Rats Orally Administered with Ethyl Alcohol for a Prolonged Time Show Histopathology of the Epididymis and Seminal Vesicle Together with Changes in the Luminal Metabolite Composition.

Prolonged ethanol (EtOH) consumption is associated with male infertility, with a decreased spermatogenesis rate as one cause. The defective maturation and development of sperm during their storage in the cauda epididymis and transit in the seminal vesicle can be another cause, possibly occurring before the drastic spermatogenesis disruption. Herein, we demonstrated that the cauda epididymis and seminal vesicle of rats, orally administered with EtOH under a regimen in which spermatogenesis was still ongoing, showed histological damage, including lesions, a decreased height of the epithelial cells and increased collagen fibers in the muscle layer, which implicated fibrosis. Lipid peroxidation (shown by malondialdehyde (MDA) levels) was observed, indicating that reactive oxygen species (ROS) were produced along with acetaldehyde during EtOH metabolism by CYP2E1. MDA, acetaldehyde and other lipid peroxidation products could further damage cellular components of the cauda epididymis and seminal vesicle, and this was supported by increased apoptosis (shown by a TUNEL assay and caspase 9/caspase 3 expression) in these two tissues of EtOH-treated rats. Consequently, the functionality of the cauda epididymis and seminal vesicle in EtOH-treated rats was impaired, as demonstrated by a decreases in 1H NMR-analyzed metabolites (e.g., carnitine, fructose), which were important for sperm development, metabolism and survival in their lumen.

Momordica charantia fruit extract with antioxidant capacity improves the expression of tyrosine-phosphorylated proteins in epididymal fluid of chronic stress rats.

OBJECTIVE: Although the protective effects of Momordica charantia L. (MC) extract on chemical-induced testicular damage have been studied, the preventive effects of MC extract on functional proteins in the epididymis under chronic stress have never been reported. This study investigated the protective effects of MC fruit extract on protein secretion, especially tyrosine-phosphorylated proteins, in the epididymis of rats exposed to chronic unpredictable stress (CUS). METHODS: Total phenolic compounds (TPC), total flavonoid compounds (TFC) and antioxidant capacities of MC extract were measured. Adult male rats were divided into 4 groups: control group, CUS group, and 2 groups of CUS that received different doses of MC extract (40 or 80 mg/kg). In treated groups, rats were given MC daily, followed by induction of CUS (1 stressor was randomly applied from a battery of 9 potential stressors) for 60 consecutive days. Plasma corticosterone and testosterone levels were analyzed after the end of experiment. Expressions of heat-shock protein 70 (HSP-70) and tyrosine-phosphorylated proteins present in the fluid of the head and tail of the epididymis were quantified using Western blot. RESULTS: MC extract contained TPC of (19.005 ± 0.270) mg gallic acid equivalents and TFC of (0.306 ± 0.012) mg catechin equivalents per gram, and had 2,2-diphenyl-1-picrylhydrazyl antioxidant capacity of (4.985 ± 0.086) mg trolox equivalents per gram, radical 50% inhibitory concentration of (2.011 ± 0.008) mg/mL and ferric reducing antioxidant power of (23.697 ± 0.819) µmol Fe(II) per gram. Testosterone level in the epididymis was significantly increased, while the corticosterone level was significantly improved in groups treated with MC extract, compared to the CUS animals. Particularly, an 80 mg/kg dose of MC extract prevented the impairments of HSP-70 and tyrosine-phosphorylated protein expressions in the luminal fluid of the epididymis of CUS rats. CONCLUSION: MC fruit extract had antioxidant activities and improved the functional proteins secreted from the head and tail of the epididymis. It is possible to develop the MC fruit extract as a male fertility supplement for enhancing functional sperm maturation in stressed men.

Protective effects of Thai Mucuna pruriens (L.) DC. var. pruriens seeds on sexual behaviors and essential reproductive markers in chronic unpredictable mild stress mice.

Background and aims: Chronic stress is a major common cause of male infertility. Many species of velvet beans are shown to be rich in l-DOPA. In Thai folklore medicine, seeds of Mucuna pruriens (L.) DC. var. pruriens (Thai Mhamui or T-MP) have been used for treating erectile dysfunction. This study aimed to determine l-DOPA levels in T-MP seed extract and investigate its preventive on sexual behaviors and reproductive parameter damages including essential proteins in chronic unpredictable mild stress (CUMS) mice. Experimental procedure: Mice were divided into 4 groups: (I) control, (II) CUMS, (III) T-MP300 + CUMS, and (IV) T-MP600 + CUMS. Groups I and II received DW while groups III and IV were pretreated with the seed extracts (300 and 600 mg/kg BW) for 14 consecutive days before co-treatment with a randomly different CUMS/day (from 12 mild stressors) for 43 days. Results and conclusion: T-MP seed extract contained l-DOPA approximately 10% of total dried weight. A dose of 600 mg/kg improved sexual performances and degenerative seminiferous epithelium in CUMS mice. Sperm qualities and testosterone level were elevated while corticosterone was decreased in co-treatment groups. T-MP-CUMS cotreated groups also improved expressions of AKAP4, AR, and TyrPho proteins in testis, epididymis, and sperm. T-MP increased StAR and CYP11A1 expressions in testis. It also suppressed testicular apoptosis via decreased expressions of Hsp70, caspases 3, and 9. T-MP seeds containing l-DOPA could improve sexual behaviors and essential reproductive proteins caused by CUMS. Section: Natural Products. Taxonomy classification by evise: Traditional Herbal Medicine; Animal Model; Histopathology.

Seed extract of Thai Mucuna pruriens (L.) DC. var. pruriens enhances sexual performance and improves male reproductive damages in ethanol-induced rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Thai Mucuna pruriens (L.) DC. var. pruriens (T-MP) has been traditionally used in treating depressive disorders, dysuria and enhancing male sexual desire. Although T-MP seed is demonstrated to have antioxidant capacity, its aphrodisiac and protective tissue damage properties have never been documented. Recently, ethanol (Eth) is known to cause sexual behavior dysfunction and damage reproductive system. This study aimed to investigate the protective effects of T-MP seed extract on sexual behavior dysfunction and reproductive damages in male rats admisted with Eth. MATERIALS AND METHODS: T-MP possessing antioxidant activity was determined for L-DOPA content using NMR analysis. Thirty-six male rats were divided into four groups (9 animals/group). Control rats received DW and the ethanol (Eth) group was given with Eth (3 g/kgBW; 40%v/v). In preventive groups (T-MP150 + Eth and MP300 + Eth groups), animals were treated with T-MP extract at a dose of 150 and 300 mg/kgBW before Eth administration for consecutive 56 days. Sexual behaviors including mounting frequency (MF), intromission frequency (IF), mounting latency (ML), intromission latency (IL), ejaculation latency (EL), post-ejaculatory interval (PEI), and ejaculation frequency (EF) were evaluated. Epididymal sperm quality and daily sperm production (DSP) were examined. Testicular histology was observed using Masson's trichrome staining. The malondialdehyde (MDA) levels and expressions of androgen receptor (AR), heat shock protein 70 (HSP70), steroidogenic acute regulatory (StAR), and tyrosine-phosphorylated (TyrPho) proteins in testis were also determined. RESULTS: T-MP extract contained L-DOPA and improved sexual behaviors including increased MF and IF and decreased ML and IL in Eth treated rats. Significantly, sperm quality, DSP, and testicular histopathology observed in Eth group were improved after T-MP treatment. T-MP also decreased the testicular MDA levels. Additionally, T-MP could correct testicular functional proteins of AR and StAR except HSP70 expression in Eth group. Expressions of TyrPho proteins in testicular and sperm lysates were improved in co-administered groups. CONCLUSIONS: T-MP seed extract possessing L-DOPA could enhance the sexual behaviors and protect reproductive damages via improvement of testicular functional proteins.

Seed extract of Thai Mucuna pruriens reduced male reproductive damage in rats induced by chronic stress.

CONTEXT: Thai Mucuna pruriens (L.) DC. var. pruriens (Fabaceae) (TMP) is known to enrich reproduction but preventive effects on stress related adverse reproductive parameters are not documented. OBJECTIVE: This study investigates the protective property of TMP seed extract on reproductive damage under chronic stress (CS). MATERIALS AND METHODS: Male Sprague-Dawley rats were divided into four groups. The control and CS groups received distilled water, whereas the pre-treated rats received the aqueous TMP seed extract at doses of 150 and 300 mg/kg BW for 20 days before co-treatments with CS induction (immobilization and forced swimming) for 81 days. Serum was used to determine the cortisol and testosterone levels. Histology of testis and epididymis was observed with localization of androgen receptor (AR). Sperm parameters and the expression of steroidogenic acute regulatory (StAR), cytochrome P450 family 11 subfamily a member 1 (CYP11A1), AR, HSP70, caspases (3 and 9) and tyrosine phosphorylation (TyrPho) proteins were investigated. RESULTS: TMP extract improved cortisol level (0.84 ± 0.02 µg/dL) and protected against the damage of reproductive tissues and sperm parameters (count [49.78 ± 3.74 million sperm/mL], viability [90.01 ± 1.17%] and precocious acrosome reaction [1.38 ± 0.48%]). Expression of testicular StAR, CYP11A1, AR and HSP70 proteins was improved. Caspase expression was decreased in treated rats. TMP increased AR expression in CS sperm. Moreover, TyrPho protein expression was corrected after TMP administration. CONCLUSIONS: TMP seed protected against adverse reproductive parameters in CS via improvements of functionally testicular markers and reductions of apoptotic proteins. It is possible to develop the TMP beans as an alternative medicine in treating of male subfertility caused by CS.

The effect of Dolichandrone serrulata (wall. ex DC.) Seem. flower extract containing antioxidant capacity and terpenoids on the male reproductive system.

Although the fruit extract of Dolichandrone genus was shown to inhibit spermatogenesis, the reproductive toxicity of Dolichandrone serrulata flowers (DSFs) is not documented. Recent study aimed to evaluate the sub-chronic toxicity of DSF on male reproductive system. Antioxidant capacity and total phenolic contents of DSF extract were determined using Folin-Ciocalteu's, 2,2-diphenyl-1-picrylhydrazyl and ferric reducing antioxidant power assays. The terpenoid components were determined using nuclear magnetic resonance spectrum. Adult male rats were treated orally with DSF (100, 300 or 600 mg/kg) for 48 days. Histopathology of testis and epididymis was observed. Sperm concentration, viability, acrosome status and morphology were also examined. Expressions of heat shock protein 70 (Hsp70), tyrosine-phosphorylated (TyrPho) proteins, androgen receptor (AR) and steroidogenic acute regulatory (StAR) protein in testis were investigated. Results showed that DSF contained phenolic compounds and terpenoids (phytoandrogens; rengyolone and cleroindicin B). No reproductive histopathology was observed in DSF-treated rats. Although DSF decreased the serum testosterone level, the sperm qualities were not affected. Particularly, sperm concentration of DSF-treated animals was significantly increased. DSF changed the testicular TyrPho proteins but the expression of AR, StAR or Hsp70 was not altered. In conclusion, DSF possesses antioxidant capacity with no toxicity on male reproductive system.

Localization (and profiles) of tyrosinephosphorylated proteins in female reproductive organs of adult rats.

OBJECTIVE: Tyrosine phosphorylation is an essential process in many biological systems, including the male reproductive system. The presence of tyrosine-phosphorylated (TyrPho) proteins has been well documented in male reproductive organs, but research in fertile females is still limited. METHODS: The ovary, oviduct, and uterus of adult female Sprague-Dawley rats in the estrus phase were used to localize TyrPho proteins using an immunohistochemical technique. These proteins were separated and their expression patterns were examined by sodium dodecyl sulfatepolyacrylamide gel electrophoresis and Western blot analysis, respectively. RESULTS: TyrPho proteins were localized in the cytoplasm of the oocyte except the antral fluid; in the granulosa cells, theca cells, and stromal cells of the ovary; at the apical surface of oviductal epithelial cells; and in the basal epithelium and submucosa of the uterine wall. Moreover, we found that 72-, 43-, and 28-kDa TyrPho proteins were localized in the ovary, while 170-, 55-, and 43-kDa proteins were localized in the oviduct. In the uterus, we detected four major bands, corresponding to 61-, 55-, 54-, and 43-kDa TyrPho proteins. CONCLUSION: Given that these TyrPho proteins were found in major reproductive organs in the estrus phase, these proteins may play important roles in female fertility.

Effect of ethanol on the changes in testicular protein expression in adult male rats.

Ethanol consumption is a major cause of male infertility, but the exact mechanism is still largely unknown. This study attempted to investigate the effect of ethanol on sperm morphology, acrosome reaction status and the alteration of the testicular protein expressions. Fourteen male rats were divided into control and ethanol groups (n = 7/each group). Ethanol-treated rats received ethanol (5 g/kg, 40% v/v) via oral gavage for consecutive 14 days. Testosterone hormone, sperm parameters, and testicular and epididymal histopathologies were evaluated. In addition, the expressions of testicular proteins including androgen receptor (AR), heat shock protein 70 (HSP70), steroidogenic acute regulatory protein (StAR) and tyrosine-phosphorylated (TyrPho) proteins were investigated. The results showed that ethanol significantly increased percentage of abnormal sperm morphology and acrosome-reacted spermatozoa. Some seminiferous and ductus epididymal histopathologies were observed in ethanol-treated rats. Significantly, ethanol reduced serum testosterone and expressions of testicular AR and TyrPho proteins. However, the overexpression of StAR and HSP70 proteins in ethanol testis was found. It was concluded that the changes in testicular protein expressions may be involved in mechanism of male infertility caused from ethanol consumption.

Evaluation of antioxidant capacity and reproductive toxicity of aqueous extract of Thai Mucuna pruriens seeds.

OBJECTIVE: In traditional medicine, the seeds of Thai Mucuna pruriens (T-MP) are used to treat male dysuria and are believed to enhance fertility. However, information pertaining to the toxicity of T-MP and its interaction with other properties is limited. This study was thus conducted to evaluate the antioxidant capacity and subacute toxicity of T-MP in the reproductive system. METHODS: Total phenolic content and antioxidant capacity of T-MP seed extract were determined using total phenolic content, 2,2-diphenyl-1-picrylhydrazyl and ferric reducing antioxidant power assays. Male and female adult rats were treated orally with T-MP at a dosage of 150 or 300 mg/kg body weight for 14 consecutive days. Sex hormones and functional parameters in the liver and kidney were evaluated. Histopathology of all tissue was conducted using Masson's trichrome staining. Sperm parameters, including concentration, morphology, acrosome reaction status and DNA damage, were also examined. Expression of tyrosine phosphorylated protein (TyrPho), androgen receptor and A-kinase-anchoring protein 4 (AKAP4) were investigated using the Western blot technique. RESULTS: T-MP seed extract contained phenolic compounds and exhibited high antioxidant capacity with no toxicity at the tested doses. It did not affect liver or kidney function parameters in the male rats, but increased estradiol, aspartate aminotransferase and alanine aminotransferase levels in the females. Additionally, it decreased serum progesterone and alkaline phosphatase levels in female rats. Serum and intratesticular testosterone levels were significantly lower in male rats that received a high dosage of T-MP. Histopathological changes were not observed in any tissue treated with T-MP. T-MP also significantly increased sperm concentration (but did not affect sperm parameters), and enhanced testicular TyrPho protein and androgen receptor and expression of AKAP4 in sperms. CONCLUSION: T-MP seed extract exhibited antioxidant capacity and was not harmful to reproductive tissues. It also had a phytoestrogenic effect on females and increased the expression of testicular and sperm markers of male fertility.

Changes of sperm quality and hormone receptors in the rat testis after exposure to methamphetamine.

Methamphetamine (METH) is known to damage neurons and induce psychosis. It can also induce apoptosis in seminiferous tubules and affect sperm quality. The present study was carried out to investigate the effect of a rat model of METH addiction on sperm quality and expression of progesterone receptors (PR) and estrogen receptors (ER) in the testis. Sperm quality parameters including sperm motility, sperm morphology and sperm concentration were examined. Protein and gene expressions PR, ERα and ERß were studied using immunohistochemistry and reverse transcriptase-polymerase chain reaction, respectively. The percentages of normal sperm motility and normal sperm morphology were significantly decreased in animals receiving METH, especially in escalating dose (ED METH) and escalating dose-binge (ED-binge METH) groups when compared with control. In addition, sperm concentrations in ED METH and ED-binge METH groups were numerically decreased. PR, ERα and ERß immunoreactive cells were significantly decreased in spermatogonia, spermatogenic cells and especially in Sertoli cells in all METH-treated groups. Furthermore, messenger RNA expression of PR, ERα and ERß were also significantly decreased in all METH-treated animals. These results indicate that METH can induce abnormal sperm quality. These changes of sperm quality may relate to the reduction of PR, ERα and ERß expressions in male germ cells and Sertoli cells which are essential for spermatogenesis and development of sperm.