Abstract Prothioconazole (PTC), a widely used triazole fungicide, and its metabolite, prothioconazole-desthio (dPTC), have been frequently detected in environmental matrices, raising significant concerns regarding their potential adverse effects on mammalian reproductive health. While melatonin has demonstrated protective properties across multiple physiological systems, its efficacy in mitigating PTC- and dPTC-induced reproductive toxicity remains poorly understood. This study investigated the reproductive effects of PTC and dPTC in male mice and evaluated the potential protective effects of melatonin. PTC and dPTC exposure induced reproductive dysfunction, characterized by reduced sperm counts, disrupted spermatogenesis, altered HPG axis hormone secretion, elevated oxidative stress, and compromised BTB integrity. Melatonin co-treatment significantly attenuated these adverse effects, restoring sperm parameters, normalizing HPG axis function, mitigating oxidative stress, and preserving BTB integrity, possibly via the modulation of the MAPK/ERK signaling pathway. These findings underscore the potential reproductive toxicity of PTC/dPTC and propose melatonin as a promising therapeutic approach to alleviate the adverse reproductive impacts induced by triazole fungicides.

1 Title: Spike development inhibition in the ftin mutant is associated with multiple phenotypes 2 and regulated by multiple biological pathways 3 Yongsheng Zheng , Jinpeng Zhang, Cheng Liu, Han Zhang, Xiajie Ji, Mumu Wang, Hui wang, Rongzhi Zhang, Ruyu Li 4 , Weihua Liu 5 1 Institute of Crop Sciences, National Key Facilities for Crop Gene Resources and Genetic Improvement, Chinese Academy of 6 Agricultural Sciences, Beijing 100081, P. R. China 7 2 Crop Research Institute, Shandong Academy of Agricultural Sciences, Jinan 250100, P. R. China 8 ∗Co-first author; Corresponding author: Professor Weihua Liu, Institute of Crop Sciences, Chinese Academy of Agricultural 9 Sciences, Beijing 100081, P. R. China, e-mail: liuweihua@caas.cn; ∗∗ Additional corresponding author: Professor Ru-Yu Li, 10

OBJECTIVE:Nonsyndromic clefts of the lip and/or palate (NSCL/P) are one of the most common polygenic diseases. Recently, many studies focused on the association between CRISPLD2 polymorphisms and NSCL/P risk. However, some studies have shown opposite results. In this study, meta-analysis was used to confirm whether CRISPLD2 polymorphism was associated with NSCL/P, and the possible mechanism between CRISPLD2 and NSCL/P was explored.METHODS:Relevant studies were conducted on PubMed, Ovid, EBSCO, CINAHL, FMRS, Web of Science, CNKI, and Wanfang databases from their inception up to June 31, 2016. Review Manager 5.0.24 was used to analyze whether CRISPLD2 polymorphism was involved in NSCL/P by pooling odds ratios (ORs) and 95% confidence intervals (CIs). Potential publication bias was evaluated by visual inspection of the funnel plot.RESULTS:CRISPLD2 rs4783099 was associated with cleft lip and/or palate (CL/P) statistically (OR = 3.18, P < .01). Compared to genotype TT, genotypes CC and CT were correlated significantly (OR = 2.04, P = .04) with CL/P. No evidence showed an association between genetic variation at the CRISPLD2 locus and cleft palate only (CP).CONCLUSION:The polymorphism of CRISPLD2 rs4783099 is correlated with an increased risk of CL/P.

We have shown Bisphenol A (BPA) acts as an androgen receptor (AR) antagonist in the previous study. However, the mechanisms underlying anti-androgenic effects of BPA remain unclear. The objective of this study was to explore whether the AR signaling was involved in AR antagonism of BPA. The Cell Counting Kit-8 (CCK-8) assay and Real-Time Cell Analysis (RTCA) iCELLigence system were applied to analyze the mouse Sertoli cell TM4 proliferation. The mammalian two-hybrid assays were performed to investigate the effects of BPA on the AR amino- and carboxyl-terminal regions (N/C) interaction and the interactions of the AR with steroid receptor coactivator-1 (SRC-1), co-repressors including silencing mediator for thyroid hormone receptors (SMRT) and nuclear receptor co-repressor (NCoR). BPA exposure resulted in decreased TM4 cell proliferation. BPA inhibited the AR N/C interaction significantly. Furthermore, BPA enhanced the interactions of AR-SMRT and AR-NCoR significantly. In conclusion, these data suggest BPA inhibits Sertoli cell proliferation due to its anti-androgenic actions. The mechanisms responsible for AR antagonism of BPA involve inhibiting the AR N/C interaction and enhancing the interactions of AR-SMRT and AR-NCoR. The data uncover novel anti-androgenic mechanisms by which BPA antagonizes AR signaling, contributing to Sertoli cell proliferation suppression and male reproductive toxicology.

Abstract The insecticide cypermethrin has been considered as an endocrine-disrupting chemicals (EDCs) with anti-androgenic activity by interfering with interleukin-6 (IL-6) – induced ligand-independent AR signaling. The purpose of this study was to clarify whether the signal transducer and activator of transcription 3 (STAT3) was involved in the antagonism effect of cypermethrin. In this study, the Western blot was to test the level of STAT3 phosphorylation and the mammalian two-hybrid assay was developed to assess the AR–STAT3 interaction. The date showed that IL-6 increased the phosphorylation level of STAT3 and enhanced the AR–STAT3 interaction. Cypermethrin did not affect the phosphorylation level of STAT3 induced by IL-6, while suppressed the AR-STAT3 interaction induced by IL-6 significantly at the concentration of 10−5 M (p < 0.05). The study indicates cypermethrin inhibits IL-6–induced AR signaling by suppressing the interaction between the AR and STAT3. We provide a novel mechanism of cypermethrin-mediated antagonism on IL-6–induced AR activation associated with STAT3.

Background The androgen receptor (AR) can be stimulated by interleukin-6 (IL-6) in the absence of androgens to induce prostate cancer progression. The purpose of this study was to investigate whether the co-activator steroid receptor coactivator-1 (SRC-1) and co-repressor silencing mediator for retinoid and thyroid hormone receptors (SMRT) are involved in IL-6-induced AR activation. Methods The effects of IL-6 on LNCaP cell proliferation were monitored using real-time cell analysis (RTCA) iCELLigence system. The impacts of IL-6 on the association of the AR with SRC-1 and SMRT were investigated using the mammalian two-hybrid assay. Results IL-6 increased the proliferation of LNCaP cells with maximal induction at 50 ng/mL. The AR-SRC-1interaction was enhanced by IL-6, with maximal induction at the concentration of 50 ng/mL ( P <0.05). IL-6 decreased theAR-SMRT interaction and a marked reduction was detected at 50 ng/mL ( P <0.05). Conclusions IL-6 enhances LNCaP cells proliferation, which suggests that IL-6 might cause AR-positive prostate cancer growth through activation of the AR. The mechanism of IL-6-inducedARactivation is mediated through enhancing AR-SRC-1 interaction and inhibiting AR-SMRT interaction. We have shown a significant role for SRC-1 and SMRT in modulating IL-6-induced AR transactivation.

目的 研究氯氰菊酯(CYP)对白介素-6(IL-6)作用下前列腺特异性抗原(PSA)基因mRNA表达的效应.方法 将LNCaP细胞接种于6孔板中培养24 h,以含0、25、50、100 mg/L质量浓度IL-6的培养基培养24 h后收获细胞,提取总RNA,以荧光定量RT-PCR检测PSA mRNA相对表达水平.以50 mg/L的IL-6和10-7、10-6、10-5 mol/L的CYP共同培养LNCaP细胞24 h,收获细胞,提取总RNA,以荧光定量RT-PCR检测PSA mRNA相对表达水平.结果 IL-6能够促进LNCaP细胞PSA mRNA的表达水平,50 mg/L的IL-6对诱导作用最强(P＜0.05);PSA mRNA的相对表达水平随着氯氰菊酯浓度的增加而降低,各实验组与对照组相比,差异均有统计学意义(P值均＜0.05).结论 实验条件下,氯氰菊酯可以抑制IL-6作用下LNCaP细胞PSA mRNA的表达.

It is hypothesized that the pesticide cypermethrin may induce androgen receptor (AR) antagonism via ligand-independent mechanisms. The Real-Time Cell Analysis (RTCA) iCELLigence system was used to investigate the inhibitory effect of cypermethrin on interleukin-6 (IL-6)-induced ligand-independent LNCaP cell growth. Then, the mammalian two-hybrid assays were applied to clarify whether the mechanism of IL-6-induced AR antagonism of cypermethrin was associated with the interactions of the AR and co-activator steroid receptor co-activator-1 (SRC-1) and co-repressor silencing mediator for retinoid and thyroid hormone receptors (SMRT). Cypermethrin inhibited the LNCaP cell growth induced by IL-6. The interactions of AR-SRC-1 and AR-SMRT mediated by IL-6 were suppressed by cypermethrin. The results indicate that the IL-6-mediated AR antagonism induced by cypermethrin is related to repress the recruitment of co-regulators SRC-1 and SMRT to the AR in a ligand-independent manner. Inhibition of the interactions of AR-SRC-1 and AR-SMRT mediated by IL-6 contributes to the AR antagonism induced by cypermethrin.

To identify whether androgen receptor (AR) antagonism by cypermethrin involves interleukin-6 (IL-6)-induced ligand-independent AR signaling, we have developed the AR reporter gene assay. The reporter gene plasmid pMMTV-chloramphenicol transferase (CAT) was transfected into LNCaP cells. IL-6 increased expression of MMTV-CAT significantly (P<0.05). Cypermethrin decreased CAT reporter expression induced by IL-6 (50 ng/ml), and the significant inhibition was detected at 10(-5)M (P<0.05). IL-6 induces ligand-independent activation of AR. Cypermethrin exhibits inhibitory effects on IL-6-induced ligand-independent AR signaling. We provide a novel insight into cypermethrin-mediated antagonism of the IL-6-mediated ligand-independent activation of the AR.

The 15-day intact adult male assay was used to evaluate effects of isoflurane on the testes and sexual hormones. Forty adult male Sprague-Dawley rats were divided into five groups exposed to air containing 0, 50, 300, 1800 or 10,800 ppm isoflurane. After the treatments, serum was collected for the hormones assay. The right testis was to be used for daily sperm production. The left testis was processed for histopathology and electron microscopy observation. Daily sperm productions were significantly decreased at doses of 300, 1800 and 10,800 ppm. Impaired seminiferous tubules were noted at doses of 300, 1800 and 10,800 ppm. Ultrastructural changes included nucleus agglutination of spermatocytes, big lipid drops and autophagosome in cytoplasm. The serum follicle-stimulating hormone and testosterone concentrations reduced significantly at doses of 1800 and 10,800 ppm. Isoflurane induced impairments of seminiferous tubules and spermatogenesis. The testicular damages caused by isoflurane can be related to the imbalances in the sexual hormones.