphological relationship of infundibular ligament and suspensory ligament with penile radix, and the feature of crus penis with relation to the deep penile artery.</p><p><b>RESULTS</b>The average length of the penile shaft was 8.13 cm, the penile radix was 7.67 cm and the crus penis was 5.96 - 5.98 cm. The deep penile artery penetrated into the crus penis at its middle 1/3. The infundibular ligament attached to superficial fascia of the penis and extended downward to the scrotal septum to constitute the suspensory structure for both of them. The suspensory ligament attached to the dorsal deep fascia of the penis. Becoming thicker, the rear part of the suspensory ligament connected firmly to the pubic arcuate ligament to constitute a part of suspensory mechanism for the urethra. There was a part of cavernous body, which was free from either ligament or bony attachment, between the penile radix and the crus penis, where the dorsal artery and nerve of penis turned around from the ventral to the dorsal aspect of the penis and the penile dorsal vain penetrated the urogenital septum, draining into intrapelvic venous plexus.</p><p><b>CONCLUSIONS</b>The divisional measurement of the penis length, the recognition of the suspensory ligaments and the anatomic feature of the crus penis with relation to the deep penile artery are all of significant importance to improve the operation of phalloplasty.</p>