Miki Yoshiike

Spermatogenic ability is different among males in different Y chromosome lineage

AbstractIt is a controversial question whether sperm concentrations in humans are changing. Several researchers have reported on environmental factors affecting sperm quality, but the influence of genetic factors is still not fully understood. In this study, we examined the relationship between Y chromosome haplotypes and sperm concentration in fertile males. In addition, we determined the haplotypes of azoospermic patients. The results show that the mean sperm concentration correlates with Y chromosome type. Moreover, the occurrence of azoospermia is related to one particular Y chromosome lineage. Thus, males with a certain haplotype are at a disadvantage for fathering children. The difference of spermatogenic ability among men is important not only in pursuing male competition as in the past but also as relates to the future of modern human males.

Therapeutic Effects of MicroRNA-582-5p and -3p on the Inhibition of Bladder Cancer Progression

Many reports have indicated that the abnormal expression of microRNAs (miRNAs) is associated with the progression of disease and have identified miRNAs as attractive targets for therapeutic intervention. However, the bifunctional mechanisms of miRNA guide and passenger strands in RNA interference (RNAi) therapy have not yet been clarified. Here, we show that miRNA (miR)-582-5p and -3p, which are strongly decreased in high-grade bladder cancer clinical samples, regulate tumor progression in vitro and in vivo. Significantly, the overexpression of miR-582-5p or -3p reduced the proliferation and invasion of UM-UC-3 human bladder cancer cells. Furthermore, transurethral injections of synthetic miR-582 molecule suppressed tumor growth and metastasis in an animal model of bladder cancer. Most interestingly, our study revealed that both strands of miR-582-5p and -3p suppressed the expression of the same set of target genes such as protein geranylgeranyltransferase type I beta subunit (PGGT1B), leucine-rich repeat kinase 2 (LRRK2) and DIX domain containing 1 (DIXDC1). Knockdown of these genes using small interfering RNA (siRNA) resulted in the inhibition of cell growth and invasiveness of UM-UC-3. These findings uncover the unique regulatory pathway involving tumor suppression by both strands of a single miRNA that is a potential therapeutic target in the treatment of invasive bladder cancer.

Physiological roles of semenogelin I and zinc in sperm motility and semen coagulation on ejaculation in humans

At ejaculation, human sperm are considered to be mechanically trapped and become immotile in the semen coagulum by binding to semenogelins (Sgs) from the seminal vesicle and zinc ions from the prostate. However, the physiological combined roles of the protein and heavy metal on sperm motility are unknown. Here, we have first demonstrated that Sg I alone, which does not form the semen coagulum without zinc, is an inhibitor of the motility of intact human sperm at physiological concentration. On the other hand, zinc ions alone had no effect on sperm motility, but confer recovery of sperm motility that has been inhibited by Sg I at a concentration equal to or less than 1 mg/ml. These observations suggest that the roles played by Sg I and zinc on sperm motility are not mechanical but physiological. Quartz crystal microbalance analysis suggests that the sperm extract first bind to Sg I and then zinc ions which subsequently increase the protein accumulation, suggesting that Sgs inhibit sperm motility by directly binding to the sperm surface. Further accumulation of Sg I mediated by zinc ions may entrap the quiescent sperm at semen ejaculation.

Xenografting of testicular tissue from an infant human donor results in accelerated testicular maturation

BACKGROUND Grafting of testicular tissue into immunodeficient mice has been used to differentiate the neonatal testes from different animal species up to the level of complete spermatogenesis; however, this approach has not been successful for human testicular tissue. The aim of this study was to evaluate the capacity for differentiation of infant human testicular tissue grafts. METHODS AND RESULTS Testicular tissue from a 3-month-old patient with testicular cancer was grafted into immunodeficient nude mice. At the time of grafting, A spermatogonia were the only germ cells present in the testicular tissue. B spermatogonia and first spermatocytes were observed at 7 months and 1 year after grafting, respectively. Positive immunostaining with antibodies against BOULE and CDC25A suggested that spermatocytes in the graft were not arrested but in meiosis. Furthermore, ultrastructural and immunohistochemical analyses showed that the onset of both Sertoli cell maturation and partial differentiation of Leydig cells preceded the appearance of spermatocytes. Differentiation of testicular cells was accelerated compared with in vivo development. CONCLUSIONS Spermatogenesis in the xenograft of infant human testicular tissues proceeded successfully from the stage of spermatogonial stem cells until pachytene spermatocyte formation. The differentiation of Sertoli cells and Leydig cells was reproduced in a manner similar to that in normal testicular development. Grafting of infant human testicular tissue may be a powerful tool to examine the early period of human spermatogenesis and may pave the way for fertility preservation among infant patients.

Semen quality of 1559 young men from four cities in Japan: a cross-sectional population-based study

Objectives To provide information of semen quality among normal young Japanese men and indicate the frequency of reduced semen quality. Design Cross-sectional, coordinated studies of Japanese young men included from university areas. The men had to be 18–24 years, and both the man and his mother had to be born in Japan. Background information was obtained from questionnaires. Standardised and quality-controlled semen analyses were performed, reproductive hormones analysed centrally and results adjusted for confounding factors. Setting Four study centres in Japan (Kawasaki, Osaka, Kanazawa and Nagasaki). Participants 1559 men, median age 21.1 years, included during 1999–2003. Outcome measures Semen volume, sperm concentration, total sperm count, sperm motility, sperm morphology and reproductive hormone levels. Results Median sperm concentration was 59 (95% CI 52 to 68) million/ml, and 9% and 31.9% had less than 15 and 40 million/ml, respectively. Median percentage of morphologically normal spermatozoa was 9.6 (8.8 to 10.3)%. Small, but statistically significant, differences were detected for both semen and reproductive hormone variables between men from the four cities. Overall, the semen values were lower than those of a reference population of 792 fertile Japanese men. Conclusions Assuming that the investigated men were representative for young Japanese men, a significant proportion of the population had suboptimal semen quality with reduced fertility potential, and as a group they had lower semen quality than fertile men. However, the definitive role—if any—of low semen quality for subfertility and low fertility rates remain to be investigated.

Semen quality of fertile Japanese men: a cross-sectional population-based study of 792 men

Objectives To establish a base line for future studies on temporal trends, to describe potential geographical differences in semen quality and reference values for studies of men from the general population. Design Cross-sectional study of fertile men from four areas in Japan. Inclusion criteria were: age 20–45 years at the time of invitation, and both the man and his mother had to be born in Japan. Additionally, the current pregnancy of the female partner had to be achieved by normal sexual relations without any fertility treatment. Setting Four Japanese study centres at urban areas located in Sapporo, Osaka, Kanazawa and Fukuoka. Participants 792 men, median age 31.4 years, included from 1999 to 2002. Outcome measures Semen volume, sperm concentration, total sperm count, sperm motility and sperm morphology. Results Semen volumes, percentages of motile spermatozoa and morphologically normal spermatozoa differed slightly between the four groups, whereas no differences in sperm concentrations or total sperm counts were found. In total, 1.2% of men had a sperm concentration below 5 million/ml, 2.1% below 10 million/ml, 3.5% below 15 million/ml and 16.3% below 40 million/ml. For morphology, 14.7% had less than 5% normal spermatozoa. Reproductive hormone levels varied significantly, however, only little from a biological point of view. Conclusions This is the first cross-sectional study on semen quality covering fertile men from the major regions of Japan. It showed that semen quality of fertile Japanese men is comparable to that of the best in European regions. The results may serve as reference values for studies of men from the general population.

A dot-blot-immunoassay for semen identification using a polyclonal antibody against semenogelin, a powerful seminal marker

Among various seminal plasma proteins, semenogelin (Sg), produced in the seminal vesicle, has been considered a candidate for demonstrating the presence of semen. Sg consists of two proteins, one 52 kDa (Sg-I) in size, and the other a mixture of 71 and 76 kDa proteins (Sg-II). Recombinant Sg-I and Sg-II proteins were obtained using a baculovirus system and then injected into a rabbit to produce the respective antibodies [Characterization of recombinant precursor proteins of the human seminal plasma sperm motility inhibitor synthesized in insect cells, Int. J. Mol. Med. 2 (1998) 693]. When liquefied seminal plasma was immunoblotted with the anti-Sg-I and Sg-II antibodies, the anti-Sg-II antibody identified a wider range of the polypeptides originating from Sg than did the anti-Sg-I antibody. A dot-blot-immunoassay using anti-Sg-II antibody revealed a clear immunoreactive spot even when the semen was diluted 6400-fold. However, this assay showed that the Sg antigen was undetectable in saliva, urine, vaginal secretions, sweat, nasal secretions and serum. To determine the stability of Sg antigenic activity, filter paper with dried semen stains were kept at 37, 4 and 22 degrees C for 1, 6 and 18 months, respectively, and the Sg antigenic activity was examined. The activity was detectable in an area not less than 0.5 cm x 0.5 cm under all of the above environmental conditions during each period. Finally, semen was mixed with saliva or blood at various volumetric ratios, and used as a source of dried stains. The Sg antigenic activity was detectable in the stains until the ratio of semen to saliva or blood reached 1:8. These results suggest that Sg may be useful as a marker for semen identification.

An association study of four candidate loci for human male fertility traits with male infertility

STUDY QUESTION Are the four candidate loci (rs7867029, rs7174015, rs12870438 and rs724078) for human male fertility traits, identified in a genome-wide association study (GWAS) of a Hutterite population in the USA, associated with male infertility in a Japanese population? SUMMARY ANSWER rs7867029, rs7174015 and rs12870438 are significantly associated with the risk of male infertility in a Japanese population. WHAT IS KNOWN ALREADY Recently, a GWAS of a Hutterite population in the USA revealed that 41 single-nucleotide polymorphisms (SNPs) were significantly correlated with family size or birth rate. Of these, four SNPs (rs7867029, rs7174015, rs12870438 and rs724078) were found to be associated with semen parameters in ethnically diverse men from Chicago. STUDY DESIGN, SIZE, DURATION This is a case-control association study in a total of 917 Japanese subjects, including 791 fertile men, 76 patients with azoospermia and 50 patients with oligozoospermia. PARTICIPANTS/MATERIALS, SETTING, METHODS Azoospermia was diagnosed on the basis of semen analysis (the absence of sperm in ejaculate), serum hormone levels and physical examinations. Oligozoospermia was defined as a sperm concentration of <20 × 10(6)/ml. We excluded patients with any known cause of infertility (i.e. obstructive azoospermia, varicocele, cryptorchidism, hypogonadotropic hypogonadism, karyotype abnormalities or complete deletion of AZF a, b or c). The SNPs rs7867029, rs7174015, rs12870438 and rs724078 were genotyped using DNA from peripheral blood samples and either restriction fragment length polymorphism PCR or TaqMan probes. Genetic associations between the four SNPs and male infertility were assessed using a logistic regression analysis under three different comparative models (additive, recessive or dominant). MAIN RESULTS AND THE ROLE OF CHANCE The genotypes of all four SNPs were in Hardy-Weinberg equilibrium in the fertile controls. The SNPs rs7867029 and rs7174015 are associated with oligozoospermia [rs7867029: odds ratio (OR) = 1.70, 95% confidence interval (CI) = 1.07-2.68, P = 0.024 (log-additive); rs7174015: OR = 6.52, 95% CI = 1.57-27.10, P = 0.0099 (dominant)] and rs12870438 is associated with azoospermia (OR = 10.90, 95% CI = 2.67-44.60, P = 0.00087 (recessive)] and oligozoospermia [OR = 8.54, 95% CI = 1.52-47.90, P = 0.015 (recessive)]. The association between rs7174015 and oligozoospermia under a dominant model and between rs12870438 and azoospermia under additive and recessive models remained after correction for multiple testing. There were no associations between rs724078 and azoospermia or oligozoospermia. LIMITATIONS, REASONS FOR CAUTION Even though the sample size of case subjects was not very large, we found that three SNPs were associated with the risk of male infertility in a Japanese population. WIDER IMPLICATIONS OF THE FINDINGS The three infertility-associated SNPs may be contributing to a quantitative reduction in spermatogenesis. STUDY FUNDING/COMPETING INTERESTS This study was supported in part by the Ministry of Health and Welfare of Japan (1013201) (to T.I.), Grant-in-Aids for Scientific Research (C) (23510242) (to A.Ta.) from the Japan Society for the Promotion of Science, the European Union (BMH4-CT96-0314) (to T. I.) and the Takeda Science Foundation (to A.Ta.). None of the authors has any competing interests to declare.

Pyrethroid insecticide exposure and semen quality of young Japanese men.

The present study aimed at assessing the relationship between exposure to pyrethroid insecticides and semen quality in 323 university students recruited in a population-based manner in Metropolitan Tokyo. Urinary concentrations of pyrethroid insecticide metabolite, 3-phenoxybenzoic acid (3-PBA), were measured by LC/MS/MS and semen parameters were measured by following internationally harmonized protocols. Median urinary 3-PBA concentration was 0.641 ng/mL (specific gravity-adjusted, n=322). Median values of semen volume, sperm concentration, motility, total number of sperm, and total number of motile sperm were 2.5 mL, 56×10(6)/mL, 61%, 141×10(6), and 82×10(6), respectively. Urinary concentration of 3-PBA was not selected as significant in multiple regression models indicating, in contrast to previous findings, that environmental exposure to pyrethroid insecticides did not affect semen quality. This inconsistency may be related to exposure to different pyrethroid insecticides and/or levels of exposure as well as to survey design (hospital- vs population-based subject recruitment).

Functional implications of membrane modification with semenogelins for inhibition of sperm motility in humans.

Semenogelin I and II (Sgs) are the major component of human semen coagulum. The protein is rapidly cleaved after ejaculation by a prostate-specific antigen, resulting in liquefaction of the semen coagulum and the progressive release of motile spermatozoa. Sgs inhibit human sperm motility; however, there is currently no information on its effect on the sperm membrane. This study investigated the role of Sgs on human sperm motility through regulation of membrane potential and membrane permeability. Fresh semen samples were obtained from normozoospermic volunteers, and studies were conducted using motile cells selected using the swim-up method. Sgs changed the characteristics of sperm motion from circular to straightforward as evaluated by a computer-assisted motility analyzer, and all parameters were decreased more than 2.5 mg/mL. The results demonstrate that Sgs treatment immediately hyperpolarized the membrane potential of swim-up-selected sperm, changed the membrane structure, and time-dependently increased membrane permeability, as determined through flow cytometric analysis. The biphasic effects of Sgs were time- and dose-dependent and partially reversible. In addition, a monoclonal antibody against Sgs showed positive binding to cell membrane proteins in fixed cells, observed with confocal fluorescence microscopy. These results demonstrate that Sgs modifies the membrane structure, indirectly inhibiting motility, and provides suggestions for a therapy for male infertility through selection of a functional sperm population using Sgs.