[Combined effects of aluminium and immobilization stress on the reproductive system of male rats].

The evaluation of the isolated and combined effects of aluminum and immobilization stress on the reproductive system of male laboratory rats has been performed. We have established the influence of these factors on spermatogenesis and spermiogenesis: the development of degenerative processes in the testes, on specific hormonal status, apoptosis, increasing abnormal forms of epididymal sperms and reducing their mobility, and on the growth of early embryonic losses. Neither synergistic nor antagonistic effect in the combined aluminum and the stress action has been found.

Adjudin targeting rabbit germ cell adhesion as a male contraceptive: a pharmacokinetics study.

Adjudin (1-(2,4-dichlorobenzyl)-1H-indazole-3-carbohydrazide; formerly called AF-2364) has been shown to inhibit spermatogenesis by disrupting anchoring junctions at the Sertoligerm cell interface. This, in turn, leads to germ cell loss from the seminiferous epithelium, and transient infertility. Adjudin's efficacyin inhibiting spermatogenesis, the recovery of spermatogenesis after cessation of the drug, and side effects were examined in adult male Japanese rabbits. The pharmacokinetics profiles of adjudin in rabbits after oral administration and after intravenous injection were compared. Rabbits received 25 mg/kg adjudin once weekly for 4 consecutive weeks either by intravenous injection or by gavage. Vehicle-treated rabbits were used as controls. At 1, 2, 3, 4, and 8 weeks after treatment, testes were removed for microscopic examination to assess the status of spermatogenesis. Four weeks after intravenous cessation of adjudin, the recovery of spermatogenesis also was monitored. Blood was withdrawn after first administration to measure plasma concentrations of adjudin by high-performance liquid chromatography. Four weeks after intravenous treatment, examination of testis sections showed rapid exfoliation of elongated/elongating spermatids and the presence of large multinucleated cells; more than 95% of germ cells were absent from the seminiferous epithelium. Intravenous treatment showed a more severe disturbance of spermatogenesis compared with gavage treatment, which was correlated with bioavailability of the drug. The areas under the curve for intravenous injection and gavage were 20.11 +/- 1.90 and 2.23 +/- 0.45 mg x h x L(-1), respectively. These results illustrate the potential of adjudin as a male contraceptive, and the efficacy is associated with the bioavailability of the drug.

Progress and prospects in male hormonal contraception.

PURPOSE OF REVIEW: Testosterone functions as a contraceptive by suppressing the secretion of luteinizing hormone and follicle-stimulating hormone from the pituitary. Low concentrations of these hormones deprive the testes of the signals required for spermatogenesis and results in markedly decreased sperm concentrations and effective contraception in a majority of men. Male hormonal contraception is well tolerated and acceptable to most men. Unfortunately, testosterone-alone regimens fail to completely suppress spermatogenesis in all men, meaning that in some the potential for fertility remains. RECENT FINDINGS: Because of this, novel combinations of testosterone and progestins, which synergistically suppress gonadotropins, have been studied. Two recently published testosterone/progestin trials are particularly noteworthy. In the first, a long-acting injectable testosterone ester, testosterone decanoate, was combined with etonogestrel implants and resulted in 80-90% of subjects achieving a fewer than 1 million sperm per milliliter. In the second, a daily testosterone gel was combined with 3-monthly injections of depot medroxyprogesterone acetate producing similar results. SUMMARY: Testosterone-based hormone combinations are able to reversibly suppress human spermatogenesis; however, a uniformly effective regimen has remained elusive. Nevertheless, improvements, such as the use of injectable testosterone undecanoate, may lead to a safe, reversible and effective male contraceptive.

Gamendazole, an orally active indazole carboxylic acid male contraceptive agent, targets HSP90AB1 (HSP90BETA) and EEF1A1 (eEF1A), and stimulates Il1a transcription in rat Sertoli cells.

Gamendazole was recently identified as an orally active antispermatogenic compound with antifertility effects. The cellular mechanism(s) through which these effects occur and the molecular target(s) of gamendazole action are currently unknown. Gamendazole was recently designed as a potent orally active antispermatogenic male contraceptive agent. Here, we report the identification of binding targets and propose a testable mechanism of action for this antispermatogenic agent. Both HSP90AB1 (previously known as HSP90beta [heat shock 90-kDa protein 1, beta]) and EEF1A1 (previously known as eEF1A [eukaryotic translation elongation factor 1 alpha 1]) were identified as binding targets by biotinylated gamendazole (BT-GMZ) affinity purification from testis, Sertoli cells, and ID8 ovarian cancer cells; identification was confirmed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry and Western blot analysis. BT-GMZ bound to purified yeast HSP82 (homologue to mammalian HSP90AB1) and EEF1A1, but not to TEF3 or HBS1, and was competed by unlabeled gamendazole. However, gamendazole did not inhibit nucleotide binding by EEF1A1. Gamendazole binding to purified Saccharomyces cerevisiae HSP82 inhibited luciferase refolding and was not competed by the HSP90 drugs geldanamycin or novobiocin analogue, KU-1. Gamendazole elicited degradation of the HSP90-dependent client proteins AKT1 and ERBB2 and had an antiproliferative effect in MCF-7 cells without inducing HSP90. These data suggest that gamendazole may represent a new class of selective HSP90AB1 and EEF1A1 inhibitors. Testis gene microarray analysis from gamendazole-treated rats showed a marked, rapid increase in three interleukin 1 genes and Nfkbia (NF-kappaB inhibitor alpha) 4 h after oral administration. A spike in II1a transcription was confirmed by RT-PCR in primary Sertoli cells 60 min after exposure to 100 nM gamendazole, demonstrating that Sertoli cells are a target. AKT1, NFKB, and interleukin 1 are known regulators of the Sertoli cell-spermatid junctional complexes. A current model for gamendazole action posits that this pathway links interaction with HSP90AB1 and EEF1A1 to the loss of spermatids and resulting infertility.

A novel potent indazole carboxylic acid derivative blocks spermatogenesis and is contraceptive in rats after a single oral dose.

Women have historically been the focus for development of new contraceptive methods. The National Institutes of Health, World Health Organization, and Institute of Medicine have stressed the need to develop nonhormonal, nonsteroidal male contraceptive agents. We report results from initial dose-ranging studies of a new indazole carboxylic acid analogue, gamendazole. An infertility rate of 100% was achieved in seven out of seven proven-fertile male rats 3 wk after a single oral dose of 6 mg/kg of gamendazole. Fertility returned by 9 wk in four of seven animals, with typical numbers of normal-appearing conceptuses. A fertility rate of 100% returned in four of six animals that became infertile at a single oral dose of 3 mg/kg of gamendazole. No differences in mating behavior were observed in either of the gamendazole-treated groups versus the control (vehicle-only) group. In the animals that showed reversible infertility, a transient increase in circulating FSH levels coincided with an initial decline in inhibin B levels after administration of gamendazole, but no other significant changes in circulating reproductive hormones were observed. Gamendazole inhibited production of inhibin B by primary Sertoli cells in vitro with a median inhibitory concentration of 6.8 thorn+/- 3.0 (SEM) (3/4)x 10(-10) M, suggesting that Sertoli cells are a primary target. A biotinylated gamendazole analogue revealed cytoplasmic and perinuclear binding of gamendazole in primary Sertoli cells. Gamendazole represents the most potent new oral antispermatogenic indazole carboxylic acid to date. Our results, however, demonstrate that additional dose-finding studies are required to improve reversibility and widen the therapeutic window before more detailed drug development of this potential nonhormonal male contraceptive agent can occur.

[Hormonal contraception in men]. / Hormonale anticonceptie bij de man.

Over the past few decades, female hormonal contraception has been seen to be very successful. However, this has still not resulted in a hormonal contraceptive for men. Certain injectable combinations ofandrogens and progestagens have been found to suppress spermatogenesis. All combinations that have been tested so far suffer from a relative lack of efficacy, a long lag time to achieve azoospermia, requiring the user to undergo one or more semen analyses, a moderate user friendliness, and concerns about the long-term safety and reversibility. It is not to be expected that male hormonal contraception will become a serious alternative to the already existing female equivalent during the coming 5 years.

Intratesticular injection of a zinc-based solution as a contraceptive for dogs.

The objective of the present study was to evaluate, by light and transmission electron microscopy, the efficacy of a single intratesticular injection of a novel zinc-based solution, as a contraceptive for male dogs. Fifteen mongrel dogs were assigned to three groups (five dogs/group). Group 1, the control group, which consisted of animals ranging from 8 mo to 4 yr, was injected with saline solution. Group 2, which consisted of animals ranging from 8 mo to 1 yr old and Group 3, animals ranging from 2 to 4 yr old, were injected with a zinc-based solution (0.2-1.0mL; volume based on testicular width). There were no histopathological changes detected in testes from control dogs. Histological examination of treated groups revealed degeneration, vacuolation, fewer germ cells, formation of multinucleated giant cells, and a lack of elongated spermatids in atrophic seminiferous tubules. Leydig cells had varying degrees of lipid degeneration and necrosis. The majority of seminiferous tubules in all zinc-treated dogs were lined only by Sertoli cells, which were vacuolated. Ultrastructure of testis of treated groups had degenerate Sertoli and Leydig cells, characterized by numerous mitochondria with the lack of a matrix and agglomeration of lysosomal bodies. The cytoplasm of elongated spermatids was characterized by tubules of hyperplastic and hypertrophic smooth endoplasmic reticulum and numerous Golgi apparati. Round spermatids in Golgi phase had lysis of acrosomal vesicles. The degree of histological changes suggested irreversibility. In conclusion, intratesticular injection of a zinc-based solution effectively impaired spermatogenesis.

7alpha-methyl-19-nortestosterone (MENT) vs testosterone in combination with etonogestrel implants for spermatogenic suppression in healthy men.

Testosterone with a progestogen can suppress spermatogenesis for contraception. The synthetic androgen 7alpha-methyl-19-nortestosterone (MENT) may offer advantages because it is resistant to 5alpha-reduction and is therefore less active at the prostate. This study aimed to investigate MENT implants in combination with etonogestrel on spermatogenesis, gonadotropins, and androgen-dependent tissues in comparison with a testosterone/etonogestrel regimen. Healthy men (n = 29) were recruited and randomized to receive 2 etonogestrel implants with either 600-mg testosterone pellets repeated every 12 weeks or 2 MENT implants for up to 48 weeks. Testosterone concentrations in the testosterone group remained in the normal range. Subjects with 2 MENT implants showed peak MENT levels at 4 weeks with testosterone concentrations of 2 nmol/L. Sperm concentrations fell rapidly to less than 1 x 10(6)/mL at 12 weeks in 8 of 10 subjects in the MENT group and 13 of 16 subjects in the testosterone group with equally suppressed gonadotropins. Thereafter, suppression was not maintained in the MENT group, and 6 men noted loss of libido. Fourteen men completed 48 weeks of testosterone treatment, and all became azoospermic. Hemoglobin concentrations rose, and high density lipoprotein-cholesterol (HDL-C) fell in both groups. The MENT group showed a fall in prostate-specific antigen with no change in bone mass. MENT with a progestogen can achieve rapid suppression of spermatogenesis similar to testosterone, but this promising result was not sustained due to a decline in MENT release from the implants. This dose of testosterone, compared with previous studies using a lower dose with a higher dose of etonogestrel, had nonreproductive side effects without any increase in spermatogenic suppression. These data indicate the importance of the doses of progestogen and testosterone for optimum spermatogenic suppression while minimizing side effects.

Preliminary investigation of the nasal delivery of liposomal leuprorelin acetate for contraception in rats.

The purpose of the study was to investigate the nasal route as a non-invasive alternative for delivery of leuprorelin acetate (leuprolide acetate, LEU) and to achieve an effective concentration of leuprorelin acetate in blood after nasal administration for contraception in rats. The plain drug solution, physical mixture (plain drug along with constituents of liposomes), or drug encapsulated in either neutral or charged liposomes containing 5 microg leuprorelin acetate were administered to rats through the nasal route. The plain drug solution was administered subcutaneously (s.c.). Simultaneous evaluation was performed on the influence of a mucoadhesive agent (chitosan) on nasal absorption of the plain drug and the liposome-encapsulated drug. Blood samples were taken at regular time intervals and subjected to luteinising hormone (LH) analysis using a specific immunoassay kit. The plasma luteinising hormone concentration vs time data of nasal and subcutaneous treatments were plotted and compared with that of subcutaneous administration. Relative percentage of bioavailability (F) for nasal treatments was calculated from plasma concentration vs time plots. Sperm count and fertility performance studies were carried out for selected formulations in rats. Neutral liposomes (LLEU) and negatively-charged liposomes (LLEUn) showed higher relative percentage of bioavailability (F 27.83 and 21.30%, respectively) as compared with the plain drug and the physical mixture (F 10.89 and 10.96%, respectively) after nasal administration. Hence, work on neutral liposomes was continued. F was further improved after incorporation of chitosan i.e. 10.89 to 49.13% for plain leuprorelin acetate and 27.83 to 88.90% for liposomal leuprorelin acetate formulations. Liposomal chitosan formulation administered nasally and leuprorelin acetate solution subcutaneously achieved complete azoospermia. No implantation sites were observed after the mating of female rats with treated males. It was observed that in the treated female rats, the estrous cycles ceased with the same formulations from the first treatment cycle. The findings of these investigations demonstrated that the bioavailability of the nasally-administered liposomal leuprorelin acetate with chitosan formulation was comparable with that of the subcutaneously administered drug. Complete contraception was obtained in male and female rats that had been treated with either the nasally administered liposomal leuprorelin acetate with chitosan or the subcutaneously administered drug.

Immune complex deposition in adult male Sprague-Dawley rats chronically immunized with GnRH.

PROBLEM: This study was undertaken to evaluate whether the anti-GnRH antibodies and immune complexes (IC) generated by immunization with GnRH-TT cause cellular damage within the animal. METHOD OF STUDY: Chronic immunization of rats with GnRH-TT injected i.m. was followed by tissue/organ analysis for immune complex deposition by immunofluorescence microscopy. Two groups were studied: (1) those immunized throughout the experiment until their ultimate demise, and (2) those given a chance to recover from the effects of chronic immunization before final analysis. RESULTS: GnRH-TT was effective in stopping spermatogenesis, which resumed after withdrawal of the immunogen. Most tissues from chronically immunized animals were not significantly different than controls, however the kidneys of treated animals exhibited a higher accumulation of IC. Despite increased IC deposition, pathologic effects were not detected at the cellular level. CONCLUSIONS: GnRH-TT is an effective immunocontraceptive although the accumulation of glomerular IC represents a potential deleterious side effect.

Oral desogestrel with testosterone pellets induces consistent suppression of spermatogenesis to azoospermia in both Caucasian and Chinese men.

BACKGROUND: Effective hormonal male contraception requires a high prevalence of spermatogenic suppression, which has proved particularly difficult in Caucasian populations. We have investigated the combination of oral desogestrel with depot testosterone in Caucasian and Chinese men. METHOD: Thirty men in Edinburgh and 36 men in Shanghai received 150 or 300 microg desogestrel p.o. daily for 24 weeks with 400 mg testosterone pellets s.c. on day 1 and at 12 weeks. RESULTS: Eight men withdrew before completing 24 weeks treatment. Testosterone concentrations remained within the normal range. Spermatogenesis was profoundly suppressed in all men. Azoospermia was achieved by a higher proportion of men in the 300 microg desogestrel group: 28/28 men versus 22/31 men (P < 0.05). All Caucasian men in the 150 microg group achieved sperm concentrations of < 1 x 10(6)/ml whereas three men in the Shanghai group maintained sperm concentrations of > 3 x 10(6)/ml. Fifteen men continued on this regimen for a subsequent 24 weeks: all remained azoospermic for the duration of treatment. High-density lipoprotein cholesterol fell by 15% in Caucasian men, but was unchanged in the Chinese men; both groups showed some weight gain. CONCLUSION: This combination of oral desogestrel with depot testosterone maintains physiological testosterone concentrations with consistent suppression of spermatogenesis to azoospermia in both Caucasian and Chinese men and therefore has many of the properties necessary for a contraceptive preparation for men.

Male contraceptive research steps back into spotlight.

PIP: The goal of male hormonal contraception is to block spermatogenesis by suppressing the secretion of pituitary gonadotropins. Prospects are good for the development of such a male contraceptive, with at least one hormonal contraceptive for men potentially available within 5-7 years. Finding an acceptable, reversible, and preferably long-lasting hormonal contraceptive for men is a top priority of the World Health Organization (WHO). Considerable evidence suggests that an androgen, with or without a progestogen, can provide effective contraception and is well tolerated. Schering and Organon are planning to collaborate with WHO in developing hormonal contraception for men. Fred Wu of the University of Manchester, UK, will soon begin a trial comparing the effectiveness of new injectable testosterone undecanoate and testosterone buciclate formulations, with or without synthetic progestogens.^ieng

Establishing the minimum effective dose and additive effects of depot progestin in suppression of human spermatogenesis by a testosterone depot.

Hormonally induced azoospermia induced by weekly im injections of testosterone enanthate provides effective and reversible male contraception, but more practical regimens are needed. Given our previous findings that six 200-mg pellets implanted subdermally produced more stable, physiological T levels and reduced the delivered T dose by more than 50% while maintaining equally effective suppression of sperm output with fewer metabolic side-effects than weekly 200-mg testosterone enanthate injections, we sought in this study to determine 1) whether further dose-sparing could be achieved by lower testosterone doses while maintaining efficacy and 2) the efficacy of adding a depot progestin to a suboptimally suppressive depot testosterone dose as a model depot progestin/androgen combination male contraceptive. Healthy volunteers were randomized into groups (n = 10) who received either of two lower T doses (two or four 200-mg T pellets) or four 200-mg T pellets plus a single im injection of 300 mg depot medroxyprogesterone acetate (DMPA). Two T pellets (400 mg, 3 mg/day) had a negligible effect on sperm output. Four T pellets (800 mg, 6 mg/day) suppressed sperm output between the second to fourth postimplant months; output returned to normal by the seventh postimplant month, although only 4 of 10 men became azoospermic or severely oligozoospermic (< 3 mol/L/mL). The addition of a depot progestin markedly increased the extent, but not the rate, of sperm output suppression, with 9 of 10 becoming azoospermic and 10 of 10 becoming severely oligozoospermic. There were no serious adverse effects during the study. Plasma total and free testosterone levels remained within the eugonadal range at all times with each treatment. Plasma epitestosterone was suppressed by all 3 regimens, consistent with a dose-dependent inhibition of endogenous Leydig cell steroidogenesis. Plasma LH and FSH measured by a two-site immunoassay were suppressed in a dose-dependent fashion by T and further suppressed by the addition of DMPA. Sex hormone-binding globulin levels were decreased by DMPA, but not by either T dose. Prostate-specific antigen and lipids (total, low or high density lipoprotein cholesterol, and triglycerides) were not significantly changed in any group. Thus, a depot testosterone preparation with zero order release must be delivered at between 6-9 mg/day to provide optimal (but not uniform) efficacy at inducing azoospermia. The addition of a single depot dose of a progestin to a suboptimal testosterone dose (6 mg/day) markedly enhances the extent, but not the rate, of spermatogenic suppression, with negligible biochemical androgenic side-effects. These findings provide a basis for the use of a progestin/androgen combination depot for hormonal male contraception.

PIP: Clinical research conducted in Australia suggests that a progestin-androgen combination depot has potential for hormonal male contraception. The authors' previous research had indicated that 6 200-mg testosterone enanthate pellets implanted subdermally produced substantial reductions over injections in the delivered testosterone dose while maintaining equally effective suppression of spermatogenesis with few metabolic side effects. The present study sought to determine whether lower testosterone doses would maintain efficiency and to assess the efficacy of adding a depot progestin to a suboptimally suppressive depot testosterone dose (6 mg/day). 10 volunteers received either 2 or 4 200-mg testosterone pellets or 4 200-mg pellets plus a single intramuscular injection of 300-mg depot medroxyprogesterone acetate (DMPA). The testosterone implants alone achieved inadequate suppression of spermatogenesis for a male contraceptive; 400 mg of testosterone (3 mg/day) had a negligible effect on sperm output, while 800 mg (6 mg/day) produced azoospermia or severe oligozoospermia in only 4 of 10 men. However, the addition of DMPA markedly increased the extent, but not the rate, of sperm output suppression: azoospermia was achieved in 9 men and oligozoospermia in all 10 subjects, and sperm suppression persisted for 3 months. Epitestosterone concentrations, used as a marker of Leydig cell steroidogenesis, were decreased in a time- and dose-dependent manner, reaching castrate levels in the combined group. Plasma luteinizing hormone and follicle-stimulating hormone levels were suppressed in a dose-dependent fashion by testosterone and further suppressed by the addition of DMPA. Sex hormone-binding globulin levels were decreased by DMPA, but not by either testosterone dose. Prostate-specific antigens and lipids were not significantly altered by any regimen. There were no discontinuations or reports of side effects.

Haemostatic effects of supraphysiological levels of testosterone in normal men.

The effects of exogenous testosterone on the haemostatic system were studied in a group of 32 healthy men undergoing a clinical trial of hormonal male contraception. The men received 200 mg testosterone oenanthate (TE) weekly i.m., and plasma samples were taken pretreatment, at defined time points up to 52 weeks of treatment, and 4 and 8 weeks after discontinuing TE. This dose of TE caused a 2-fold increase in trough plasma testosterone levels. TE caused a fall in plasma fibrinogen concentration after 16 weeks of treatment. This was sustained for the duration of TE treatment and recovered to pretreatment levels during the recovery phase. There was also a sustained fall in the level of C4b binding protein which showed a rebound to levels above pretreatment during recovery. Levels of antithrombin III and prothrombin fragment F1.2 rose initially during TE treatment, and levels of protein C, protein S (free) and plasminogen activator inhibitor fell, but the concentrations of these factors all returned to pretreatment levels during continued treatment. There was no change in the plasma concentrations of beta-thromboglobulin, tissue plasminogen activator, protein S (total), or D-dimer. There was a sustained increase in haemoglobin concentration and haematocrit, without any change in platelet count. The observed changes were consistent with mild activation of the haemostatic system during initial treatment with testosterone. After several months the raised activation markers had returned to pretreatment levels indicating that a new equilibrium had been established which did not appear to be prothrombotic.(ABSTRACT TRUNCATED AT 250 WORDS)

Vasectomía con esterilización temprana mediante nonoxinol intraductal / Vasectomy with early sterilization by means of intraductal nonoxinol injection

Se realizó un estudio prospectivo y comparativo de 36 pacientes que fueron sometidos a vasectomía. Se dividieron en dos grupos: grupo testigo A, 20 pacientes , y grupo de estudio B, 16 pacientes. El promedio de edad fue de 33.9 y 36.5 años, respectivamente. A todos los pacientes se les realizó espermatobioscopia basal, cuyo resultado se encontró dentro de límites normales. A los pacientes del grupo B se les aplicó nonoxinol intraductal durante la intervención con la finalidad de reducir el tiempo de fertilidad en el periodo postoperatorio. A todos los pacientes se les efectuó espermatobioscopia de control a la semana de la operación; los resultados obtenidos muestran, en el grupo A, persistencia del potencial de fertilidad, y en el grupo B, ocho pacientes con necrospermia total y, de los ocho restantes, 5 con inmovilidad de 100 por ciento y tres con oligospermia y astenospermia graves, lo que señala un potencial fértil nulo. LOs efectos colaterales consistieron en sensación de micción inmediata tras la aplicación del fármaco, dolor suprapúbico y datos de cistitis aguda, todos ellos transitorios. Se concluye que el uso del nonoxinol intraductal durante la vasectomía garantiza la esterilidad temprana