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Oligo-astheno-teratozoospermia ( OAT), a condition that includes low sperm number, low sperm motility and abnormal sperm morphology, is the commonest cause of male infertility. Because genetic analysis is frequently impeded by the infertility phenotype, the genetic basis of many of OAT conditions has been hard to verify. Here, we show that deficiency of ORP4, a sterol-binding protein in the oxysterol-binding protein ( OSBP)-related protein family, causes male infertility due to severe OAT in mice. In ORP4-deficient mice, spermatogonia proliferation and subsequent meiosis occurred normally, but the morphology of elongating and elongated spermatids was severely distorted, with round-shaped head, curled back head or symplast. Spermatozoa derived from ORP4-deficient mice had little or no motility and no fertilizing ability in vitro. In ORP4-deficient testis, postmeiotic spermatids underwent extensive apoptosis, leading to a severely reduced number of spermatozoa. At the ultrastructural level, nascent acrosomes appeared to normally develop in round spermatids, but acrosomes were detached from the nucleus in elongating spermatids. These results suggest that ORP4 is essential for the postmeiotic differentiation of germ cells. [ABSTRACT FROM AUTHOR]

During the last decade, evidence-based medicine has been described as a paradigm shift in clinical practice, and as 'the conscientious, explicit, and judicious use of current best evidence in making decisions about the care of individual patients'. Appropriate statistical methods for analyzing data are critical for the correct interpretation of the results in proof of the evidence. However, in the medical literature, these statistical methods are often incorrectly interpreted or misinterpreted, leading to serious methodological errors and misinterpretations. This review highlights several important aspects related to the design and statistical analysis for evidence-based reproductive medicine. First, we clarify the distinction between ratios, proportions, and rates, and then provide a definition of pregnancy rate. Second, we focus on a special type of bias called 'confounding bias', which occurs when a factor is associated with both the exposure and the disease but is not part of the causal pathway. Finally, we present concerns regarding misuse of statistical software or application of inappropriate statistical methods, especially in medical research. [ABSTRACT FROM AUTHOR]

Purpose: Zona pellucida (ZP)-free eggs are often used for studies such as evaluating the interaction of sperm-oolemma. To acquire ZP-free eggs, the most commonly used methods employ acidified Tyrode's solution, enzymatic digestion with a trypsin-like enzyme, or mechanical methods using micropipettes. However, acidified Tyrode's solution and trypsin-like enzymes often damage the oolemma, especially when many eggs are treated at once for mass sample analyses. The mechanical method requires skill, and it is time-consuming to prepare many ZP-free eggs. Therefore, in this study, to establish an easy, reliable method for preparing ZP-free eggs, we examined the ZP digestion method originally reported by Zuccotti et al. (J Reprod Fertil 93:515-520, ) that uses collagenase. Methods: Mouse unfertilized eggs were treated with collagenase and acidified Tyrode's solution to compare the ZP-free rates, the effect on the oolemma, and the two-cell development rates of ZP-free eggs by in vitro fertilization. The effects on the oolemma were gauged by observing the polarity of the transmembrane protein localization of enhanced green fluorescence protein tagged CD9 protein (CD9-EGFP) and using differential interference contrast microscopy. Results: Collagenase removed the ZP and the cumulus cells from the cumulus oocyte complex. The collagenase method had no influence on the localization of CD9-EGFP, resulting in a high two-cell development rate. Additionally, the collagenase method could exclude low quality eggs with hardened ZP, since collagenase could not digest the hardened ZP. Conclusions: The one-step collagenase method is an easy preparation method for large numbers of high-quality ZP-free eggs. [ABSTRACT FROM AUTHOR]

Aim: l-Carnitine, an essential cofactor for mitochondrial, β-oxidation of long-chain fatty acids, is known to play important roles in sperm maturation and metabolism when spermatozoa pass and acquire motility in the epididymis. We reported that obstructive azoospermia occurred in the epididymis in the juvenile visceral steatosis (JVS) mice, which are OCTN2 dysfunction mice caused by mutations in the gene encoding OCTN2, have been used for animal models of primary systemic carnitine deficiency. The aim of present study is to investigate the expression of OCTN2 protein in the mouse epididymis and its relation between the localization of OCTN2 and obstructive azoospermia in JVS mice as animal models for human male infertility. Methods: Animals used in this study were wild-type (C57BL/6 J) mice ( n = 4) and JVS mice ( n = 4). We made a specific polyclonal antibody against OCTN2 and examined immunohistochemically the localization of OCTN2 in the mouse epididymis. Results: OCTN2 was localized on the apical membrane of the principal cells of distal corpus and cauda epididymides. Immunocytochemistry demonstrated that OCTN2 was localized on the surface of microvillus upon the principal cells. In JVS mice, immunoreactivity started in a region immediately distal to where the sperm obstruction occurred. Conclusions: Our results suggest that OCTN2 functions as a carnitine transporter between the epithelium and the lumen in distal corpus and cauda epididymides and provides a clue as to why obstructive azoospermia is induced in distal parts of epididymis. [ABSTRACT FROM AUTHOR]

Background: The oviductal isthmus is known to act as a sperm reservoir in several mammalian species including mice, but it is still unclear how sperm are released from the reservoir after ovulation. Recently, nitric oxide (NO) was reported to have important roles as a mediator in various sperm functions, including hyperactivation and capacitation. Therefore, we have investigated the change of the activity of nitric oxide synthase (NOS) of sperm of the isthmus in relation to ovulation under in vivo fertilization conditions. Methods and Results: The sperm were collected from the isthmus and uterus of the female mated before or after ovulation. The NOS activity change was evaluated by using the β-nicotinamide adenine dinucleotide phosphate-diaphorase staining method, and sperm NOS activity was quantified by using NIH image software. The results showed that, in the reservoir, the peak intensity of sperm NOS activity was higher after ovulation (135.5 ± 22.4) than before ovulation (102.7 ± 15.5; P ≤ 0.05). After ovulation, the number of free sperm in the isthmus increased, and these sperm expressed strong NOS activity. Conclusion: The change of sperm NOS activity is related to their release from the epithelium of the oviductal reservoir. (Reprod Med Biol 2003; 2: 75–81). [ABSTRACT FROM AUTHOR]