Vas deferens, a site of male contraception: an overview

The vas deferens is a site which can be exploited for male contraception without undue side effects. The only ef-fective technique available for male contraception is vasectomy, being practiced world wide, despite that it is a perma-nent surgical procedure and its successful reversal is not assured. Although no-scalpel vasectomy minimizes surgicalprocedures, the fate of its reversal is akin to that of vasectomy. Several occlusive and non-occlusive vasal procedureswhich claim to be reversible without surgical intervention, possess more disadvantages than advantages. Vas occlusionwith plug, ' Shug' or medical grade silicone rubber, although claimed to produce reversible azoospermia without affect-ing spermatogenesis, requires skilled microsurgery for their implantation and later removal. RISUG^R, a non-scleroticpolymer styrene maleic anhydride (SMA), could be more advantageous than vasectomy and other vas occlusive proce-dures in that it could be a totally non-invasive procedure by 'no-scalpel injection' and 'non-invasive reversal'. It isclaimed to offer long-term contraception without adverse side effects and also to be possible as a male spacing methodby repeated vas occlusion and non-invasive reversal. The drug is currently under multicentre Phase Ⅲ clinical trial.

Development of methods of male contraception: impact of the World Health Organization Task Force

Objective: To give an historical record of the research of the World Health Organization (WHO) Task Force to develop methods of male contraception; to examine the social, political, medical, pharmaceutical, funding, and other factors that influenced progress; and to suggest reasons why such methods are only now becoming available. Design: Review of basic and clinical research over 30 years. Setting: Task force of a multinational agency and collaborating agencies. Conclusion(s): Through the involvement of many international scientists, the WHO Task Force has uniquely contributed to the exploratory phases of the research in male contraception and by its multicenter contraceptive efficacy studies has accelerated progress towards the ideal hormonal method. Despite an adverse climate involving social and political attitudes, funding constraints, and pharmaceutical industry hesitations, WHO formed coalitions with governments and international agencies to sustain research with results that apply to men in culturally diverse populations and thereby to influence activities across the whole range of global reproductive health and family planning.

Predictors for partial suppression of spermatogenesis of hormonal male contraception

Aim： To analyze factors influencing the efficacy of hormonal suppression of spermatogenesis for male contraception. Methods： A nested case-control study was conducted, involving 43 subjects, who did not achieve azoospermia or severe oligozoospermia when given monthly injections of 500 mg testosterone undecanoate （TU）, defined as partial suppressors compared with 855 subjects who had suppressed spermatogenesis （complete suppressors）. Sperm density, serum testosterone, luteinizing hormone （LH） and follicle stimulating hormone （FSH） concentrations at the baseline and the suppression phase were compared between partial and complete suppressors. Polymorphisms of androgen receptor （AR） and three single nucleotide variants and their haplotypes of FSH receptor （FSHR） genes determined by polymerase chain reaction （PCR） and DNA sequencing technique were compared between 29 partial and 34 complete suppressors. Results： Baseline serum LH level was higher and serum LH as well as FSH level during the suppression phase was less suppressed in partial suppressors. Additionally, in a logistic regression analysis larger testis volume, higher serum FSH concentrations alone, or interaction of serum LH, FSH, testosterone and sperm concentrations were associated with degree of suppression. The distribution of polymorphisms of AR or FSH receptor genes did not differ between partial and complete suppressors. In cases with incomplete FSH suppression （FSH 〉 0.2 IU/L）, the chances of reaching azoospermia were 1.5 times higher in the subjects with more than 22 CAG triplet repeats. Conclusion： Partial suppression of spermatogenesis induced by 500 mg TU monthly injections is weakly influenced by hormonal and clinical features but not polymorphism in AR and FSHR genes.

Development of Newer Approaches to Male Contraception: Prospects of Availability for Mass Application in the Near Future

Progress in development of a simple, effective reversible male contraceptive has been difficult due to the indispensable role of testosterone in spermatogenesis and maintenance of secondary sexual characters. Efforts are in progress to develop methods, which do not interfere with testosterone production. These include blockade of FSH action by immunization against FSH or FSHR, and interfere with sperm maturation by interfering with estrogen action or by immunization against epididymal or sperm specific proteins. An evaluation of these various approaches for practical application is also presented.

Low dose gossypol for male contraception

Aim: To ascertain whether the side effects of gossypol, hypokalemia and irreversibility, could be avoided on dose re-duction. Methods: Seventy-seven male volunteers were divided into 3 groups: control (22 cases), 10 mg gossypol(29 cases) and 12.5 mg (26 cases). Serum levels of testosterone, FSH and LH were measured by RIA and potassiumby flame photometry. Spema counts and motility were examined before and regularly after treatment for the evaluationof contraceptive efficacy. Results: The average sperm density and motility started to decrease significantly by theend of month 2 of medication and gradually reached the infertility levels ( < 4 million /mL) in both treated groups. Af-ter that the 10 mg group was asked to take the same dose every other day for up to a total observation period of 16-18months for the maintenance of infertility. Subjects in the 12.5 mg group did not take gossypol any more so as to ob-serve the length of the loading dose required, but in a few, a maintenance dose of 12.5 mg every other day was insti-tuted for a few more months. In both treated groups, none of the spouses was pregnant during the maintenance dose pe-riod. Serum levels of potassium, FSH, LH and testosterone were not significantly changed and not a single volunteercomplained of myoasthenia. After cessation of drug administratioin, the semen data returned to pretreatment levels.Conclusion: A regimen with 10 or 12.5 mg of gossypol as the daily loading dose and 35 or 43.75 mg as the week-ly maintenance dose could induce infertility in male volunteers without developing hypokalemia or irreversibility.(Asian J Androl 2000 Dec; 2: 283-287)

Male contraception:prospects for the new millennium

Effective regulation of human fertility has global consequences in terms of resource depletion, pollution and pover-ty. Current family planning services predominantly target a female clientele with few significant developments in malefertility regulation for over a century. The last two decades have witnessed a gathering interest, initially from the scien-tific community, and laterally from industry, in the development of safe, reliable, reversible methods of contraceptionfor men. This review summarises the methods of male fertility regulation which are currently available and critically ex-amines the published data on novel developments in male hormonal contraception which offer the potential of improvedcontraceptive choice for all in new millennium. (Asian J Androl 2000; 2: 3 - 12)

Hormonal contraception for human males:prospects

Development of an ideal hormonal contraceptive for man has been the goal of several research workers during thepast few decades. Suppression of pituitary gonadotropic hormones, which in turn would inhibit spermatogenesis whilemaintaining normal libido and potentia has been the approach for a contraceptive agent. Intramuscularly administeredand orally active testosterone or testosterone in combination with progesterone have been shown to cause inhibition ofspermatogenesis resulting in azoospermia in normal men. Similarly testosterone has been used in combination with go-nadotropin releasing hormone antagonists and agonists to inhibit pituitary gonadotropic hormone release. Immunologicalapproach to neutralize the circulating levels of follicle stimulating hormone has also been shown to cause inhibition ofspermatogenesis. The available literature shows that testosterone causes reversible azoospermia without any significantside effects in Asian population effectively and appears to be a promising chemical for control of fertility in man.

Contraceptive effect of Curcuma longa (L.) in male albino rat

Aim: To study the contraceptive effect of the crude extracts of Curcuma longa in male albino rats. Methods: Rats were fed orally with Curcuma longa aqueous and 70 % alcoholic extract for 60 days (500 mg·kg-1· day-1). Results: A reduction in sperm motility and density was observed in both the treated groups. Conclusion: Curcuma longa may have affected the androgen synthesis either by inhibiting the Leydig cell function or the hypo-thalamus pituitary axis and as a result, spermatogenesis is arrested.

The preparation and application of N-terminal 57 amino acid protein of the follicle-stimulating hormone receptor as a candidate male contraceptive vaccine

Follicle-stimulating hormone receptor （FSHR）, which is expressed only on Sertoli cells and plays a key role in spermatogenesis, has been paid attention for its potential in male contraception vaccine research and development. This study introduces a method for the preparation and purification of human FSHR 57-amino acid protein （FSHR-57aa） as well as determination of its immunogenicity and antifertility effect. A recombinant pET-28a（＋）-FSHR-57aa plasmid was constructed and expressed in Escherichia coil strain BL21 StarTM （DE3） and the FSHR-57aa protein was separated and collected by cutting the gel and recovering activity by efficient refolding dialysis. The protein was identified by Western blot and high-performance liquid chromatography analysis with a band of nearly 7 kDa and a purity of 97.4%. Male monkeys were immunized with rhFSHR-57aa protein and a gradual rising of specific serum IgG antibody was found which reached a plateau on day 112 （16 weeks） after the first immunization. After mating of one male with three female monkeys, the pregnancy rate of those mated with males immunized against FSHR-57aa was significantly decreased while the serum hormone levels of testosterone and estradiol were not disturbed in the control or the FSHR-57aa groups. By evaluating pathological changes in testicular histology, we found that the blood-testis barrier remained intact, in spite of some small damage to Sertoli cells. In conclusion, our study demonstrates that the rhFSHR-57aa protein might be a feasible male contraceptive which could affect sperm production without disturbing hormone levels.

Regulation of fertilization in male rats by CatSper2 knockdown

Interest in ion channels as drug targets for contraception has grown with the realization that certain ion channel subunits are located exclusively in sperm. Selective knockdown of ion channel subunits can lead to infertility without ill effects, and selective inhibitors and/ or openers of these ion channels could interfere with sperm function. In this study, in vivo electmporation （EP） and rete testis microinjection-mediated plasmid DNA were adopted to silence CatSper2 expression, which is essential in sperm hyperactivation. The results showed that high transfection efficiency and expression were achieved by plasmid DNA that was directly injected into the rete testis. As a result of the expression of CatSper2 being blocked, the treatment group showed significantly lower （P〈0.05） hyperactivation rate, fertilization rate in vitro, migration motility in viscoelastic solution and intracellular Ca2＋ peak. The low hyperactivation and fertilization rates lasted for 60 days. Meanwhile, analysis of the sperm survival rate and testis histology indicated that in vivo EP had no significant effect on the function of the testis, spermatogenesis or sperm activity. The present study demonstrated that it was feasible to achieve male contraception by silencing the expression of CatSper2, the key protein involved in sperm hvoeractivation.

Does ethnicity matter in male hormonal contraceptive efficacy？

The development of male hormonal contraception has progressed significantly during the last three decades. The ultimate goal is to produce an effective, safe and reversible male method of contraception that are within reach of and can be used by all men globally. This review aims to outline the recent developments in male hormonal contraception with special emphasis on how ethnicity influences acceptability, extent of sperm suppression, and rate of recovery of spermatogenesis. Baseline differences in testicular histomorphology and testosterone metabolism between East Asian and Caucasian men have been reported, but whether this contributes significantly to varying degrees of sperm suppression in response to exogenous testosterone therapy is less known. Testosterone alone male hormonal contraceptive regimens are effective and applicable for East Asian men, and less so for Caucasians. Combinations of progestins with androgens are sufficient to optimize effectiveness of suppression and applicability to all ethnicities. New compounds such as steroidal or non-steroidal selective androgen receptor modulators with dual androgenic and progestational activities are potential compounds for further development as male hormonal contraceptive methods. At the present time, combined androgen and progestin contraceptive regimens appear to be effective, safe, reversible and convenient to use for all men with ethnic, cultural and environmental differences. Further refinements on the hormonal agent, methods of delivery, and dose optimization of the androgen relative to the progestin are necessary. This goal mandates further investment and large clinical trials in multiethnic populations to better define safety and efficacy.

Safety evaluation of long-term vas occlusion with styrene maleic anhydride and its non-invasive reversal on accessory reproductive organs in langurs

Aim:To evaluate the safety of the long term vas occlusion with styrene maleic anhydride (SMA) and its non-invasive reversal at the level of accessory reproductive glands (ARGs) in langurs.Methods:The morphology of seminal vesicle and ventral prostate was evaluated by light as well as transmission electron microscopy.Serum clinical chemistry and urine albumin were evaluated in an autoanalyzer using reagent kits.Fructose,acid phosphatase and zinc in the seminal plasma were evaluated spectrophotometrically according to the WHO manual.Serum testosterone, prostate specific antigen and sperm antibodies were evaluated by enzyme-linked immunosorbent assays (ELISA) using reagent kits and hematology was estimated according to standard procedures.Results:The morphological features and secretory activity of the seminal vesicle and prostate were normal as evidenced by the presence of well- developed mitochondria,rough endoplasmic reticulum,Golgi bodies,secretory granules and normal nuclear charac- teristics throughout the course of investigation.Serum testosterone and prostate specific antigen remained unaltered and serum antisperm antibodies level presented negative titres.Urine albumin was nil.Total red blood corpuscles (RBC),white blood corpuscles (WBC),hemoglobin (Hb) and red cell indices,serum protein,glucose,cholesterol, creatinine,creatine kinase (CK),serum glutamate oxalate transaminase (SGOT),serum glutamate pyruvate transami- nase (SGPT),lactate dehydrogenase (LDH),bilirubin,urea,triglycerides and high-density lipoprotein (HDL) did not show appreciable changes following vas occlusion and after its non-invasive reversal.Although fructose,acid phos- phatase (ACP) and zinc in the seminal plasma showed a significant reduction following vas occlusion,it could not be related to the morphology of seminal vesicle and prostate.Conclusion:SMA vas occlusion and its non-invasive reversal do not damage the accessory reproductive organs.

RNA Interference-Mediated Downregulation of sAC Gene Inhibits Sperm Hyperactivation in Male Rats(Rattus norvegicus)

Hyperactivation is one of the most critical parts for fertilization, cAMP generated by soluble adenylyl cyclase （sAC） is necessary to activate sperm and is a prerequisite for sperm hyperactivation. The aim of this study is to investigate the function of sAC in hyperactivation in male rats. Four siRNAs of sAC gene were designed and separately transformed into rat sperm using electrotransformation method. Cultured for 12 and 24 h, physiological and biochemical indexes of these sperm were analyzed, and the expressions of some hyperactivation-related genes were detected using real-time PCR. We demonstrated 26.3-30.8% and 49.1-50.5% reduction in sAC at the protein by Western blot and mRNA levels by real-time PCR, respectively. The results showed that two siRNAs, Actb-717 and Actb-4205, were the best RNAi sites for silencing sAC. The VCL （curvilinear velocity） and ALH （amplitude of lateral head displacement） of RNA interference （RNAi）-transfected sperm were reduced, cAMP and protein phosphorylation in RNAi transfected sperm were also decreased. The hyperactivation-related genes, such as CatSper2, LDHC and PKA, were downregulated in the sperm, which sAC was knockdown. These findings demonstrated that sAC might play a critical role in cAMP signaling in the rat sperm hyperactivation, and downregulated sAC gene might prevent the expression of these hyperactivation-ralated genes resulting in sperm dysfunction. These findings suggest that these hyperactivation-ralated genes and sAC are functionally related in sperm hyperactivation and sAC falls into an expanding group of sperm proteins that appear to be promising targets for the development of male contraceptives.

More than eight years＇ hands-on experience with the novel long-acting parenteral testosterone undecanoate

Testosterone （T） as a compound for treatment of T deficiency has been available for almost 70 years, but the pharmaceutical formulations have been less than ideal. Traditionally, injectable T esters have been used for treatment, but they generate supranormal T levels shortly after the 2-3 weekly injection interval. T levels then decline very rapidly, becoming subnormal during the days preceding the next injection. The rapid fluctuations in plasma T are subjectively experienced as disagreeable. T undecanoate （TU） is a new injectable T preparation with a considerably better pharmacokinetic profile. After two initial injections separated by a 6-week interval, the following intervals between two injections are generally 12 weeks, eventually amounting to a total of four injections per year. Plasma T levels with this preparation are nearly always in the range of normal men, as are its metabolic products estradiol and dihydrotestosterone （DHT）. It reverses the effects of hypogonadism on bone and muscle and metabolic parameters, and on sex functions. It is suitable for male contraception. Its safety profile is excellent because of the continuous normalcy of plasma T levels. No polycythemia has been observed and no adverse effects on lipid profiles. Prostate safety parameters are well within reference limits. TU is a valuable treatment option of androgen deficiency.

Sperm motility inhibitory effect of the benzene chromatographic fraction of the chloroform extract of the seeds of Carica papaya in langur monkey, Presbytis entellus entellus

Aim： To assess the contraceptive efficacy of the benzene chromatographic fraction of the chloroform extract of the seeds of Carica papaya in langur monkeys. Methods： The test substance was given p.o. to five monkeys at 50 mg/kg body weight/day for 360 days. Control animals （n = 3） received olive oil as vehicle. Sperm parameters as per World Health Organization standards, sperm functional tests, morphology of testis and epididymis, haematology, clinical biochemistry, serum testosterone and libido were evaluated. Following completion of 360 days treatment the animals were withdrawn from the treatment and the recovery pattern was assessed by semen analysis and sperm functional tests. Results： Total inhibition of sperm motility was observed following 60 days of treatment that continued until 360 days study period. Sperm count, percent viability and percent normal spermatozoa showed a drastic decline following 30 days of treatment. Sperm morphology showed predominant mid piece abnormalities. Sperm functional tests scored in sterile range. Histology and ultrastructure of testis revealed vacuolization in the Sertoli cells and germ cells. Loss of cytoplasmic organelles was evident in spermatocytes and round spermatids. Histology and ultrastructure of epididymis of treated animals were comparable to those of control animals. Hematological and serum clinical parameters and testosterone levels fluctuated within the control range throughout the study period. Recovery was evident following 60--120 days of treatment withdrawal. Conclusion- The results suggest that the benzene chro- matographic fraction of the chloroform extract of the seeds of Carica papaya shows contraceptive efficacy without adverse toxicity, mediated through inhibition of sperm motility. （Asian JAndro12008 Mar; 10： 298-306）

Histological study of vas deferens following intravasallaser irradiation

<abstract>Aim: To study the histologic changes of the vas deferens following Nd: YAG laser irradiation. Methods: Intravasal laser irradiation was given to (i) 52 segments of rabbit (laser dosage: 2 seconds at 40 W-50 W) and 16 segments of human (3 seconds at 45 W-55 W) vas deferens in vitro, (ii) 25 rabbit vasa (2 seconds-2.5 seconds at 40 W-45 W) in vivo and (iii) 2 human vasa (3 seconds at 55W) in vivo. Segments of vasa were removed from the in vivo irradiated vasa deferentia 15 days-180 days (rabbit) or 15 days (man) after the exposure. All vas segments were embedded in methacrylate resin. Serial sections (thickness 25μm-30μm) were obtained and observed under a light microscope. Results: (i) Laser-induced damage reached the muscularis layer in 27 % and 94 % of the rabbit and human vas segments in vitro, respectively, (ii) Fourteen of the 25 in vivo rabbit vasa were completely occluded by fibrous tissue and the longer the time interval after treatment, the more likely was the vas occluded. Those unoccluded vasa had either a normal histology or a mucosal damage, (iii) One in vivo human vas was almost completely occluded by the fibrous tissue but the other had a relatively large lumen packed with sperm granulomatous tissue and partial destruction of the smooth muscle layer. Conclusion: Laser irradiation can induce long-term vas occlusion; for rapid occlusion, laser doses just completely destroying the mucosal layer will be advisable.

Preparation and immunogenicity of tag-free recombinant human eppin

Human epididymal protease inhibitor （eppin） may be effective as a male contraceptive vaccine. In a number of studies, eppin with an engineered His6-tag has been produced using prokaryotic expression systems. For production of pharmaceutical-grade proteins for human use, however, the His6-tag must be removed. This study describes a method for producing recombinant human eppin without a His6-tag. We constructed plasmid pET28a （＋）-His6-tobacco etch virus （TEV）-eppin for expression in Escherichia coli. After purification and refolding, the fusion protein His6-TEV-eppin was digested with TEV protease to remove the His6-tag and was further purified by NTA-Ni2＋ affinity chromatography. Using this procedure, 2 mg of eppin without a His6-tag was isolated from 1 I of culture with a purity of 〉95%. The immunogenicity of the eppin was characterized using male Balb/c mice.

Study on Differences in Spermatogenic Suppression between Azoospermic and Oligozoospermic Responders Treated with Levonorgestrel Implant Plus Testosterone Undecanoate Injectable in Chinese Men

Objective To investigate possible causes resulting in the differences in the spermatogenesis suppression on individual treated with levonorgestrel (LNG) implants and testosterone undecanoate (TU) injectableMethods Totally 21 Chinese male volunteers were given treatment with LNG implants (four rods, 75 mg/rod) and intramuscular injection of TU (500 mg,bimonthly for 3 times). According to the effects of treatment, they were divided into two groups, namely, azoospermia group (group A) and oligozoospermia group (group O). Then seminal FSH, LH, T and estradiol (E2) were determined by immunoenzymetric assay, while seminal and serum dihydrotachysterol (DHT) and serum sex hormone binding globulin (SHBG) were by radioimmunoassay, and seminal transferrin (Tf) by scatter turbidimetry assay.Results Seminal FSH, LH and serum DHT, SHBG, FTI (T/SHBG ×100) levels were significantly lower in group A than in group O, while higher seminal concentrations ofE2 were observed in azoospermia group.Conclusion The differences in the spermatogenic suppression in Chinese men might be attributed to different rate of peripheral androgen metabolism, variations in serum SHBG levels, 5á-reductase activity and individual aromatase activity during LNG plus TU administration. In addition, seminal sex hormones might be more sensitive indexes to assess the extent of feedback inhibition on hypothalamus-pituitary-testis with exogenous testosterone plus progestogen in the efficacy hormone male contraceptive trials.

Differences in reproductive endocrinology between Asian men and Caucasian men——a literature review

This review provides an overview of the literature on aspects of reproductive endocrinology wherein Asian menmay differ from Caucasian, notably, prostatic nioplasm and the sensivity to pharmacological regimens of male contra-ception. Both genetic and environmental factors, such as nutrition, might be relevant. Asian men residing in Asia seemto be relatively protected from clinical prostatic nioplasm while the prevalence of preclinical prostatic nioplasm is notdifferent. Migration to an area with a higher prevalence reduces this difference but does not undo it. With regard toprostatic nioplasm the following factors have been considered as relevant in Asian men: 1) a reduction in 5α-reductaselevel, 2) decreased levels of androgenic ketosteroid precursors of 5α-reduced androgen metabolites, 3) the decreasedpresence of a P53 mutation, 4) a higher CAG-repeat length of the androgen receptor, 5) a possible higher level ofphysical activity, 6) differences in sexual activity. Furthermore, Asian men respond to a higher degree with azoosper-mia in response to contraceptive steroids. Possible explanations offered for the more pronounced response to contracep-tive steroids are: 1) differences in testicular structure and decreased spermatogenic potential, 2) an earlier and moremarked suppression in LH secretion by exogenous androgens. The differences may be due to genetical and/or environ-mental factors influencing the peripheral testosterone metabolism. Dietary factors such as the higher intake of phytoe-strogens in Asians might exert effects on 5α-reductase activity and/or on sex hormone binding globulin (SHBG) levels,thus having an impact on the biological efficay of circulating androgens. ( Asian J Androl 2000; 2: 13 - 20)

Non-invasive reversal of intraluminal vas deferens polymer injection-induced azoospermia——technology

Aim: The rationale and technique underlying a novel concept of non-invasive removal of an intravasal vas deferens poly-meric contraceptive drug to reverse drag injection-induced azoospermia are explained. Thus the conventional methods ofsurgical exploration to remove vas deferens plugs and intravasal injection of solvents to flush out contraceptive drugs are tobe replaced by steps which will be readily accepted by subjects. Methods: The approach is based upon the non-invasiveapplication of specific forces to various segments of the vas deferens so that non-sclerosing and non-tissue-adherent com-pounds, in particular styrene maleic anhydride (SMA) can be expelled. Forces are generated by palpation; percuta-neous electrical stimulation; vibration application; and percussion. The forces help to propel the intravasal polymer to-wards the ejaculatory duct for expulsion during ejaculation. All aspects of the total technique are clinically acceptable,simple, atraumatic, unlikely to cause pain and discomfort even without tranquilizers, local or general anaesthetics. Theprocedure may be repeated several times in different sittings spaced apart by about one week to achieve adequate plug ex-pulsion. Results: Model experiments demonstrated the feasibility of the concept. The polymer was nonadherent andcould be moved within the vas deferens by the application of specific forces. Sufficient removal was possible to enablespermatic fluid to be transported along a region previously occupied by the polymer. A corroborating subhuman primatestudy by an independent investigator has shown that the semen profile becomes normal following the reversal. Conclu-sion: Adoption of the new technique may provide a means of non surgical restoration of normal semen profile after a pe-riod of fertility control obtained by intravasal drag injection. (Asian J Androl 1999 Sep ; 1: 131 - 134)