Epimedium polysaccharide alleviates adenine-induced oligoasthenozoospermia in mice by inhibiting ferroptosis and inflammation through TLR4/NF-κB and SLC7A11/GPX4 axis.

Nurture vs nature: how can we optimize sperm quality?

Icariin alleviates the injury of Sertoli cell junction function by upregulating PKR pathway via ERα/c-fos signaling in aged mice

Ginsenoside Rb1 combined with Lycium barbarum polysaccharide alleviate the Tripterygium wilfordii polyglycoside-induced oligoasthenozoospermia in mice by inhibiting ZnT3-mediated oxidative stress response.

Hyperuricemia (HUA) is a known factor contributing to testicular spermatogenic dysfunction. Shenling Baizhu San (SLBZS), a traditional Chinese medicine compound, has demonstrated efficacy in reducing uric acid levels. However, its specific impact on testicular spermatogenic function in mice with HUA remains unclear. To investigate the impact and mechanism of SLBZS on testicular spermatogenic function in HUA mice. A hyperuricemia-induced spermatogenic dysfunction model was created by administering intraperitoneal injections of potassium oxonate (600mg/kg/d) for seven days. Following model establishment, 48 Balb/c mice were randomly divided into six groups: control, model, low-dose SLBZS (5.04g/kg/d), medium-dose SLBZS (10.07g/kg/d), high-dose SLBZS (20.14g/kg/d), and febuxostat (10mg/kg/d). All groups, except the control, underwent model induction, followed by specific interventions. Subsequent analyses included serum uric acid levels, testicular and epididymal indices, histopathological assessments, sperm quality, oxidative stress and inflammation markers, and the expression of proteins related to apoptosis and inflammation signaling pathways. SLBZS markedly enhanced sperm quality, testicular and epididymal indices, and serum uric acid levels in mice, while ameliorating histopathological lesions in testicular tissue. Additionally, SLBZS significantly reduced oxidative stress, serum inflammation markers, and testicular cell apoptosis, with the high-dose group showing superior effects compared to the febuxostat group. Further investigation revealed that SLBZS inhibited the expression and phosphorylation of proteins in the MAPK/NF-κB pathway and suppressed the expression of proteins in the NLRP3 inflammasome pathway. SLBZS potentially modulates the MAPK/NF-κB and NLRP3 inflammasome signaling pathways, thereby suppressing inflammatory responses and enhancing spermatogenesis in the testes of HUA mice.

Background: Oligoasthenozoospermia (OA) is a common cause of male infertility. Modified Liuwei Dihuang Decoction (MLWDH) is an improved version of Liuwei Dihuang Decoction (LWDH), a traditional Chinese medicine prescription, which has demonstrated significant therapeutic effects against OA. This study aims to evaluate the protective effects of MLWDH against OA and elucidate its underlying molecular mechanisms. Methods: The constituents of MLWDH were identified via UPLC-HRMS and compound databases (TCMSP, HERB). Network pharmacology analysis was conducted to predict potential therapeutic targets and associated signaling pathways. In vivo, a CP-induced mouse model of OA was established to evaluate the therapeutic efficacy of MWDH by assessing testicular and epididymal indices, sperm quality, histopathological changes and serum hormone levels. Oxidative stress markers, including MDA, SOD, GSH and NO, were measured using commercial assay kits. The underlying molecular mechanisms, particularly those related to oxidative stress and inflammation (PI3K, Akt, Nrf2, Keap1, HO-1, NQO1, NF-κB, TNF-α, IL-6), were further elucidated by RT-qPCR, Western blot, and immunofluorescence. Results: A total of 345 major bioactive compounds were identified in MLWDH. Network pharmacology and molecular docking analyses indicated that MLWDH exerts its effects primarily through the PI3K/AKT signaling pathway. MLWDH administration in vivo significantly improved sperm count, motility, and morphology, while also increasing serum levels of testosterone, FSH, and LH. Moreover, MLWDH significantly mitigated oxidative damage, as evidenced by decreased MDA concentrations and elevated levels of GSH, NO and SOD. Mechanistic investigations further substantiated that MLWDH enhanced PI3K/AKT/Nrf2 signaling while inhibiting NF-κB signaling in OA mice. Conclusions: Our findings suggest that MLWDH ameliorates OA in a preclinical mouse model by improving sperm quality and testicular function, potentially via activation of the PI3K/AKT/Nrf2 signaling pathway and the inhibition of NF-κB signaling, thereby alleviating oxidative stress and inflammatory responses.

Supplementation of Epimedium polysaccharide (EPS) improves goat semen characteristics following cryopreservation

PurposeOne of the most common types of male infertility is recognized as oligoasthenozoospermia (OA), characterized by low sperm count and quality in males. As a traditional Chinese medicine (TCM), Cuscutae Semen-Mori Fructus coupled-herbs (CSMFCH) has been known to act a curative effect on OA for thousands of years. Nevertheless, the substantial basis and molecular mechanism of CSMFCH in treating OA remain elusive.MethodsHerein, an integrated approach, including network pharmacology, molecular docking, and experiment validation, was utilized to reveal the new candidate active component and mechanism of CSMFCH in treating OA.ResultsThe results show that kaempferol is the most significant bioactive component of CSMFCH on OA. The mechanism and targets of CSMFCH against OA are relevant to hormone regulation, oxidant stress, and reproductive promotion. In order to validate network pharmacology results, molecular docking and experiment validation were conducted. In detail, molecular docking was employed to verify the strong binding interactions between kaempferol and the core targets. UHPLC-Q-Orbitrap-MS was used to identify kaempferol in the CSMFCH extract. In vitro and in vivo experiments further proved CSMFCH and kaempferol could enhance the mouse Leydig (TM3) and mouse Sertoli (TM4) cell viability, improve the male reproductive organ weights, sperm quality, and decrease testis tissue damage in the OA mouse model induced by CP.ConclusionOur results not only identify the new candidate active component of CSMFCH in treating OA but also provide new insights into the mechanisms of CSMFCH against OA.

This study aims to reveal the mechanism of prenatal stress in affecting the testicular development of offspring rats and the intervention effects of Zuogui Pills via connexin 43(Cx43). Forty pregnant SD rats were randomized into a blank control group, a mo-del group, a high-dose(18.9 g·kg~(-1)) Zuogui Pills group, a low-dose(9.45 g·kg~(-1)) Zuogui Pills group, and a vitamin E(1.44 mg·kg~(-1)) group. The other groups except the blank control group was subjected to chronic unpredictable mild stress for the modeling of prenatal stress. The model was evaluated by sucrose preference test, open field test, and enzyme-linked immunosorbent assay(ELISA) of the glucocorticoid level. ELISA was employed to measure the thyroxine 4(T4), testosterone(T), and follicle-stimulating hormone(FSH) levels to assess kidney deficiency. Hematoxylin-eosin(HE) staining was employed to evaluate the status of testicular germ cells. An automatic sperm analyzer was used to measure the sperm quality. Immunofluorescence double staining was employed to detect the expression of Cx43 and follicle-stimulating hormone receptor(FSHR) in the testes of offspring rats. The mRNA and protein levels of Cx43, FSHR, phosphatidylinositol 3-kinase(PI3K), and protein kinase B(Akt) were determined by real-time fluorescence quantitative polymerase chain reaction and Western blot, respectively. Prenatal stress induced testicular development disorders in offspring rats. The HE staining results showed that on the day of birth, the model group had reduced seminiferous tubules in the testes, elevated FSH level in the serum, and lowered Cx43 level in the testicular tissue. Male offspring rats of 60 days old had reduced testicular spermatogenic function, decreased sperm quality, elevated FSH level and lowered T level in the serum, and down-regulated protein and mRNA levels of Cx43, FSHR, PI3K, and Akt in the testicular tissue. Zuogui Pills alleviated the abnormal development and dysfunction of testicles in the offspring rats caused by prenatal stress. In summary, Zuogui Pills may weaken the effects of prenatal stress on testicular development and spermatogenic function of offspring rats by activating the PI3K/Akt pathway to regulate Cx43 expression in the testicular tissue.

Yttrium nitrate activates the oxidative stress-mediated NF-κB pathway to induce testicular inflammatory response and reduce sperm quality in mice.

To investigate the serum levels of reproductive hormones in men with azoospermia and explore the correlation between abnormal reproductive hormones and the results of testicular biopsy and seminiferous extraction. The serum levels of luteinizing hormone (LH), follicle stimulating hormone (FSH), testosterone (T), and pituitary prolactin (PRL), were measured by Radio immunoassay. Testicular biopsy was used to determine whether patient's had spermatozoa. Spearman's correlation analysis was used to analyse the correlation between LH, FSH, testicular volume, and spermatogenic function. Receiver operator characteristic (ROC) curves were used to evaluate the efficacy and significance of LH and FSH for the detection of spermatozoa and copy number variation sequencing (CNV-seq). Compared with the control group, the serum levels of FSH and LH in the azoospermia group were significantly up-regulated (p < 0.001). Prolactin (PRL) and testosterone levels did not differ significantly between the two groups (p > 0.05). The serum levels of FSH and LH were negatively correlated with testicular spermatogenic function and testicular volume. The areas under the ROC curves for determining FSH and LH potency by testicular biopsy with or without spermatozoa were 0.640 and 0.622, respectively. The areas under the ROC curves for the potency of FSH and LH, as judged by CNV detection results, were 0.523 and 0.534, respectively. Serum levels of FSH and LH are associated with azoospermia and may be of predictive significance in the clinical setting for the acquisition of spermatozoa by testicular biopsy.

Following testicular ischemia, the return of blood circulation promotes reactive oxygen species formation. By damaging cellular components such as proteins, DNA and lipids, reactive oxygen species negatively affect testicular spermatogenic function. Numerous plant species, particularly those within the Oleaceae family, contain oleanolic acid as a principal active ingredient. Extensive research has confirmed oleanolic acid's efficacy in exerting antioxidant action. We examined the therapeutic potential of oleanolic acid in mitigating testicular damage induced by ischemia-reperfusion in rats. The study included three groups, each comprising twenty male rats: a sham group, an ischemia-reperfusion group, and an ischemia-reperfusion group treated with oleanolic acid (30 mg/kg). Left testicular torsion of 720 degrees counterclockwise, maintained for 2 hours, induced testicular ischemia-reperfusion injury. After surgical detorsion of the left testis, the ischemia-reperfusion + oleanolic acid group was treated immediately with a single 30 mg/kg dose of oleanolic acid via intraperitoneal injection. Multiple analytical procedures were performed on testicular tissues collected from the three rat groups. Biochemical measurements encompassed both nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity (critical for reactive oxygen species production) and malondialdehyde concentration (a reactive oxygen species indicator). We used hematoxylin-eosin staining for the evaluation of spermatogenic function in testicular tissue. Relative to the sham group, the ischemia-reperfusion group exhibited significantly elevated NADPH oxidase activity and malondialdehyde levels in ipsilateral testes, accompanied by impaired spermatogenic function (p<0.05). Oleanolic acid intervention effectively suppressed oxidative stress markers (NADPH oxidase activity and malondialdehyde levels) in ipsilateral testes, relatively enhancing spermatogenic capacity (p<0.05). Overall, oleanolic acid enhances testicular spermatogenic function by lowering NADPH oxidase activity and curbing reactive oxygen species formation.

The study aims to elucidate the changes in testicular spermatogenic function in high-fat diet (HFD)-induced obese rats and to evaluate the protective effects of metformin intervention. Male Sprague-Dawley rats (n = 18) were randomly divided into a control group (standard diet), an HFD group, and a metformin group (HFD + metformin at 100 mg/kg, once daily by oral gavage). After 8 weeks, rats were euthanized, and the weights of body and testes were measured. Testis and epididymis were dissected and hematoxylin-eosin-stained for histopathological examination and semen parameter analysis. Blood samples were collected for assessment of sex hormones and metabolic parameters (serum glucose, insulin, and leptin). Spermatogenic cell apoptosis was accessed by TUNEL. Compared with the control group, the final body weight and weight gain were significantly higher in HFD rats, while the testicle weight and coefficients were lower. In HFD rats, metformin treatment induced weight loss and increased testicle weight (P < 0.05). In HFD rats, obvious pathological changes in the testicular tissue were characterized by small, atrophic, and distorted seminiferous tubules and destroyed basement membrane. Metformin treatment protected against the HFD-induced decrease in the number of spermatogonia, Sertoli cells, and Leydig cells (P < 0.05); ameliorated the HFD-induced increases in serum glucose, insulin, leptin, and estrogen; and decreased serum testosterone (P < 0.05) and reduced the rate of testicular cell apoptosis in obese male rats. Finally, metformin significantly improved semen parameters (including concentration, viability, motility, and normal morphology) in HFD rats (P < 0.05). HFD-induced obesity in rats results in detrimental effects on spermatogenesis, semen quality, endogenous hormones, and testicular cell apoptosis. Metformin intervention improved the semen parameters, possibly due to its effects on weight loss, increased testicular weight, reduced testicular cell apoptosis, and resulted in restoration of hormonal homeostasis and correction of metabolic disorder.

Zinc (Zn) is an important trace element in the human body and plays an important role in growth, development, and male reproductive functions. Marginal zinc deficiency (MZD) is common in the human population and can cause spermatogenic dysfunction in males. Therefore, the aim of this study was to investigate methods to improve spermatogenic dysfunction caused by MZD and to further explore its mechanism of action. A total of 75 4-week-old male SPF ICR mice were randomly divided into five groups (control, MZD, MZD + ZnY2, MZD + ZnY4, and MZD + ZnY8, 15 mice per group). The dietary Zn content was 30mg/kg in the control group and 10mg/kg in the other groups. From low to high, the Zn supplementation doses administered to the three groups were 2, 4, and 8mg/kg·bw. After 35days, the zinc content, sperm quality, activity of spermatogenic enzymes, oxidative stress level, and apoptosis level of the testes in mice were determined. The results showed that MZD decreased the level of Zn in the serum, sperm quality, and activity of spermatogenic enzymes in mice. After Zn supplementation, the Zn level in the serum increased, sperm quality was significantly improved, and spermatogenic enzyme activity was restored. In addition, MZD reduced the content of antioxidants (copper-zinc superoxide dismutase (Cu-Zn SOD), metallothionein (MT), and glutathione (GSH) and promoted malondialdehyde (MDA) production. The apoptosis index of the testis also increased significantly in the MZD group. After Zn supplementation, the level of oxidative stress decreased, and the apoptosis index in the testis was reduced. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) showed that the expression of B-cell lymphoma-2 (Bcl-2) mRNA and Bcl-2/BCL2-associated X (Bax) in the control group decreased in testicular cells, and their expression was restored after Zn supplementation. The results of this study indicated that Zn supplementation can reduce the level of oxidative stress and increase the ability of testicular cells to resist apoptosis, thereby improving spermatogenic dysfunction caused by MZD in mice.

Mechanism of action of Wuzi Yanzong pill in the treatment of oligoasthenozoospermia in rats determined via serum metabolomics

Diabetes mellitus (DM), a high incidence metabolic disease, is related to the impairment of male spermatogenic function. Spermidine (SPM), one of the biogenic amines, was identified from human seminal plasma and believed to have multiple pharmacological functions. However, there exists little evidence that reported SPM’s effects on moderating diabetic male spermatogenic function. Thus, the objective of this study was to investigate the SPM’s protective effects on testicular spermatogenic function in streptozotocin (STZ)-induced type 1 diabetic mice. Therefore, 40 mature male C57BL/6 J mice were divided into four main groups: the control group (n = 10), the diabetic group (n = 10), the 2.5 mg/kg SPM-treated diabetic group (n = 10) and the 5 mg/kg SPM-treated diabetic group (n = 10), which was given intraperitoneally for 8 weeks. The type 1 diabetic mice model was established by a single intraperitoneal injection of STZ 120 mg/kg. The results showed that, compare to the control group, the body and testis weight, as well the number of sperm were decreased, while the rate of sperm malformation was significantly increased in STZ-induced diabetic mice. Then the testicular morphology was observed, which showed that seminiferous tubule of testis were arranged in mess, the area and diameter of which was decreased, along with downregulated anti-apoptotic factor (Bcl-2) expression, and upregulated pro-apoptotic factor (Bax) expression in the testes. Furthermore, testicular genetic expression levels of Sertoli cells (SCs) markers (WT1, GATA4 and Vimentin) detected that the pathological changes aggravated observably, such as the severity of tubule degeneration increased. Compared to the saline-treated DM mice, SPM treatment markedly improved testicular function, with an increment in the body and testis weight as well as sperm count. Pro-apoptotic factor (Bax) was down-regulated expression with the up-regulated expression of Bcl-2 and suppression of apoptosis in the testes. What’s more, expression of WT1, GATA4, Vimentin and the expressions of glycolytic rate-limiting enzyme genes (HK2, PKM2, LDHA) in diabetic testes were also upregulated by SPM supplement. The evidence derived from this study indicated that the SMP’s positive effect on moderating spermatogenic disorder in T1DM mice’s testis. This positive effect is delivered via promoting spermatogenic cell proliferation and participating in the glycolytic pathway’s activation.