Obesity is a systemic metabolic disease that can induce male infertility or subfertility through oxidative stress. The aim of this study was to determine how obesity impairs sperm mitochondrial structural integrity and function, and reduces sperm quality in both overweight/obese men and mice on a high‐fat diet (HFD). Mice fed the HFD demonstrated higher body weight and increased abdominal fat content than those fed the control diet. Such effects accompanied the decline in antioxidant enzymes, such as glutathione peroxidase (GPX) and catalase and superoxide dismutase (SOD) in testicular and epidydimal tissues. Moreover, malondialdehyde (MDA) content significantly increased in sera. Mature sperm in HFD mice demonstrated higher oxidative stress, including increased mitochondrial reactive oxygen species (ROS) levels and decreased protein expression of GPX1, which may impair mitochondrial structural integrity and reduce mitochondrial membrane potential (MMP) and ATP production. Moreover, cyclic AMPK phosphorylation status increased, whereas sperm motility declined in the HFD mice. Clinical studies demonstrated that being overweight/obese reduced SOD enzyme activity in the seminal plasma and increased ROS in sperm, accompanied by lower MMP and low‐quality sperm. Furthermore, ATP content in the sperm was negatively correlated with increases in the BMI of all clinical subjects. In conclusion, our results suggest that excessive fat intake had similar disruptive effects on sperm mitochondrial structure and function, as well as oxidative stress levels in humans and mice, which in turn induced lower sperm motility. This agreement strengthens the notion that fat‐induced increases in ROS and impaired mitochondrial function contribute to male subfertility.

Background: Recently, human infertility incidence is increasing in obese women causing it to become an emerging global health challenge requiring improved treatment. There is extensive evidence that obesity caused female reproductive dysfunction is accompanied by an endocrinological influence. Besides, systemic and tissue-specific chronic inflammatory status are common characteristics of obesity. However, the underlying molecular mechanism is unclear linking obesity to infertility or subfertility. Methods: To deal with this question, we created an obese mouse model through providing a high fat diet (HFD) and determined the fertility of the obese mice. The morphological alterations were evaluated in both the reproductive glands and tracts, such as uterus, ovary and oviduct. Furthermore, to explore the underlying mechanism of these functional changes, the expressions of pro-inflammatory cytokines as well as the activations of MAPK signaling and NF-κB signaling were detected in these reproductive tissues. Results: The obese females were successful construction and displayed subfertility. They accumulated lipid droplets and developed morphological alterations in each of their reproductive organs including uterus, ovary and oviduct. These pathological changes accompanied increases in pro-inflammatory cytokine expression levels of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in all of these sites. Such effects also accompanied increases in nuclear factor kappa B (NF-kB) expression and mitogen-activated protein kinase (MAPK) signaling pathway stimulation based on uniform time dependent increases in the NF-κB (p-NF-κB), JNK (p-JNK), ERK1/2 (p-ERK) and p38 (p-p38) phosphorylation status. Conclusions: These HFD-induced increases in pro-inflammatory cytokine expression levels and NF-κB and MAPKs signaling pathway activation in reproductive organs support the notion that increases of adipocytes resident and inflammatory status are symptomatic of female fertility impairment in obese mice.

Chemotherapy and radiation are more effective in wild-type (WT) p53 tumors due to p53 activation. This is one rationale for developing drugs that reactivate mutant p53 to synergize with chemotherapy and radiation. Zinc metallochaperones (ZMC) are a new class of mutant p53 reactivators that restore WT structure and function to zinc-deficient p53 mutants. We hypothesized that the thiosemicarbazone, ZMC1, would synergize with chemotherapy and radiation. Surprisingly, this was not found. We explored the mechanism of this and found the reactive oxygen species (ROS) activity of ZMC1 negates the signal on p53 that is generated with chemotherapy and radiation. We hypothesized that a zinc scaffold generating less ROS would synergize with chemotherapy and radiation. The ROS effect of ZMC1 is generated by its chelation of redox active copper. ZMC1 copper binding (K_Cu) studies reveal its affinity for copper is approximately 10⁸ greater than Zn²⁺. We identified an alternative zinc scaffold (nitrilotriacetic acid) and synthesized derivatives to improve cell permeability. These compounds bind zinc in the same range as ZMC1 but bound copper much less avidly (10⁶- to 10⁷-fold lower) and induced less ROS. These compounds were synergistic with chemotherapy and radiation by inducing p53 signaling events on mutant p53. We explored other combinations with ZMC1 based on its mechanism of action and demonstrate that ZMC1 is synergistic with MDM2 antagonists, BCL2 antagonists, and molecules that deplete cellular reducing agents. We have identified an optimal Cu²⁺:Zn²⁺ binding ratio to facilitate development of ZMCs as chemotherapy and radiation sensitizers. Although ZMC1 is not synergistic with chemotherapy and radiation, it is synergistic with a number of other targeted agents.

Background: Overwhelming evidences suggest oxidative stress is a major cause of sperm dysfunction and male infertility. Zinc is an important non-enzymatic antioxidant with a wide range of biological functions and plays a significant role in preserving male fertility. Notably, zinc trafficking through the cellular and intracellular membrane is mediated by specific families of zinc transporters, i.e., SLC39s/ZIPs and SLC30s/ZnTs. However, their expression and function were rarely evaluated in the male germ cells. The aim of this study is to determine and characterize the crucial zinc transporter responsible for the maintenance of spermatogenesis. Methods: The expression patterns of all 14 ZIP members were characterized in the mouse testis. qRT-PCR, immunoblot and immunohistochemistry analyses evaluated the ZIP12 gene and protein expression levels. The role of ZIP12 expression was evaluated in suppressing the sperm quality induced by exposure to an oxidative stress in a spermatogonia C18–4 cell line. Zip12 RNAi transfection was performed to determine if its downregulation altered cell viability and apoptosis in this cell line. An obese mouse model fed a high-fat-diet was employed to determine if there is a correlation between changes in the ZIP12 expression level and sperm quality. Results: The ZIP12 mRNA and protein expression levels were higher than those of other ZIP family members in both the mouse testis and other tissues. Importantly, the ZIP12 expression levels were very significantly higher in both mice and human spermatogonia and spermatozoa. Moreover, the testicular ZIP12 expression levels significantly decreased in obese mice, which was associated with reduced sperm zinc content, excessive sperm ROS generation, poor sperm quality and male subfertility. Similarly, exposure to an oxidative stress induced significant declines in the ZIP12 expression level in C18–4 cells. Knockdown of ZIP12 expression mediated by transfection of a ZIP12 siRNA reduced both the zinc content and viability whereas apoptotic activity increased in the C18–4 cell line. Conclusions: The testicular zinc transporter ZIP12 expression levels especially in spermatogonia and spermatozoa are higher than in other tissues. ZIP12 may play a key role in maintaining intracellular zinc content at levels that reduce the inhibitory effects of rises in oxidative stress on spermatogonia and spermatozoa viability during spermatogenesis which help counteract declines in male fertility.

Asthenozoospermia is one of the leading causes of male infertility owing to a decline in sperm motility. Herein, we determined if there is a correlation between RNASET2 content on human spermatozoa and sperm motility in 205 semen samples from both asthenozoospermia patients and normozoospermia individuals. RNASET2 content was higher in sperm from asthenozoospermia patients than in normozoospermia individuals. On the other hand, its content was inversely correlated with sperm motility as well as progressive motility. Moreover, the inhibitory effect of RNASET2 on sperm motility was induced by incubating normozoospermic sperm with RNase T2 protein. Such treatment caused significant declines in intracellular spermatozoa PKA activity, PI3K activity and calcium level, which resulted in severely impaired sperm motility, and the sperm motility was largely rescued by cAMP supplementation. Finally, protein immunoprecipitation and mass spectrometry identified proteins whose interactions with RNASET2 were associated with declines in human spermatozoa motility. AKAP4, a protein regulating PKA activity, coimmunoprecipated with RNASET2 and they colocalized with one another in the sperm tail, which might contribute to reduced sperm motility. Thus, RNASET2 may be a novel biomarker of asthenozoospermia. Increases in RNASET2 can interact with AKAP4 in human sperm tail and subsequently reduce sperm motility by suppressing PKA/PI3K/calcium signaling pathways.

Background: To determine if overweight/obese pregnant women have altered microRNA expression patterns in fetal umbilical cord blood that may affect the development of offspring. Methods: Umbilical cord blood samples were obtained from the fetuses of 34 overweight/obese and 32 normal-weight women after delivery. Next generation sequencing (NGS) analyzed their miRNA expression patterns. miRanda and TargetScan databases were used to predict the miRNAs’ target genes followed by analyses of Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) to perform function grouping and pathway analyses. qRT-PCR verified the identity of differentially expressed miRNAs that were revealed in the NGS results. Results: There was a positive correlation between newborn body weight and pregestational BMI of pregnant individuals (r = 0.48, P < 0.001). One hundred and eight miRNAs were differentially expressed between the normal and overweight/obese groups, which target genes were enriched in the metabolic pathway. Five C19MC miRNAs (miR-516a-5p, miR-516b-5p, miR-520a-3p, miR-1323, miR-523-5p) were upregulated in the overweight/obese group. Target enrichment analysis suggests their involvement in post-embryonic development, lipid and glucose homeostasis, T cell differentiation and nervous system development. Conclusions: C19MC miRNA expression upregulation in the fetal circulation during the gestation of overweight/obese pregnant women may contribute to altered multisystem metabolic pathway development in their offspring.