Epididymal Sperm Count Research Articles

Retinoic Acid Regulates Spermiogenesis Via Hoxb1 and Shh Signaling in Testicular Germ Cells.

Retinoic acid (RA) regulates all four major events in spermatogenesis; spermatogonial differentiation, meiotic entry, spermiogenesis, and spermiation. For the pre-meiotic phase, Sertoli cells are the source of RA and for the post-meiotic phase, pachytene spermatocytes are the source of RA. While the entire spermatogenic process is regulated by RA, how each of these phases is regulated by RA remains completely unknown. Homeobox B1 (Hoxb1) has two retinoic acid response elements (RARE) upstream and downstream of the gene. In this study, we investigated if RA facilitates spermatogenesis by its action on Hoxb1. The expressions of the Hoxb1 and Sonic hedgehog (Shh) genes were analyzed in the post-natal mouse testes and the testicular localizations of Hoxb1, Shh and Gli1 were analyzed by immunohistochemistry in the adult rat testis. To delineate the signaling mechanisms, Hoxb1 expression was altered in vitro and in vivo using retinoic acid and miR-361-3p. Finally, the levels of miR-361-3p and HOXB1 were analyzed in infertile human sperm samples. Hoxb1 and Shh gene expressions were found to be low in the testis of post-natal Swiss mice of 7, 14, 28, 35, and 60days, after which the expressions of both spiked. Immunohistochemistry in the adult mouse testis showed the expressions of Hoxb1, Shh, and Gli1 in the elongating spermatids. Exposure of GC2 cells to RA and in vivoIP RA injection upregulated Hoxb1 andShh signaling in the testiswith increased expressions of Shh, Gli1, and Hdac1. Retinoic acid administration in Swiss mice compromised sperm production and reduced epididymal sperm count. The analysis of infertile human semen samples revealed an increased level of HOXB1 and a decreased level of miR-361-3p as compared to fertile controls. We conclude that retinoic acid regulates late stage of spermatogenesis (spermiogenesis) by affecting Hoxb1 and Shh signaling.

CircCamsap1 is dispensable for male fertility in mice.

Circular RNAs (circRNAs) are a large class of RNAs present in mammals. Among these, circCamsap1 is a well-acknowledged circRNA with significant implications, particularly in the development and progression of diverse tumors. However, the potential consequences of circCamsap1 depletion in vivo on male reproduction are yet to be thoroughly investigated. The presence of circCamsap1 in the mouse testes was confirmed, and gene expression analysis was performed using reverse transcription quantitative polymerase chain reaction. CircCamsap1 knockout mice were generated utilizing the CRISPR/Cas9 system. Phenotypic analysis of both the testes and epididymis was conducted using histological and immunofluorescence staining. Additionally, fertility and sperm motility were assessed. Here, we successfully established a circCamsap1 knockout mouse model without affecting the expression of parental gene. Surprisingly, male mice lacking circCamsap1 (circCamsap1-/-) exhibited normal fertility, with no discernible differences in testicular and epididymal histology, spermatogenesis, sperm counts or sperm motility compared to circCamsap1+/+ mice. These findings suggest that circCamsap1 may not play an essential role in physiological spermatogenesis. Nonetheless, this result also underscores the complexity of circRNA function in male reproductive biology. Therefore, further research is necessary to elucidate the precise roles of other circRNAs in regulating male fertility.

Effects of intratesticular injection of hypertonic mannitol and saline on the quality of donkey sperm, indicators of oxidative stress and testicular tissue pathology

ObjectivesThe aim of the present study was to examine donkey sperm quality after intratesticular injection of hypertonic mannitol (HM) and saline (HS).MethodsRandomly assigned to five treatment groups were 15 adult male donkeys: (1) Control group (no treatment), (2) Surgery group (surgical castration for testosterone control), (3) NS group (normal saline intratesticular injection), (4) HS group (hypertonic saline), and (5) HM group. We injected 20 mL per testicle. We took 5 mL blood from all donkeys before injection. Castration was performed under general anesthesia 60 days later. Samples included blood and testicular tissue. Total motility (TM), progressive motility (PM), movementy features, DNA damage, morphology, viability, and plasma membrane functionality were evaluated. Hormone analyses, histomorphometric studies and oxidative stress indices including total antioxidant capacity (TAC), glutathione peroxidase (GPx), glutathione (GSH), superoxide dismutase (SOD), malondialdehyde (MDA), and NADP+/NADPH were evaluated. Apoptosis, pyroptosis-related Bax, Caspase-1, GSDMD, and Bcl-2 expression were also assessed.ResultsIn HS and HM groups, testosterone, epididymal sperm count, motility, viability, and plasma membrane functionality dropped while sperm DNA damage increased. HS and HM groups had significantly lower histomorphometric parameters, TAC, GPx, SOD, GSH, and Bcl-2 gene expression. MDA, NADP+/NADPH, Bax, Caspase-1, and GSDMD gene expression were substantially higher in the HS and HM groups than in the control group.ConclusionsToxic effects of hypertonic saline and mannitol on reproductive parameters were seen following, hence, they might be considered as a good chemical sterilizing treatment in donkeys.

Evaluation of Spermatogenic Activity of Polyherbal Compound Musalyadi Churna and its Protective Effect Against Nicotine-Induced Testicular Degeneration in Sprague-Dawley Rats.

The ancient science of life known as Ayurveda uses a variety of formulations or herbal compounds to treat a wide range of illnesses. Male infertility accounts for half of all infertility worldwide, and numerous etiological variables play a role. A pharmacologically active tobacco component called nicotine has a negative impact on male fertility and the reproductive system. Musalyadi Churna (MSC) is one of the polyherbal compound formulations mentioned in Ayurveda Classics for aphrodisiac therapy and recommended for the treatment of male infertility. The current study aims to evaluate the spermatogenic potential of MSC along with its protective effect against testicular degeneration induced by nicotine in rat models. Sixty adult Sprague-Dawley male rats were used in the experiment, which were divided into ten groups of six rats each. Nicotine was administered orally as disease control. The standard and vehicle control were also included along with study drug groups in which MSC was administered orally to the rats. The study was conducted for 70 days for each group with assessment of body weight and food consumption. After treatment, rats were weighed and sacrificed for evaluation of parameters like organ weight, epididymal sperm count, and sperm morphology along with histological assessment. The data were statistically evaluated using analysis of variance test and Bonferroni multiple-comparison range test to compare different treatment groups. In the present study, MSC possesses substantial spermatogenic efficacy, and the result indicates that the study drug possesses a protective effect against nicotine-induced impaired testicular functions in rats, but further research to elucidate its exact mechanism of action is essential.

Effects of Perilla frutescens Var. Acuta in Busulfan-Induced Spermatogenesis Dysfunction Mouse Model.

The leaves of Perilla frutescens var. acuta (PFA) are generally reported to have antioxidant, anti-allergic, anti-inflammatory, and antitumor effects and commonly used as a traditional medicine in East Asia. This study aimed to investigate the protective effect and antioxidant activity of PFA on busulfan-induced testicular dysfunction, histological damage, oxidative stress (OS), sperm quality, and hormone levels using a mouse model. C57BL/6 male mice were divided into four groups: control, busulfan-only treated, and varying concentrations of PFA (100 and 200 mg/kg) with busulfan. In the busulfan group, 40 mg/kg of busulfan was intraperitoneally injected to induce azoospermia. Mice were orally administered PFA for 35 consecutive days after busulfan administration. Samples were collected and assessed for testis/body weight, testicular histopathology, sperm quality, serum hormone levels, and OS to evaluate the effects of PFA treatment on spermatogenesis dysfunction induced by busulfan. The busulfan-induced testicular dysfunction model showed reduced testis weight, adverse histological changes, significantly decreased sex hormones and sperm quality, and attenuated OS. These results indicate that PFA treatment significantly increased testis weight, testis/body weight, epididymal sperm count, motility, and testosterone level compared with busulfan alone. PFA treatment also attenuated the busulfan-induced histological changes. Furthermore, compared with mice treated with busulfan alone, PFA supplementation upregulated the testicular mRNA expression of the antioxidant enzymes superoxide dismutase 1 (Sod1) and glutathione peroxidase 1 (Gpx1), with a decrease in malondialdehyde (MDA) production and an increase in SOD and GPx activities. This study shows that PFA exerts a protective effect against testicular damage by attenuating OS induced by busulfan. Our results suggest that PFA is a potentially relevant drug used to decrease the side effects induced by busulfan on testicular function and sperm during cancer chemotherapy.

SP-Max-A herbomineral formulation attenuates busulfan-induced oligospermia in mice by preventing loss of reproductive hormones

SP-Max herbal capsule formulation contains Withania somnifera, Asparagus recemosus, Mucuna pruriens, Chlorophytum arundinaceum, Ipomoea digitata, and Dioscorea bulbifera which are reported in the ‘Ayurveda’, an Indian Traditional System of medicine as aphrodisiacs. The present study focused on the effect of herbomineral formulation, SP-Max in the treatment of oligospermia. Oligospermia was induced in male Swiss Albino mice by a single intraperitoneal injection of busulfan at a dose of 45 mg/kg. SP-Max herbomineral formulation was given at various doses of 130, 270, and 390 mg/kg for 45 days. Treatment with SP-Max herbomineral formulation at 130, 270 and 390 mg/kg doses significantly improved the sperm count, sperm motility and viability (p < 0.001). SP-Max treatment at a dose of 390 mg/kg significantly prevented the loss of anti-oxidant enzymes in testicular cells. SP-Max prevented the reduction in the level of testosterone, luteinizing hormone, and follicle-stimulating hormone. Histological findings showed that SP-Max treatment prevented degeneration of spermatid, interstitial cells, and Sertoli cells of the testes and also improved epididymal sperm count. High dose of SP-Max treatment i.e 390 mg/kg found to be more effective. Results showed that SP-Max herbomineral formulation is an effective treatment option for oligospermia by decreasing free radical damage to the testes and improving the levels of reproductive hormones.

Epigenetic transgenerational effects of PM2.5 collected from southern Taiwan on sperm functions and DNA methylation in mouse offspring

During respiration, particulate matter with a diameter of 2.5 µm or less (PM2.5) suspended in the atmosphere enters the terminal alveoli and blood. PM2.5 particles can attach to toxic substances, resulting in health problems. Limited information is available regarding the effects of prenatal exposure to water-soluble PM2.5 (WS-PM2.5) and water-insoluble PM2.5 (WI-PM2.5) on male reproduction. In addition, whether exposure to these particles has transgenerational effects remains unknown. We investigated whether prenatal exposure to WS-PM2.5 and WI-PM2.5 disrupts sperm function in generations F1, F2, and F3 of male mice. Pregnant BALB/c mice were treated using intratracheal instillation on gestation days 7, 11, and 15 with 10 mg of a water extract or insoluble PM2.5. On postnatal day 105, epididymal sperm count, motility, morphology, mitochondrial membrane potential (MMP), reactive oxygen species (ROS) production, the sperm chromatin DNA fragmentation index (DFI), and testicular DNA methyltransferase (Dnmt) levels were evaluated in all generations. Whole-genome bisulfite sequencing was used to analyze the DNA methylation status of generation F3. According to the results, exposure to WS-PM2.5 affected sperm morphology, ROS production, and mean DFI in generation F1; ROS production and mean DFI in generation F2; and sperm morphology and MMP in generation F3. Similarly, exposure to WI-PM2.5 affected sperm morphology, ROS production, mean DFI, %DFI, and Dnmt1 expression in generation F1; sperm morphology, MMP, and ROS production in generation F2; and sperm morphology, ROS, and %DFI in generation F3. Two hypermethylated genes, PRR16 and TJP2, were observed in the WS-PM2.5 and WI-PM2.5 groups, two hypomethylated genes, NFATC1 and APOA5, were observed in the WS-PM2.5 group, and two hypomethylated genes, ZFP945 and GSE1, were observed in the WI-PM2.5 group. Hence, prenatal exposure to PM2.5 resulted in transgenerational epigenetic effects, which may explain certain phenotypic changes in male reproduction.

Rutin attenuated hexachlorobenzene-induced testicular injury via regulation of oxidative stress, steroidogenic enzymes and apoptotic process in male rats

Hexachlorobenzene (HCB) is a pesticide that is constantly produced as a by-product in many industrial processes. Rutin (RUT) is a flavonoid found in many plants with numerous biological and pharmacological properties. However, there is no scientifical information on the impact of RUT on HCB-induced reproductive toxicity in male rats. Thus, we investigated the effect of rutin on HCB-induced reproductive toxicity. Rats were divided into four groups consisting of control administered with corn oil, HCB alone (16 mg/kg), RUT alone (100 mg/kg) and HCB + RUT that were treated orally by gavage for 28 days. Hexachlorobenzene caused a significant reduction in epididymal sperm count, sperm progressive motility, viability, number of hypo-osmotic tail swelled spermatozoa and increased sperm morphological abnormalities (head, midpiece and tail). Furthermore, HCB decreased the activities of gamma glutamyl transferase, lactate dehydrogenase, sorbitol dehydrogenase, superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase, levels of glutathione and total protein with increased malondialdehyde level. Additionally, HCB increased the levels of total cholesterol, low density lipoprotein, triglycerides, follicle stimulating hormone, luteinizing hormone but decreased high density lipoproteins, serum testosterone, steroidogenic enzymes (3β-hydroxysteroid dehydrogenase and 17β-hydroxysteroid dehydrogenase), B-cell lymphoma and increased pro-apoptotic markers (Bcl-2 associated protein (Bax), caspases-3 and-9) alongside histological and histomorphometrical abnormalities. Nevertheless, RUT prevented all the aberrations caused by HCB. Our result showed that HCB induced reproductive toxicity in rats and RUT protected against HCB-induced r reproductive toxicity.

Combination Therapy with Enalapril and Paricalcitol Ameliorates Streptozotocin Diabetes-Induced Testicular Dysfunction in Rats via Mitigation of Inflammation, Apoptosis, and Oxidative Stress.

As the impacts of diabetes-induced reproductive damage are now evident in young people, we are now in urgent need to devise new ways to protect and enhance the reproductive health of diabetic people. The present study aimed to evaluate the protective effects of enalapril (an ACE inhibitor) and paricalcitol (a vitamin D analog), individually or in combination, on streptozotocin (STZ)-diabetes-induced testicular dysfunction in rats and to identify the possible mechanisms for this protection. This study was carried out on 50 male Sprague-Dawley rats; 10 normal rats were allocated as a non-diabetic control group. A total of 40 rats developed diabetes after receiving a single dose of STZ; then, the diabetic rats were divided into four groups of equivalent numbers assigned as diabetic control, enalapril-treated, paricalcitol-treated, and combined enalapril-and-paricalcitol-treated groups. The effects of mono and combined therapy with paricalcitol and enalapril on testicular functions, sperm activity, glycemic state oxidative stress, and inflammatory parameters, as well as histopathological examinations, were assessed in comparison with the normal and diabetic control rats. As a result of diabetes induction, epididymal sperm count, sperm motility, serum levels of testosterone, follicle-stimulating hormone (FSH) as well as luteinizing hormone (LH), and the antioxidant enzyme activities, were significantly decreased, while abnormal sperm (%), insulin resistance, nitric oxide (NO), malondialdehyde (MDA), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) were significantly increased, along with severe distortion of the testicular structure. Interestingly, treatment with paricalcitol and enalapril, either alone or in combination, significantly improved the sperm parameters, increased antioxidant enzyme activities in addition to serum levels of testosterone, FSH, and LH, reduced insulin resistance, IL-6, and TNF-α levels, and finally ameliorated the diabetes-induced testicular oxidative stress and histopathological damage, with somewhat superior effect for paricalcitol monotherapy and combined therapy with both drugs compared to monotherapy with enalapril alone. Monotherapy with paricalcitol and its combination therapy with enalapril has a somewhat superior effect in improving diabetes-induced testicular dysfunction (most probably as a result of their hypoglycemic, antioxidant, anti-inflammatory, and anti-apoptotic properties) compared with monotherapy with enalapril alone in male rats, recommending a synergistic impact of both drugs.

Cylicins are a structural component of the sperm calyx being indispensable for male fertility in mice and human.

Cylicins are testis-specific proteins, which are exclusively expressed during spermiogenesis. In mice and humans, two Cylicins, the gonosomal X-linked Cylicin 1 (Cylc1/CYLC1) and the autosomal Cylicin 2 (Cylc2/CYLC2) genes, have been identified. Cylicins are cytoskeletal proteins with an overall positive charge due to lysine-rich repeats. While Cylicins have been localized in the acrosomal region of round spermatids, they resemble a major component of the calyx within the perinuclear theca at the posterior part of mature sperm nuclei. However, the role of Cylicins during spermiogenesis has not yet been investigated. Here, we applied CRISPR/Cas9-mediated gene editing in zygotes to establish Cylc1- and Cylc2-deficient mouse lines as a model to study the function of these proteins. Cylc1 deficiency resulted in male subfertility, whereas Cylc2-/-, Cylc1-/yCylc2+/-, and Cylc1-/yCylc2-/- males were infertile. Phenotypical characterization revealed that loss of Cylicins prevents proper calyx assembly during spermiogenesis. This results in decreased epididymal sperm counts, impaired shedding of excess cytoplasm, and severe structural malformations, ultimately resulting in impaired sperm motility. Furthermore, exome sequencing identified an infertile man with a hemizygous variant in CYLC1 and a heterozygous variant in CYLC2, displaying morphological abnormalities of the sperm including the absence of the acrosome. Thus, our study highlights the relevance and importance of Cylicins for spermiogenic remodeling and male fertility in human and mouse, and provides the basis for further studies on unraveling the complex molecular interactions between perinuclear theca proteins required during spermiogenesis.

Antioxidant, anti-inflammatory, and anti-apoptotic effects of genkwanin against aflatoxin B1-induced testicular toxicity

Aflatoxin B1 (AFB1) is the most hazardous aflatoxin that causes significant damage to the male reproductive system. Genkwanin (GNK) is a bioactive flavonoid that shows antioxidant and anti-inflammatory potential. Therefore, the current study was planned to evaluate the effects of GNK against AFB1-induced testicular toxicity. Forty-eight male rats were distributed into four groups (n = 12 rats). AFB1 (50 μg/kg) and GNK (20 mg/kg) were administered to the rats for eight weeks. Results of the current study revealed that AFB1 exposure induced adverse effects on the Nrf2/Keap1 pathway and reduced the expressions and activities of antioxidant enzymes. Additionally, it increased the levels of oxidative stress markers. Furthermore, expressions of steroidogenic enzymes were down-regulated by AFB1 intoxication. Besides, AFB1 exposure reduced the levels of gonadotropins and plasma testosterone, which subsequently reduced the epididymal sperm count, motility, and hypo-osmotic swelled (HOS) sperms, while increasing the number of dead sperms and causing morphological anomalies of the head, midpiece, and tail of the sperms. In addition, AFB1 decreased the activities of testicular function marker enzymes and the levels of inflammatory markers. Moreover, it severely affected the apoptotic profile by up-regulating the expressions of Bax and Casp3, while down-regulating the Bcl2 expression. Besides, AFB1 significantly damaged the histoarchitecture of testicular tissues. However, GNK treatment reversed all the AFB1-induced damages in the rats. Taken together, the current study reports the potential use of GNK as a therapeutic agent to prevent AFB1-induced testicular toxicity due to its antioxidant, anti-inflammatory, and anti-apoptotic properties.

Assessment of reproductive toxicity of Cyfluthrin and Pestban either individually or combined in adult male albino rats

Many studies on individual pesticide risk assessments are available, but the toxicity of combined usage is still to estimate. So, the current study investigated reproductive toxicity induced by exposure to cyfluthrin (CYF) and pestban (PES) and their mixture in adult male albino rats. Forty adult male albino rats were randomized into four groups. All treatments were given daily by oral gavage for 60 days. Group I (control group): this group included 16 rats, divided into two equal subgroups: subgroup Ia (negative control) and subgroup Ib (a vehicle control), in which each rat received 2 ml of corn oil. Group II: CYF group (15.6 mg/kg). Group III; PES group (7.45 mg/kg). Group IV: CYF + PES. Individual CYF and PES exposure significantly decreased testicular weight, serum testosterone level, and epididymal sperm count when compared to the control group. These biochemical changes were confirmed by histological and ultra-structural disarray and reduced immunoreactions of Melan-A (also known as MART-1, Melanoma Antigen Recognized by T-cells), but mutual exposure to both pesticides resulted in a highly significant difference compared to other treated groups. Co-administration of CYF and PES aggravated testicular toxicity, exhausting the endogenous antioxidant status, and down-regulating the immune expression of Melan-A. So, mixing both components can intensify the damaging effects of each compound on testes.

Protective effect of didymin against 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin-induced reproductive toxicity in male rats.

2, 3, 7, 8-Tetrachlorodibenzo-p-dioxin (TCDD) is one of the most potent environmental toxicants, which causes oxidative stress and adversely affects the male reproductive system. The current study aimed to evaluate the ameliorative role of didymin (DDM) against TCDD-induced testicular toxicity. Forty-eight male Sprague-Dawley rats were divided into four equal groups (n=12). (i) Control group, (ii) TCDD-induced group was provided with 10 μg/kg/day of TCDD, (iii) TCDD + DDM group received 10 μg/kg/day of TCDD and 2 mg/kg/day of DDM, and (iv) DDM-treated group was administered with 2 mg/kg/day of DDM. After 56 days of treatment, biochemical, steroidogenic, hormonal, spermatogenic, apoptotic, and histopathological parameters were estimated. TCDD affected the biochemical profile by reducing the activities of antioxidant enzymes, while increasing the levels of malondialdehyde (MDA) and reactive oxygen species (ROS). Furthermore, it decreased the expressions of steroidogenic enzymes, 3β-hydroxysteroid dehydrogenase (HSD), 17β-HSD, steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (CYP11A1), and 17α-hydroxylase/17, 20-lyase (CYP17A1), as well as reduced the levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and plasma testosterone. Besides, epididymal sperm count, viability, and motility were decreased, while sperm morphological anomalies were increased. Moreover, TCDD altered the apoptotic profile by up-regulating the expressions of Bax and caspase-3, while downregulated the Bcl-2 expression. Additionally, histopathological damages were prompted due to TCDD administration. However, DDM restored all the TCDD-induced damages owing to its antioxidant, anti-apoptotic, and androgenic potential. Our data suggested that DDM might play its role as a therapeutic agent against TCDD-prompted testicular toxicity.

Deep-frying palm olein oil-fried street falafel induces testicular toxicity in rats

Falafel is the most common Egyptian street food, and deep-frying palm oil is a commonly used for frying it. The main objective of this research was to evaluate the effect of deep-frying palm olein oil-fried street falafel on testes of Wistar rats. Twenty-one adult male Wistar rats were randomly divided into three equal groups and given treatment as follows: control group received only distilled water, fresh palm olein oil (FPO) group received FPO (1 ml/100 g BW/day) and deep-frying palm olein oil (DPO) group received DPO (1 ml/100 g BW/day) orally for 28 days. Serum level of testosterone, testicular tissue oxidative status, and sperm characteristics were determined. Testicular specimens were processed for histopathological examination. The results revealed that DPO group showed a significant (p<0.01) decrease in serum testosterone with significant (p<0.01) reduction of testicular glutathione, superoxide dismutase, and glutathione peroxidase, whereas testicular malondialdehyde was markedly (p<0.001) raised. There were significant decrease in epididymal sperm count (p<0.01), sperm progress motility (p<0.001), and increase abnormal sperm rate (p<0.001) in DPO group. Testicular histology in DPO group showed structural abnormalities which are compatible with lipid peroxidation and antioxidant deficiency. In Conclusion, deep-frying palm olein oil that used for the frying falafel induces testicular abnormalities in rats.

Effect of sunflower seed husk and rape straw in total mixed ration during peri-puberty on testis development in Hu lambs

Sunflower seed husk (SH) and rape straw (RS) are widely used in mutton sheep husbandry in the Hexi Corridor of China. This study explored whether or not a pre-puberty lamb diet consisting of sunflower SH and RS affects ram testis development. Fifty 60-day-old healthy Hu ram lambs (average body weight of 22.3 ± 0.1 kg) were randomly divided into two groups. In each treatment group, 25 lambs were allocated to five pens (five lambs per pen). The SH group received a diet containing 20 % SH, and the RS group received a diet containing 20 % RS. After a 77-day feeding trial, three lambs, except for the heaviest and lightest in each pen, were humanely slaughtered and investigated. Results revealed that the two groups had no difference in body weight (39.23 ± 0.52 kg vs. 38.53 ± 0.46 kg, P = 0.321). However, the testis weight (288.74 ± 23.45 g vs. 215.18 ± 17.96 g), volume (294.29 ± 25.39 mL vs. 208.67 ± 16.63 mL), epididymis weight (38.38 ± 1.44 g vs. 33.86 ± 1.37 g), testicular index (0.74 ± 0.06 vs 0.56 ± 0.05), and epididymis sperm count (4.88 ± 0.92 ×109/g vs. 2.27 ± 0.69 ×109/g) in the SH group were significantly higher than those in the RS group (P < 0.05). SH reduced the mtDNA relative copy number (P < 0.01) in the testis and increased the total antioxidant capacity (3.41 ± 0.94 U/mg prot vs. 1.54 ± 0.38 U/mg prot, P = 0.05). RNA sequencing results showed 50 differentially expressed genes, with 27 genes upregulated and 23 genes downregulated in the SH group compared with the RS group. KEGG analysis revealed that the pathways related to polyunsaturated fatty acid metabolism, and vitamin digestion and absorption were enriched. The findings suggest that feeding SH in the early reproductive development stage of lambs is more beneficial for testis development and spermatogenesis compared with feeding RS.

Fenofibrate and Diosmetin in a rat model of testicular toxicity: New insight on their protective mechanism through PPAR-α/NRF-2/HO-1 signaling pathway

One of the most significant chemotherapeutic side effects of cisplatin (Cis) that limits its use and efficacy is testicular toxicity. Thus, the objective of the present study was to investigate the possible ameliorative effect of Fenofibrate (Fen), Diosmetin (D), and their combination against cis-mediated testicular damage. Fifty-four adult male albino rats were randomly allocated into nine groups (6 rats each): Control group, Fen (100 mg/kg), D20 (20 mg/kg), D40 (40 mg/kg), Cis group (7 mg/kg), Cis +Fen group (7 mg/kg+100 mg/kg), Cis+D20 group (7 mg/kg+20 mg/kg), Cis+D40 group (7 mg/kg+40 mg/kg), Cis+Fen+D40 treated group (7 mg/kg+100 mg/kg+40 mg/kg). Relative testicular weight, epididymal sperm count and viability, serum testosterone level, testicular oxidative stress indices, mRNA expression of peroxisome proliferator-activated receptor alpha (PPAR-α), nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1), histopathological, and immunohistochemical alterations were assessed. Our results revealed that cis administration induced testicular oxidative and inflammatory damage as indicated by a substantial reduction in relative testicular weight, sperm parameters, serum testosterone levels, the antioxidant enzyme activity of catalase, and Johnson's histopathological score, PPAR-α/NRF-2/HO-1 and proliferating cell nuclear antigen (PCNA) immunoexpression with marked increment in malondialdehyde (MDA), Cosentino's score, nuclear factor kappa B (NF-κβ p65), interleukin (IL)− 1β and caspase 3 in testicular tissue. Interestingly, Fen and D diminished the harmful effects of cis on testes via upregulation of the antioxidant activities and downregulation of lipid peroxidation, apoptosis, and inflammation. Moreover, the combination therapy Fen/D40 also exhibited a more pronounced enhancement of previous markers than either treatment alone. In conclusion, because of their antioxidant, anti-inflammatory, and anti-apoptotic properties, cotreatment with Fen or D or their combination could be beneficial in reducing the harmful impacts of cis on testicular tissue, particularly in patients that receive cis chemotherapy.

The possible protective effect of the nuclear factor kappa B inhibitor pyrrolidine dithiocarbamate on tramadol-induced testicular damage in rats

BackgroundTramadol provokes oxidative stress that gives rise to apoptosis with alterations in the cellular structure and adversely influences male fertility. A specific inhibitor of nuclear factor kappa B (NF-κB), pyrrolidine dithiocarbamate (PDTC), has observable antioxidant and anti-inflammatory characteristics and enhances the improvement of organs damage caused by various agents. The impact of PDTC on testicular damage caused by tramadol has not been previously examined.ObjectiveThis study was designed to investigate the potential impact of pyrrolidine dithiocarbamate on testicular damage provoked by chronic tramadol usage.Materials and methodsForty healthy adult male albino rats were included in this study. Rats were randomly and equally divided into 4 groups: group (I), control group; group (II), pyrrolidine dithiocarbamate (PDTC) group; group (III), tramadol (Tr) group; and group (IV), Tr + PDTC group. This study measured serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), and free testosterone levels. Testicular malondialdehyde (MDA), interleukin-1β (IL-1β), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) levels, and superoxide dismutase (SOD), glutathione peroxidase (GPX), and caspase-8 and caspase-3 activities were also detected. Immuno-histochemical evaluation of inducible NOS (iNOS) expression in testicular tissue along with histopathological examination of testicular tissue has been studied.ResultsTramadol caused a significant reduction in serum FSH, LH and testosterone levels, epididymal sperm count, and motility, as well as testicular GPX and SOD activities. On the other hand, a significant elevation of testicular MDA, IL-1β, IL-6, and TNF-α levels and caspase-8 and caspase-3 activities were found. However, PDTC administration with tramadol showed significantly increased sperm production and motility and alleviated tramadol-induced disturbance in other measured parameters in the Tr + PDTC group compared to the Tr group. Moreover, co-administration of PDTC with tramadol significantly alleviated the histopathological structure of testicular tissue and the increased iNOS expressions noticed in the tramadol-treated group.ConclusionConsidering the protective effects of PDTC against the reproductive toxicity induced by tramadol, this compound can be used as a possible protective and treating target for tramadol-induced reproductive toxicity.

Radical scavenging activity of Chlorophytum borivilianum L. root extract and its protective role in cauda epididymal sperm integrity in Mus musculus after gamma irradiation.

Background: Chlorophytum borivilianum L. is a recognized herbal medicine for the management of impotency in South Asian countries. In Ayurveda, it is used for the management of multiple health conditions, including diabetes, infection, and cardiovascular diseases. Parts of the plant have been used as excellent antioxidants and scavengers of free radicals. Since oxidative stress plays an important role in spermatogenesis and fertility in male populations, this study evaluated the role of ethanolic extract of C. borivilianum roots in epididymal sperm maturation against adversities posed by ionizing gamma irradiation. Materials and methods: Antioxidant potential of C. borivilianum root extract (CRE) was evaluated through DPPH (2,2-diphenylpicrylhydrazyl) and NO (nitric oxide) scavenging assays. Four groups of healthy Swiss albino mice were constituted, which were labeled as follows: Group I: sham control, Group II: 7-day pre-treatment with 50mg/kg CRE, Group III: 6Gy irradiation without pre-treatment, and Group IV: 7-day pre-treatment with 50mg/kg CRE and 6Gy irradiation on day 7. Swiss albino mice were observed for 30 days and later sacrificed to evaluate sperm quality parameters. Results: CRE showed a remarkable antioxidant potential with IC50 values of 46.37μg/ml and 98.39μg/ml for DPPH and NO, respectively. A significant decline (p < 0.001) in cauda epididymal sperm count, motility, and viability was observed in Group III animals. Group IV also showed a substantial decline (p < 0.01) in all three parameters compared to Group I; nonetheless, these were significantly higher than Group III. Morphological alterations indicated a coiled and bent tail, with the presence of cytoplasmic droplets in Group III, which declined substantially in Group IV. The ultrastructure of sperm indicated higher curvature of hook in Group III than Group IV, indicating specific interferences in the sperm maturation process. Conclusion: It was concluded that pre-treatment with 50mg/kg body weight of CRE could protect sperm during epididymal maturation against oxidative stress.

Therapeutic effect of gossypetin against paraquat-induced testicular damage in male rats: a histological and biochemical study.

Paraquat (PQ) is an organic compound, which is commonly used as a herbicide in the agriculture sector, and it is also known to stimulate critical damages in the male reproductive system. Gossypetin (GPTN) is one of important members of the flavonoid family, which is an essential compound in flowers and calyx ofHibiscus sabdariffawithpotential pharmacological properties. The current investigation was aimed to examine the ameliorative potential of GPTN against PQ-instigated testicular damages. Adult male Sprague-Dawley rats (n = 48) were distributed into four groups: control, PQ (5mg/kg), PQ + GPTN (5mg/kg + 30mg/kg respectively), and GPTN (30mg/kg). After 56days of treatment, biochemical, spermatogenic indices, hormonal, steroidogenic, pro-or-anti-apoptotic, and histopathological parameters were estimated. PQ exposure disturbed the biochemical profile by reducing the activities of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx), and glutathione reductase (GSR), while it increased the concentration of reactive oxygen species (ROS) and malondialdehyde (MDA) level. Furthermore, PQ exposure decreased the sperm motility, viability, number of hypo-osmotic tail swelled spermatozoa, and epididymal sperm count; additionally, it increased sperm morphological (head mid-piece and tail) abnormalities. Moreover, PQ lessened the follicle-stimulating hormone (FSH), luteinizing hormone (LH), and plasma testosterone levels. Besides, PQ-intoxication downregulated the gene expression of steroidogenic enzymes (StAR, 3β-HSD, and 17β-HSD) and anti-apoptotic marker (Bcl-2), whereas upregulated the gene expression of apoptotic markers (Bax and Caspase-3). PQ exposure led to histopathological damages in testicular tissues as well. Nonetheless, GPTN inverted all the illustrated impairments in testes. Taken together, GPTN could potently ameliorate PQ-induced reproductive dysfunctions due to its antioxidant, androgenic, and anti-apoptotic potential.

Mechanistic insight into the protective effects of fisetin against arsenic-induced reproductive toxicity in male rats

Arsenic is one of the most hazardous environmental contaminants, which adversely affects the dynamics of male reproductive system. Fisetin (FIS) is a bioactive flavonoid, which is known to exert strong antioxidative effects. Therefore, the current research was planned to evaluate the alleviative efficacy of FIS against arsenic-induced reproductive damages. Forty-eight male albino rats were divided into 4 groups (n = 12), which were treated as follows: (1) Control, (2) Arsenic-intoxicated group (8 mg kg−1), (3) Arsenic + FIS-treated group (8 mg kg−1 + 10 mg kg−1), and (4) FIS-treated group (10 mgkg−1). After 56 days of treatment, the biochemical, lipidemic, steroidogenic, hormonal, spermatological, apoptotic and histoarchitectural profiles of rats were analyzed. Arsenic intoxication reduced the enzymatic activities of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GSR), in addition to glutathione (GSH) level. Conversely, the levels of thiobarbituric acid reactive substance (TBARS) and reactive oxygen species (ROS) were increased. Moreover, it escalated the level of low-density lipoprotein (LDL), triglycerides and total cholesterol, while declining the level of high-density lipoprotein (HDL). Furthermore, steroidogenic enzymes expressions, 3β-hydroxysteroid dehydrogenase (HSD), 17β-HSD, steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (CYP11A1) and 17α-hydroxylase/17, 20-lyase (CYP17A1), were found to be reduced, which brought down the level of testosterone. Besides, the levels of gonadotropins (LH and FSH) were decreased. Additionally, a decline in sperm mitochondrial membrane potential (MMP), motility, epididymal sperm count and hypo-osmotic swelling (HOS) coil-tailed sperms was observed, whereas the dead sperms and structural damages (head, midpiece and tail) of sperms were escalated. Moreover, arsenic exposure up-regulated the mRNA expressions of apoptotic markers, namely Bax and caspase-3, whereas lowered the expression of anti-apoptotic marker, Bcl-2. In addition, it induced histoarchitectural changes in testes of rats. However, FIS treatment resulted in remarkable improvements in testicular and sperm parameters. Therefore, it was inferred that FIS could serve as a therapeutic candidate against arsenic-generated male reproductive toxicity attributing to its anti-oxidant, anti-lipoperoxidative, anti-apoptotic, and androgenic efficacy.