Interactions of the male contraceptive target EPPIN with semenogelin-1 and small organic ligands.

Novel male contraceptives will promote gender equality in sharing contraceptive responsibility. The sperm-associated protein epididymal protease inhibitor (EPPIN) is a promising target for non-hormonal male contraception. EPPIN interacts with the semen coagulum protein semenogelin-1 (SEMG1) on the sperm surface, leading to transient inhibition of sperm motility after ejaculation. Small organic molecules targeting EPPIN's SEMG1-binding are under development as male contraceptives. Here, we combined computational approaches to uncover key aspects underlying EPPIN binding to SEMG1 and small organic ligands. We generated a human EPPIN model showing a typical arrangement of the WFDC (Whey-acid four disulfide core)-type and Kunitz-type domains, connected by a hinge region. Determining the EPPIN model's intrinsic motion by molecular dynamics simulations and normal mode analysis revealed a conformation, presenting a binding pocket that accommodates SEMG1Glu229-Gln247, EP055, and EP012. EPPIN's residues Phe63 and Lys68 (WFDC domain), Asp71 (hinge region), and Asn113, Asn114, and Asn115 (Kunitz domain) were identified as hot spots for SEMG1, EP055, and EP012 binding. Moreover, hydrophobic and hydrophilic residues in the WFDC and Kunitz domains allow plasma membrane anchoring, orienting the EPPIN binding pocket to the solvent. Targeting EPPIN's essential residues for its biomolecular interactions may improve the rational design of EPPIN ligands as spermiostatic compounds.

Advances in non-hormonal male contraception targeting sperm motility.

BACKGROUND: The high rates of unintended pregnancy and the ever-growing world population impose health, economic, social, and environmental threats to countries. Expanding contraceptive options, including male methods, are urgently needed to tackle these global challenges. Male contraception is limited to condoms and vasectomy, which are unsuitable for many couples. Thus, novel male contraceptive methods may reduce unintended pregnancies, meet the contraceptive needs of couples, and foster gender equality in carrying the contraceptive burden. In this regard, the spermatozoon emerges as a source of druggable targets for on-demand, non-hormonal male contraception based on disrupting sperm motility or fertilization. OBJECTIVE AND RATIONALE: A better understanding of the molecules governing sperm motility can lead to innovative approaches toward safe and effective male contraceptives. This review discusses cutting-edge knowledge on sperm-specific targets for male contraception, focusing on those with crucial roles in sperm motility. We also highlight challenges and opportunities in male contraceptive drug development targeting spermatozoa. SEARCH METHODS: We conducted a literature search in the PubMed database using the following keywords: 'spermatozoa', 'sperm motility', 'male contraception', and 'drug targets' in combination with other related terms to the field. Publications until January 2023 written in English were considered. OUTCOMES: Efforts for developing non-hormonal strategies for male contraception resulted in the identification of candidates specifically expressed or enriched in spermatozoa, including enzymes (PP1γ2, GAPDHS, and sAC), ion channels (CatSper and KSper), transmembrane transporters (sNHE, SLC26A8, and ATP1A4), and surface proteins (EPPIN). These targets are usually located in the sperm flagellum. Their indispensable roles in sperm motility and male fertility were confirmed by genetic or immunological approaches using animal models and gene mutations associated with male infertility due to sperm defects in humans. Their druggability was demonstrated by the identification of drug-like small organic ligands displaying spermiostatic activity in preclinical trials. WIDER IMPLICATIONS: A wide range of sperm-associated proteins has arisen as key regulators of sperm motility, providing compelling druggable candidates for male contraception. Nevertheless, no pharmacological agent has reached clinical developmental stages. One reason is the slow progress in translating the preclinical and drug discovery findings into a drug-like candidate adequate for clinical development. Thus, intense collaboration among academia, private sectors, governments, and regulatory agencies will be crucial to combine expertise for the development of male contraceptives targeting sperm function by (i) improving target structural characterization and the design of highly selective ligands, (ii) conducting long-term preclinical safety, efficacy, and reversibility evaluation, and (iii) establishing rigorous guidelines and endpoints for clinical trials and regulatory evaluation, thus allowing their testing in humans.

"Uma pílula difícil de engolir?" Representações sociais do anticoncepcional masculino no Facebook / "A hard pill to swallow?" Social representations of the male contraceptive pill on Facebook

Resumo A pílula anticoncepcional feminina é um dos principais métodos contraceptivos utilizados na atualidade; já a pílula anticoncepcional masculina, apesar de ter sido pensada há mais de 50 anos, ainda não é comercializada. O objetivo do presente trabalho é compreender aspectos psicossociais da dinâmica da sociedade em relação a novas formas contraceptivas e examinar se novas formas contraceptivas direcionadas ao corpo do homem seriam acatadas. Em março de 2018, a revista Superinteressante postou em sua página do Facebook uma reportagem sobre uma pílula anticoncepcional masculina que foi bem-sucedida em sua primeira fase de teste. A postagem gerou grande comoção e surgiram muitos comentários expressando diferentes opiniões sobre o assunto. Foram submetidos à análise de conteúdo temático-categorial 294 comentários feitos nessa postagem. Os resultados revelam que esse método contraceptivo masculino aparece inscrito em um sistema representacional que salienta discussões sobre: efeitos colaterais da medicalização, a liberdade feminina, a ampliação da responsabilidade masculina na contracepção/cuidado com os filhos, a desconfiança em relação à postura feminina e outros aspectos que remetem a possíveis avanços científicos e sociais. Os dados analisados não podem ser generalizados, mas apontam para a aceitação de novas formas contraceptivas direcionadas ao corpo masculino.

ABSTRACT The female contraceptive pill is one of the main contraceptive methods used today, while the male contraceptive pill, despite being thought of more than 50 years ago, is not yet commercialized. This work aims to understand psychosocial aspects of the dynamics of society in relation to new forms of contraception and to examine whether new forms of contraception directed to the male body would be accepted. In March 2018, the magazine Superinteressante posted on Facebook a report about a male contraceptive pill that was successful in its first test phase ; the post generated great commotion and many comments emerged expressing different opinions on the subject. 294 comments made on this post were submitted to thematic-categorical content analysis. The results indicate that this male contraceptive method appears inscribed in a representational system that highlights discussions about: side effects of medicalization, female freedom, the expansion of male responsibility in contraception/child care, distrust of the female posture and other aspects that refer to possible scientific and social advances. The analyzed data cannot be generalized, but point to the acceptance of new forms of contraception aimed at the male body.

Dissecting EPPIN protease inhibitor domains in sperm motility and fertilizing ability: repercussions for male contraceptive development.

EPPIN (epididymal protease inhibitor) is a mammalian conserved sperm-binding protein displaying an N-terminal WFDC (whey-acidic protein four-disulfide core) and a C-terminal Kunitz protease inhibitor domains. EPPIN plays a key role in regulating sperm motility after ejaculation via interaction with the seminal plasma protein SEMG1 (semenogelin-1). EPPIN ligands targeting the SEMG1 binding site in the Kunitz domain are under development as male contraceptive drugs. Nevertheless, the relative contributions of EPPIN WFDC and Kunitz domains to sperm function remain obscure. Here, we evaluated the effects of antibodies targeting specific epitopes in EPPIN's WFDC (Q20E antibody, Gln20-Glu39 epitope) and Kunitz (S21C and F21C antibodies, Ser103-Cys123 and Phe90-C110 epitopes, respectively) domains on mouse sperm motility and fertilizing ability. Computer-assisted sperm analysis showed that sperm co-incubation with S21C antibody (but not F21C antibody) lowered progressive and hyperactivated motilities and impaired kinematic parameters describing progressive (straight-line velocity; VSL, average path velocity; VAP and straightness; STR) and vigorous sperm movements (curvilinear velocity; VCL, amplitude of lateral head movement; ALH, and linearity; LIN) compared with control. Conversely, Q20E antibody-induced milder inhibition of progressive motility and kinematic parameters (VAP, VCL and ALH). Sperm co-incubation with S21C or Q20E antibodies affected in vitro fertilization as revealed by reduced cleavage rates, albeit without changes in capacitation-induced tyrosine phosphorylation. In conclusion, we show that targeting specific epitopes in EPPIN Kunitz and WFDC domains inhibits sperm motility and capacitation-associated events, which decrease their fertilizing ability; nevertheless, similar observations in vivo remain to be demonstrated. Simultaneously targeting residues in S21C and Q20E epitopes is a promising approach for the rational design of EPPIN-based ligands with spermostatic activity.

Influence of Echeveria gibbiflora DC aqueous crude extract on mouse sperm energy metabolism and calcium-dependent channels.

ETHNOPHARMACOLOGY RELEVANCE: In traditional Mexican medicine, Echeveria gibbiflora DC has been used as a vaginal post-coital rinse to prevent pregnancy. The aqueous crude extract (OBACE) induces sperm immobilization/agglutination and a hypotonic-like effect, likely attributed to the high concentration of calcium bis-(hydrogen-1-malate) hexahydrate [Ca2+ (C4H5O5)2•6H2O]. Likewise, OBACE impedes the increase of [Ca2+]i during capacitation. AIM OF THE STUDY: Evaluate the effect of OBACE on sperm energy metabolism and the underlying mechanism of action on sperm-specific channel. MATERIAL AND METHODS: In vitro, we quantified the mouse sperm immobilization effect and the antifertility potential of OBACE. The energetic metabolism status was also evaluated by assessing the ATP levels, general mitochondrial activity, mitochondrial membrane potential, and enzymatic activity of three key enzymes of energy metabolism. Furthermore, the effect of the ion efflux of Cl- and K+, as well as the pHi, were investigated in order to elucidate which channel is suitable to perform an in silico study. RESULTS: Total and progressive motility notably decreased, as did fertility rates. ATP levels, mitochondrial activity and membrane potential were reduced. Furthermore, the activities of the three enzymes decreased. Neither Cl- or K+ channels activities were affected at low concentrations of OBACE; nevertheless, pHi did not alkalinize. Finally, an in silico analysis was performed between the Catsper channel and calcium bis-(hydrogen-1-malate) hexahydrate, which showed a possible blockade of this sperm cation channel. CONCLUSION: The results were useful to elucidate the effect of OBACE and to propose it as a future male contraceptive.

Quem tomará a "pílula masculina"? Reflexões sobre a construção do usuário de contraceptivos para homens / ¿Quién tomará la "píldora masculina"? Reflexiones sobre la construcción del usuario de anticonceptivos para hombres / Who will take the "male pill"? Reflections on the construction of the contraceptive user for men

Resumo O objetivo do artigo é analisar como é concebido o usuário de um anticoncepcional masculino por um ator relevante no campo, a organização não governamental estadunidense Male Contraception Initiative. A partir de uma abordagem socioantropológica, utilizamos a técnica de análise de documentos para compreender como ali se configuram a aceitação e o perfil dos possíveis consumidores de tal tecnologia. Observamos que a construção do homem usuário se realiza com base em uma concepção igualitária dos direitos reprodutivos, enfatizando o direito de os homens - assim como as mulheres - terem autonomia e controle sobre a própria fertilidade. Há uma expansão do perfil do usuário em relação a tentativas anteriores de disponibilização de uma "pílula masculina", mas, ao enfocar a noção dos homens como sujeitos de direitos reprodutivos, tal construção pode gerar conflitos com questões centrais dos movimentos feministas.

Resumen El objetivo del artículo es analizar cómo el usuario de un anticonceptivo masculino es concebido por un actor relevante en el campo, la ONG estadounidense Male Contraception Initiative. A partir de un enfoque socioantropológico, utilizamos la técnica de análisis de documentos para comprender cómo la ONG configura la aceptación y los consumidores de tal tecnología. Observamos que la construcción del hombre usuario se realiza con base en una concepción igualitaria de los derechos reproductivos, enfatizando el derecho de los hombres - así como las mujeres - tener autonomía y control sobre la propia fertilidad. Hay una expansión del perfil del usuario en relación a intentos anteriores de disponibilización de una "píldora masculina", pero, al enfocar la noción de los hombres como sujetos de derechos reproductivos, tal construcción puede generar conflictos con cuestiones centrales de los movimientos feministas.

Abstract This article analyzes how the American NGO Male Contraception Initiative - a relevant actor in the field - conceives the male contraceptive user. The ways this NGO is shaping both acceptance and consumers of that technology are examined from a social anthropological perspective and through document analysis. We observe that the construction of the male user is based on a concept of equality of reproductive rights, emphasizing the right of men as well as women to have autonomy and control over their own fertility. While there is an expansion of the user's profile compared to previous attempts to make a "male pill" available, a closer look on the notion of men as subject of reproductive rights reveals that such a construction may cause conflicts with central issues of feminist movements.

Effect of the indenopyridine RTI-4587-073 (l) on feline testicle.

The aim of this study was to describe the seminal, histomorphological and hormonal effects of the oral indenopyridine RTI-4587-073(l) on feline testicle. Clinical side effects were also recorded. Sixty testicles of 30 adult cats that had been treated (d 0) with RTI-4587-073(l) 12.5 mg/kg PO and randomly hemiorchiectomized twice on: day -14 (n = 8), 6 h (n = 6), 12 h (n = 8), 24 h (n = 6), day 7 (n = 8), day 14 (n = 6), day 21 (n = 6), day 35 (n = 6) or day 42 (n = 6) were studied. Before each hemiorchiectomy, fecal samples for testosterone (T) measurement were collected and the testes were grossly and ultrasound examined. This indenopyridine did not cause changes in testicular weight (P > 0.1), volume (P > 0.1), echostructure, gonadosomatic index (P > 0.1), fecal T concentrations (P > 0.1), nor clinical side effects. A severe disorganization of the cytoarchitecture of the seminiferous epithelium, sloughed cells and fluid, were observed in the 6 h samples up to a maximum at 24 h. Tubular diameter (P < 0.01) increased twice, during the first 24 h and on d 35. Germinal epithelium achieved its minimum height on d 14 to rapidly recover thereafter. This treatment caused a significant decrease in the volume of all the seminiferous cell components, except spermatogonias. All histotological parameters normalized by the end of the study. It was concluded that RTI-4587-073(l) severely disrupted spermatogenesis during the first 24 h after treatment returning to normality in approximately one spermatic cycle without clinical side effects.

Relación entre el conocimiento y la intención de uso del anticonceptivo oral masculino (DMAU) en estudiantes varones en edad fértil de una universidad del sur de Chile / Relationship between the knowledge and the intention of use of the male oral contraception (DMAU) in students male in the fertile age of a university of the south of Chile

A lo largo de la historia las mujeres han sido las responsables de la reproducción biológica, han cargado mayormente con la responsabilidad de la anticoncepción (1); surge la necesidad de incorporar a hombres en salud reproductiva con métodos modernos, efectivos y reversibles, como la Píldora Anticonceptiva Oral Masculina, que se encuentra en fase II de investigación, y así participar activamente en la planificación familiar. OBJETIVO: Establecer la relación entre el conocimiento e intención de uso de la Píldora Anticonceptiva Oral Masculina, en varones en edad fértil de una Universidad del Sur de Chile, durante segundo semestre del 2018. MATERIAL Y MÉTODO: Investigación Cuantitativa, correlacional, no experimental y transversal, en 301 estudiantes regulares, se autoadministra instrumento: "Encuesta Anticonceptivos Orales en Varones en Edad Fértil", validado por medio de comité de expertos, con Coeficiente de Alpha de Cronbach de 0,81. RESULTADOS: Respecto al nivel de conocimiento acerca de la Píldora, sólo un 16% tuvo un puntaje mayor o igual a 17 puntos considerándose como alto conocimiento. Respecto de intención de uso, un 58% de los estudiantes varones tiene una alta intención de uso, además de que el 42% estaría dispuesto a utilizar este método anticonceptivo de forma diaria. CONCLUSIÓN: No existe una relación entre el nivel de conocimiento y la intención de uso de la píldora, pues independiente del nivel de conocimiento siempre se mantuvo una alta intención de uso investigación y así participar activamente en la planificación familiar

Throughout history, women have been responsible for biological reproduction, they have been mostly responsible for contraception (1); There is a need to incorporate men into reproductive health with modern, effective and reversible methods, such as the Male Oral Contraceptive Pill, which is in phase II of research, and thus actively participate in family planning. OBJECTIVE: To establish the relationship between the knowledge and intention to use the Male Oral Contraceptive Pill, in men of childbearing age of a University of Southern Chile, during the second half of 2018. MATERIAL AND METHOD: Quantitative, correlational, non-experimental research and Transversal, in 301 regular students, self-administered instrument: "Survey of Oral Contraceptives in Men of Fertile Age", validated through an expert committee, with Cronbach's Alpha Coefficient of 0.81. RESULTS: Regarding the level of knowledge about the Pill, only 16% had a score greater than or equal to 17 points, considered ashigh knowledge. Regarding intention to use, 58% of male students have a high intention to use, in addition to 42% would be willing to use this method of contraception on a daily basis. CONCLUSION: There is no relationship between the level of knowledge and the intention to use the pill, because independent of the level of knowledge, a high intention to use research was always maintained and thus actively participate in family planning.

Reestablishment of sperm quality after long-term deslorelin suppression in tomcats.

The aim of the study was to determine the time after treatment with a 4.7 mg deslorelin implant until Tomcat spermatogenesis activity was restored, and seminal parameters reached pre-implant values. Tomcats (n = 6) were randomly assigned to one of two treatments. Three cats (n = 3) received a deslorelin implant (4.7 mG; Suprelorin®, Virbac, France) in the interscapular subcutaneous region whereas three (n = 3) received no implant and served as control group. Semen samples were collected by electroejaculation every 4 wk from 3 mo before treatment (pretreatment samples) until reestablishment of pre-treatment sperm quality, 32 mo post-implant insertion (PI). Each semen sample was assessed for motility, velocity, concentration, total sperm count, viability, acrosome integrity, plasma membrane integrity and sperm morphology. After semen collection, testicular volume and presence/absence of penile spines were recorded. Additionally, blood samples were taken to measure testosterone concentration. An increase in sperm concentration and total sperm count was present 1 mo PI despite of an abrupt decrease in serum testosterone concentrations after 2-4 weeks. This initial stimulatory effect was followed by a decrease in seminal parameters, reduction of testicular volume and disappearance of penile spines 2 mo PI. A single Suprelorin® 4.7 mg implant suppressed sperm production for 22-25 months. No clinically side effect was observed during the study period. All toms returned to their initial seminal quality 23-28 months after treatment. Therefore, we conclude that Suprelorin® 4.7 mg is a safe option for reversible reproduction control during long periods in tomcats.

Influencia de los factores socioculturales del varón para la aceptación del uso de métodos anticonceptivos en la pareja puérpera. Instituto Nacional Materno Perinatal - enero 2016 / Influence of the socio-cultural factors of the male for the acceptance of the use of contraceptive methods in the puerperal couple. National Institute Maternal Perinatal - January 2016

Objetivo: Determinar la influencia de los factores socioculturales del varón asociados a la aceptación del uso de métodos anticonceptivos en la pareja puérpera del Instituto Nacional Materno Perinatal en enero del 2016. Material y Métodos: Estudio de tipo observacional, analítico de casos y controles, que contó con una muestra de 118 varones que visitaban a sus parejas hospitalizadas en el servicio de puerperio, divididos en dos grupos: 59 varones que aceptan el uso de métodos anticonceptivos por sus parejas (grupo caso) y 59 varones que no acepten el uso de métodos anticonceptivos por sus parejas (grupo control), con un muestreo no probabilístico pareado por fecha de parto. Para el análisis descriptivo de las variables cualitativas se estimaron frecuencias absolutas y relativas y para las variables cuantitativas, medidas de tendencia central y promedios. Además se utilizó la prueba Chi-cuadrado (p<0.05, significativo) y el Odds Ratio (OR). Resultados: No se encontraron factores sociales asociados a la aceptación de los varones para el uso de métodos anticonceptivos por sus parejas. Los factores culturales asociados a la aceptación de los varones para el uso de métodos anticonceptivos por sus parejas fueron el grado de instrucción superior (p=0.009), el nivel de conocimiento alto (p=0.000) y medio (p=0.000) y las creencias correctas sobre métodos anticonceptivos (p=0.000 superior (OR=0.37) y con creencias correctas (OR=0.06) tienen mayor probabilidad de aceptar el uso de métodos anticonceptivos, mientras que los varones con conocimiento). Asimismo, se observó que los varones con nivel de instrucción alto (OR=0.13) tienen mayor probabilidad de no aceptar el uso de métodos anticonceptivos por sus parejas. Conclusión: La influencia de los factores socioculturales del varón asociados a la aceptación del uso de métodos anticonceptivos en la pareja de las puérperas del Instituto Nacional Materno Perinatal son las creencias y el nivel de conocimientos sobre métodos...

Objective: To determine the factors associated sociocultural acceptance of postpartum use of modern contraceptive methods by men who visit your partner in the service of the Maternal puerperal National Perinatal Institute in January 2016. Material and methods: Study observational, with prospective design, analytical case-control studies, which included a sample of 118 men who visited their partners hospitalized in service puerperium divided into two groups: 59 men who accept the use of modern contraceptive methods by their partners (case group) and 59 men who do not accept the use of modern contraceptive methods by their partners (control group), with a non-probability sampling matched by date of birth. For the descriptive analysis of qualitative variables and absolute and relative frequencies for quantitative variables, measures of central tendency and averages were estimated. Besides the Chi-square (p<0.05, significant) and Odds Ratio (OR) test was used. Results: No social factors associated with the acceptance of men for the use of modern contraceptive methods by their partners were found. Cultural factors associated with accepting men for the use of modern contraceptive methods by their partners were the degree of higher education (p = 0.009), the highest level of knowledge (p = 0.000) and medium (p = 0.000) and the correct beliefs about modern contraceptive methods (p = 0.000). It was also noted that men with higher level of education (OR = 0.37) and with correct beliefs (OR = 0.06) were more likely to accept the use of contraceptive methods, while men with high knowledge (OR = 0.13) have more likely not accept the use of contraceptive methods by their partners. Conclusions: The main sociocultural factors associated with acceptance of men to use contraception in the postpartum women in the National Maternal Perinatal Institute are the beliefs and level of knowledge of modern contraceptive methods.

Mechanisms involved in reproductive damage caused by gossypol in rats and protective effects of vitamin E.

BACKGROUND: Gossypol is a chemical present in the seeds of cotton plants (Gossypium sp.) that reduces fertility in farm animals. Vitamin E is an antioxidant and may help to protect cells and tissues against the deleterious effects of free radicals. The aim of this study was to evaluate the mechanisms of reproductive toxicity of gossypol in rats and the protective effects of vitamin E. Forty Wistar rats were used, divided into four experimental groups (n = 10): DMSO/saline + corn oil; DMSO/saline + vitamin E; gossypol + corn oil; and gossypol + vitamin E. RESULTS: Fertility was significantly reduced in male rats treated with gossypol in that a significant decrease in epididymal sperm count was observed (P < 0.05) and the number of offspring was significantly reduced in females mated with them (P < 0.05). This dysfunction was prevented by vitamin E. Gossypol caused a significant increase in the activity of the enzymes glutathione peroxidase (P < 0.01) and glutathione reductase (P < 0.01), but vitamin E did not reduce the enzyme activities (P > 0.05). The levels of reduced glutathione and pyridine nucleotides in testis homogenate were significantly reduced by gossypol (P < 0.05 and P < 0.01, respectively) and this reduction was accompanied by increased levels of oxidized glutathione (P < 0.05). Vitamin E showed a preventive effect on the changes in the levels of these substances. Gossypol significantly increased the levels of malondialdehyde (P < 0.01), a lipid peroxidation indicator, whereas treatment with vitamin E inhibited the action of the gossypol. Vitamin E prevented a decrease in mitochondrial ATP induced by gossypol (P < 0.05). CONCLUSIONS: This study suggests that the reproductive dysfunction caused by gossypol may be related to oxidative stress and mitochondrial bioenergetic damage and that treatment with vitamin E can prevent the infertility caused by the toxin.

Mechanisms involved in reproductive damage caused by gossypol in rats and protective effects of vitamin E

BACKGROUND: Gossypol is a chemical present in the seeds of cotton plants (Gossypium sp.) that reduces fertility in farm animals. Vitamin E is an antioxidant and may help to protect cells and tissues against the deleterious effects of free radicals. The aim of this study was to evaluate the mechanisms of reproductive toxicity of gossypol in rats and the protective effects of vitamin E. Forty Wistar rats were used, divided into four experimental groups (n = 10): DMSO/ saline + corn oil; DMSO/saline + vitamin E; gossypol + corn oil; and gossypol + vitamin E. RESULTS: Fertility was significantly reduced in male rats treated with gossypol in that a significant decrease in epididy-mal sperm count was observed (P < 0.05) and the number of offspring was significantly reduced in females mated with them (P < 0.05). This dysfunction was prevented by vitamin E. Gossypol caused a significant increase in the activity of the enzymes glutathione peroxidase (P < 0.01) and glutathione reductase (P < 0.01), but vitamin E did not reduce the enzyme activities (P > 0.05). The levels of reduced glutathione and pyridine nucleotides in testis homogen-ate were significantly reduced by gossypol (P < 0.05 and P < 0.01, respectively) and this reduction was accompanied by increased levels of oxidized glutathione (P < 0.05). Vitamin E showed a preventive effect on the changes in the levels of these substances. Gossypol significantly increased the levels of malondialdehyde (P < 0.01), a lipid peroxida-tion indicator, whereas treatment with vitamin E inhibited the action of the gossypol. Vitamin E prevented a decrease in mitochondrial ATP induced by gossypol (P < 0.05). CONCLUSIONS: This study suggests that the reproductive dysfunction caused by gossypol may be related to oxidative stress and mitochondrial bioenergetic damage and that treatment with vitamin E can prevent the infertility caused by the toxin.

Gossypol promotes degeneration of ovarian follicles in rats.

The present study aimed to determine if gossypol interferes with ovarian follicles in rats. Twenty-four female Wistar rats were assigned to two equal groups: one control group and the other dosed with gossypol (25 mg/kg/day, subcutaneously) for 15 days. Ovarian follicles were histologically classified according to the stage of development and as normal or atretic. Gossypol treatment reduced the length of estrous with an increase in the duration of the diestrus phase. This compound was responsible for reduced serum levels of T4 and progesterone. Treatment with gossypol was responsible for a significant reduction in the number of normal ovarian follicles and a significant increase in the number of atretic follicles, both in all stages of development. Thus, treatment of rats with gossypol was responsible for reduction in the number of viable follicles and changes in hormone levels that resulted in interference of the estrous cycle.

Injection of a chemical castration agent, zinc gluconate, into the testes of cats results in the impairment of spermatogenesis: a potentially irreversible contraceptive approach for this species?

Male sterilization by chemical agents is a nonsurgical contraceptive approach designed to induce azoospermia and, therefore, infertility. Intratesticular injection of zinc gluconate for sterilization of dogs has been described, but its use in cats remains limited. The objective of the present study was to evaluate, by light and transmission electron microscopy, the efficacy of a single intratesticular injection of a zinc gluconate solution (Testoblock) as a sterilant for male cats. Twelve sexually mature mixed breed cats were allocated at random into two groups (control = 6; treated = 6) and given a single injection into each testis of either isotonic saline or zinc gluconate, respectively. Histopathologic and ultrastructural evaluation was assessed at 120 days postinjection. Histopathologic changes were not detected in the testes from the control group. However, histologic evaluation of the treated group revealed atrophic and dilated seminiferous tubules, a decrease in the number of germ cells, and incomplete spermatogenesis. Sertoli cells had various degrees of cytoplasmic vacuolization. Intertubular tissue revealed active fibroblasts, collagen deposition, and inflammatory cells. The diameter of seminiferous tubules, epithelial height and tubular area were reduced (P < 0.05) in the treated group compared with controls. Azoospermia occurred in 8 of the 11 treated cats (73%). Ultrastructural evaluation of Leydig cells revealed loss of nuclear chromatin, increased smooth endoplasmatic reticulum, and mitochondria degeneration. Intratesticular injection of zinc gluconate solution impaired spermatogenesis in cats and has great potential as a permanent sterilant in this species.

Intratesticular injection of a zinc-based solution for contraception of domestic cats: a randomized clinical trial of efficacy and safety.

It has been reported that a commercial zinc gluconate preparation disrupts spermatogenesis and apparently causes permanent sterilization in male dogs, but there is little information regarding similar approaches in the male cat. The objective of this study was to evaluate zinc gluconate as a permanent contraceptive for domestic male cats. Sixteen sexually mature mixed breed cats were allocated at random, by replicate, into two groups and given a single injection into each testis of either isotonic saline or zinc gluconate, respectively. Clinical and reproductive parameters were assessed immediately before injection and after 60 and 120 days. On day 120 the testis size of treated cats was decreased (P<0.05). Azoospermia occurred in 8/11 (73%) cats, and penile spines were decreased in 6/11 (55%) and absent in 4/11 (36%) cats, and there were substantial reductions in male behavior. However, plasma testosterone concentrations (single samples collected at each assessment) were not significantly different between treated and control cats at any time point. Although additional studies are warranted, intratesticular injection of zinc gluconate might have potential as a permanent contraceptive for cats.

Oral administration of an anti-inflammatory does not compromise the efficacy of intra-testicular injection of zinc gluconate as a contraceptive for dogs.

The objective was to determine whether the efficacy of zinc gluconate (Testoblock(®)) as a chemical contraceptive in male dogs was compromised in the presence of metamizole sodium (a nonsteroidal antiinflammatory/analgesic agent). Ten sexually mature mongrel dogs were assigned to two groups, a control group (n = 5) and a treated group (n = 5). Testoblock(®), a proprietary zinc-gluconate (13.1 mg zinc/ml) solution in a physiological vehicle, was injected into each testis (0.2-1.0 ml/testis, based on testis width). Half of the dogs (treated group) were also given metamizole sodium (also known as sodium dipyrone) orally (25mg/kg three times a day for 2 days), starting 2-3 h after testis injection. A physical examination and assessment of testis width, hematology, clinical chemistry (hepatic and renal function) and semen characteristics, were done immediately after treatment and then every 2 months for 180 days. There was no post-treatment scrotal biting or licking, although there was transient testicular swelling in both control and treated dogs during the first 3 days after injection. At 60 days after injection, all dogs were azoospermic. At 120 and 180 days, seven dogs had azoospermia and the remaining three (two control and one treated) had apparent aspermia (no ejaculate could be collected). There were no significant differences between groups for clinical findings or any aspect of hematology, renal, or hepatic function. In conclusion, giving metamizole sodium concurrent with an intra-testicular injection of a zinc-based solution did not interfere with chemical sterilization and it improved animal welfare.

A single administration of the GnRH antagonist acyline inhibits basal and GnRH-stimulated serum testosterone concentrations in male dogs.

The objective of this study was to describe testosterone (T) response to GnRH challenge in antagonist-treated dogs over a 30-day period. Eight mongrel dogs were randomly assigned to either the GnRH antagonist acyline 330 µg/kg sc (ACY; n = 4) or a placebo group (PLA; n = 4). The dogs were serially challenged with the GnRH agonist, buserelin 0.2 µg/kg sc on days -1, 1, 3, 7, 10, 14, 21 and 30. On these days, blood samples for T determinations were collected before (-30 min) and 60, 120 and 180 min after the agonist injection. Basal (-30 min) and post-GnRH agonist stimulation T values were compared by anova for repeated measures. Before treatments (day -1), there were no differences in basal T serum concentrations between groups (p > 0.1). After treatments, basal T showed a significant interaction between treatment and day (p < 0.05). Furthermore, when both groups were analysed independently, basal T varied in the ACY (p < 0.01) but not in the PLA group (p > 0.1). On day -1, before treatments, the stimulation tests had only a time effect (p = 0.05) although on days 1 (p < 0.01), 3 (p < 0.01), 7 (p < 0.01), 10 (p < 0.01) and 14 (p < 0.05), the response to the agonist differed between groups, becoming similar on days 21 (p > 0.05) and 30 (p > 0.05). It was concluded that, in dogs, a single administration of the GnRH antagonist prevented canine gonadal axis to physiologically respond to agonistic challenge during 14 days.

Permanent contraception of dogs induced with intratesticular injection of a Zinc Gluconate-based solution.

The objective was to evaluate the efficacy of a single intratesticular injection of a Zinc Gluconate-based solution to induce sterility in male dogs. Fifteen pubertal mongrel dogs (8 mo to 4 y old) were assigned to two groups; Control dogs (n = 5) received a single injection of an isotonic saline solution into each testis, whereas Treated dogs (n = 10), were given Testoblock, a proprietary zinc-gluconate (13.1 mg zinc/ml) solution in a physiological vehicle. The volume of saline or Testoblock injected varied from 0.2 to 1.0 ml/testis (based on testis width). Physical examination, testis width, hematology, clinical chemistry (hepatic and renal function), plasma testosterone concentration, semen characteristics, and libido, were assessed until castration (150 d after treatment). In Treated dogs, testis width increased (approximately 20%) relative to that in Control dogs, but subsequently was not significantly different from Controls (group × time interaction, P < 0.0001). For all dogs, values for hematology and clinical chemistry consistently remained within reference ranges. Although plasma testosterone concentrations decreased over time (P < 0.006), there was only a tendency for an effect of group (P < 0.09), and libido was not significantly affected. By 63 d after Testoblock treatment, eight Treated dogs were azoospermic, whereas the remaining two were oligozoospermic (<10 × 10(6) sperm/ml). We concluded that intratesticular injection of the Zinc Gluconate-based chemical sterilant Testoblock has considerable potential to induce permanent contraception in male dogs.