Effect of B9 and B12 vitamin intake on semen parameters and fertility of men with MTHFR polymorphisms.

The methylenetetrahydrofolate reductase (MTHFR) gene codes a crucial enzyme which involve in folate metabolism. The effect of MTHFR gene polymorphisms on male fertility status is uncertain and controversial. We evaluated the effect of B vitamin family intake on total homocysteine content and semen parameters of men with MTHFR gene polymorphisms. MTHFR genotypes frequency and serum total homocysteine concentration were measured among 280 men with impaired spermatogenesis (asthenospermia, oligospermia, severe oligospermia and azoospermia) and 85 control participants. B vitamin family dietary intakes were assessed using a semi-quantitative food-frequency questionnaire. In addition, concentrations of vitamins B9 and B12 were evaluated in serum samples of some participants (n = 60). We observed significantly higher frequency of TC or TT genotypes in C677T polymorphism among oligospermic, severe oligospermic and azoospermic men. CC genotype of A1298C polymorphism was significantly higher only in azoospermic men. Also, we observed critical effect of vitamin B9 and B12 intake on decreasing of total homocysteine and improving of semen parameters among the men with T allele of MTHFR C677T polymorphism. Our investigation showed that sufficient consumption of vitamins B9 and B12 influences sperm parameters of men with different MTHFR polymorphisms, especially genotypes with T allele.

Protective features of resveratrol on human spermatozoa cryopreservation may be mediated through 5' AMP-activated protein kinase activation.

Biochemical and physical modifications during the freeze-thaw process adversely influence the restoration of energy-dependent sperm functions required for fertilization. Resveratrol, a phytoalexin, has been introduced to activate 5' AMP-activated protein kinase which is a cell energy sensor and a cell metabolism regulator. The cryoprotection of resveratrol on sperm cryoinjury via activation of AMP-activated protein kinase also remains to be elucidated. Our aim, thus, was to investigate: (i) the presence and intracellular localization of AMP-activated protein kinase protein; (ii) whether resveratrol may exert a protective effect on certain functional properties of fresh and post-thaw human spermatozoa through modulation of AMP-activated protein kinase. Spermatozoa from normozoospermic men were incubated with or without different concentrations of Compound C as an AMP-activated protein kinase inhibitor or resveratrol as an AMP-activated protein kinase activator for different lengths of time and were then cryopreserved. AMP-activated protein kinase is expressed essentially in the entire flagellum and the post-equatorial region. Viability of fresh spermatozoa was not significantly affected by the presence of Compound C or resveratrol. However, although Compound C caused a potent inhibition of spermatozoa motility parameters, resveratrol did not induce negative effect, except a significant reduction in motility at 25 µm for 1 h. Furthermore, resveratrol significantly increased AMP-activated protein kinase phosphorylation and mitochondrial membrane potential and decreased reactive oxygen species and apoptosis-like changes in frozen-thawed spermatozoa. Nevertheless, it was not able to compensate decreased sperm viability and motility parameters following cryopreservation. In contrast, Compound C showed opposite effects to resveratrol on AMP-activated protein kinase phosphorylation, reactive oxygen species, apoptosis-like changes, mitochondrial membrane potential, and motility parameters. These findings, although preliminary, suggest that resveratrol-induced improvement of cryopreserved sperm functions may be mediated through activation of AMP-activated protein kinase, indicating the importance of AMP-activated protein kinase activity for human spermatozoa functions. Further investigations are required to elucidate the mechanism by which resveratrol ameliorates oxidative stress-mediated damages in an AMP-activated protein kinase-dependent mechanism.

Association of G/T(rs222859) polymorphism in Exon 1 of YBX2 gene with azoospermia, among Iranian infertile males.

Animal model studies have shown that MSY2 gene has a potential role in spermatogenesis. Some mutations on this gene have been proposed to be associated with human male infertility. In this study, polymorphisms of exon 1 of YBX2 gene have been investigated. A total of 276 men were evaluated. They included 96 men with normal spermatogenesis, 60 men with nonobstructive azoospermia, 60 men with oligospermia and 60 men with asthenospermia. We extracted DNA from blood and testis tissues of samples, and analysed polymorphisms of exon 1 by sequencing method. Moreover, YBX2 gene expression was studied by real-time PCR on blood and testis tissue of samples. Sequencing results showed that among the studied polymorphisms, frequency of TT genotype in rs222859 polymorphism was significantly higher in azoospermic patients compared to control group (P < 0.001). Azoospermic men exhibited significant underexpression of YBX2 gene in blood and testis samples in comparison with controls, oligosperm and asthenosperm samples (P < 0.001), but there was no significant difference in gene expression of YBX2 gene in blood and testis tissues of azoospermic men, with and without mutation (P > 0.05). According to our results, the alterations of this gene might be involved in azoospermia among Iranian population.

Frequency of sex chromosomal disomy in spermatozoa of normal and oligozoospermic Iranian patients and its effects on fertilisation and implantation rates after ICSI.

Chromosomal aneuploidy is a well-known phenomenon in human gametes including spermatozoa. Success rate of fertilisation and implantation in subfertile patients with male factor has always been shown to be very low. We tried to relate the possible impact of sex chromosomal aneuploidy in spermatozoa used for intracytoplasmic sperm injection (ICSI) on fertilisation and implantation rate. To evaluate the frequency of disomy for X and Y chromosomes in sperm samples retrieved from normal and oligozoospermic individuals, primed in situ labelling (PRINS) technique was used. Following ICSI, the rate of eight-cell embryos for each category was determined and followed up for successful implantation. Results showed a statistically significant higher frequency of disomy for all chromosomes under study in spermatozoa of oligozoospermic patients compared with normal men (P<0.01). The rate of eight-cells embryo formation was significantly lower than in normal group (P<0.01). The number of embryos transferred for both groups were nearly similar. Implantation rate for oligozoospermic patients was much lower than that of the normal group but was not significantly different (P>0.05). These results demonstrate that men especially with severe oligozoospermia have an elevated risk for chromosome abnormalities in their spermatozoa. These abnormalities might affect fertilisation and pre-embryo formation with less impact on implantation.

Leber hereditary optic neuropathy: do folate pathway gene alterations influence the expression of mitochondrial DNA mutation?

BACKGROUND: Leber hereditary optic neuropathy (LHON) is an inherited form of bilateral optic atrophy leading to the loss of central vision. The primary cause of vision loss is mutation in the mitochondrial DNA (mtDNA), however, unknown secondary genetic and/or epigenetic risk factors are suggested to influence its neuropathology. In this study folate gene polymorphisms were examined as a possible LHON secondary genetic risk factor in Iranian patients. METHODS: Common polymorphisms in the MTHFR (C677T and A1298C) and MTRR (A66G) genes were tested in 21 LHON patients and 150 normal controls. RESULTS: Strong associations were observed between the LHON syndrome and C677T (P= 0.00) and A66G (P= 0.00) polymorphisms. However, no significant association was found between A1298C (P =0.69) and the LHON syndrome. CONCLUSION: This is the first study that shows MTHFR C677T and MTRR A66G polymorphisms play a role in the etiology of the LHON syndrome. This finding may help in the better understanding of mechanisms involved in neural degeneration and vision loss by LHON and hence the better treatment of patients.

Differences between the radiation hybrid and genetic linkage maps of bovine chromosome 5 resolved with a quasi-phylogenetic method of analysis.

Two major differences were detected in gene order between the radiation hybrid map and the genetic linkage map of bovine Chromosome (Chr) 5, and these were resolved by analyzing the raw radiation hybrid data by a quasi-phylogenetic method. Seventeen loci were typed on the new cattle whole genome radiation hybrid panel. Most of these loci are framework loci and include AGLA293, BM315, BM6026, BP1, BZRP, CD9, CSSM22, CSSM34, CYP2D@, ETH2, ETH10, ETH152, IGF1, LALBA, SLC2A3, SYT1, and TPI1. BP1 was found to be closer to the centromere than either BM6026 or SYT1 with two standard computer software packages for analyzing radiation hybrid panel data. This is inconsistent with any of the genetic linkage maps as well as their consensus. CYP2D@ was placed between ETH2 and BZRP, and this is also inconsistent with the genetic linkage maps, since CYP2D@ should be the most telomeric of the loci tested in this study. Resolution was reached by analyzing the raw radiation hybrid data for clones that bind some but not all of the loci, and the binding pattern was more consistent with the linkage maps and less consistent with the software-generated radiation hybrid map. The comparative mapping data confirm the relative inversion of gene order of SYT1 compared with humans and mice. A non-polymorphic fragment for CD9 indicates the conservation of gene order for three loci located on human Chr 12p. The genes of bovine Chr 5 conserved on human Chr 12p are located separately from the genes conserved on human Chr 12q. It is recommended that the raw data for radiation hybrid maps be made publicly available so that conflicts in gene order can be evaluated explicity.

Comparative mapping of genes from human chromosome 12 by genetic linkage mapping in cattle.

Loci from human chromosome 12 were mapped in cattle to compare the gene order between species. Polymorphisms were detected in cattle in six loci that had been mapped with high precision in humans. Four of these loci, LALBA, SLC2A3, SYT1, and TPI1, mapped to bovine chromosome 5, and one, PLA2G1B, mapped to bovine chromosome 17. The sixth locus, SLC2A3L, due to a fragment produced by the SLC2A3 primers, maps to the telomeric region of BTA18. The differences in gene order between human chromosome 12 and cattle chromosome 5, when these loci are added to others already mapped in cattle, show evidence of significant rearrangement in gene order requiring several evolutionary events. There is also evidence in cattle chromosome 5 of the interspersal of material conserved on human chromosome 22 into the material conserved on human chromosome 12, consistent with ZOOFISH analyses. This analysis indicates that the larger block near the centromere is conserved on the long arm of human chromosome 12 and the smaller block near the telomere is conserved as part of the short arm of human chromosome 12. The level of variation detected in the amplified cattle DNA was approximately 1 variant per 464 nucleotides of haploid DNA using single-strand conformation polymorphism analysis. This corresponds to a per individual level of 1 variant per 1, 961 nucleotides of haploid DNA. This confirms lower genetic variability in cattle compared to humans but indicates the potential for millions of single nucleotide polymorphisms in cattle.

Report of the first workshop on the genetic map of bovine chromosome 1.

A report of the first workshop on the genetic map of bovine chromosome 1 (BTA1) is presented. Five laboratories contributed 31,962 informative meioses from 70 loci. Thirty-two loci which had been typed by at least two laboratories were used to construct a framework genetic map with a likelihood ratio support of at least 1000:1 for locus order. The resulting sex-averaged framework map contained 26 loci and spanned 163.6 CM. The lengths of the female and male maps were 159.5 CM and 165.3 CM, respectively, and there was evidence for an expansion in the telomeric one-third of the male map. Of the four cases where order for closely linked loci differed among the maps produced for each of the contributing laboratories, a consensus order was obtained for three in the framework map. The average genetic distance between framework loci on the sex-averaged map was 6.3 CM.