Archives of andrology最新文献

Conventional semen analysis is rapidly losing its place as the gold standard of diagnosis and the cornerstone of treating the infertile male in modern times. Recent technology allows scientists to analyze sperm fertilizing potential and subsequent embryonic growth by studying factors that have previously escaped traditional parameters. It has become increasingly evident that nuclear DNA arrangement is essential to the fertilizing potential of sperm. A vast array of tests are now available to examine the genetic makeup of individual spermatozoa, ranging the entire gamut from simple bench top assays performed routinely to complex flow cytometric assays requiring highly-skilled technologists. Future research to compare these new tests to those more commonly in use, correlating them with reproductive outcome promises to fill the current void in the field of male infertility, paring innovative diagnostic (and prognostic) technological standards to the already existing sophisticated assortment of successful treatment modalities.

cAMP responsive element modulator (CREM) activator isoforms are involved in mammalian spermatogenesis and spermiogenesis. CREM proteins are highly expressed in postmeiotic germ cells of rodents and primates. Homozygous CREM inactivated mice exhibit round spermatid maturation arrest. The lack of CREM expression at both the mRNA and protein levels is associated with spermatid maturation arrest in infertile patients. Using real-time RT-PCR, we have examined the levels of CREM activator isoform mRNAs: CREMtheta1, CREMtheta2 and CREMt2 + Ex-gamma in gametogenic and interstitial cell fractions from normal human testis, in homogenized tissue samples from spermatogenic arrest and from testicular tumors. We have shown for the first time the presence of CREM activator isoform containing exon gamma (CREMtau2 + Exgamma) in normal human spermatogenesis. Among the three CREM isoforms, CREMtheta1 was expressed in its highest level in the male gonads. In comparison, CREMtheta2 mRNA was significantly less suggesting that the P3 promoter is much more active in human testis than the P4 promoter. Minimal-nill levels of mRNA for either of the CREM activator isoforms were detected in lymphocytes or in gonadal tissues from patients with SCOS (Sertoli Cell Only Syndrome). This data underlines the significance of CREMtheta1 isoform in the regulation of transcription during post-meiotic germ cell differentiation.

Deregulation of sperm nuclear protamine ratio (P1/P2) has been shown to correlate with male factor infertility in humans, but the cause of this abnormal protein expression has yet to be identified. Recent studies have shown that there is little genetic variability in the coding regions of either of the protamine gene sequences. However, these studies did not investigate the 5' or 3' non-coding regions of these genes for mutations that might account for changes in the transcriptional or translational regulation of the protamines. In an effort to determine if genetic variation in these non-coding regions may account for aberrant protamine expression, we have sequenced the 5' and 3' untranslated regions (UTRs) of both protamine 1 (P1) and protamine 2 (P2) genes in a population of infertile men with protamine deregulation, men presenting for infertility work-up with normal protamine ratios, and a population of unrelated, fertile men from the Utah Genetic Reference Project (UGRP). This analysis has identified 14 single nucleotide polymorphisms (SNPs), of which 13 were novel SNPs in the UTRs of P1 and P2, and verified the existence of a variable length repeat (VLR), GAn, in the P2 5' region. The SNP frequencies and VLR allelic frequencies did not achieve statistical significance between the populations, however, one of the SNPs identified in the 3' UTR of protamine 2 was found at a low frequency in the abnormal protamine patients, but was completely absent in men with verified normal protamine ratio and donors of known fertility. In conclusion, a number of SNPs have been reported in the protamine genes and the untranslated regions, however, these gene variants do not appear to be responsible for protamine deficiency. Hence, the underlying cause for aberrant protamine expression may possibly be due to abnormalities in candidate spermatogenic transcriptional/translational regulators, post-translational modifiers, or as-of-yet unidentified factors affecting the testicular environment.

The existence of a complex population of mRNAs in human sperm is well documented but their role is not completely elucidated. Evidence for a latent transcriptional capacity and/or a potential translation de novo in mature spermatozoa from fertile men has been provided and is helpful in understanding the final steps of sperm maturation (capacitation and/or the acrosome reaction). Spermatogenesis is controlled by gonadotrophins and testosterone, their effects are modulated by locally-produced factors that include estrogens derived from the irreversible transformation of androgens by aromatase. The data demonstrating an additional source of estrogens in rat germ cells along with several studies showing a decreased sperm motility in men deficient in aromatase has led to the further explanation of the expression of aromatase in ejaculated spermatozoa from fertile men. A significant decrease in the amount of aromatase transcripts in the immotile sperm fraction was recorded. In addition, the levels of transcripts encoding for proteins involved in either nuclear condensation protamines 1 and 2 (Prm1 and Prm2) or in capacitation endothelial nitric oxide synthase (eNOS) and neuronal nitric oxide sythase (nNOS) and c-myc were compared in low and high motile sperm prepared from the same sample. No significant change in the ratio of c-myc/Prm2 between the two populations of spermatozoa was observed. Conversely the amount of Prm1 mRNA was significantly higher in the low motile fraction than in the high motile fraction; in most of the high motile sperm samples analyzed, eNOS and nNOS transcripts were undetectable, whereas they were observed in low motile sperm. Moreover, a partial or complete disappearance of c-myc transcripts was observed after capacitation. Analysis of the mRNA profile in human ejaculated sperm could be helpful either as a diagnostic tool to evaluate the male gamete quality and/or as a prognostic value for fertilization and embryo development.

This study tested a method for maintaining human spermatozoa without freezing for subsequent use in intracytoplasmic sperm injection (ICSI). We demonstrated that human sperm stored in electrolyte-free solution maintain their motility and viability for at least 4 and 6 weeks, respectively. We also have shown that preserved spermatozoa are fully functional in ICSI. Sperm chromosome analysis after injection of human sperm into mouse oocytes revealed that two weeks of storage does not negatively affect sperm DNA integrity. A mouse model was used to analyze the ability of preserved sperm to participate in normal embryogenesis. Mouse sperm preserved in electrolyte-free solution in a similar manner as human sperm maintained motility for up to 3 weeks. When mouse spermatozoa stored for 1 week were injected into the oocytes by ICSI, they yielded normal blastoctysts and normal viable fetuses. The results of the study bear significance for human assisted reproduction technologies (ART) and provide clinicians and infertile patients with a new method that can simplify sperm preparation for ICSI, assisting men who are unable to provide semen on the day of assisted fertilization.

Seminal vesicle cysts with ipsilateral renal agenesis is rare. When the patient is symptomatic, surgical treatment may be necessary. However, the seminal vesicle is difficult to access surgically, and current transurethral or open surgical approaches have inherent shortcomings. The laparoscopic techniques developed in the last decade may overcome the difficulties in the surgical treatment of seminal vesicle pathology. In this study we report a patient diagnosed with left seminal vesicle cyst and ipsilateral renal agenesis who was managed successfully through the laparoscopic approach. The patient was a 41-year-old who suffered from perineal pain and intermittent hemospermia for 20 years. Ultrasonography and computerized tomography, CT, indicated a cyst of the left seminal vesicle and an absent left kidney. The total laparoscopic operation time was 90 minutes and the estimated blood loss was 80 ml. With a follow-up of 13 months, the patient had total relief of his preoperative symptoms without complication.

Despite significant advances in the treatment of infertility via assisted reproductive technology (ART), the underlying causes of idiopathic male infertility still remain unclear. Accumulating evidence suggests that disorders associated with testicular gene expression may play an important role in male infertility. To be able to fully study the molecular mechanisms underlying spermatogenesis and fertilization, it is necessary to manipulate gene expression in male germ cells. Since there is still no reliable method of recapitulating spermatogenesis culture, the development of alternative transgenic approaches is paramount in the study of gene function in testis and sperm. Established methods of creating transgenic animals rely heavily upon injection of DNA into the pronucleus or the injection of transfected embryonic stem cells into blastocysts to form chimeras. Despite the success of these two approaches for making transgenic and knockout animals, concerns remain over costs and the efficiency of transgene integration. Consequently, efforts are in hand to evaluate alternative methodologies. At present, there is much interest in developing approaches that utilize spermatozoa as vectors for gene transfer. These approaches, including testis mediated gene transfer (TMGT) and sperm mediated gene transfer (SMGT), have great potential as tools for infertility research and in the creation of transgenic animals. The aim of this short review is to briefly describe developments in this field and discuss how these gene transfer methods might be used effectively in future research and clinical arenas.

Finasteride is a specific inhibitor of the 5alpha reductase enzyme originally approved for the treatment of benign prostatic hypertrophy and also for the treatment of androgenetic alopecia (AGA) in men at a dose of 1 mg/day. We report on three cases of young men recruited at our Centre for Male Infertility who had used finasteride for five years. Semen quality was investigated by light microscopy to evaluate sperm concentration and motility. Sperm morphology was performed by transmission electron microscope (TEM) and the data were analyzed. The presence of Y microdeletions was investigated by PCR. Meiotic segregation was explored by fluorescence in situ hybridization (FISH). Patient 1 was azoospermic, patients 2 and 3 showed a normal sperm concentration and severely reduced progressive motility. TEM analysis revealed altered sperm morphology consistent with necrosis and FISH data revealed elevated diploidy and sex chromosome disomy frequencies. This examination was repeated 1 year after the men had suspended the use of finasteride, without receiving any other treatment. A recovery of spermatogenetic process was observed. Motility and morphology improved whereas the meiotic pattern did not change presenting elevated diploidy and sex chromosome disomy frequency.

Mammalian spermatozoa acquire the capacity for motility and fertilization during the transit through the epididymis under the control of different factors, such as cAMP, intracellular pH, intracellular calcium and phosphorylation of sperm proteins. As the acquisition of functional competence including gaining motility during epididymal transit occurs in the complete absence of contemporaneous gene transcription and translation on the part of the spermatozoa, it is widely accepted that post-translational modifications are the only means by which spermatozoa can acquire functionality. Serine-threonine protein phosphatase 1 (PP1) together with their testis/sperm-specific interacting proteins might be involved in this regulatory mechanism. PP1alpha, PP1beta/delta, PP1gamma1 and PP1gamma2 are all expressed in the testis whereas PP1gamma2 is the only isoform expressed on spermatozoa. I2, I3, sds22, 14-3-3 and hsp90 are associated with PP1gamma2 in spermatozoa located on the sperm head and tail. Activity of PP1gamma2 and the binding pattern to these regulatory proteins changes in spermatozoa recruited from the caput and those from the cauda part of the epididymis. In this review, we summarize the possible roles of PP1 on spermatozoa during spermatogenesis and flagellar motility control. We suggest that PP1 might take part in the inhibition of the sperm motility activation by interacting with AKAPs and CAMKII. A hypothesized signaling pathway of mammalian sperm motility activation and PP1's function has been proposed.