Wenhao Tang

Mechanistic insights into acephalic spermatozoa syndrome–associated mutations in the human SUN5 gene

Acephalic spermatozoa syndrome has been reported for many decades; it is characterized by very few intact spermatozoa and tailless sperm heads in the semen and causes severe male infertility. The only gene in which mutations have been found to be associated with this syndrome encodes Sad1 and UNC84 domain–containing 5 (SUN5), a testis-specific nuclear envelope protein. The functional role of SUN5 has been well-studied in mouse models, but the molecular basis for the pathogenic effects of mutations in the human SUN5 gene remains elusive. Here, we report a new SUN5 mutation (c.475C→T; p.Arg159*), and explore the pathogenic effects of all known SUN5 mutations on acephalic spermatozoa syndrome. Using an artificial splicing system, we found that the intronic mutation affects the splicing of SUN5 mRNA, yielding a premature stop codon that results in a truncated SUN5 protein. We also found that SUN5 interacts with the coupling apparatus protein DnaJ heat shock protein family (Hsp40) member B13 (DNAJB13) during spermatogenesis, and the substitutions in the SUN5 SUN domain impair its interaction with DNAJB13. Furthermore, we observed that many SUN5 mutations affect the secondary structure of the protein and influence its folding and cellular localization. In summary, our findings indicate an interaction of SUN5 with DNAJB13 during spermatogenesis, provide mechanistic insights into the functional role of this interaction in sperm head–tail integration, and elucidate the molecular etiology of acephalic spermatozoa syndrome–associated SUN5 mutations.

Sodium-Hydrogen-Exchanger expression in human sperm and its relationship with semen parameters

PurposeSperm-specific sodium-hydrogen exchanger (sNHE) is essential to maintain sperm normal function in mice; however, its role in human sperm has not been clarified to date. The aim of this study is to investigate the expression pattern of sNHE in human spermatozoa and its relationship with sperm functional parameters.MethodSemen samples from 68 asthenozoospermic and 61 normozoospermic men were analyzed for sperm concentration, motility, and acrosome reaction, and high motile spermatozoa were collected by swim-up method. The expression of sNHE in spermatozoa was detected by Western blot and immunofluorescence staining. The relationship between sNHE expression and sperm parameters was assessed.ResultsWe identified sNHE is mainly localized to the principal piece of the human sperm tail. The expression of sNHE was positively correlated with sperm concentration, total number, and progressive motility. Moreover, sNHE expression was upregulated in swim-up sperm and associated with most of sperm motility parameters including straight line velocity and curvilinear velocity. Our results also showed that sNHE expression is decreased in sperm from patients with asthenozoospermia compared with that from normal controls. However, no correlation was found between sNHE expression and acrosome reaction in spermatozoa.ConclusionsThe expression pattern of sNHE suggested that this protein may be involved in the regulation of sperm motility, and aberration of its expression in sperm may contribute to the pathogenesis of asthenozoospermia.

Multiple factors affecting surgical outcomes and patency rates in use of single-armed two-suture microsurgical vasoepididymostomy: a single surgeon's experience with 81 patients.

Vasoepididymostomy (VE), as the most challenging procedure in microsurgeries, is often carried out with a double-armed two-suture technique. In this study, we evaluated the efficacy and safety of the single-armed two-suture VEs on humans and studied the factors that could possibly affect the patency rates. From July 2012 to July 2013, we reviewed 81 patients with consecutive primary epididymal obstruction who underwent single-armed two-suture longitudinal intussusception microsurgical VEs by a single surgeon, Kai Hong (KH). At the same time, we analyzed seven factors that possibly related to the patency rates. With the single-armed technique, a total of 81 men underwent the microsurgical VEs. Data on 62 patients were completely recorded. 19 patients were lost to follow-up. Mean age was 31 years old. Mean follow-up time was 8.8 (2-17) months. The patency rate was 66.1% (41/62). Natural pregnancy rate was 34.1% (14/41). Overall pregnancy rate was 22.6% (14/62). No severe surgical complications were noted. With logistic regression test analysis, there were two factors related to a higher patency rate: anastomosis sites (P = 0.035) and motile sperm found in the epididymal fluid (P = 0.006). Motile sperm found in the epididymal fluid were associated with a higher patency rate (OR = 11.80, 95% CI = 1.79, 77.65). The single-armed two-suture longitudinal VE technique is feasible for microsurgical practice. The patency and pregnancy rates are comparable to the doubled-armed technique. Anastomosis sites and motile sperm found in the epididymal fluid were the most two important factors related to higher patency.

Identification and Characterization of Xlr5c as a Novel Nuclear Localization Protein in Mouse Germ Cells

Background Spermatogenesis is the complex process by which diploid stem cells generate haploid germ cells in gamete production. Members of the Xlr (X-chromosome linked, lymphocyte regulated) superfamily play essential roles in spermatogenesis. The expression, localization and role in spermatogenesis of one such member, Xlr5c, has not been reported previously. Methodology/Principal Findings Xlr5c mRNA and protein levels in murine testes and other tissues were investigated using RT-PCR and Western blotting. Xlr5c was abundantly transcribed in mouse testes, particularly during the early stages of spermatogenesis and throughout prophase I in the nuclei of spermatocytes. Xlr5c was specifically localized at synaptonemal complexes(SCs) region in preleptotene and pachytene spermatocytes, as was the homologous Xlr protein Sycp3. Conclusions/Significance These results suggest that Xlr5c was abundantly transcribed in germ cells, localized at SCs region, where it may play a potential role during the early stages of spermatogenesis. Identification and characterization of this novel testis protein may offer a new perspective for understanding of the molecular mechanisms involved in germ cell differentiation.

Effect of male body mass index on live-birth sex ratio of singletons after assisted reproduction technology

OBJECTIVEnTo determine the effect of male body mass index (BMI) on the probability of achieving a live birth and the sex ratio of singletons at birth after IVF and intracytoplasmic sperm injection (ICSI) treatment.nnnDESIGNnA retrospective cohort study.nnnSETTINGnUniversity-affiliated infertility center.nnnPATIENT(S)nPatients seeking infertility treatment who received IVF or ICSI treatment with autologous oocytes from January 2009 to December 2013.nnnINTERVENTION(S)nNone.nnnMAIN OUTCOME MEASURE(S)nLive-birth sex ratio of singletons at birth stratified by male BMI and adjusted by parental age, parental BMI, type of infertility, parity, embryo culture media, and cause of infertility.nnnRESULT(S)nA total of 8,490 couples undergoing IVF or ICSI treatment resulted in 39.12% live births and gave birth to 2,377 live birth singletons and 943 twins. There was no significant difference in the live birth rate between groups stratified by BMI. The probability of live births for overweight and obese groups were not decreased compared with the normal-weight group; similar null findings existed in the IVF and ICSI subgroups. Of note, the sex ratio of offspring in the overweight and obese male groups was significantly higher than in the normal-weight group (1.27 vs. 1.07). Male BMI was significantly associated with sex ratio of singletons after adjusting for confounders. In twins, incidences of twins with male-male infants in the overweight/obese group were not different from the normal-weight group.nnnCONCLUSION(S)nIncreased male BMI has no effect on live birth success, but has an increased probability of giving birth to male singletons.

A high performance triboelectric generator for harvesting low frequency ambient vibration energy

We present a novel triboelectric generator for vibration energy harvesting based on the mass production manufacture of flexible printed circuit (FPC). 10 pairs of friction surfaces were integrated on the zigzag-shaped FPC structure, which served as an elastic spring in the mass-spring system. This design makes the triboelectric generator simple and easy to be stimulated. Using industrial FPC manufacture makes the fabrication efficient, low-cost and with high yield. Load circuits can be integrated with the generator without additional assembly. The generator can be operated within a large frequency range from 1 Hz to 200 Hz. Low resonant frequency of 16 Hz and wide bandwidth of 37 Hz were achieved. The maximum effective output power of 77.3 μW was obtained with an optimal load of 10 MΩ at 16 Hz with an oscillation amplitude of 2 mm.

Single-Cell RNA Sequencing Analysis Reveals Sequential Cell Fate Transition during Human Spermatogenesis

Spermatogenesis generates mature male gametes and is critical for the proper transmission of genetic information between generations. However, the developmental landscapes of human spermatogenesis remain unknown. Here, we performed single-cell RNA sequencing (scRNA-seq) analysis for 2,854 testicular cells from donors with normal spermatogenesis and 174 testicular cells from onexa0nonobstructive azoospermia (NOA) donor. A hierarchical model was established, which was characterized by the sequential and stepwise development of three spermatogonia subtypes, seven spermatocyte subtypes, and four spermatid subtypes. Further analysis identified several stage-specific marker genes of human germ cells, such as HMGA1, PIWIL4, TEX29, SCML1, and CCDC112. Moreover, we identified altered gene expression patterns in the testicular somatic cells of one NOA patient via scRNA-seq analysis, paving the way for further diagnosis of male infertility. Our work allows for the reconstruction of transcriptional programs inherent to sequential cell fate transition during human spermatogenesis and hasxa0implications for deciphering male-related reproductive disorders.

Effects of maternal acrolein exposure during pregnancy on testicular testosterone production in fetal rats

Acrolein has been reported to have diverse toxic effects on various organs, including the reproductive system. However, little is known regarding the effects of maternal acrolein exposure on testicular steroidogenesis in male offspring. The present study investigated the effects of acrolein on fetal testosterone production and associated genes. Pregnant Sprague-Dawley rats were intraperitoneally injected with vehicle (normal saline) or 1, 2 or 5 mg/kg acrolein from gestational day (GD) 14–20, and fetal testes were examined on GD 21. Fetal body and testicular weights were markedly reduced in pups following exposure to high doses of acrolein (5 mg/kg) in late pregnancy. Notably, in utero exposure of 5 mg/kg acrolein significantly decreased the testicular testosterone level and downregulated the expression levels of steroidogenic acute regulatory protein (StAR) and 3β-hydroxysteroid dehydrogenase (3β-HSD), whereas the levels of other steroidogenic enzymes, including scavenger receptor class B, cholesterol side-chain cleavage enzyme and steroid 17 alpha-hydroxylase/17,20 lyase, were unaffected. Furthermore, the 3β-HSD immunoreactive area in the interstitial region of the fetal testes was reduced at a 5 mg/kg dose, whereas the protein expression levels of 4-hydroxynonenalwere dose-dependently increased following maternal exposure to acrolein. mRNA expression levels of insulin-like factor 3, a critical gene involved in testicular descent, were unaltered following maternal acrolein exposure. Taken together, the results of the present study suggested that maternal exposure to high doses of acrolein inhibited fetal testosterone synthesis, and abnormal expression of StAR and 3β-HSD may be associated with impairment of the steroidogenic capacity.

Trim27 interacts with Slx2, is associated with meiotic processes during spermatogenesis.

ABSTARCT Formation of the XY body is believed to prevent recombination between X and Y chromosomes during meiosis. We recently demonstrated that SYCP3-like X-linked 2 (Slx2) could be involved in synaptonemal complex formation as well as XY body maintenance during meiosis. In order to further investigate the role and composition of XY body protein complexes in meiotic processes and spermatogenesis, a yeast 2-hybrid screening was performed, and the tripartite motif protein 27(Trim27) was found to interact with Slx2 and co-localized in the XY body. Trim27 has a tripartite motif (TRIM) consisting of a RING finger, B-box and coiled-coil domains, and is a transcriptional regulator that is expressed in various tumor cell lines. In this study, we showed that Slx2 and Trim27 were highly expressed in meiosis of mouse testis. And the Slx2/Trim27 interaction was confirmed in vivo by co-immunoprecipitation and mammalian 2-hybrid interaction assays. Moreover, cytoimmuno localization experiments revealed that Slx2/Trim27 was co-localized to the XY body of spermatocytes during meiosis, and immunohistochemical results revealed co-localization of Trim27 and γ-H2AX in the XY body of primary spermatocytes in the mouse testis. Trim27 may therefore be a transcriptional regulation protein connecting Slx2 and γ-H2AX, thereby promoting the formation of a more potent XY body protein complex in meiotic processes and spermatogenesis. In conclusion, Trim27 connecting Slx2 may regulate meiotic processes in multiple ways by influencing XY body formation and germ cell proliferation during spermatogenesis.

A clinical trial on the consistency of bilateral testicular tissue histopathology and Johnsen score: single side or bilateral side biopsy?

To evaluate and compare left and right testicular tissue histopathology and Johnsen score, and to investigate the necessity for bilateral testicular biopsy. We recruited180 patients with non-obstructiveazoospermia (NOA) on testicular biopsy who had undergonetesticular sperm aspiration (TESA). Pathological sections of testicular tissue were diagnosed by specially-assigned doctors, who evaluated pathological findings, determined the Johnsen score and confirmed for the presence or absence of sperm. Sperm positive rates for left and right testicular histopathology were 55.0% and 51.7% respectively, and the proportion of Johnsen scores≥8 for left and right testes were 53.3% and 50.0%, respectively. Cohen kappa values revealed that the identification of sperm in bilateral testicular samples was not consistent and was related to random effects; Optimized cut-off value for bilateral testicular volume was 11ml (Johnsen score ≥8), and optimized cut-off values of E2 on left and right testes were 144.5pmol/L and 133.5 pmol/L (Johnsen score≤7). However, age, serum prolactin (PRL), follicle stimulating hormone (FSH), luteinizing hormone (LH) and total testosterone (TT) levels were not accurate predictors for the existence of testicular sperm. There was nostatistical significance between left and right testicular histopathology in terms of sperm positive rates or Johnsen score; the Johnsen score were caused entirely by random effects and a score from one side could not represent the other side. Therefore, we recommend that both testes need to undergo surgery when NOA patients undergo testicular biopsy or sperm retrieval.