Enhancement of Frozen-Thawed Human Sperm Quality with Zinc as a Cryoprotective Additive.

BACKGROUND Cryopreservation preserves male fertility, crucial in oncology, advanced age, and infertility. However, it damages sperm motility, membrane, and DNA. Zinc (Zn), an antioxidant, shows promise in improving sperm quality after thawing, highlighting its potential as a cryoprotectant in reproductive medicine. MATERIAL AND METHODS Gradient concentration of ZnSO4 (0, 12.5, 25, 50, and 100 µM) was added in the Glycerol-egg yolk-citrate (GEYC) cryopreservative medium as an extender. Alterations in sperm viability and motility parameters after cryopreservation were detected in each group. Sperm plasma membrane integrity (PMI), acrosome integrity (ACR), DNA fragment index (DFI), and changes in sperm mitochondrial function were examined, including: mitochondrial potential (MMP), sperm reactive oxygen species (ROS), and sperm ATP. RESULTS We found that 50 µM ZnSO4 was the most effective for the curvilinear velocity (VCL) and the average path velocity (VAP) of sperm after cryo-resuscitation. Compared to the Zn-free group, sperm plasma membrane integrity (PMI) was increased, DNA fragmentation index (DFI) was decreased, reactive oxygen species (ROS) was reduced, and mitochondrial membrane potential (MMP) was increased after cryorevival in the presence of 50 µM ZnSO4. CONCLUSIONS Zn ion is one of the antioxidants in the cell. The results of our current clinical study are sufficient to demonstrate that Zn can improve preserves sperm quality during cryopreservation when added to GEYC. The addition of 50 µM ZnSO4 increased curve velocity, mean path velocity, sperm survival (or plasma membrane integrity), and mitochondrial membrane potential while reducing ROS production and DNA breaks compared to GEYC thawed without ZnSO4.

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.

Liver transcriptome analysis and cortisol immune-response modulation in lipopolysaccharide-stimulated in channel catfish (Ictalurus punctatus).

Channel catfish (Ictalurus punctatus) is an important aquaculture species in China. In channel catfish, diseases such as haemorrhagic, sepsis and tail-rot disease are all caused by bacteria as general in China. Most of the pathogenic bacteria are Gram-negative bacteria. Liver transcriptome analysis of the co-injection of cortisol and lipopolysaccharide (LPS) was performed in this study. Preliminary evidence from the results suggest that after the emergence of immune stress, cortisol will up-regulate the complement cascade pathway, down-regulate the coagulation cascade pathway, down-regulate the platelet activation pathway, down-regulate antigen presentation pathway, and show complex regulation relationship to inflammatory factors. At 12 h, the number of differential genes regulated by cortisol was about half less than the number of differential genes regulated by LPS. At 24 h, there was no significant difference between the number of differential genes regulated by cortisol and LPS, but the types of differential genes vary widely. KEGG enrichment analysis found that cortisol regulated LPS-stimulated immune responses mainly focus on cytokines, complement and coagulation cascades pathways, antigen presentation pathways, haematopoiesis, and inflammation. It is suggested that there may be some strategic choice in the regulation of immune response by cortisol. These results will help understand the pathogenesis and host defence system in bacterial disease caused by Gram-negative bacteria.

[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.

[Status quo of autologous sperm preservation in the human sperm bank in Beijing].

OBJECTIVE: To comprehensively analyze the status quo of autologous sperm preservation in the human sperm bank in Beijing and better utilize the existing resources for the preservation of male fertility. METHODS: We retrospectively analyzed the geographical data and semen quality of 251 males with autologous sperm preservation in the Human Sperm Bank, Science and Technology Research Institute, National Health and Family Planning Commission of China from July 2006 to December 2016. RESULTS: The rate of autologous sperm preservation in the Human Sperm Bank was as low as 8.76% between July 2006 and December 2010 but increased annually by 119% on average from 2011 to 2013. Of the 251 males involved, 204 (81.27%) were aged 20－39 years, 175 (69.72%) had bachelor's or master's degree, 223 (88.84%) had no child, 69 (27.49%) got less than 10 tubes of semen samples frozen, and 26 (10.36%) had their semen samples cryopreserved only once. The utilization rate of the cryopreserved sperm was only 5.58 % (n = 14). The main reason for autologous sperm preservation was carcinoma (55.78% ï¼»n = 140ï¼½), including blood cancer (22.31% ï¼»n = 66ï¼½), testicular cancer (13.15% ï¼»n = 33ï¼½) and other cancers (16.33% ï¼»n = 41ï¼½). Compared with the non-cancer males, the cancer patients had a significantly reduced mean sperm concentration (90.45 vs 60.53 ×106/ml, P < 0.05), total sperm count (311.3 vs 175.8 ×106, P < 0.05), percentage of progressively motile sperm (PMS) (49.21% vs 43.55%, P < 0.05) and recovery rate of PMS (68.13% vs 52.17%, P < 0.05). In the subgroups of testicular, blood and other cancers, the sperm concentration averaged 37.68, 57.98 and 90.69 ×106/ml, the semen volume 2.73, 2.82 and 3.41 ml, the total sperm count 93.29, 158.41 and 349.49 ×106, the percentage of PMS 45.32%, 43.47% and 44.49%, and the recovery rate of PMS 48.32%, 50.07% and 61.09%, respectively, the sperm concentration and total sperm count significantly lower in the testicular cancer patients than in the other two groups (P < 0.05). CONCLUSIONS: The number of the cases of autologous sperm preservation in Beijing is increasing year by year, and the majority of them are cancer patients. As most of the cancer patients have missed the best period for sperm preservation, sperm bank workers should endeavor to increase the public awareness of autologous sperm preservation.

[L-carnitine protects the motion parameters and mitochondrial function of human sperm in cryopreservation].

OBJECTIVE: To study the effects of L-carnitine (LC) on cryopreserved human sperm. METHODS: Ten semen samples were collected from normal sperm donors, each divided into six groups, fresh ejaculate (FE), non-LC cryopreservation (non-LC), and cryopreservation with LC at 1 mmol/L (LC-1), 2.5 mmol/L (LC-2), 5 mmol/L (LC-3) and 10 mmol/L (LC-4), respectively. The optimal concentration of LC was identified based on the motility and motion parameters of the post-thaw sperm. The plasma membrane integrity (PMI) of the sperm was assessed by eosin-nigrosin staining, their mitochondrial membrane potential (MMP) monitored by JC-1 assay, and the level of sperm ROS measured by the fluorescent probe DCFH-DA, followed by analysis of the mechanisms of LC protecting sperm against cryopreservation injury. RESULTS: Compared with the sperm in the FE group, the post-thaw sperm in the non-LC and LC groups showed significantly decreased progressive motility, average path velocity (VAP), straight line velocity (VSP) and curvilinear velocity (VCP) (P < 0.05). In comparison with the non-LC group, the LC-3 group exhibited a remarkably higher percentage of progressively motile sperm (ï¼»41.9 ± 4.6ï¼½ vs ï¼»47.0 ± 4.3ï¼½%, P = 0.0261) and VAP (ï¼»34.9 ± 2.6ï¼½ vs ï¼»38.9 ± 4.2ï¼½ µm/s, P = 0.0152), indicating that the optimal concentration of LC was 5 mmol/L. Both PMI and MMP were significantly lower in the non-LC than in the FE group (ï¼»52.7 ± 5.7ï¼½ vs ï¼»75.5 ± 5.4ï¼½%, P < 0.01 and ï¼»44.5 ± 3.5ï¼½ vs ï¼»57.3 ± 4.4ï¼½%, P < 0.01), but higher in the LC groups (ï¼»70.1 ± 8.2ï¼½% and ï¼»50.3 ± 3.4ï¼½%) than in the non-LC group (P < 0.01 and P < 0.05). The level of sperm ROS, however, was markedly higher in the non-LC than in the FE group (ï¼»12.5 ± 3.9ï¼½ vs ï¼»6.8 ± 2.4ï¼½, P < 0.01) but lower in the LC groups (ï¼»8.4 ± 5.3ï¼½%) than in the non-LC group (P = 0.05). CONCLUSIONS: L-carnitine can improve the motility and motion parameters of cryopreserved human sperm by reducing sperm ROS, enhancing sperm mitochondrial membrane potential and protecting the sperm plasma membrane.

[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.

Seminal plasma microbiomes, sperm parameters, and cryopreservation in a healthy fertile population.

Background: Recent advances in microbiome research have revealed the presence of diverse microbial communities in human tissues previously thought to be sterile. The present study delves into the emerging field of seminal plasma microbiomics, examining the relationship between semen microbes and semen parameters and post-freezing tolerance. Methods: The study involved a cohort of healthy fertility males and microbial genome analysis using 16S rRNA to characterize the microbial diversity of seminal plasma. Microbial diversity analysis identified unique amplicon sequence variants (ASVs) and genera dominant in seminal plasma. Spearman's correlation coefficient was used to assess the relationship between flora and semen parameters. A paired t-test was used to compare the changes in microbiome expression in seminal plasma before and after cryo-resuscitation. Results: The relevant results show that the top five phyla in terms of abundance of seminal plasma microbiome were Firmicutes, Bacteroidota, Proteobacteria, Actinobacteriota, and Campylobacterota. Spearman correlation analysis highlighted the association between specific microbial species and semen parameters, between Porphyromonas_asaccharolytica and sperm concentration. Microbial changed significantly after cryo-resuscitation, affecting taxonomic units such as Campylobacter and Muribaculaceae, and KEGG enrichment analyses, suggesting that metabolic pathways are associated with sperm freezing. Eubacterium_coprostanoligenes and Eptoniphilus_duerdenii exhibited a potential impact, while Orynebacterium_tuberculostearicum demonstrated a positive correlation with the recovery rate of progressive motile sperm. Conclusion: The semen of normal fertile individuals contains a microflora component that is closely related to semen quality, including the sperm's ability to withstand freezing.

Combined Analysis of the Transcriptome, Proteome and Metabolome in Human Cryopreserved Sperm.

PURPOSE: This study aimed to identify the altered pathways and genes associated with freezing damage in human sperm during cryopreservation by multiomics analysis. MATERIALS AND METHODS: Fifteen fresh human semen samples were collected for transcriptomic analysis, and another 5 fresh human semen samples were obtained for metabolomic analysis. For each semen sample, 1 mL was cryopreserved, and another 1 mL was left untreated for paired design. The results were then combined with previously published proteomic results to identify key genes/pathways. RESULTS: Cryopreservation significantly reduced sperm motility and mitochondrial structure. Transcriptomic analysis revealed altered mitochondrial function, including changes in tRNA-methyltransferase activity and adenosine tri-phosphate/adenosine di-phosphate transmembrane transporter activity. Metabolomic analysis showed that the citrate cycle in mitochondria was significantly altered. Combining transcriptomic, proteomic, and metabolomic analyses revealed 346 genes that were altered in at least two omics analyses. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that metabolic pathways were significantly altered and strongly associated with mitochondria. Five genes were altered in all three omics analyses: COL11A1, COL18A1, LPCAT3, NME1, and NNT. CONCLUSIONS: Five genes were identified by multiomics analysis in human cryopreserved sperm. These genes might have specific functions in cryopreservation. Explorations of the functions of these genes will be helpful for sperm cryopreservation and sperm motility improvement or even for reproduction in the future.

Novel biallelic variants in DNAH1 cause multiple morphological abnormalities of sperm flagella with favorable outcomes of fertility after ICSI in Han Chinese males.

BACKGROUND: Multiple morphological abnormalities of sperm flagella is an idiopathic asthenoteratozoospermia characterized by absent, short, coiled, angulation, and irregular-caliber flagella. Genetic variants of DNAH1 gene have been identified as a causative factor of multiple morphological abnormalities of sperm flagella and intracytoplasmic sperm injection is an available strategy for infertile males with dynein axonemal heavy chain 1 defects to conceive. OBJECTIVES: To identify novel variants and candidate mutant hotspots of DNAH1 gene related to multiple morphological abnormalities of sperm flagella and male infertility in humans. MATERIALS AND METHODS: The DNAH1 variants were identified by whole exome sequencing and confirmed with Sanger sequencing. Papanicolaou staining, scanning and transmission electron microscopy, and immunostaining were performed to investigate the morphological and ultrastructural characteristics of spermatozoa. Intracytoplasmic sperm injection was applied for the assisted reproductive therapy of males harboring biallelic DNAH1 variants. RESULTS: We identified 18 different DNAH1 variants in 11 unrelated families, including nine missense variants (p.A2564T, p.T3657R, p.G1862R, p.L2296P, p.T4041I, p.L611P, p.A913D, p.R1932Q, p.R2356W) and nine loss-of-function variants (c.2301-1G>T, p.Q1518*, p.R1702*, p.D2845Mfs*2, p.P3909Rfs*33, p.Q4040Dfs*33, p.Q4058*, p.E4060Pfs*61, p.V4071Cfs*54). A total of 66.7% (12/18) of the identified variants were novel. Morphological analysis based on Papanicolaou staining and scanning electron microscopy demonstrated the typical multiple morphological abnormalities of sperm flagella characteristics of dynein axonemal heavy chain 1-deficient spermatozoa. Immunostaining further revealed the absence of inner dynein arms but not outer dynein arms, which induced a general ultrastructural disorganization, such as the loss of central pair and mis-localization of the microtubule doublets and outer dense fibers. To date, seven affected couples have accepted the intracytoplasmic sperm injection treatment, and three of them have given birth to five healthy babies. DISCUSSION AND CONCLUSION: These findings further expand the variant spectrum of DNAH1 gene related to multiple morphological abnormalities of sperm flagella and male infertility in humans, thus providing new information for the molecular diagnosis of asthenoteratozoospermia. The favorable fertility outcomes of intracytoplasmic sperm injection will facilitate the genetic counseling and clinical treatment of infertile males with multiple morphological abnormalities of sperm flagella in the future.

Insufficiency of Mrpl40 disrupts testicular structure and semen parameters in a murine model.

Approximately 31% of patients with 22q11.2 deletion syndrome (22q11.2DS) have genitourinary system disorders and 6% of them have undescended testes. Haploinsufficiency of genes on chromosome 22q11.2 might contribute to the risk of 22q11.2DS. In this study, we used mice with single-allele deletion in mitochondrial ribosomal protein L40 (Mrpl40 +/- ) as models to investigate the function of Mrpl40 in testes and spermatozoa development. The penetrance of cryptorchidism in Mrpl40 +/- mice was found to be higher than that in wild-type (WT) counterparts. Although the weight of testes was not significantly different between the WT and Mrpl40 +/- mice, the structure of seminiferous tubules and mitochondrial morphology was altered in the Mrpl40 +/- mice. Moreover, the concentration and motility of spermatozoa were significantly decreased in the Mrpl40 +/- mice. In addition, data-independent acquisition mass spectrometry indicated that the expression of genes associated with male infertility was altered in Mrpl40 +/- testes. Our study demonstrated the important role of Mrpl40 in testicular structure and spermatozoa motility and count. These findings suggest that Mrpl40 is potentially a novel therapeutic target for cryptorchidism and decreased motility and count of spermatozoa.

Determination of Parabens and Their Metabolites in Seminal Plasma from Chinese Men by Ultra High Performance Liquid Chromatography Tandem Mass Spectrometry (UPLC-MS/MS).

Parabens are endocrine-disrupting chemicals (EDCs) that have estrogen-like activities and may cause male reproductive disorders. Here, we developed a method for the simultaneous determination of four parabens (MeP, EtP, n-PrP, n-BuP) and two metabolites (4-HB and 3,4-DHB) in human seminal plasma by UPLC-MS/MS. The method was used to analyze 144 seminal plasma samples from Chinese males. MeP, EtP, n-PrP, and 4-HB were the dominant compounds. MeP, EtP, and n-PrP were significantly correlated to each other. In addition, 4-HB was significantly correlated to MeP, EtP, n-PrP, and 3,4-DHB, respectively. The results provide direct evidence that parabens and their metabolites are widely distributed in the male reproductive system. The study presents the paraben metabolites levels in human seminal plasma for the first time.

Reproductive performance and semen characteristics of pond-reared and wild-caught large-sized male broodstock of the Chinese mitten crab Eriocheir sinensis.

There has been a recent emphasis on production of large-sized Eriocheir sinensis broodstock. In China, aquaculturists generally prefer wild-caught (WC) crabs from the Yangtze River as broodstock because offspring performance is superior to that of pond-reared (PR) broodstock. Currently, however, there is a ban on fishing in the Yangtze River, and effects on E. sinensis breeding have not been ascertained. There was comparison in the present study of reproductive performance and semen characteristics of male broodstock of PR and WC groups. After copulation, sperm quantity in the vas deferens of crabs in specimens of both groups was large, although there was a consistent decrease in vaso-somatic index. Although sperm density of PR crabs was less, that of WC specimens remained relatively constant. Specimens of neither group, however, had changes in the hepatopancreas index or condition factor, and sperm survival was close to 100%. Although the acrosome reaction was detected in response to cold-temperature induction, there were differences in extent of reaction to cold temperatures. Importantly, in as many as 98% of sperm from female spermathecae, the reaction was completed, which was considerably greater than 15% for sperm of males post-mating. It is concluded there was no difference between PR and WC crabs with respect to reproductive performance or semen characteristics, and, notably, sperm from PR crabs were of sufficient quality for use in E. sinensis aquaculture enterprises. Accordingly, it is predicted the Yangtze River fishing ban would only have a limited effect on supply of male E. sinensis broodstock.

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.

Glycolysis metabolic changes in sperm cryopreservation based on a targeted metabolomic strategy.

Our aim was to identify potential metabolomic pathway changes in the sperm cryopreservation process and to find new markers of human sperm freezability. Targeted metabolomic experiments were used to identify the quantitative metabolomic compound characterization of human sperm cryopreservation. A KEGG pathway analysis was used for these deregulated compounds. A total of 16 significantly deregulated compounds was identified between fresh and post-thawed sperm; of these, 7 were downregulated and 9 were upregulated in the frozen-thawed group. A bioinformatics analysis revealed that metabolic pathways play an important role in cryopreservation, including the citrate cycle (TCA cycle), glycolysis or gluconeogenesis, glyoxylate and dicarboxylate metabolism, pyruvate metabolism and galactose metabolism. We used immunoblotting and immunofluorescence to analyze the expression and localization of the three key enzymes in glycolysis. The glycolytic metabolic changes were noted in sperm cryopreservation. HK2 expression levels in fresh sperm were significantly higher than the levels in freeze-thawed sperm.