Association of TUSC1 and DPF3 gene polymorphisms with male infertility.

PURPOSE: Recently, genome-wide association studies of a Hutterite population in the USA revealed that five single nucleotide polymorphisms (SNPs) with a significant association with sperm quality and/or function in ethnically diverse men from Chicago were significantly correlated with family size. Of these, three SNPs (rs7867029, rs7174015, and rs12870438) were found to be significantly associated with the risk of azoospermia and/or oligozoospermia in a Japanese population. In this study, we investigated whether the rs10966811 (located in an intergenic region between the TUSC1 and IZUMO3 genes) and rs10129954 (located in the DPF3 gene) SNPs, previously related to family size, are associated with male infertility. In addition, we performed association analysis between rs12348 in TUSC1 and rs2772579 in IZUMO3 and male infertility. METHODS: We genotyped 145 patients with infertility (including 83 patients with azoospermia and 62 with oligozoospermia) and 713 fertile controls by PCR-RFLP technique for polymorphism. Because rs10966811 has no restriction sites, the SNP rs12376894 with strong linkage disequilibrium was selected as an alternative to rs10966811. RESULTS: There was a statistically significant association between rs12376894 proxy SNP of rs10966811 and oligozoospermia. Also, a statistically significant association between rs10129954 and azoospermia, and oligozoospermia was observed. When we assessed the relationship between rs12348 in TUSC1 and rs2772579 in IZUMO3 and male infertility traits, we found that rs12348 in TUSC1 was significantly associated with azoospermia and oligozoospermia, but rs2772579 in IZUMO3 was not associated with male infertility. CONCLUSION: We found that the polymorphisms in TUSC1 and DPF3 displayed strong associations with male infertility.

Erratum: An independent validation study of three single nucleotide polymorphisms at the sex hormone-binding globulin locus for testosterone levels identified by genome-wide association studies.

[This corrects the article DOI: 10.1093/hropen/hox002.][This corrects the article DOI: 10.1093/hropen/hox002.].

An independent validation study of three single nucleotide polymorphisms at the sex hormone-binding globulin locus for testosterone levels identified by genome-wide association studies.

STUDY QUESTION: Are the single nucleotide polymorphisms (SNPs) rs2075230, rs6259 and rs727428 at the sex hormone-binding globulin (SHBG) locus, which were identified by genome-wide association studies (GWASs) for testosterone levels, associated with testosterone levels in Japanese men? SUMMARY ANSWER: The SNP rs2075230, but not rs6259 and rs727428, is significantly associated with testosterone levels in Japanese men. WHAT IS ALREADY KNOWN: Previous GWASs have revealed that rs2075230 is associated with serum testosterone levels in 3495 Chinese men and rs6259 and rs727428 are associated with serum testosterone levels in 3225 men of European ancestry. STUDY DESIGN SIZE AND DURATION: This is an independent validation study of 1687 Japanese men (901 in Cohort 1 and 786 in Cohort 2). PARTICIPANTS/MATERIALS SETTING AND METHOD: Cohort 1 (20.7 ± 1.7 years old, mean ± SD) and Cohort 2 (31.2 ± 4.8 years) included samples obtained from university students and partners of pregnant women, respectively. The three SNPs were genotyped using either TaqMan probes or restriction fragment length polymorphism PCR. Blood samples were drawn from the cubital vein of the study participants in the morning, and total testosterone and SHBG levels were measured using a time-resolved immunofluorometric assay. Association between each SNP and testosterone levels was evaluated by meta-analysis of the two Japanese male cohorts. MAIN RESULTS AND THE ROLE OF CHANCE: The age of the two cohorts was significantly different (P < 0.0001). We found that rs2075230 was significantly associated with serum testosterone levels (ß STD = 0.15, P = 7.2 × 10-6); however, rs6259 and rs727428 were not (ß STD = 0.17, P = 0.071; ß STD = 0.082, P = 0.017, respectively), after adjusting for multiple testing in a combined analysis of two Japanese male cohorts. Moreover, rs2075230, rs6259 and rs727428 were significantly associated with high SHBG levels (ß STD = 0.22, P = 3.4 × 10-12; ß STD = 0.23, P = 6.5 × 10-6 and ß STD = 0.21, P = 3.4 × 10-10, respectively). LARGE SCALE DATA: Not applicable. LIMITATIONS REASONS FOR CAUTION: This study had differences in the age and background parameters of participants compared to those observed in previous GWASs. In addition, the average age of participants in the two cohorts in our study also differed from one another. Therefore, the average testosterone levels, which decrease with age, between studies or the two cohorts were different. WIDER IMPLICATIONS OF THE FINDINGS: The three SNPs have a considerable effect on SHBG levels and hence may indirectly affect testosterone levels. STUDY FUNDING/COMPETING INTERESTS: This study was supported partly by the Ministry of Health and Welfare of Japan (1013201) (to T.I.), Grant-in-Aids for Scientific Research (C) (26462461) (to Y.S.) and (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.). There are no conflicts of interest to declare.

A replication study of a candidate locus for follicle-stimulating hormone levels and association analysis for semen quality traits in Japanese men.

In men, follicle-stimulating hormone (FSH) acts on the seminiferous tubules and enhances spermatogenesis. Recently, a candidate locus (rs2414095) for FSH levels was identified by a genome-wide association study (GWAS) in Chinese men. The rs2414095 single-nucleotide polymorphism (SNP) is found on the third intron of the cytochrome P450, family 19, subfamily A, peptide 1 (CYP19A1) gene encoding an aromatase. In the present study, we performed a replication study in 1687 Japanese men (901 from cohort 1 and 786 from cohort 2) to assess whether this SNP is associated with circulating FSH levels. Furthermore, we investigated whether the rs2414095 SNP is correlated with semen quality traits in 2015 Japanese men (1224 from cohort 1 and 791 from cohort 2). The rs2414095 SNP was significantly associated with circulating FSH levels (ßSTD=0.15, P=9.7 × 10-5), sperm concentration (ßSTD=0.073, P=0.032) and total sperm number (TSN) (ßSTD=0.074, P=0.027) in a combined analysis of the two Japanese male cohorts. We successfully replicated, in Japanese men, the results of the previous GWAS for the rs2414095 SNP in Chinese men, and found that the rs2414095 SNP was related with sperm production.

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.

Lack of replication of four candidate SNPs implicated in human male fertility traits: a large-scale population-based study.

STUDY QUESTION: Are the four candidate loci (rs7867029, rs12870438, rs7174015 and rs724078) for human male fertility traits, identified in a genome-wide association study (GWAS) of a Hutterite population in the USA, associated with semen quality traits in a Japanese population? SUMMARY ANSWER: The four single nucleotide polymorphisms (SNPs) rs7867029, rs12870438, rs7174015 and rs724078 have no association with semen parameters in a meta-analysis of two Japanese male cohorts. WHAT IS KNOWN ALREADY: Four (rs7867029, rs12870438, rs7174015 and rs724078) of the SNPs associated with family size or birth rate in the GWAS of a Hutterite population in the USA were associated with semen parameters in ethnically diverse men from Chicago, USA. STUDY DESIGN, SIZE, DURATION: This is a replication study in a total of 2015 Japanese subjects, including 791 fertile men and 1224 young men from the general population. PARTICIPANTS/MATERIALS, SETTING, METHODS: We performed a replication study in two cohorts to assess whether the SNPs rs7867029, rs12870438, rs7174015 and rs724078 are associated with sperm concentration, semen volume, total sperm numbers, total motile sperm numbers or sperm motility. The rs12870438 SNP was detected by restriction fragment length polymorphism PCR while rs7174015, rs724078 and rs7867029 SNPs were genotyped using TaqMan probes. MAIN RESULTS AND THE ROLE OF CHANCE: This study indicated that none of the four SNPs rs7867029, rs12870438, rs7174015 and rs724078 displayed a significant association with semen parameters in the meta-analysis of two Japanese male cohorts. LIMITATIONS, REASONS FOR CAUTION: Only four SNPs identified in the Hutterite GWAS were examined for associations with semen quality traits in a Japanese population. In addition, the linkage disequilibrium structures around the testing markers were different between ethnic groups. WIDER IMPLICATIONS OF THE FINDINGS: Locus mapping studies using a set of tagging SNPs across the loci will be necessary in populations with larger sample sizes in order to understand the contribution of specific genes to semen quality. STUDY FUNDING/COMPETING INTEREST S: 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.

Y chromosome gr/gr subdeletion is associated with lower semen quality in young men from the general Japanese population but not in fertile Japanese Men.

Several case-control studies have investigated whether Y chromosome haplogroups or deletions are associated with spermatogenic failure. However, the relationships between Y chromosome haplogroups or deletions and semen quality in general population have not been elucidated. In this study, we assessed relationships between Y chromosome haplogroups or deletions and semen parameters in 791 fertile Japanese men and 1221 young men from the general Japanese population. We found that the haplogroup D2 (M55 lineage) was significantly associated with lower semen parameters, especially total motile sperm count (P = 0.00051, beta = -0.097), in men from the general population but not in fertile men. In addition, we found that the gr/gr subdeletion was associated with semen quality and in particular, strongly associated with decreased sperm motility (P = 0.00041, beta = -3.14) and total motile sperm count (P = 0.00031, beta = -0.099) in men from the general population but not in fertile men. The combined analysis of fertile Japanese men and men from the general Japanese population showed that the haplogroup D2 (M55 lineage) and the gr/gr subdeletion were strongly associated with reduced sperm motility (P = 0.00056, beta = -2.71, and P = 7.7 × 10(-5), beta = -3.05, respectively) and that haplogroup O2b1 was strongly associated with elevated sperm motility (P = 0.00089, beta = 2.94). These observations add further support for the view that the gr/gr subdeletion diminishes sperm motility that consequently may result in male infertility.

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.

Single nucleotide polymorphism analysis of the follicle-stimulating hormone (FSH) receptor in Japanese with male infertility: identification of codon combination with heterozygous variations of the two discrete FSH receptor gene.

AIMS: Dysfunction of the FSH receptor (FSHR) may be involved in some form of male infertility with azoospermia or oligozoospermia. We assessed the discrete codon combination with homo/heterozygous variation of the exon 10 in the FSHR gene. METHODS: The genotype of codon 307 and codon 680 were analysed in 352 patients with idiopathic male infertility and 145 men with proven fertility. RESULTS AND CONCLUSION: There was no significant difference in the distributions of each homozygous codon 307 or 680 between these two groups as reported in the literature. However, the population with heterozygous combinations Thr/Ala (codon 307) and Ser/ Asn (codon 680) comprised 26% (38/146) and 44.9% (157/343) in subjects with proven fertility and idiopathic infertile men, respectively. Moreover, the heterozygous genotype Thr/Ala-Ser/Asn was significantly increased in infertile patients compared with the controls. This finding showed that the combination of heterozygous FSHR can be responsible for male infertility.

Androgen-independent growth in LNCaP cell lines and steroid uridine diphosphate-glucuronosyltransferase expression.

AIM: To investigate the mechanism of androgen-independent growth of prostate cancer after androgen ablation in LNCaP cells and the effect of glucuronidation activity. METHODS: To establish androgen-independent growth in prostate cancer LNCaP-SF, continuous passage was performed in androgen-stripped medium and the cells were evaluated for glucuronidation activity. The expression vector of antisense uridine diphosphate glucuronosyl-transferase (UGT) 2B15 cDNA was also constructed and evaluated. RESULTS: LNCaP-SF lead to a higher expression in UGT2B15 and their glucuronidation activity is 2.5 times higher than that of LNCaP cells. Significantly fewer LNCaP and LNCaP-SF than control were transfected with the antisense UGT2B15 cDNA, suggesting that UGT2B15 plays an important part in the glucuronidation activity of androgens in both cells. CONCLUSION: The alteration of UGT2B15 expression in LNCaP-SF cells is proposed as a biological characteristic involved in the growth of hormone-refractory prostate cancer.

Differential expression of 17beta-hydroxysteroid dehydrogenase isozyme genes in prostate cancer and noncancer tissues.

BACKGROUND: The adrenal steroids dehydroepiandrosterone and androstenediones are converted into active androgen testosterone in prostatic tissues. Different 17beta-hydroxysteroid dehydrogenase (17betaHSD) isozymes are characterized by either oxidation or reduction reactions. These redox reactions represent an important step in both biosynthesis and metabolism of androgens. This study presents the differential expression of 17betaHSD isozyme genes in cancerous and noncancerous prostate tissues of in vivo samples. METHODS: Thirty-four fresh specimens of transrectal prostatic needle biopsy were obtained; 11 were pathologically diagnosed as adenocarcinoma and 23 as without malignancy. The gene expression levels of five isozymes (type 1-5) of 17betaHSD were evaluated. The quantification of gene expression was assessed by means of the real-time polymerase chain reaction. RESULTS: The expression levels of the type 3 17betaHSD gene with malignancy were significantly higher than those in prostatic tissues without malignancy, and those of type 2 17betaHSD with malignancy were significantly lower than those in nonmalignant tissues. There were no significant differences in 17betaHSD type 1, type 4, and type 5 gene expression in cancerous and noncancerous tissues. CONCLUSION: Our results suggest that 17betaHSD type 2 and type 3 play an important role in the conversion of adrenal steroids into potential androgens in prostate cancer tissue.