Shifu Hu

Effect of transient scrotal hyperthermia on sperm parameters, seminal plasma biochemical markers, and oxidative stress in men

In this experimental prospective study, we aimed to analyze the effect of transient scrotal hyperthermia on the male reproductive organs, from the perspective of sperm parameters, semen plasma biochemical markers, and oxidative stress, to evaluate whether different frequencies of heat exposure cause different degrees of damage to spermatogenesis. Two groups of volunteers (10 per group) received testicular warming in a 43°C water bath 10 times, for 30 min each time: group 1: 10 consecutive days; group 2: once every 3 days. Sperm parameters, epididymis and accessory sex gland function, semen plasma oxidative stress and serum sex hormones were tested before treatment and in the 16-week recovery period after treatment. At last, we found an obvious reversible decrease in sperm concentration (P = 0.005 for Group 1 and P= 0.008 for Group 2 when the minimums were compared with baseline levels, the same below), motility (P = 0.009 and 0.021, respectively), the hypoosmotic swelling test score (P = 0.007 and 0.008, respectively), total acrosin activity (P = 0.018 and 0.009, respectively), and an increase in the seminal plasma malondialdehyde concentration (P = 0.005 and 0.017, respectively). The decrease of sperm concentration was greater for Group 2 than for Group 1 (P = 0.031). We concluded that transient scrotal hyperthermia seriously, but reversibly, negatively affected the spermatogenesis, oxidative stress may be involved in this process. In addition, intermittent heat exposure more seriously suppresses the spermatogenesis compared to consecutive heat exposure. This may be indicative for clinical infertility etiology analysis and the design of contraceptive methods based on heat stress.

Transient scrotal hyperthermia affects human sperm DNA integrity, sperm apoptosis, and sperm protein expression

This prospective randomized clinical study is aimed to evidence the reproductive impairment of frequent scrotal heat exposure. A total of 20 normozoospermic subjects were randomly divided into two groups to undergo testicular warming in a 43 °C water bath 10 times, for 30 min each time; the subjects in group 1 underwent testicular warming for 10 consecutive days and those in group 2 once every 3 days. Sperm chromatin structure assay (SCSA), sperm mitochondrial membrane potential (MMP), apoptosis, and seminal plasma‐soluble Fas (sFas) were analyzed before treatment and every 2 weeks after, for a total of 10 times. In group 1, some critical proteins involved in heat stress, hypoxia, structure, and function of sperm mitochondria and flagella were evaluated before hyperthermia and 2, 6, 10, and 16 weeks after hyperthermia. Both groups showed a reversible increase in the proportion of spermatozoa with a disrupted MMP (both p < 0.05 when the minimums were compared with baseline levels, the same below), sperm apoptosis (both p < 0.01) and high DNA stainability (both p < 0.05). The sFas concentration in both groups showed no obvious changes except one: the value at week 2 was significantly increased over baseline in group 1 (p = 0.036). The level of Bcl‐2 decreased significantly at weeks 6 and 10 (p = 0.017 and 0.05, respectively) and recovered to baseline at week 16. Proteins involved in heat stress and mitochondria functions were up‐regulated, whereas in flagella structure and function was down‐regulated (all p < 0.05). This study demonstrated that transient and frequent scrotal hyperthermia severely and reversibly damaged spermatogenesis, consecutive heat exposure had more serious effects than intermittent exposure, whereas intermittent exposure led to a later recovery of sperm damage.

Copper nanoparticle-induced ovarian injury, follicular atresia, apoptosis, and gene expression alterations in female rats

Numerous studies have reported the accumulation of copper nanoparticles (Cu NPs) in organs and the corresponding damage, although whether Cu NPs can be translocated to the ovaries and their ovarian toxicity are still unknown. In this study, three groups of female rats were injected with 3.12, 6.25, or 12.5 mg/kg Cu NPs for 14 consecutive days. The pathological changes, hormone levels, apoptosis and apoptotic proteins, oxidative stress, and gene expression characteristics in the ovaries were then investigated. The results demonstrated that the Cu NPs exhibited obvious accumulation in the rat ovaries, leading to ovarian injury, an imbalance of sex hormones, and ovarian cell apoptosis. Cu NP exposure activated caspase 3, caspase 8, caspase 9, and tBid, decreased the protein levels of Bcl-2, increased the expression levels of the proteins Bax and cytochrome c, and promoted malondialdehyde (MDA) accumulation and superoxide dismutase (SOD) reduction. Furthermore, gene microarray analysis showed that Cu NPs (12.5 mg/kg/d) caused 321 differentially expressed genes. Of these, 180 and 141 genes were upregulated and downregulated, respectively. Hsd17b1, Hsd3b1, Hsd3b6, and Hsd3b were involved in steroid and hormone metabolism, whereas Mt3 and Cebpb were associated with apoptosis. Overall, these findings provide strong evidence that Cu NPs trigger both intrinsic and extrinsic apoptotic pathways and regulate key ovarian genes in oxidative stress-mediated ovarian dysfunction.

Alterations in the endometrium of rats, rabbits, and Macaca mulatta that received an implantation of copper/low-density polyethylene nanocomposite.

A copper/low-density polyethylene nanocomposite (nano-Cu/LDPE), a potential intrauterine device component material, has been developed from our research. A logical extension of our previous work, this study was conducted to investigate the expression of plasminogen activator inhibitor 1 (PAI-1), substance P (SP), and substance P receptor (SP-R) in the endometrium of Sprague Dawley rats, New Zealand White rabbits, and Macaca mulatta implanted with nano-Cu/LDPE composite. The influence of the nano-Cu/LDPE composite on the morphology of the endometrium was also investigated. Animals were randomly divided into five groups: the sham-operated control group (SO group), bulk copper group (Cu group), LDPE group, and nano-Cu/LDPE groups I and II. An expression of PAI-1, SP, and SP-R in the endometrial tissues was examined by immunohistochemistry at day 30, 60, 90, and 180 postimplantation. The significant difference for PAI-1, SP, and SP-R between the nano-Cu/LDPE groups and the SO group (P<0.05) was identified when the observation period was terminated, and the changes of nano-Cu/LDPE on these parameters were less remarkable than those of the Cu group (P<0.05). The damage to the endometrial morphology caused by the nano-Cu/LDPE composite was much less than that caused by bulk copper. The nano-Cu/LDPE composite might be a potential substitute for conventional materials for intrauterine devices in the future because of its decreased adverse effects on the endometrial microenvironment.

Expression patterns of p38αMAPK during follicular development in the ovaries of neonatal rats

The p38αMAPK signaling pathway plays a critical role in female reproduction, but an understanding of its expression in rats remains elusive. This study was carried out to investigate the temporal and spatial expression of p38αMAPK and p-p38αMAPK. Ovarian tissue samples were collected from 2-, 4-, 8-, 12-, 16-, 20- and 30-day-old female rats. Western blotting was used to examine the relative expression of p38αMAPK and p-p38αMAPK in ovarian tissue, and subcellular localization was examined using immunohistochemistry of the rat ovaries at different ages of postpartum. The immunohistochemical results showed that p38αMAPK and p-p38αMAPK were widely expressed in the rat ovaries, mainly localized in the follicle cells and granulosa cells. The expression of p38αMAPK was relatively stable for the different stages of oocytes, whereas the expression of p-p38αMAPK gradually increased. At different stages of granulosa cells, the expression of p38αMAPK was also relatively stable, and the p-p38αMAPK expression showed an upward trend during follicular development. Western blotting revealed that the expression of p38αMAPK in the ovaries was relatively stable, where as p-p38αMAPK expression initially exhibited an increasing trend and subsequently decreased, with a maximum at day 20. The expression patterns of p38αMAPK and p-p38αMAPK in the rat ovaries indicate their possible involvement in folliculogenesis. Taken together, the stage- and cell-specific expression of p-p38αMAPK in rat ovaries indicated that p-p38αMAPK might play a vital role during rat follicular development.

Antifertility effectiveness of a novel copper-containing intrauterine device material and its influence on the endometrial environment in rats

This study was designed to investigate the antifertility effectiveness of a novel copper-containing intrauterine device material containing a composite of micro-copper (Cu), low-density polyethylene (LDPE), and methyl vinyl silicone rubber (MVQ) and its effects on the endometrial environment in rats. The contraceptive effectiveness was examined 12 days after pregnancy. The pathological changes; factors associated with bleeding, pain, and inflammation in the endometrium; and the surface condition of the implants were investigated after insertion for 90 days. Furthermore, the release rate of copper ions in simulated uterine solution (SUS) was investigated for 270 days. The contraceptive effectiveness was 100% in both the bulk Cu and micro-Cu/LDPE/MVQ groups, and that in the LDPE/MVQ group was 30%. On day 90 after insertion, histopathological observation and the ultrastructural changes in the endometrium showed that the damage caused by bulk Cu was much more severe than that caused by the Cu/LDPE/MVQ microcomposite and that the surface of the latter was much smoother than that of the former. Furthermore, compared with the sham-operated control group, the concentrations of tissue plasminogen activator and prostaglandin E2 were significantly increased 90 days after insertion in all of the experimental groups except for the LDPE/MVQ group (P < 0.05), and the parameters in the Cu/LDPE/MVQ group were significantly lower than those in the Cu group (P < 0.05). In addition, the expression levels of matrix metalloproteinase 9, metalloproteinase 1 tissue inhibitor, plasminogen inhibitor 1, CD34, vascular endothelial growth factor, substance P, and substance P receptor in the endometrium in all of the experimental groups were significantly lower than those in the Cu group 90 days after insertion (P < 0.05). The results of this study indicate that micro-Cu/LDPE/MVQ exhibits satisfactory contraceptive efficacy and causes fewer side effects than Cu.

Studies of acute and subchronic systemic toxicity associated with a copper/low-density polyethylene nanocomposite intrauterine device

Introduction The physiologic safety of devices and materials intended for clinical implantation should be evaluated. This study, a logical extension of our previous work, aimed to investigate the safety of a novel contraceptive device, the copper/low-density polyethylene nanocomposite intrauterine device (nano-Cu/LDPE IUD), through studies of its potential toxicity after acute and subchronic administration in mice and rats. Methods For the acute toxicity study, single 50 mL/kg doses of nano-Cu/LDPE IUD extracts were administered to mice via intravenous or intraperitoneal injection. General behavioral adverse effects, mortality, and body weights were evaluated for up to 72 hours. In the 13-week subchronic toxicity study, the nano-Cu/LDPE composite with 10-fold higher than the standard clinical dose was implanted subcutaneously into the dorsal skin of Wistar rats. The control group underwent a sham procedure without material insertion. Results During all acute study observation times, the biologic reactions of the mice in the nano-Cu/LDPE group did not differ from those observed in the control group. The groups did not differ statistically in terms of body weight gain, and no macroscopic changes were observed in any organs. In the subchronic study, no clinical signs of toxicity or mortality were observed in either the nano-Cu/LDPE or control group during the 13-week period. The nano-Cu/LDPE composite did not cause any alterations in body weight, food consumption, hematologic and biochemical parameters, or organ weight relative to the control for any observed sample group. Histopathologic examinations of the organs revealed normal architecture, indicating that the inserted material did not cause morphologic disturbances in the rats. Conclusion Overall, the results indicate that the nano-Cu/LDPE IUD did not induce systemic toxicity under experimental conditions of the recommended standard practices, suggesting that the novel material IUD is safe and feasible for future contraceptive applications.

Effect of a novel copper-containing intrauterine device material on the endometrial environment in rabbits

OBJECTIVE(S) This study aimed to determine whether intrauterine placement of a novel composite material [copper (Cu) microparticles, low-density polyethylene, and methyl vinyl silicone rubber (Cu/LDPE/MVQ)] could prevent pregnancy in rabbits, and to evaluate the effects of Cu/LDPE/MVQ on the endometrial environment. STUDY DESIGN Eighty sexually mature female rabbits were randomly divided into four groups (n=20 each group): control (sham-operated), LDPE/MVQ, Cu/LDPE/MVQ microcomposite, and bare Cu. Ten rabbits from each implant-bearing group were randomly selected for a mating experiment beginning 30 days after insertion. Pregnancy outcomes were observed 15 days after mating. Factors associated with endometrial bleeding and inflammation in the remaining rabbits in each group, and the surface conditions of the implants, were investigated 90 days post-insertion. RESULTS The Cu (0 embryo) and Cu/LDPE/MVQ (0 embryo) groups had significantly fewer embryos than the LDPE/MVQ (1.0±0.6 embryos, p<.05) and sham-operated groups (4.1±1.3 embryos, p<.05). Compared with bare Cu, the Cu/LDPE/MVQ composite material was associated with considerable reductions in injuries and factors associated with abnormal endometrial bleeding and inflammation, such as matrix metalloproteinase 9 (MMP9) and prostaglandin E2 (PGE2). Additionally, the surface of implanted Cu/LDPE/MVQ remained much smoother than that of implanted bare Cu. CONCLUSION(S) This novel Cu-containing intrauterine device material exhibits a similar effect in prevention of pregnancy with bare copper, and lower levels of inflammatory markers. IMPLICATIONS This study demonstrates the potential of the novel Cu/LDPE/MVQ microcomposite material as a future substitute for conventional intrauterine device materials.

Isobaric tags for relative and absolute quantification-based proteomic analysis of testis biopsies in rhesus monkeys treated with transient scrotal hyperthermia

This study aimed to examine the cellular and molecular events that occur in rhesus monkey testes after scrotal hyperthermia. Eight male adult rhesus monkeys were subjected to scrotal hyperthermia at 43°C for 30 min daily for 6 consecutive days. Sperm concentration, reproductive hormones, and testis histology were examined before hyperthermia (day 0), and at 8, 15, 30, 45, 60, 75, and 90 days after the initiation of hyperthermia. iTRAQ-based proteomic analysis was conducted on testicular tissues collected on days 0, 8, and 60 to identify differentially expressed proteins at the early and recovery stages of testicular damage. The sperm concentration was significantly decreased at days 30 and 45 after treatment (p < 0.01) and recovered to baseline at day 60. When compared with day 0, 101 and 24 differentially expressed proteins were identified at days 8 and 60 after heat treatment, respectively. The molecular functions of the differentially expressed proteins at day 8 were mainly nucleic acid binding, unfolded protein binding, nucleotide binding, and nucleoside phosphate binding. Spliceosome was enriched as the most significant pathway at day 8. CIRBP, PSIP1, Sam68, and Decorin were validated and found to be consistent with the proteomic data, indicating the reliability of the proteomic profiles identified in this study. In summary, we suggest that the proteins identified in this study may play important roles in heat-induced spermatogenic impairment. Some of these proteins, such as CIRBP, PSIP1, Sam68, and Decorin, may be early molecular targets responsible for spermatogenesis suppression induced by heat treatment.