Bisphenol F (BPF), a widespread environmental contaminant and a major substitute for the restricted bisphenol A (BPA), has raised increasing concerns regarding its potential male reproductive health risks, yet its underlying mechanisms remain poorly understood. This study investigates the mechanisms underlying BPF-induced testicular damage, focusing on the interplay among gut microbiota (GM) dysbiosis, histidine metabolism disruption, and ferroptosis. Using a mouse model exposed to BPF (50, 100, and 200 mg/kg/day) for 28 days, we observed significant testicular pathology, including seminiferous tubule atrophy, vacuolation, and blood-testis barrier (BTB) impairment. Metagenomic and metabolomic analyses revealed GM dysbiosis and suppressed intestinal histidine metabolism, accompanied by decreased abundance of beneficial taxa (e.g., Bacteroides, Ligilactobacillus) and increased potential pathobionts (e.g., Akkermansia, Mucispirillum). BPF exposure was associated with reduced testicular histidine levels and decreased expression of the histidine transporter-related marker LAT1, suggesting impaired histidine availability and a possible alteration in LAT1/CD98-mediated transport; however, direct inhibition of LAT1/CD98 transport activity was not experimentally demonstrated. BPF exposure was accompanied by ferroptosis-related alterations in the testes, including mitochondrial damage, iron accumulation, lipid peroxidation, and downregulation of the xCT-GSH-GPX4 antioxidant axis. In vitro experiments using mouse Sertoli cells (mSCs) confirmed BPF-induced ferroptosis, which was mitigated by the exogenous histidine supplementation. Histidine administration in vivo ameliorated testicular damage, restored BTB integrity, and reversed ferroptotic markers. Our findings support a working model in which a GM-histidine-testis axis may contribute to BPF-induced reproductive toxicity, while further functional studies are required to establish direct causality and transporter-level mechanisms.

Repeat breeding syndrome (RBS), characterized by repeated artificial insemination failure, is associated with ovarian dysfunction in ruminants and causes economic losses. Environmental endocrine-disrupting chemicals (EDCs), such as bisphenol F (BPF), may contribute to RBS, but the relationship between BPF exposure and RBS in cows remains unclear. In this study, we conducted an epidemiological investigation and clinical examinations in 111 adult Simmental cows, including 7 cows diagnosed with RBS, from two local farms, and measured the BPF levels in the drinking water, feed, and the serum samples. We further evaluated the protective effects of phillyrin, a bioactive compound derived from Forsythia suspensa, against BPF-induced ovarian toxicity using cow granulosa cells (CGCs). RBS mainly occurred in young cows with two or more parities. RBS cows showed signs of potential liver dysfunction, kidney injury, anemia, and fecal abnormalities suggestive of altered gut health, together with higher serum BPF levels and lower estradiol levels. Detectable BPF contamination in the drinking water and feed suggested environmental exposure as a possible contributor to RBS in cows. In vitro, BPF induced inflammatory and apoptotic responses in CGCs by upregulating pro-inflammatory and pro-apoptotic mediators, whereas phillyrin pretreatment markedly alleviated these effects. These findings suggest that BPF exposure might be associated with RBS in cows and phillyrin could be used as a promising natural agent for mitigating BPF-related reproductive toxicity.

Serotonin (5-hydroxytryptamine, 5-HT) is a neurotransmitter with emerging roles in mammalian embryonic development. However, its epigenetic effects on early porcine embryos remain poorly understood. This study investigated the impact of 5-HT on histone acetylation and its regulatory mechanisms during early porcine embryo development. Using parthenogenetic embryos, we treated blastocyst, 4-cell, and 2-cell stage embryos with 5-HT and analyzed histone acetylation levels (H3K9Ac and H4K16Ac) and the expression of histone-modifying enzymes (HDAC1, HDAC2, HDAC3, and KAT8). Our results revealed that 5-HT significantly increased histone acetylation at the 4-cell stage but decreased it at the blastocyst stage. These changes were associated with altered expression of histone deacetylases (HDAC2 and HDAC3) and acetyltransferase (KAT8). Specifically, 5-HT upregulated KAT8 and downregulated HDAC2 and HDAC3 at the 4-cell stage, leading to elevated histone acetylation. Conversely, at the blastocyst stage, 5-HT increased HDAC1 and HDAC3 expression while reducing KAT8 activity, resulting in decreased histone acetylation. These findings demonstrate that 5-HT modulates histone acetylation by regulating the balance between histone acetyltransferases and deacetylases, thereby influencing gene expression and embryonic development in pigs. This study provides new insights into the epigenetic role of 5-HT in early embryogenesis and its potential applications in reproductive technologies.

In mammals, the ovum is one of the largest cells with a relatively small ratio of surface area to cytoplasm and unique biological characteristics. The quantity of ova in the ovary is very limited and will decreases dramatically along with female development. Ovarian tissue cryopreservation (OTC) significantly contributes to the preservation of genetic material from the endangered and rare species, animals with special economic value, emotional pets, and excellent police/military dogs. Additionally, it offers the potential for long-term storage of female germplasm resources. Consequently, it is crucial to cryopreserve ovarian tissue and oocytes. During the cryopreservation process, cryoprotectant agents are vital in reducing the fatal or reversible cryogenic damage. In recent years, slow freezing and vitrification are the main methods of mammalian OTC. Currently, challenges of mammalian OTC include the damage of ovarian cells (particularly oocytes) caused by freezing techniques and the subsequent development of transplanted ovarian tissue after freezing and thawing. Thus, further research is required to optimize and enhance the freezing technology or develop novel freezing equipment to improve the efficiency of mammalian OTC. This review encompasses the cryopreservation progress of mammalian ovarian tissue and its prospects.

The cryopreservation of testicular tissues meets the demands for the germplasm preservation of humans and animals. Previously, we reported on the cryopreservation of bovine testicular tissues. To further evaluate the viability of these tissues, subcutaneous xenotransplantation of the frozen–thawed calf testicular tissues was performed with castrated nude mice as the recipients. After 28 days (D28), the survival and development of the grafts were examined. The grafts from 1-day-old (D1) calf testes were recovered and angiogenesis around the grafts was observed. Histologically, the seminiferous cords in the grafts were well maintained and capillaries in the interstitium were observed. Quantitative real-time PCR (qRT-PCR) analysis showed that the grafts expressed germline genes Gfrα-1, C-kit, and Sycp3 and somatic genes Sox9, Acta2, and Star. The expressions of C-kit, Sox9, Acta2, and Star were higher in 28D grafts than those in 1D and 30-day-old (30D) calf testicular controls. Together, we initially demonstrate that cryopreserved calf testicular tissues retain their viability and developmental capacity after xenotransplantation.

Oxidative stress and disruption of blood‒testis barrier permeability are considered key factors in the pathogenesis of testicular inflammation, degeneration, and functional impairment, which play crucial roles in male infertility. Antimicrobial peptides (AMPs) are internationally recognized as some of the most promising alternatives to antibiotics. However, the molecular mechanisms by which AMPs regulate oxidative stress and the blood‒testis barrier in the testis are still poorly understood. In this study, we orally administered 0.5 mg/kg antimicrobial peptide MPX (MPX) to mice for 20 and 40 days and evaluated its effects on Lipopolysaccharide LPS-induced testicular oxidative stress and blood‒testis barrier repair, and elucidateed the pharmacokinetics of MPX in mice. The experiment was divided into six groups, control, LPS, MPX, MPX + LPS, Polymyxin and Polymyxin + LPS, respectively. The results showed that oral administration of MPX effectively increased testicular Glutathione (GSH), Total superoxide dismutase (T-SOD), and Catalase (CAT) levels and reduced Nitric oxide (NO) and Malondialdehyde (MDA) levels in the testes and Lactate dehydrogenase (LDH) levels in serum; these findings were consistent with the oxidative stress parameters in the liver. MPX significantly upregulated the expression of Kelch-like ECH-associated protein 1 (Keap1), Nuclear factor erythroid 2-related factor 2 (Nrf2), and Glutamate cysteine ligase, modifier (GLCM) in the testes while downregulating the expression of Glutamate cysteine ligase, catalytic (GCLC) and Inducible nitric oxide synthase (iNOS), thus exerting a regulatory effect on oxidative stress. MPX also effectively increased sperm count and motility and counteracted the LPS-induced blood‒testis barrier damage, and its molecular mechanism involved upregulating the expression of Slug, which subsequently promoted high expression of Claudin, Occludin, Zonula occludens-1 (ZO-1), N-cadherin, and E-cadherin in the testes. After intragastric administration of FITC-MPX for 30 min, FITC-MPX was mainly distributed in the stomach and thoracic cavity, then showed multi-tissue distribution after 30 min. The fluorescence signal could be detected in the testis 1 h later, which confirmed that MPX had testicular targeting. Moreover, both intraperitoneal and intravenous injection of FITC-MPX also confirmed its testicular targeting ability. In conclusion, this study systematically evaluated the long-term effects of the orally administered antimicrobial peptide MPX on oxidative stress and the blood‒testis barrier in the male reproductive system. This study laid the foundation for the antimicrobial peptide MPX to be used in the treatment of male testicular inflammatory diseases.

Optimizing the cultivation system is crucial for tissue culture. The culture of cryopreserved testicular tissues is of great importance for the germplasm preservation of endangered animals and especially to ensure high-quality and high-output livestock. In this study, we compared two cultivation systems (Agarose-Supported system and Direct Adherent system) by evaluating their effects on tissue morphology, cell proliferation, apoptosis, gene expression, and endocrine function in cryopreserved testicular tissues from 30-day-old calves. The testicular tissues were cultured for 18 and 27 days with three biological replicates per group, aiming to identify which system better supports tissue preservation, cellular viability, and spermatogenic differentiation. This allowed us to clarify how different cultivation systems influence the structural maintenance and developmental potential of immature bovine testicular tissues. Histological and gene expression analyses revealed that the Agarose-Supported system better preserved the seminiferous cord architecture and supported the development of the seminiferous epithelium compared to the Direct Adherent system. The Agarose system significantly reduced the apoptosis and enhanced the expression of some key genes, including spermatogonial stem cell (SSC) markers (GFRα-1, UCHL1), meiotic marker (SYCP3), mature sperm marker (CRISP1), and testicular somatic cell markers (STAR, SOX9, ACTA2). The Agarose-Supported system also benefited spermatogenic differentiation and testosterone secretion. These findings demonstrate that the Agarose-Supported system facilitates the in vitro development of spermatogenic cells and Leydig cells in post-cryopreserved immature bovine testicular tissues.

Pluripotent stem cells (PSCs) can differentiate into three germ layers and diverse autologous cell lines. Since cattle are the most commonly used large domesticated animals, an important food source, and bioreactors, great efforts have been made to establish bovine PSCs (bPSCs). bPSCs have great potential in bovine breeding and reproduction, modeling in vitro differentiation, imitating cancer development, and modeling diseases. Currently, bPSCs mainly include bovine embryonic stem cells (bESCs), bovine induced pluripotent stem cells (biPSCs), and bovine expanded potential stem cells (bEPSCs). Establishing stable bPSCs in vitro is a critical scientific challenge, and researchers have made numerous efforts to this end. In this review, the category of PSC pluripotency; the establishment of bESCs, biPSCs, and bEPSCs and its challenges; and the application outlook of bPSCs are discussed, aiming to provide references for future research.

Bacterial orchitis has a very high percentage of reproductive diseases in males. Therefore, specific medicines are urgently needed to prevent and treat orchitis. Antimicrobial peptides (AMPs) are widely recognized as one of the most promising substitutes for antibiotics. However, it is unknown whether AMPs play a key role in regulating the inflammation of male reproductive system. In the current study, mice were orally administered MPX (a member of Mastoparan family of AMPs, 0.5mg/kg) for 20 and 40 days to evaluate the effect of AMPs on lipopolysaccharide (LPS)-induced testicular inflammation. We found that MPX could effectively improve the physiological function of the testes; upregulate the expression of androgen receptor (AR), inhibin subunit beta B (IBP) and SRY-related HMG box protein (SOX9); and promote sperm quantity and quality. Pathological analysis revealed that MPX significantly prevented LPS-induced inflammatory injury in mouse testes by decreasing the expression of inflammatory cytokines, such as interleukin-1β (IL-1β), IL-6, IL-18, tumor necrosis factor-α (TNF-α), and C-C motif chemokine ligand 2 (CCL2). These results suggest that MPX regulates the testicular inflammatory response by inhibiting the TLR4/NF-kappaB and p65 signaling pathway. Moreover, long-term feeding of MPX could significantly upregulate Bcl-2 and downregulate Bax and Caspase-3, thereby inhibiting LPS-induced apoptosis of testicular cells. In addition, long-term treatment with MPX could regulate LPS-induced pneumonia and enteritis. In summary, this research provides a detailed evaluation of the impact of long-term MPX feeding on inflammation in the male reproductive system, laying the groundwork for the development of specific medicines.

While high-fat diet (HFD)-induced obesity is a major threat to global public health, the effect of HFD on cognition and insulin signaling during ageing remains controversial. The aim of this study was to characterize the dynamic alterations in cognition and cerebral insulin signaling during 6-month HFD consumption, and to investigate the potential therapeutic target and optimal timing to rescue obesity-related cognitive deficits. In the present study, impaired memory retention induced by 2-month HFD was recovered after 4 months on HFD. Prolonged (6-month) HFD did not further enhance tau hyperphosphorylation and β-amyloid deposition, which was consistent with the alleviation of memory retention. In brain insulin signaling, 2-month HFD increased IRS-1 and p-IRS-1(Ser307)/IRS-1, while decreasing pAKT(Ser473)/AKT, PI3K and mTOR; 4-month HFD decreased IRS-1 and pAKT(Ser473)/AKT, while increasing AKT; 6-month HFD increased IRS-1, pAKT(Ser473)/AKT, and mTOR, while decreasing p-IRS-1(Ser307)/IRS-1, PI3K and AKT. Notably, bioinformatic analysis revealed a rhythmic process presented only in 4-month HFD group, with Srebf1 emerging as a link between circadian rhythms and insulin signaling pathway. These results suggest that prolonged HFD prevents further cognitive decline and the progression of Alzheimer’s disease (AD)-related pathologies during ageing. Moreover, there may be a window for recovery, in which Srebf1 acts as a self-recovery switch to address obesity-related cognitive disorders in elders.

Bacterial testicular inflammation is one of the important causes of male infertility. Using plant-derived compounds to overcome the side effects of antibiotics is an alternative treatment strategy for many diseases. Schizandrin B (SchB) is a bioactive compound of herbal medicine Schisandra chinensis which has multiple pharmacological effects. However its effect and the mechanism against testicular inflammation are unknown. Here we tackled these questions using models of lipopolysaccharide (LPS)-induced mice and -Sertoli cells (SCs). Histologically, SchB ameliorated the LPS-induced damages of the seminiferous epithelium and blood-testicular barrier, and reduced the production of pro-inflammatory mediators in mouse testes. Furthermore, SchB decreased the levels of pro-inflammatory mediators and inhibited the nuclear factor kB (NF-κB) and MAPK (especially JNK) signaling pathway phosphorylation in LPS-induced mSCs. The bioinformatics analysis based on receptor prediction and the molecular docking was further conducted. We targeted androgen receptor (AR) and illustrated that AR might bind with SchB in its function. Further experiments indicate that the AR expression was upregulated by LPS stimulation, while SchB treatment reversed this phenomenon; similarly, the expression of the JNK-related proteins and apoptotic-related protein were also reversed after AR activator treatment. Together, SchB mitigates LPS-induced inflammation and apoptosis by inhibiting the AR-JNK pathway.

To obtain high-quality bovine oocytes, the effects of vitamin C (VC) on the IVM of bovine oocytes and early embryo development were investigated. The results showed the following. (1) The IVM medium containing 50 µg/mL VC improved the oocyte maturation rate but did not affect the parthenogenetic embryo development. (2) The IVC medium containing 20 µg/mL VC improved the cleavage rate of the IVF embryos and enhanced the mRNA transcriptions of pluripotency gene Oct4, Sox2, Cdx2, and Nanog in the blastocysts but had no effects on the blastocyst rate. (3) Combining supplementation of 50 µg/mL VC in IVM medium + 20 µg/mL VC in IVC medium (named as VC 50/20, similar hereinafter) elevated the cleavage rate of IVF embryos and enhanced the mRNA expressions of Oct4, Sox2, Cdx2, and Nanog in the blastocysts. (4) Combination of VC 0/20 and VC 50/20 enhanced the transcription of anti-apoptotic gene Bcl-2 and VC 50/0 weakened the transcription of pro-apoptotic gene Bax, while VC 0/40 and VC 0/60 increased Bax expression and diminished the Bcl-2/Bax ratio in blastocysts. Together, employing 50 µg/mL VC improves the IVM of bovine oocytes and combination of VC 50/20 potentially changes bovine embryo quality by enhancing the expressions of the pluripotency genes and regulating the expressions of apoptosis-related genes.

Spermatogonial stem cells (SSCs) possess the characteristics of self-renewal and differentiation, as well as the ability to generate functional sperm. Their unique stemness has broad applications in male infertility treatment and species preservation. In rodents, research on SSCs has been widely reported, but progress is slow in large livestock such as cattle and pigs due to long growth cycles, difficult proliferation in vitro, and significant species differences. Previously, we showed that histone 3 (H3) lysine 9 (K9) trimethylation (H3K9me3) is associated with the proliferation of bovine SSCs. Here, we isolated and purified SSCs from calf testicular tissues and investigated the impact of different H3K9me3 levels on the in vitro proliferation of bovine SSCs. The enriched SSCs eventually formed classical stem cell clones in vitro in our feeder-free culture system. These clones expressed glial cell-derived neurotrophic factor family receptor alpha-1 (GFRα1, specific marker for SSCs), NANOG (pluripotency protein), C-KIT (germ cell marker), and strong alkaline phosphatase (AKP) positivity. qRT-PCR analysis further showed that these clones expressed the pluripotency genes NANOG and SOX2, and the SSC-specific marker gene GFRα1. To investigate the dynamic relationship between H3K9me3 levels and SSC proliferation, H3K9me3 levels in bovine SSCs were first downregulated using the methyltransferase inhibitor, chaetocin, or transfection with the siRNA of H3K9 methyltransferase suppressor of variegation 3-9 homologue 1 (SUV39H1). The EDU (5-Ethynyl-2′-deoxyuridine) assay revealed that SSC proliferation was inhibited. Conversely, when H3K9me3 levels in bovine SSCs were upregulated by transfecting lysine demethylase 4D (KDM4D) siRNA, the EDU assay showed a promotion of cell proliferation. In summary, this study established a feeder-free culture system to obtain bovine SSCs and explored its effects on the proliferation of bovine SSCs by regulating H3K9me3 levels, laying the foundation for elucidating the regulatory mechanism underlying histone methylation modification in the proliferation of bovine SSCs.

Abstract: To obtain bovine oocytes in high quality, effects of vitamin C (VC) on in vitro maturation (IVM) of bovine oocytes and early embryo development were investigated. The results showed that: 1) The IVM medium containing 50 µg/mL VC improved the oocyte maturation rate, while no effects on the development of parthenogenetic embryos; 2) The in vitro culture (IVC) medium containing 20 µg/mL VC increased the cleavage rate of the in vitro fertilized (IVF) embryos, enhanced the mRNA levels of pluripotency gene Oct4, Sox2, Cdx2 and Nanog in the blastocysts, but no effect on the blastocyst rate; 3) Combing supplemention of 50 µg/mL in IVM medium + 20 µg/ml VC in IVC medium (VC 50/20) increased the cleavage rate of IVF embryos and enhanced the mRNA levels of Oct4, Sox2, Cdx2 and Nanog in the blastocysts; 4) Combination of VC 0/20 and VC 50/20 enhanced the expression of anti-apoptotic gene Bcl-2 and VC 50/0 weakened the expression of pro-apoptotic gene Bax, while VC 0/40 and VC 0/60 increased Bax expression and decreased the Bcl-2/Bax ratio in blastocysts. Together, VC supplementation at tuned concentration as well its combination in IVM and IVC media improves the developmental rates of bovine oocytes and embryos.

睾丸支持细胞(Sertoli cells,SCs)和精原干细胞(spermatogonial stem cells,SSCs)发生相应变化才能维持正常精子发生.因此,探讨生精上皮发育中两者的关系对深入了解精子发生有重要意义.基于前期基础,分别取青春期前(出生后5,10 d)、青春期(15,20 d)、近性成熟(25,30 d)及性成熟后(35,40,50,70 d)的昆明雄性小鼠,采用本室已建立的曲细精管漂浮免疫荧光染色全铺片法,测量了小鼠各阶段曲细精索/管的直径;以GATA4为SCs标记、胶质细胞源神经营养因子家族受体α-1(GFRα-1)为SSCs分子标记,显示生精上皮中的SCs和SSCs并分析其数量变化.结果显示,随日龄增加其曲细精索/管直径显著增大,特别是在性成熟前;5 d小鼠的GATA4+SCs数量最少,随后快速上升,15 d时达到最高且显著高于其他各阶段(P＜0.05),之后呈波动下降趋势且趋于相对稳定;GFRα-1+SSCs数量也随日龄增大而增多,15 d时达到最高且显著高于其他各阶段(P＜0.01),随后在20～30 d有所下降,35 d后急剧下降且处于相对稳定状态.提示小鼠SCs与SSCs数量之间基本呈正相关,SCs在维持SSCs的增殖、分化及正常精子发生方面有重要作用.本研究对探讨其他动物和人的精子发生机理有一定借鉴意义.

Spermatogonial stem cells (SSCs) are the only primitive spermatogonial cells in males that can naturally transmit genetic information to their offspring and replicate throughout their lives. Phospholipase D family member 6 (PLD6) has recently been found to be a surface marker for SSCs in mice and boars; however, it has not been validated in cattle. The results of reversed transcription-polymerase chain reaction (RT-PCR) and quantitative real-time PCR (qRT-PCR) found that the relative expression of the PLD6 gene in the testicular tissues of two-year-old Simmental calves was significantly higher than that of six-month-old calves. Immunofluorescent staining further verified the expression of PLD6 protein in bovine spermatogenic cells like germ cell marker DEAD box helicase 4 (DDX4, also known as VASA). Based on multiple bioinformatic databases, PLD6 is a conservative protein which has high homology with mouse Q5SWZ9 protein. It is closely involved in the normal functioning of the reproductive system. Molecular dynamics simulation analyzed the binding of PLD6 as a phospholipase to cardiolipin (CL), and the PLD6-CL complex showed high stability. The protein interaction network analysis showed that there is a significant relationship between PLD6 and piwi-interacting RNA (piRNA) binding protein. PLD6 acts as an endonuclease and participates in piRNA production. In addition, PLD6 in bovine and mouse testes has a similar expression pattern with the spermatogonium-related genes VASA and piwi like RNA-mediated gene silencing 2 (PIWIL2). In conclusion, these analyses imply that PLD6 has a relatively high expression in bovine testes and could be used as a biomarker for spermatogenic cells including SSCs.

Feeder cells are essential to derive pluripotent stem cells (PSCs). Mouse embryonic fibroblasts (MEF) are widely used as feeder to generate and culture embryonic stem cells (ESCs) and induced PSCs (iPSCs) in many species. However it may not be suitable for livestock ESCs/iPSCs due to interspecies difference. Previously we derived bovine iPSCs from bovine Sertoli cells using MEF feeder. Here we compared the effects of MEF feeder and bovine embryonic fibroblasts (BEF) feeder on the maintenance of bovine iPSC pluripotency and morphology as well their contributions to the naive-like conversion, based on a naive medium (NM). The results showed successful conversion of the primed bovine iPSCs to naive-like state within 3-4 days both on MEF feeder and BEF feeder in NM (termed as MNM and BNM respectively). These naive-like iPSCs showed normal karyotype. There were more iPSC colonies under BNM condition than MNM condition. Epigenetically, histone modification H3K4 was upregulated, while H3K27 was downregulated in the naive-like iPSCs. We further analyzed the naive markers and differentiation po-tential both in vitro and in vivo of these cells, which were all reserved throughout the maintenance. Together, bovine naive-like iPSCs can be generated both on MEF and BEF feeder in NM condition. The BNM condition is able to sustain the pluripotency and differentiation potential of the naive-like bovine iPSCs, and improve the conversion efficiency.(c) 2022 Elsevier Inc. All rights reserved.

Epigenetic mechanisms play an important role in oogenesis and early embryo development in mammals. Dimethyl sulfoxide (DMSO) is frequently used as a solvent in biological studies and as a vehicle for drug therapy. Recent studies suggest that DMSO detrimentally affects porcine embryonic development, yet the mechanism of the process in parthenogenetically activated porcine embryos has not been reported. In this study, we found that treatment of embryos with 1.5% DMSO significantly decreased the cleavage and blastocyst rates, total cell number of blastocysts and the anti-apoptotic gene BCL-2 transcription level; however, the percentage of apoptotic cells and the expression levels of the pro-apoptotic gene BAX were not changed. Treatment with DMSO significantly decreased the expression levels of DNMT1 , DNMT3a , DNMT3b , TET1 , TET2 , TET3 , KMT2C , MLL2 and SETD3 in most of the stages of embryonic development and increased 5-mC signals, while the staining intensity for 5-hmC had no change in porcine preimplantation embryos from 2-cell to the blastocyst stages. Meanwhile, DMSO decreased the level of H3K4me3 during the development of parthenogenetically activated porcine embryos. After treatment with DMSO, expression levels of the pluripotency-related genes POU5F1 and NANOG decreased significantly (P <0.01), whereas the imprinted gene H19 did not change (P >0.05). In conclusion, these results suggest that DMSO can affect genome-wide DNA methylation and histone modification by regulating the expression of epigenetic modification enzymes, and DMSO also influences the expression level of pluripotent genes. These dysregulations lead to defects in embryonic development.

Staphylococcus aureus is a common pathogen that can cause pneumonia and a variety of skin diseases. Skin injuries have a high risk of colonization by S. aureus , which increases morbidity and mortality. Due to the emergence of multidrug-resistant strains, antimicrobial peptides are considered to be among the best alternatives to antibiotics due to their unique mechanism of action and other characteristics. MPX is an antibacterial peptide extracted from wasp venom that has antibacterial activity against a variety of bacteria. This study revealed that MPX has good bactericidal activity against S. aureus and that its minimum inhibitory concentration (MIC) is 0.08 μM. MPX (4×MIC) can kill 99.9% of bacteria within 1 h, and MPX has good stability. The research on the bactericidal mechanism found that MPX could destroy the membrane integrity, increase the membrane permeability, change the membrane electromotive force, and cause cellular content leakage, resulting in bactericidal activity. Results from a mouse scratch model experiment results show that MPX can inhibit colonization by S. aureus , which reduces the wound size, decreases inflammation, and promotes wound healing. This study reports the activity of MPX against S. aureus and its mechanism and reveals the ability of MPX to treat S. aureus infection in mice, laying the foundation for the development of new drugs for bacterial infections.