Journal of andrology最新文献

The central theme of the XXth North American Testis Workshop, “Testicular Function: Levels of Regulation,” reflected the many recent discoveries of new and complex levels of regulation of testicular functions. These included regulation of testicular development and the initial formation of the testis and male germ cells, as well as the subsequent differentiation of key components of the testis, including Sertoli cells, Leydig cells, and spermatogenic cells. Many different regulatory mechanisms responsible for these differentiative functions were explored. The roles of genes encoding key regulatory proteins, as well as signal transduction mechanisms, RNA-processing mechanisms, regulation by small noncoding RNAs, and mechanisms governing self-renewal and/or differentiation of spermatogonial stem cells were all discussed in this light. The workshop, which was held at the Hyatt Regency Philadelphia at Penn's Landing in Philadelphia, Pennsylvania, on April 1–4, 2009, featured 15 invited talks and 6 short talks selected from abstracts submitted for the 2 poster sessions. Manuscripts from 12 of the invited talks are presented in this volume. They are organized into 4 parts: “Regulation of Testis Development,” “Regulation of Testis Function,” “Regulation of Germ Cell Development,” and “Regulation of Gamete Development and Function.”

“Part 1: Regulation of Testis Development” features 4 articles. The first, by Nel-Themaat et al, describes an elegant and cutting-edge approach to the study of differentiation of the testis involving expression of fluorescent markers in specific cell types within the developing testis as a means to visually follow the dynamics of testicular development and differentiation. This summary is preceded by a brief review of past studies aimed at elucidating the cellular dynamics associated with testicular development. The use of transgenes that differentially mark individual testicular cell types in conjunction with culture of developing testes and time-lapse imaging facilitates unprecedented insight into the developmental dynamics of testicular differentiation. This approach allows the investigators to understand aspects of testicular development that cannot be ascertained in any other way. In addition, this provides a very visual and, hence, very instructive tool that is attractive to both experts and nonexperts alike and should stimulate all readers to want to learn more about the genesis of the testis. The second article in this part is by Barsoum and Yao, and is focused on the origins of Leydig cells in the fetal testis. The authors note that these cells arise after the initial appearance of Sertoli cells induced by expression of the testis-determining Sry gene, and suggest that the origin of Leydig cells is likely based on or regulated by factors derived from Sertoli cells. They cite evidence suggesting that this process depends on a balance between differentiation-promoting and -suppressing mechanisms, such a

Three sperm-counting methods were compared within and between 3 centers to determine the sensitivity and reproducibility of assessing low sperm concentrations. Two methods were performed by phase contrast microscopy with and without centrifugation, and 1 method was performed by fluorescence microscopy (using the DNA stain Hoechst 33342) without centrifugation. Semen samples were serially diluted in fluorescent dye-containing fixative, and sperm concentrations were assessed in duplicate in the central field (100 nL) of reusable Neubauer chambers (phase contrast microscopy), in the whole field of disposable 25-μL Leja chambers (fluorescence microscopy), and in wet preparations (up to 1950 microscopic fields) of the pellet obtained after centrifugation at 3000 × g for 15 minutes (phase contrast microscopy). Agreement among the 3 participating centers was good, with lower limits of quantification (the concentrations for which counting errors [the standard error of the number of spermatozoa counted expressed as a percentage of the count] are ≤20%) determined to be 150 000 /mL for the Neubauer chamber (phase contrast microscopy) and 500/mL for the Leja chamber (fluorescence microscopy). These are equivalent to 300 000 /mL and 1000 /mL for undiluted semen. The centrifugation method consistently, seriously, and significantly underestimated mean sperm concentration compared with the other 2 methods by an average of 49%. In conclusion, the accurate measurement of low sperm counts is facilitated by the use of large-volume chambers and fluorescence microscopy, and this permits the definition of lower limits of sperm concentrations for azoospermic samples.

The absence of spermatozoa from the ejaculate has always been an important criterion for diagnosing infertility, for proving success of vasectomy, and currently for determining the efficacy of hormonal contraception. Its assessment, however, has never been easy, for reasons relating to the methodology and counting errors at very low sperm concentrations. Despite calls for a change in the definition of azoospermia to include its etiology, treatment, and prognosis (Sharif, 2000; Ezeh and Moore, 2001), in the andrology laboratory it remains a description of the semen analyzed, that is, the absence of spermatozoa from an ejaculate (World Health Organization [WHO], 1999). However, given the problems of measuring low sperm numbers, it is appropriate to reassess its definition in statistical terms and provide the sensitivity of methods routinely used to assess this condition so that the diagnoses and prognoses alluded to above can be performed from good evidence.

It is generally accepted that “should only a few or no spermatozoa be seen at initial evaluation, the sample must be centrifuged and the sediment examined for spermatozoa. The term azoospermia can only be used if no spermatozoa have been found in the sediment” (Eliasson, 1981). Where centrifugation has been used to concentrate the fe

ABSTRACT: Previous studies showed that interleukin-6 (IL-6) was expressed in human Leydig and Sertoli cells and that it inhibited sperm motility. The aim of this study was to compare the expression of IL-6, IL-6R, and GP130 in ejaculated spermatozoa between normozoospermic and asthenozoospermic men. Human spermatozoa in the semen were purified by Percoll gradient technique to separate the seminal plasma and other round cells. RT-PCR, immunocytochemistry, and Western blot were used to detect the expression of IL-6, IL-6R, and GP130 in spermatozoa. With RT-PCR, only GP130 mRNA but not IL-6 and IL-6R mRNA was expressed in human ejaculated spermatozoa. The expression of GP130 mRNA was significantly lower in asthenozoospermic men than in normozoospermic men. The protein expression of GP130 was further confirmed by both immunocytochemistry and Western blot. Again, GP130 protein levels were significantly lower in asthenozoospermic men than in normozoospermic men. The results suggested that the decreased expression of GP130 in ejaculated spermatozoa could be associated with low sperm motility in asthenozoospermic men.

ABSTRACT: Recently, we reported that male accessory sex gland (ASG) secretions protect sperm genomic integrity by demonstrating that DNA damage was more extensive in sperm not exposed to the secretions. The present study was conducted to find out if ASGs secrete the main antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase (GPx or GSH-Px), and catalase (CAT) and if the most abundant one, SOD, can protect those sperm that were not exposed to ASG secretions against NADPH-induced oxidative stress. Four experimental groups of male golden hamsters were used: intact animals with proven fertility, animals with all major ASGs removed (TX), animals that were bilaterally vasectomized, and sham-operated controls. SOD, CAT, and GPx activities were measured in secretions from all 5 ASGs and sperm-free uterine flushing from virgin females and those mated with the experimental males. The alkaline comet assay was used to analyze DNA integrity of the TX group sperm after incubation in a medium containing 50 U/mL of SOD along with 0 to 20 mmol/L NADPH. The main antioxidant enzyme in ASGs was SOD from coagulating glands (P < .05) and GPx together with CAT from ampullary glands (P < .05). Uterine flushing of ejaculates that contained ASG secretions had more SOD and CAT activities than those with epididymal secretions alone (P < .05 and P < .001, respectively), whereas activity of GPx was the same (P > .05). Addition of SOD in vitro dose dependently decreased the incidence of single-strand DNA damage in sperm not exposed to ASG secretions incubated in the presence of 0 to 20 mmol/L NADPH (P < .001). These results indicated that, in terms of abundance, SOD was the main antioxidant enzyme secreted by male ASGs, whereas CAT was the second one. The GPx activity came from both epididymis and ASGs. We conclude that ASG secretions play a significant role in protecting sperm against oxidative stress.

ABSTRACT: The rabbit is well suited for infertility research because of the animal's size, relatively low cost, and accessibility of the genitalia. There are several studies reporting sperm count and motility in rabbits; however, no easily reproducible or inexpensive device for semen collection has been reported. Herein, we report the construction of an inexpensive, effective artificial vagina assembled from easily obtainable products that may be used to collect rabbit ejaculates. This device was used to perform 243 ejaculation trials on 17 rabbits. Eighty-six percent (209/243) of the trials resulted in rabbit mounts. Overall, 91% (191/209) of mounts resulted in successful semen collection. After 60 mounts, a 98% successful ejaculate collection rate was achieved. The cost of the reusable device is $2.64, plus silicone condoms at $7.50 each. This practical artificial vagina for semen collection is an inexpensive and effective means of obtaining semen from rabbits for andrologic study.

In a heartbeat, we are there. Twenty-five years ago, Dr Nancy Alexander, President of the American Society of Andrology (ASA), delivered a Presidential Address at the 1980 ASA Annual Meeting in Chicago where she shared with us her perceptions for the future of andrology by the year 2000. This “state-of-the-art” address, titled “Andrology in the Year 2000,” was published in its entirety in the first volume of the Journal of Andrology (J Androl. 1980;1:149–157). It's a wonderful set of predictions, and we encourage our readers to go back to this manuscript and read (or reread) her insightful comments made at a time when the field of andrology was relatively new. The focus of Dr Alexander's comments and predictions for the year 2000 was not the entire field of andrology, but rather, 2 facets that are her area of expertise: 1) advances in male contraception, and 2) basic and clinical studies on development and maintenance of male fertility. In celebration of the Silver Anniversary of the Journal of Andrology, the 2004 ASA Presidents now reflect on Dr Alexander's comments and describe how the subsequent events during the past 25 years have confirmed or changed her predictions for andrology in the year 2000.

In 1978, National Institutes of Health (NIH) funding for population research had grown considerably from the previous 15 years and reached a total of $112 million set aside for the year. The ratio of male-female reproductive system funding was approximately 1:2, which was a vast improvement over the 1:4 ratio in 1972 (Alexander, 1980). The future for male reproductive research was promising. To address the current status of NIH funding for reproductive research, we asked the National Institute of Child Health and Human Development (NICHD) of the NIH to provide data on their funding levels during the past 5 years (1999–2003) in the areas of male and female reproductive research. It is important to stress that these numbers reflect NICHD funding only and do not include male reproductive system research by other institutes such as the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), the National Institute on Aging (NIA), and the National Institute of Environmental Health Sciences (NIEHS), all of which have research programs that include the male reproductive tract. Nonetheless, the NICHD is considered the primary institute for reproductive research and can be used as an indicator of fiscal commitments to reproductive research. Three categories were defined for analysis and are shown in the Figure: 1) targeted male reproductive health (research that applies only to male reproductive health [eg, endocrine regulation of germ cell apoptosis in the male, examinations of male fertility, Sertoli cell development]), 2) targeted female reproductive health (research that applies only to female reproductive health [eg, gonadotropin secretion during lactation, progestin regulation of uterine hemostasis and an

To the Editor:

The publication by Luboshitzky et al (2002) concerning the effects of melatonin on human sperm quality deserves strong critique for a number of reasons. The authors performed a double-blind crossover study during which healthy volunteers were given either melatonin (3 mg) or a placebo for 3 months each, while between the phases, a washout phase of 2 weeks was included. At the beginning and end of the 2 phases, a total of 11 parameters (sperm and endocrine) were measured and, again, 3 and 6 months after the end of the study. According to the results (!!), volunteers were divided into 2 groups, responders (n = 2) and nonresponders (n = 6). The criterion was that both sperm concentration and sperm motility “dropped during the melatonin treatment period.” Both of these men belonged to the group to which melatonin was given in the second treatment period. The title and the conclusions of this paper are simply not justified by the data for the following reasons:

In summary, I see no evidence whatsoever for the conclusion that melatonin impairs sperm parameters in healthy men. It rather appears that an expected result influenced the way the data were handled.

To the Editor:

We appreciate the comments of Lerchl (2004) regarding the effect of exogenous melatonin on sperm quality in normal men (Luboshitzky et al, 2002). In this study, we examined the possible effect of melatonin on semen concentration, motility, and morphology in 8 healthy young men. Since we did not study fertility in the general sense, we performed one semen analysis at baseline. We fully agree that 2 semen samples are indicated for the initial evaluation of fertility. In our study, we defined a subject as a responder if his sperm concentration and motility dropped during the melatonin treatment period. The individual results of semen analysis were given in our study in Figure 1. In 2 men, we found decreases in sperm concentration and motility that were below the normal range (WHO, 1993). Since the sequence of medications in these subjects was a placebo followed by melatonin, we concluded that the decrease in semen quality was associated with melatonin administration. We also observed an increase in sperm concentration in 3 subjects during melatonin administration. These counts were within the reference range and were not associated with similar changes in sperm motility. We attributed these changes to the well-known variations between samples that exist in the same individual (WHO, 1993). The data presented by Lerchl in the figure describe the average (±SEM) values for sperm concentration for all 8 volunteers examined. It is obvious from our study that, as a group, no trend is seen during melatonin treatment.

We also determined fasting serum gonadotropins and testosterone and estradiol levels. Although testosterone is secreted in a diurnal fashion (Luboshitzky et al, 2003), a single time point in the morning is sufficient for

The 33rd American Society of Andrology Annual Meeting was held in Albuquerque, NM on April 12–15, 2008. Under the theme “Emerging Concepts and Technologies in Andrology,” the program was specifically designed and developed to appeal to the wide range of interests of ASA members, spanning both basic science and clinical medicine, and bridging bench work and patient care.

ASA Keynote Lecture. Stem Cells, Small RNAs and Self-Renewal in the Germline

Haifan Lin, Ph.D., Yale University School of Medicine

Dr Haifan Lin presented the ASA Keynote Lecture on Stem Cells, Small RNAs and Self-Renewal in the Germline. A recent focus of his research has been on small RNA-mediated epigenetic programming and translational regulation that are required for stem cell self-renewal in the germline. Piwi/argonaute genes represent the only known family of genes required for stem cell renewal in both animal and plant kingdoms. The Piwi (P-element induced wimpy testis) family binds a class of short RNAs called piRNAs (for Piwi associated RNAs) that is expressed primarily in the germline. The Drosophila protein Piwi is involved in stem cell maintenance; in piwi mutants stem cells differentiate without self-renewal. Dr Lin also reviewed the germ cell phenotypes of mutations in Piwi family members in a number of species including mouse and human. Piwi has been implicated in heterochromatin formation and epigenetic silencing of genes. piRNAs exist in large numbers, with over 60,000 species to date, and are transcribed from a limited number of regions in the genome. The precise role of piRNAs in the testis is unknown. Dr Lin's group has recently shown in Drosophila that Piwi proteins can have a role in transcriptional activation in addition to their role in transcriptional repression. His group showed that Piwi can bind to a subtelomeric heterochromatic region on chromosome 3 (known as 3R-TAS) and a piRNA uniquely mapped to 3R-TAS, leading to epigenetic activation of the 3R-TAS locus. His findings reveal an increased level of complexity of small RNA-mediated epigenetic regulation, i.e. that Piwi can exert opposite effects (activation versus repression) on different genomic regions. The physiological role(s) of the Piwi-piRNA system in stem cells is currently an active area of study.

AUA Lecture. Amniotic Cells as Stem Cell Source for Tissue Engineering

Anthony Atala, MD, Wake Forest University

Dr Atala gave an enlightened and informative plenary AUA lecture on regenerative medicine in urology and other fields. The talk began with a brief history of organ engineering beginning with kidney transplants almost 50 years ago. Dr Atala reviewed the 3 issues that have limited the field of organ and tissue regenerative medicine in the past, and explained how he and other are tackling these barriers. The first limitation is that it has historically been difficult to grow many cell types outside the body. The discovery and systematic isolation of co

ABSTRACT: Efferent ductules of the male reproductive tract contain high concentrations of estrogen receptors (ER), which are essential for the regulation of fluid reabsorption and maintenance of normal epithelial morphology. Treatments with the antiestrogen ICI 182,780 and 17β-estradiol cause a reduction in ERα expression; however, the mechanisms governing the down-regulation are undetermined. In other tissues, the ubiquitin-proteasome pathway appears to have a dominant role in regulating ERα turnover, although in the efferent ductules, an abundance of epithelial lysosomes could also participate in protein turnover. To study this activity, the expressions of proteasome, ubiquitin, and markers for the endocytotic apparatus (early endosome antigen-1 [EEA1], clusterin, and cathepsin D) were examined in rat efferent ductules and initial segment of epididymis. Distinct cellular, subcellular, and regional distributions of these proteins were observed in the epithelial cells. A gradient of proteasome, ubiquitin, EEA1, and clusterin staining was seen in the efferent ducts, which decreased 30%–41% from the proximal zone to the terminal common duct. Antiestrogen treatment resulted in significant decreases in proteasome, EEA1, and clusterin in the efferent ducts. Localization of ubiquitin-proteasome and endocytotic pathway components suggests that differential regulation is required for protein degradation and turnover in efferent ductules and head of the epididymis.