Journal of andrology最新文献

A variety of methods exist for counting sperm. Since the introduction of semen analysis, one of these methods, the hemocytometer, has been regarded as the gold standard by andrology laboratories and the World Health Organization (WHO, 1999). The flexible features of this approach, involving fixation and immobilization of sperm, dilution of highly concentrated samples, and the counting of sperm in a single plane, contribute to the accuracy of the Improved Neubauer hemocytometer and its relative ease of use. This method is strongly accepted within andrology clinics and has been clinically validated by a number of studies (Dunphy et al, 1989; Tomlinson et al, 1996, 1999; Guzick et al, 2001).

As technology and techniques improve, manufacturers are continually trying to develop newer, simpler, quicker and more accurate methods for determining sperm concentration. Busy assisted reproduction technology (ART) laboratories in particular would find a quicker yet comparatively accurate method highly desirable, since sperm counting is an essential part of the semen preparation process. Although modern methods may be faster, since unfixed, undiluted semen is used, some labs find these analyses more difficult to use and believe that the counting of motile sperm may produce erroneous results. Furthermore, the WHO states that the newly introduced methods “are convenient in that they can be used without dilution of the specimen, but that they may lack the accuracy of the hemocytometer technique especially for highly viscous and/or heterogeneous specimens. If such chambers are to be used, their adequate accuracy and precision must be established by comparison with hemocytometers” (WHO, 1999).

In addition, it is now a requirement of laboratory accreditation systems that laboratories provide clinical validation for all methods used (ie, demonstrate that they are “fit for purpose”). Currently, the only sperm counting method with a considerable body of evidence to support and clincally justify its use is the hemocytometer (Mortimer, 1994; WHO, 1999).

When using the hemocytometer, the sperm number is calculated using a fixed volume of semen under the coverslip and counting the sperm in a single plane. A significant association between pregnancy and the sperm concentration measured has consistently been shown for this method (Dunphy et al, 1989; Tomlinson et al, 1996, 1999; Guzick et al, 2001). Thus, 64% of laboratories involved in the analysis of semen use this method routinely (Keel et al, 2000).

However, despite its validity as a method, the use of hemocytometry is thought by many to be inconvenient, in that the hemocytometer must be cleaned and assembled prior to each counting event and it involves the use of dilution techniques that can introduce errors, either due to poor technique or the viscous nature of the semen itself. Mathematical mistakes can occur when applying the correction factor to determine the eventual counts, and the recom

ABSTRACT: Inconsistent results have been reported for the semen quality in HIV-infected men, due to the biases inherent in some studies. The objective of the present study was to investigate the semen parameters in HIV-1-infected patients and to compare their sperm characteristics with those of a control group of fertile, noninfected men. Factors implicated in semen alterations in HIV-1 patients were also analyzed. HIV-infected men (n = 190), of whom 91% were undergoing antiretroviral therapy, and 218 fertile men were studied. Infertility risk factors were recorded and clinical examinations were performed for both groups. Records of history of HIV infection, antiretroviral treatment, and HIV-1 RNA detection in the blood as well as HIV-1 genome detection in the semen were obtained for the infected patients. Semen volumes, percentages of progressive motile spermatozoa, total sperm counts, and polymorphonuclear cell counts were decreased, while the pH values and spermatozoa multiple anomaly indices were increased in HIV-infected patients. Even after adjustment for possible sources of bias, the decreases in semen volume and progressive motility and the increase in pH remained significant. The present study demonstrates sperm motility and ejaculate volume alterations in HIV-1-infected patients, most of whom were receiving antiretroviral therapy. In HIV-1 patients, further longitudinal studies are required to analyze the impact of treatment regimen on sperm parameter alterations.

ABSTRACT: Male hormonal contraceptive regimens function by suppressing gonadotropin secretion, resulting in a dramatic decrease in testicular androgen biosynthesis and spermatogenesis. Animal studies suggest that persistent intratesticular (iT)-androgen production has a stimulatory effect on spermatogenesis in the setting of gonadotropin suppression. We hypothesized that men with incompletely suppressed spermatogenesis (>1 000 000 sperm/mL) during male hormonal contraceptive treatment would have higher iT-androgen concentrations than men who achieved severe oligospermia (≤1 000 000 sperm/mL). Twenty healthy men ages 18–55 years enrolled in a 6-month male contraceptive study of transdermal testosterone (T) gel (100 mg/d) plus depomedroxyprogesterone acetate (300 mg intramuscularly every 12 weeks) with or without the gonadotropin releasing hormone (GnRH) antagonist acyline (300 μg/kg subcutaneously every 2 weeks for 12 weeks) were studied. During the 24th week of treatment, subjects underwent fine needle aspirations of the testes and iT-T and iT-dihydrotestosterone (iT-DHT) were measured in testicular fluid by liquid chromatography—tandem mass spectrometry. All men dramatically suppressed spermatogenesis; 15 of 20 men were severely oligospermic, and 5 of 20 suppressed to 1.5 million −3.2 million sperm per milliliter. In all subjects, mean iT-T and iT-DHT concentrations were 35 ± 8 and 5.1 ± 0.8 nmol/L. IT-androgen concentrations did not significantly differ in men who did and did not achieve severe oligospermia (P = .41 for iT-T; P = .18 for iT-DHT). Furthermore, there was no significant correlation between iT-T or iT-DHT and sperm concentration after 24 weeks of treatment. In this study of prolonged gonadotropin suppression induced by male hormonal contraceptive treatment, differences in iT-androgens did not explain differences in spermatogenesis. Additional studies to identify factors involved in persistent spermatogenesis despite gonadotropin suppression are warranted.

ABSTRACT: Two enzymes are involved in the polyol pathway: an aldose reductase that reduces glucose in sorbitol followed by its oxidation in fructose by sorbitol dehydrogenase. It has been previously shown that both enzymes are presented in the bovine epididymis, where they are associated with membranous vesicles called epididymosomes. Based on the distribution of these enzymes, it has been hypothesized that the polyol pathway can modulate sperm motility during the epididymal transit. In the present study, polyol pathway was investigated in semen and along the epididymis in humans in order to determine if sperm maturation can be associated with this sugar pathway. Western blot analysis shows that both aldose reductase and sorbitol dehydrogenase are associated with ejaculated spermatozoa and prostasomes in humans. These enzymes are also associated with epididymosomes collected during surgical vasectomy reversal. Western blot, Northern blot, and reverse transcription—polymerase chain reaction analysis show that aldose reductase and sorbitol dehydrogenase are expressed at the transcriptional and translational levels along the human epididymis. Unlike what occurs in the bovine model, distribution of these enzymes is rather uniform along the human excurrent duct. Immunohistological studies together with Western blot analysis performed on epididymosomes preparations indicate that the polyol pathway enzymes are secreted by the epididymal epithelium. These results indicate that the polyol pathway plays a role in human sperm physiology.

The greatest paradox of aerobic respiration is that oxygen, which is essential for energy production, may also be detrimental because it leads to the production of reactive oxygen species (ROS) (Saleh and Agarwal, 2002). When levels of reactive oxygen species (ROS) overwhelm the body's antioxidant defense system, oxidative stress (OS) occurs. OS is a condition in which the elevated levels of ROS damage cells, tissues, or organs (Moller et al, 1996; Sharma and Agarwal, 1996; Saleh et al, 2003).

ROS are free radicals that play a significant role in many of the sperm physiological processes such as capacitation, hyperactivation, and sperm-oocyte fusion (Aitken et al, 2004; Allamaneni et al, 2004; de Lamirande et al, 1998). However, they also trigger many pathological processes in the male reproductive system, and these processes have been implicated in cancers of the bladder and prostate, as well as in male infertility (Bankson et al, 1993; Hietanen et al, 1994; Agarwal and Saleh, 2002).

Spermatozoa are sensitive to OS because they lack cytoplasmic defenses (Donnelly et al, 1999; Saleh and Agarwal, 2002). Moreover, the sperm plasma membrane contains lipids in the form of polyunsaturated fatty acids, which are vulnerable to attack by ROS. ROS, in the presence of polyunsaturated fatty acids, triggers a chain of chemical reactions called lipid peroxidation (Agarwal et al, 1994; Kobayashi et al, 2001; Zalata et al, 2004). ROS can also damage DNA by causing deletions, mutations, and other lethal genetic effects (Moustafa et al, 2004; Tominaga et al, 2004). It is difficult to block the OS-induced injury to cells or tissues because ROS are continuously produced by cellular aerobic metabolism (Davies, 2000). Several clinical trials are currently attempting to minimize the toxic effects of OS on human spermatozoa (Agarwal et al, 2004). In this review, we highlight the various protective measures available to minimize OS-induced injury to spermatozoa.

There are two main sources of ROS in semen: leukocytes and immature spermatozoa (Garrido et al, 2004). Of these, leukocytes are considered to be the primary source (Aitken et al, 1992). Leukocytes, particularly neutrophils and macrophages, have been associated with excessive ROS production that ultimately leads to sperm dysfunction (Aitken and Baker, 1995; Aitken et al, 1997; Hendin et al, 1999; Ochsendorf, 1999; Pasqualotto et al, 2000; Saleh et al, 2002; Shalika et al, 1996; Sharma et al, 2001).

Spermatozoa produce ROS mainly when a defect occurs during spermiogenesis that results in retention of cytoplasmic droplets (Gomez et al, 1996; Zini et al, 1993). A strong positive correlation exists between immature spermatozoa and ROS production, which in turn negatively affects the sperm quality (Gil-Guzman et al, 2001; Said et al, 2004).

The two main sites of ROS production are the mitochondrion and the sperm plasma membrane. The mitochondrion is the powerhouse of respiration. Hence

Perhaps one of the most exciting and revolutionary scientific discoveries of the past 3 decades has been the development of in vitro fertilization (IVF) to treat human infertility. It is impossible to quantify its effect on numerous families since the first IVF birth in 1978 in Old-ham, England (Steptoe and Edwards, 1978). With increasing clinical utilization of assisted reproductive technologies (ART), scientists and clinicians gain insights into basic gamete and embryo biology and translate that knowledge into improving the process of IVF. Critical analyses of individual steps have improved outcomes. Attention to sperm processing and isolation to increase recovery of motile sperm and reduce sperm damage has improved fertilization rates and embryo development (Mortimer, 1994). Use of intracytoplasmic sperm injection (ICSI) has allowed fertilization even in severe cases of compromised sperm quality or number (Bonduelle et al, 1999). Finally, refinement of embryo culture has led to improved in vitro embryo development and implantation rates (Gardner and Lane, 1998; Pool, 2002). Most scientific attention, however, has focused on methodologies rather than technology development and equipment. Semen is still processed in test tubes regardless of technique, sperm are physically placed with oocytes after processing, and fertilization and embryo culture occur in culture dishes, test tubes, or both with relatively large volumes (Trounson and Gardner, 2000). With the exception of gamete and embryo micromanipulation, no technologic advancements in IVF have reached widespread use. Nevertheless, it is precisely those technologic advancements, rather than procedural or methodology changes, that have had the greatest effect on assisted reproduction.

A promising new technology, microfluidics, exists and is becoming increasingly studied. This technology shows promise as an alternative for each step in the IVF process. Microfluidics, based on physical principles of fluid behavior in a microenvironment, has been used widely in chemistry and molecular biology applications (Tomlinson et al, 1995). Currently, microfluidics is gaining interest in studies of cellular behavior and interactions (Shim et al, 2003). In this article, we introduce basics of fluid behavior at the microscale and highlight previous uses of this technology outside of the reproductive sciences. We then describe fabrication of devices and review initial studies that used microfluidics in sperm sorting and microinsemination. Last, we point out some limitations of this new technology and provide speculation on future directions and application of microfluidics in ART.

Fluid mechanics is a complex physical and mathematical science; therefore, an extensive technical description and review of fluid physics is beyond the scope and intent of this review. Instead, basic principles will be discussed that govern fluid behavior in a microenvironment, especially those aspects with a specific li

ABSTRACT: The idea that varicocele plays a detrimental role in fertility is supported by the presence of a higher frequency of affected men among the infertile population than among men with normal semen parameters. In this research we examined ejaculates from a large group of selected men affected by varicocele by light and electron microscopy. The effect of varicocele on chromosome meiotic segregation was investigated by fluorescence in situ hybridization (FISH). The potential benefits of varicocelectomy on sperm quality were evaluated by analyzing sperm characteristics before and after surgical correction of varicocele. Transmission electron microscopy (TEM) analysis, elaborated previously, showed that the incidence of immaturity, apoptosis, and necrosis was higher in the varicocele group than in controls. FISH analysis performed on sperm nuclei from selected patients with varicocele showed that the mean frequencies of disomies and diploidies were generally out of the normal range, indicating a severe disturbance in meiotic segregation. Sperm characteristics evaluated before and after varicocele repair showed a general improvement. As a consequence, the varicocele seem to affect sperm morphology and function concomitantly with meiotic segregation derangement. In consideration of these data, we suggest that TEM and FISH analyses should be performed for all varicocele patients.

The interest in possible medical interventions to promote healthy aging has been recently increasing, as the absolute number and the proportion of men over 60 years of age is expected to increase during the next few decades in various countries (Liu et al, 2004). Numerous studies have demonstrated lower concentrations of testosterone in older men (Vermeulen et al, 1972; Rubens et al, 1974; Pirke and Doerr, 1975; Baker et al, 1976; Purifoy et al, 1981; Bremner and Prinz, 1983; Tenover et al, 1987; Gray et al, 1991; Ferrini and Barrett-Connor, 1998). Serum testosterone concentrations have been shown to decrease longitudinally with age (Morley et al, 1997; Zmuda et al, 1997; Harman et al, 2001), but estimates of the rate of this fall in testosterone levels may differ substantially based on the type of data analysis (Liu et al, 2004). Aging has been reported to be associated with decreased muscle mass, muscle strength, physical performance, physical activity, bone mineral density, and libido (Davidson et al, 1983; Santavirta et al, 1992; Nguyen et al, 1996; Rantanen et al, 1998; Anonymous, 2002; Hughes et al, 2002; Liu et al, 2004). The presence of a combination of these nonspecific clinical features may indicate organic androgen deficiency. Thus, testosterone replacement therapy may be of special importance in this age group, as the anabolic effects of this hormone on muscle, fat, and bone may contribute to improvement in physical function and quality of life. However, various factors may be involved in determining the clinical significance of this age-related decline in serum testosterone, as well as the safety and benefit of testosterone replacement therapy in older men, including the rate of decrement in systemic testosterone exposure, possible reduced androgen responsiveness of older tissues, and the rising age-related background rates of certain androgen-dependent cardiovascular and prostatic disorders.

This review concentrates on the key issues associated with testosterone replacement therapy in older men, including the background for this intervention, the available testosterone formulations, and their possible adverse effects, and it also provides suggested protocols for screening and monitoring patients before and during this treatment, respectively.

A health factor—independent, age-related longitudinal decrease in serum testosterone levels has been reported (Harman et al, 2001). As there is no agreement on the definition of hypogonadism in older men, a combination of clinical signs and testosterone measurements is usually used as a tool to determine whether testosterone replacement therapy is indicated. The most easily recognized clinical signs of relative androgen deficiency in older men are a decrease in muscle mass and strength, a decrease in bone mass and osteoporosis, and an increase in central body fat. However, symptoms such as a decrease in libido and sexual desire, forgetfulness, loss of memory, difficulty in concen

ABSTRACT: To investigate the possible role of testicular orphan receptors (TR) TR2, TR3, and TR4 in the process of germ cell apoptosis in the heat-treated testis of monkey, we have examined the spatiotemporal expression of the 3 TR mRNAs in relation to p53 mRNA levels in the monkey testis by in situ hybridization and reverse transcription polymerase chain reaction techniques. The results showed that TR2 mRNA was confined to spermatocytes; TR4 and TR3 mRNAs were expressed in both spermatocytes and spermatids. The heat treatment did not change TR2 mRNA level but significantly reduced TR4 mRNA expression in spermatocytes on days 3 and 8 after the heat treatment. TR3 mRNA expression was affected by the heat treatment in a time-dependent manner, with the lowest level on day 30 after the heat shock. Low to moderate signal for p53 mRNA was detected in spermatocytes before treatment, which increased dramatically on days 3, 8, and 30 after the heat shock. The coincident expression of the testicular TR3 and p53 mRNA, spatially and time dependently, implied that the decrease in TR3 expression in the heat-treated testis might be closely related to the p53 signal pathway, whereas the temporal decrease in TR4 production in the testis at the early stage indicated that this orphan receptor might be also involved in germ cell apoptosis. The data suggest that TR3, TR4, and p53 could be important regulators of germ cell apoptosis induced by the heat treatment, whereas TR2 might not be a key regulator in this process.

ABSTRACT: The purpose of the study was to evaluate 90K/MAC-2BP, a glycoprotein member of the Scavenger Receptor Cystein Rich superfamily, in the seminal plasma of infertile male patients with male accessory gland infection in order to investigate a putative autoimmune pathogenesis. 90K seminal concentration and sperm parameters were evaluated in 50 patients with male accessory gland infection at baseline and after cycles of treatment with Levofluoxacin 500 mg daily for 15 days plus serratiopeptidase 10 mg daily for 30 days. Treatment was continued for up to 6 cycles in cases of persistant bacteriospermia and/or clinical and ejaculatory signs of the disease. Patients with persistant male accessory gland infection after 6 cycles were defined as nonresponders. The same parameters were evaluated at baseline and after a 2-month period in 30 healthy controls. Patients with male accessory gland infection showed impaired sperm parameters and had lower seminal 90K concentration compared to controls. After treatment, seminal 90K level significantly increased in patients compared to controls. Twenty-two patients responded to treatment (44%), while 28 were nonresponders (56%). No difference in pretreatment and posttreatment sperm parameters and seminal 90K was observed between the 2 subgroups. Thirteen patients (26%) had identifiable bacteriospermia: significantly less pretreatment seminal 90K was observed compared to patients without bacteriospermia. Seminal 90K is decreased in patients with male accessory gland infection, and may be restored by a treatment with quinolones. However, the clinical utility of a 90K assay in these patients remains uncertain, as its level is not predictive of response to treatment.