Improvement of Qilin pills on male reproductive function in tripterygium glycoside-induced oligoasthenospermia in rats.

This study established an oligoasthenospermic rat model using tripterygium glycosides (TGs) and investigated the mechanism by which Qilin pills (QLPs) ameliorate reproductive hypofunction. Thirty-two male Sprague Dawley rats were allocated to four equal-sized groups: (1) the control group received continuous physiological levels of saline; (2) the oligoasthenospermia model group was induced with TGs by daily intragastric administration for 28 days; (3 and 4) oligoasthenospermic rats were treated intragastrically with low dose (1.62 g kg-1  d-1 ) and high dose (3.24 g kg-1  d-1 ) of QLPs once daily for 60 days. The QLP-treated rats showed a marked increase (p < .05) in testicular mass, testicular index and semen parameters compared with the untreated rats. Histopathologically, the QLP-treated groups exhibited restored seminiferous tubules in contrast to the model group. Reactive oxygen species and malondialdehyde levels were dramatically decreased (p < .05) in the testes of the QLP-treated rats. QLP treatment partly reverted (p < .05) the circulatory levels of reproductive hormones (FSH, LH, testosterone, prolactin and SHBG) and hepatic and renal function (AST, Cr and urea). Our results showed that oral QLP treatment had a curative effect on the testicular mass, sperm quality, testicular pathomorphology, antioxidants, plasmatic hormones, and liver and renal function of rats.

[Impact of the freezing-thawing process on human sperm mitochondria].

OBJECTIVE: To study the structure and function of human sperm mitochondria before and after the freezing-thawing process. METHODS: Human sperm from healthy donors were subjected to the slow freezing-thawing process, and the sperm mitochondrion-related indexes compared before and after cryopreservation. The ultrastructural changes of the mitochondria were observed under the projection electron microscope, the mitochondrial membrane potential (MMP) and seminal adenosine triphosphate (ATP) content measured by immunofluorescence labeling and ELISA, respectively, and the sperm oxidative stress related indexes detected before and after sperm cryopreservation. RESULTS: Electron microscopy showed loose structures and widened crests of the sperm mitochondria, some with vacuole-like changes after the freezing-thawing process. The sperm after cryopreservation, compared with those before it, exhibited significantly increased contents of oxygen free radicals (ï¼»11.6 ± 3.8ï¼½% vs ï¼»9.6 ± 4.1ï¼½%, P < 0.05) and malondialdehyde (ï¼»3.2 ± 1.4ï¼½ vs ï¼»2.3 ± 1.2ï¼½ nmol/108, P < 0.05), but decreased antioxidant capacity (ï¼»0.6 ± 0.4ï¼½ vs ï¼»0.9 ± 0.4ï¼½ nmol/108, P < 0.05), superoxide dismutase activity (ï¼»0.9 ± 0.4ï¼½ vs ï¼»9.1 ± 3.9ï¼½ nmol/108, P < 0.05), MMP (ï¼»52.2 ± 6.2ï¼½% vs ï¼»55.7 ± 4.9ï¼½%, P = 0.026) and ATP production (ï¼»56.5 ± 9.0ï¼½ vs ï¼»61.3 ± 10.4ï¼½ pmol/106, P = 0.014). CONCLUSIONS: The freezing-thawing process can cause ultrastructural disorder of human sperm mitochondria, reduce their membrane potential and decrease their ATP production.

Sperm Cryopreservation for Male Cancer Patients: More than 10 Years of Experience, in Beijing China.

BACKGROUND Fertility preservation is very important for male cancer patients, especially adolescents. Unfortunately, the use of fertility preservation is very low among Chinese male cancer patients. Additionally, the cumulative rate of frozen sperm use is also low. MATERIAL AND METHODS We performed a retrospective study by collecting available information at the Human Sperm Bank, National Research Institute for Family Planning from July 2006 to December 2017 to examine the data in China. RESULTS A total 145 male cancer patients underwent sperm cryopreservation. The patients were 29.3±6.9 years old, and 6.2% (9 out of 145) of the patients were adolescents under the age of 18 years old. As of June 2018, only 9.7% (14 out of 145) of patients returned to use their cryopreserved sperm for assisted reproduction technology (ART). Of the 33 ART cycles, conceptions were achieved in 51.5% (17 out of 33), and the rate of patients who had a baby was 71.4% (10 out of 14). The data indicate men with testicular cancer or leukemia had lower total sperm counts and recovery rate of progressive sperm than did men with other types of cancer, while men with sarcoma had the lowest progressive sperm. CONCLUSIONS The physician should make an effort to promote fertility preservation for male cancer patients in China. And patients with testicular cancer and leukemia require additional attention.

Quantitative proteomic characterization of human sperm cryopreservation: using data-independent acquisition mass spectrometry.

BACKGROUND: Human sperm cryopreservation is a simple and effective approach for male fertility preservation. METHODS: To identify potential proteomic changes in this process, data-independent acquisition (DIA), a technology with high quantitative accuracy and highly reproducible proteomics, was used to quantitatively characterize the proteomics of human sperm cryopreservation. RESULTS: A total of 174 significantly differential proteins were identified between fresh and cryoperservated sperm: 98 proteins decreased and 76 proteins increased in the cryopreservation group. Bioinformatic analysis revealed that metabolic pathways play an important role in cryopreservation, including: propanoate metabolism, glyoxylate and dicarboxylate metabolism, glycolysis/gluconeogenesis, and pyruvate metabolism. Four different proteins involved in glycolysis were identified by Western blotting: GPI, LDHB, ADH5, and PGAM1. CONCLUSIONS: Our work will provide valuable information for future investigations and pathological studies involving sperm cryopreservation.

Obesity impairs male fertility through long-term effects on spermatogenesis.

OBJECTIVE: This study aimed to investigate the effect and possible underlying mechanisms of high-fat diet-induced obesity on spermatogenesis in male rats. METHODS: A total of 45 male rats were randomly divided into control (n = 15, normal diet) and obesity groups (n = 30, high-fat diet) and were fed for 16 weeks. Body weight and organ indexes were determined after sacrifice. Indicators of reproductive function, including sperm count, sperm motility, apoptosis of spermatogenic cells, and oxidative stress levels, were measured. Serum metabolic parameters and reproductive hormones were also assayed. RESULTS: Compared with the control group, epididymal sperm motility in the obese rats was significantly decreased (P < 0.01). Morphological analysis of the obesity group showed vacuolar changes in seminiferous tubules, spermatogenic cell dysfunction, and increased apoptosis of spermatogenic cells in testicular tissue (P < 0.05). The calculated free testosterone (cFT) concentration in serum was decreased (P < 0.05), whereas the serum sex hormone-binding globulin (SHBG) level was significantly increased (P < 0.01). The superoxide dismutase (SOD) concentration decreased and the malondialdehyde (MDA) concentration increased in testis tissues; however, neither changes were statistically significant (P > 0.05). RESULTS: Nutritional obesity can damage spermatogenesis in male rats due to long-term effects on spermatogenesis.

[Fertility preservation for male adolescent cancer patients].

Fertility preservation is a hotspot of research in reproductive medicine, and that of male adolescent cancer patients is drawing even more attention from reproductive and oncologic clinicians. Both cancer and its treatment can decrease semen quality and even induce irreversible damage to fertility. Sperm cryopreservation is an effective method for fertility preservation. In the past few years, marked advances have been made in the cryopreservation, transplantation, and in vitro culture of testis tissue and stem spermatogonial cells. Although still experimental, these approaches may offer some options to those with no mature sperm in the testis. Unfortunately, very few people know and participate in the studies of fertility preservation and the utilization rate of cryopreserved sperm remains low. Therefor reproductive physicians and oncologists are required to make more efforts to search for effective fertility preservation methods for male adolescent cancer patients.

[ilin Pills for oligoasthenospermia: Advances in clinical studies].

Industrialization and environmental pollution are bringing more problems to human reproduction and increasing the prevalence of male infertility. Western medicine has shown its limitations in the management of male infertility, especially that of oligoasthenospermia. Traditional Chinese medicine (TCM), however, has long and rich experiences in the treatment of oligoasthenospermia, with a large variety of medicinal prescriptions based on the TCM theories, among which Qilin Pills shows a particularly significant therapeutic effect on oligoasthenospermia, especially when combined with Western medicine. At present, published studies on Qilin Pills are mainly in the stage of clinical observation, while basic researches and studies on its relevant mechanisms are rarely seen.

[Protective effect of Qilin Pills on the reproductive function of oligoasthenospermia rats].

OBJECTIVE: To study the protective effect of Qilin Pills (QLP) on the reproductive function of rats with oligoasthenospermia (OAS) induced by tripterygium glycosides. METHODS: Twenty-eight male SD rats were randomly divided into a normal control, an OAS model control, a low-dose QLP, and a high-dose QLP group of equal number. OAS models were made in the latter three groups by intragastrical administration of tripterygium glycosides at 40 mg per kg of the body weight per day, and meanwhile the animals in the low- and high-dose QLP groups were treated with QLP at 1.62 and 3.24 g per kg of the body weight per day, respectively, while those in the OAS model group with normal saline, all for 30 consecutive days. Then all the rats were executed for obtaining the testis weight, testis viscera index, epididymal sperm concentration and motility, reproductive hormone levels, and antioxidation indexes and observation of the histomorphological changes of the testis tissue by HE staining. RESULTS: After 30 days of intervention, the low- and high-dose QLP groups, as compared with the OAS model controls, showed significantly improved epididymal sperm concentration (ï¼»14.57 ± 3.95ï¼½ and ï¼»39.71 ± 11.31ï¼½ vs ï¼»4.71 ± 1.25ï¼½ ×106/ml, P <0.05) and motility (ï¼»3.71 ± 1.11ï¼½ and ï¼»4.29 ± 1.80ï¼½ vs ï¼»0.57 ± 0.53ï¼½%, P <0.05), increased levels of sex hormone binding globulin (SHBG) (ï¼»94.83 ± 11.17ï¼½ and ï¼»88.05 ± 9.21ï¼½ vs ï¼»56.74 ± 8.29ï¼½ nmol/L, P <0.05) and free testosterone (FT) (ï¼»27.27 ± 3.63ï¼½ and ï¼»32.80 ± 2.51ï¼½ vs ï¼»22.81 ± 2.75ï¼½ nmol/L, P <0.05), decreased level of follicle-stimulating hormone (FSH) (ï¼»1.49 ± 0.62ï¼½ and ï¼»1.12 ± 0.83ï¼½ vs ï¼»1.71 ± 0.52ï¼½ mIU/ml, P <0.05), but no significant change in the total testosterone (TT) level. Meanwhile, the level of superoxide dismutase (SOD) was markedly elevated in the low- and high-dose QLP groups in comparison with the OAS model control group (ï¼»277.14 ± 15.84ï¼½ and ï¼»299.60 ± 20.83ï¼½ vs ï¼»250.04 ± 31.06ï¼½ U/ml, P <0.05) while that of reactive oxygen species (ROS) remarkably reduced (ï¼»397.61 ± 62.71ï¼½ and ï¼»376.84 ± 67.14ï¼½ vs ï¼»552.20 ± 58.07ï¼½ IU/ml, P <0.05). HE staining showed that QLP intervention significantly increased the layers and quantity of spermatogenic cells in the testicular seminiferous tubules of the OAS rats. CONCLUSIONS: QLP can effectively protect the reproductive system of oligoasthenospermia rats by raising sperm quality, elevating reproductive hormone levels, reducing oxidative stress injury, and improving histomorphology of the testis.

Lycopene inhibits apoptosis of mouse spermatocytes in varicocele via miR-23a/b-induced downregulation of PROK2.

Context The varicocele is the leading cause of male infertility and can impair sperm quality and testicular function through various mechanisms. In our previous study, we found that lycopene could attenuate hypoxia-induced testicular injury. Aims To illustrate the detailed mechanism of lycopene on spermatocytes. Methods The effect of lycopene on GC-2 cells under hypoxia were detected by flow cytometry and western blot assay. miR-seq was used to determine miRNA expression in varicocele rat model testes. The function of miR-23a/b were determined by flow cytometry and western blot assay. Key results We demonstrate that lycopene could alleviate hypoxia-induced GC-2 cell apoptosis and could elevate miR-23a/b expression of the hypoxia model in vivo and in vitro . The miR-23a and -23b mimics could reduce the hypoxia-induced GC-2 cell apoptosis. Both miR-23a and -23b could directly bind with prokineticin 2 (PROK2) mRNA and downregulate its expression. Conclusions Lycopene could attenuate hypoxia-induced spermatocyte injury through the miR-23a/b-PROK2 pathway. Implications Lycopene may be an effective treatment for varicocele to improve testicular impairment.

Human Seminal Extracellular Vesicles Enhance Endometrial Receptivity Through Leukemia Inhibitory Factor.

Seminal extracellular vesicles (EVs) contain different subgroups that have diverse effects on sperm function. However, the effect of seminal EVs-especially their subgroups-on endometrial receptivity is largely unknown. Here, we found that seminal EVs could be divided into high-density EVs (EV-H), medium density EVs, and low-density EVs after purification using iodixanol. We demonstrated that EV-H could promote the expression and secretion of leukemia inhibitor factor (LIF) in human endometrial cells. In EV-H-treated endometrial cells, we identified 1274 differentially expressed genes (DEGs). DEGs were enriched in cell adhesion and AKT and STAT3 pathways. Therefore, we illustrated that EV-H enhanced the adhesion of human choriocarcinoma JAr cell spheroids to endometrial cells through the LIF-STAT3 pathway. Collectively, our findings indicated that seminal EV-H could regulate endometrial receptivity through the LIF pathway, which could provide novel insights into male fertility.

The impact of exogenous testosterone supplementation on spermatogenesis in a rat model of oligoasthenospermia.

Oligoasthenospermia is one of the main causes of infertility in reproductive-age men. This study aimed to explore the feasibility of exogenous testosterone supplemental therapy (TST) for adult male rats with oligoasthenospermia model. The rats (n=40) were randomized equally into 4 groups: control group, model group, low-dose and high-dose groups (n=10, respectively). After establishment of an oligoasthenospermia model that was treated with glucosides of tripterygium wilfordii (GTWs), the low-dose and high-dose groups were treated with 2 testosterone undecanoate (TU) injections at doses of 7.5 mg and 15 mg for 8-week period (4-week intervals). Body weights, serum reproductive hormone levels, sperm measurements in the epididymis, and testis histology were monitored. The TU injections increased serum testosterone levels steadily. The epididymis sperm concentration and motility increased slowly in high dose group at 4-weeks whereas sperm measurements increased significantly in the TST groups at 8 weeks. In addition, exogenous TST increased the intra-testicular testosterone concentration somewhat and alleviated the testicular oxidative stress markers of Malondialdehyde (MDA) and level of GSH-PX (Glutathione Peroxidase) after 8 weeks treatment. The improvement of sperm and testicular function acted mainly by curbing mitochondrial apoptosis in the testis by modulation of Bcl-2, Bax, Caspase-3, and Caspase-9 expression. However, the results of immunohistochemistry and western blotting in the low-dose group were still lower than control values. TST at an appropriate dose within a period of 8 weeks was effective to stimulate spermatogenesis and alleviate inflammation, oxidative stress, and apoptosis through suppression of testis damage in this rat model of oligoasthenospermia.

Effects of Qilin pills on spermatogenesis, reproductive hormones, oxidative stress, and the TSSK2 gene in a rat model of oligoasthenospermia.

BACKGROUND: Qilin pills (QLPs), a classic Traditional Chinese Medicine (TCM) formula for treating male infertility, effectively improve semen quality in clinical trials. This study was designed to evaluate the effects of QLPs on spermatogenesis, reproductive hormones, oxidative stress, and the testis-specific serinekinase-2 (TSSK2) gene in a rat model of oligoasthenospermia. METHODS: Forty adult male Sprague-Dawley (SD) rats were randomly divided into four groups. The rat model with oligoasthenospermia was generated by intragastric administration of tripterygium glycosides (TGs) once daily for 4 weeks. Then, two treatment groups were given different doses (1.62 g/kg and 3.24 g/kg) of QLPs once daily for 60 days. Sperm parameters, testicular histology and reproductive hormone measurements, oxidative stress tests, and TSSK2 expression tests were carried out. RESULTS: QLPs effectively improved semen parameters and testicular histology; restored the levels of FSH, LH, PRL, fT, and SHBG; reduced the levels of oxidative stress products (ROS and MDA); increased testicular SOD activity; and restored the expression of spermatogenesis-related gene TSSK2. CONCLUSION: QLPs have a therapeutic effect on a rat model of oligoasthenospermia, and this effect is manifested as improvement of semen quality and testis histology, gonadal axis stability, decreased oxidative stress, and the regulation of testis-specific spermatogenesis-related gene TSSK2.

Qilin pills alleviate oligoasthenospermia by inhibiting Bax-caspase-9 apoptosis pathway in the testes of model rats.

At present, the treatment of oligoasthenospermia with western medicine is ineffective. Qilin pill (QLP) is a Chinese traditional medicine for treating male infertility. Recent multicenter clinical studies in China reported that QLPs markedly improved sperm quality. However, the mechanism of action of QLPs on oligoasthenospermia remains unknown. In this study, we investigated the mechanistic basis for improvement of semen parameters and reversal of testis damage by QLPs in a rat model of oligoasthenospermia induced by treatment with tripterygium glycosides (TGs) (40 mg/kg) once daily for 4 weeks. Rats were administered QLPs (1.62 g/kg or 3.24 g/kg) each day for 60 days, with untreated animals serving as controls. The concentration and motility of sperm extracted from rat epididymis were determined, whereas histopathological examination and immunohistochemical apoptosis analysis of rat testes was performed. Expression profiles of apoptosis-related genes were determined by microarray analysis; the results were validated by quantitative real-time PCR, western blotting, and immunohistochemistry. Sperm concentration and motility in the QLP treatment group were increased relative to those in control rats. Testis tissue and DNA damage were reversed by QLP treatment. The improvement function of QLPs on sperm and testis works mainly by suppressing mitochondrial apoptosis in the testis via modulation of B cell lymphoma (Bcl)-2, Bcl-2-associated X protein (Bax), cytochrome C, caspase-9 and caspase-3 expression. QLPs could improve sperm quality and testis damage in a rat model of oligoasthenospermia by inhibiting the Bax-Caspase-9 apoptosis pathway and exerting therapeutic effects.