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Atrazine (ATR) is one of the most commonly used herbicides worldwide. As an endocrine disruptor, it causes ovarian dysfunction, but the mechanism is unclear. We hypothesized that ATR could affect ovarian steroidogenesis, oxidative stress, inflammation, and apoptosis. In the current study, rats aged 28 days were treated with PMSG and HCG to obtain amounts of corpora lutea. Then, rats were injected with ATR (50 mg/kg/day) or saline (0.9%) for 7 days. Sera were collected to detect biochemical indices and progesterone (P4) level, ovaries were collected for antioxidant status, HE, qPCR, and WB analysis. Results showed that ATR exposure affected growth performance as well as serum TP, GLB, and ALB levels, increased serum P4 level and ovarian mRNA and protein levels of StAR, CYP11A1, and HSD3B. ATR treatment increased ovarian mRNA and protein levels of CREB but not PKA expression. ATR treatment increased ovarian mRNA abundances of Nrf-2 and Nqo1 , MDA level, and decreased SOD, GST, and T-AOC levels. ATR exposure increased the mRNA abundances of pro-inflammatory cytokines including Tnf-α , Il-1β , Il-6 , Il-18 , and Inos. ATR exposure increased the mRNA and protein level of Caspase 3 and the ratio of BAX/BCL-2. In conclusion, NRF-2/NQO1 signaling pathway and CREB might be involved in the regulation of ATR in luteal steroidogenesis, oxidative stress, inflammation, and apoptosis in rat ovary. [Display omitted] • ATR exposure increased serum P4 level, and luteal expressions of steroidogenic proteins and CREB. • ATR exposure triggers luteal oxidative stress and inflammatory response. • ATR exposure induces luteal cell apoptosis in pseudopregnant rats. [ABSTRACT FROM AUTHOR]

Phagocytic clearance of apoptotic germ cells (GCs), as well as residual bodies released from developing spermatids, is critical for Sertoli cells (SCs) to maintain inner environment homeostasis within the testis. However, the molecular mechanisms controlling the phagocytosis are ill defined. Here, we identify a new role for alpha‐enolase (ENO1), a key enzyme during glycolysis, as a molecule that facilitates testicular phagocytosis via transactivation of the engulfment and cell motility 1 (Elmo1) gene. Using immunohistochemistry and double‐labeling immunofluorescence, ENO1 was observed to be expressed exclusively in the nuclei of SCs and its expression correlated with the completion of SC differentiation. By incubating TM4 cells with different pharmacological inhibitors and establishing TM4Tnfr1−/− cells, we demonstrated that SC‐specific expression of ENO1 was under a delicate paracrine control from apoptotic GCs. In turn, persistent blockade of ENO1 expression by a validated small interfering RNA protocol resulted in the disturbance of spermatogenesis and impairment of male fertility. Furthermore, using ChIP, electrophoretic mobility shift and luciferase reporter assays, we showed that, in the presence of apoptotic GCs, ENO1 binds to the distal region of the Elmo1 promoter and facilitates transactivation of the Elmo1 gene. In agreement, overexpression of ELMO1 ameliorated ENO1 deficiency‐induced impairment of phagocytosis in TM4 cells. These data reveal a novel role for SC‐specific expression of ENO1 in regulating phagocytosis in testis, identify tumor necrosis factor‐α and ELMO1 as critical upstream and downstream factors in mediating ENO1 action, and have important implications for our understanding of paracrine control of SC function by adjacent GCs. [ABSTRACT FROM AUTHOR]

Zearalenone (ZEA) and Deoxynivalenol (DON) are two mycotoxins highly detected in agricultural products and feed. Both mycotoxins produce reproductive toxicity and pose a serious threat to human and animal health, among which pigs are the most sensitive animals. Sertoli cells (SCs) play an important role in spermatogenesis; however, the combined toxicity of ZEA and DON and the screening of effective protective agents remains to be determined. By studying the effects of N-acetylcysteine (NAC) on the cells exposed to 20 μM of ZEA and 0.6 μM of DON, we explored the protective mechanism of NAC (4 mM) on the cytotoxic injury of piglets SCs induced by both mycotoxins. The results showed that the combination of ZEA and DON destroy organelles and SCs structures, NAC significantly alleviates the damage caused by ZEA and DON. NAC also significantly increased the expression and distribution of zonula occludens 1 (ZO-1), decreased the relative mRNA and protein expression levels of Bax, Bid, caspase-3, and caspase-9, and increased Bcl-2 expression level and inhibited the decrease of mitochondrial membrane potential. Further, NAC also eases the cell cycle arrest and oxidative stress caused by ZEA and DON. In summary, our results show that NAC could alleviate SCs injury via reducing the oxidative damage and apoptosis caused by ZEA and DON. [ABSTRACT FROM AUTHOR]

Background. Astragalus Radix (AR)-Panax notoginseng (PN), a classical herb pair, has shown significant effects in treating diabetic nephropathy (DN). However, the intrinsic mechanism of AR-PN treating DN is still unclear. This study aims to illustrate the mechanism and molecular targets of AR-PN treating DN based on network pharmacology combined with bioinformatics. Materials and Methods. The Traditional Chinese Medicine Systems Pharmacology database was used to screen bioactive ingredients of AR-PN. Subsequently, putative targets of bioactive ingredients were predicted utilizing the DrugBank database and converted into genes on UniProtKB database. DN-related targets were retrieved via analyzing published microarray data (GSE30528) from the Gene Expression Omnibus database. Protein-protein interaction networks of AR-PN putative targets and DN-related targets were established to identify candidate targets using Cytoscape 3.8.0. GO and KEGG enrichment analyses of candidate targets were reflected using a plugin ClueGO of Cytoscape. Molecular docking was performed using AutoDock Vina software, and the results were visualized by Pymol software. The diagnostic capacity of hub genes was verified by receiver operating characteristic (ROC) curves. Results. Twenty-two bioactive ingredients and 189 putative targets of AR-PN were obtained. Eight hundred and fifty differently expressed genes related to DN were screened. The PPI network showed that 115 candidate targets of AR-PN against DN were identified. GO and KEGG analyses revealed that candidate targets of AR-PN against DN were mainly involved in the apoptosis, oxidative stress, cell cycle, and inflammation response, regulating the PI3K-Akt signaling pathway, cell cycle, and MAPK signaling pathway. Moreover, MAPK1, AKT1, GSK3B, CDKN1A, TP53, RELA, MYC, GRB2, JUN, and EGFR were considered as the core potential therapeutic targets. Molecular docking demonstrated that these core targets had a great binding affinity with quercetin, kaempferol, isorhamnetin, and formononetin components. ROC curve analysis showed that AKT1, TP53, RELA, JUN, CDKN1A, and EGFR are effective in discriminating DN from controls. Conclusions. AR-PN against DN may exert its renoprotective effects via various bioactive chemicals and the related pharmacological pathways, involving multiple molecular targets, which may be a promising herb pair treating DN. Nevertheless, these results should be further validated by experimental evidence. [ABSTRACT FROM AUTHOR]

It has been reported by us and other groups that the expression of small GTP binding protein RhoB can be induced by genotoxic stressors and glucocorticoid (GC), a stress hormone that plays a key role in stress response. Until now stress-induced genes that confer cytoprotection under stressed conditions are largely unknown. In this study, we investigated the effects and mechanism of non-genotoxic stressors, including scalding in vivo and heat stress in vitro on the expression of RhoB. We found for the first time that both scalding, which could induce typical neuroendocrine responses of acute stress and cellular heat stress significantly increased the expression of RhoB at mRNA and protein levels. Moreover, in vitro experiments in human lung epithelial cells (A549) showed that induction of RhoB by heat stress was in a glucocorticoid receptor (GR)-independent manner and through multiple pathways including stabilization of RhoB mRNA and activation of p38 MAPK. Further experiments demonstrated that up-regulation of RhoB significantly inhibited heat stress-induced apoptosis and elevated transcriptional activity of NF-κB, but did not affect the expression of Hsp70 in A549 cells. In conclusion, we showed for the first time that RhoB was up-regulated by scalding in vivo and heat stress in vitro and played an important cytoprotective role during heat stress-induced apoptotic cell death. J. Cell. Physiol. 226: 729-738, 2011. © 2010 Wiley-Liss, Inc. [ABSTRACT FROM AUTHOR]

Background/Aims: The current study aimed to investigate the role by which fibronectin 1 (FN1) influences the cell cycle, senescence and apoptosis in human glioma cells through the PI3K/ AKT signaling pathway. Methods: Differentially expressed genes (DEGs) were identified based on gene expression data (GSE12657, GSE15824 and GSE45921 datasets) and probe annotation files from Gene Expression Omnibus. The DEGs were identified in connection with gene ontology (GO) enrichment analysis and with the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. The positive expression of the FN1 protein was detected by immunohistochemistry. The glioma cell lines U251 and T98G were selected and assigned into blank, negative control (NC) and siRNA-FN1 groups. A dual luciferase reporter gene assay was used to investigate the effects of FN1 on transcriptional activity through the PI3K/AKT signaling pathway. An MTT assay was applied for the detection of cell proliferation, while flow cytometry was employed for cell cycle stage and cellular apoptosis detection. β-galactosidase staining was utilized to detect cellular senescence, a scratch test was applied to evaluate cell migration, and a transwell assay was used to analyze cell invasion. Western blotting and qRT-PCR methods were used to detect the protein and mRNA expression levels, respectively, of the FN1 gene and the related genes in the PI3K/AKT pathway (PI3K, AKT and PTEN), the cell cycle (pRb, CDK4 and Cyclin D1) and cell senescence (p16 and p21) among the collected tissues and cells. Results: GSE12657 profiling revealed FN1 to be the most upregulated gene in glioma. Regarding the GSE12657 and GSE15824 datasets, FN1 gene expression was higher in glioma tissues than in normal tissues. GO enrichment analysis and KEGG pathway enrichment analysis indicated that FN1 is involved in the synthesis of extracellular matrix (ECM) components and the PI3K/AKT signaling pathway. Verification was provided, indicating the role played by the FN1 gene in the regulation of the PI3K/AKT signaling pathway, as silencing the FN1 gene was found to inhibit cell proliferation, promote cell apoptosis and senescence, and reduce migration and invasion through the down-regulation of FN1 gene expression and disruption of the PI3K-AKT signaling pathway. Conclusion: The findings of this study provide evidence highlighting the prominent role played by FN1 in stimulating glioma growth, invasion, and survival through the activation of the PI3K/AKT signaling pathway. [ABSTRACT FROM AUTHOR]

Abstract: In the present investigation, 16 new rotundic acid (RA) derivatives modified at the C-3, C-23 and C-28 positions were synthesized. The cytotoxicities of the derivatives were evaluated against HeLa, A375, HepG2, SPC-A1 and NCI-H446 human tumor cell lines by MTT assay. Among these derivatives, compounds 4–7 exhibited stronger cell growth inhibitory than RA and compound 4 was found to be the best inhibition activity on five human tumor cell lines with IC50 <10μM. The apoptosis mechanism of compound 4 in HeLa cells was investigated by western blot analysis. The results indicated that compound 4 could induce apoptosis through increasing protein expression of cleaved caspase-3 and Bax, and decreasing protein expression of Bcl-2. In summary, the present work suggests that compound 4 might serve as an effective chemotherapeutic candidate. [Copyright &y& Elsevier]

Objectives: To examine the expression of N-Myc downstream regulated gene 2 (NDRG2) in the cryptorchid testes and its potential association with germ cell apoptosis in a rat model of cryptorchidism. Methods: Groups of pubertal Sprague-Dawley rats were subjected to surgical procedure for the induction of experimental unilateral cryptorchidism. The weight of the abdominal and scrotal testes was measured and their apoptotic cells and NDRG2 expression were examined longitudinally by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling, immunohistochemistry, and immunofluorescence analyses. The expression of NDRG2 and p53 in the purified germ cells and Leydig cells was determined by Western blot assay. Results: Compared with the control scrotal testes, the mean weights of the abdominal testes were significantly reduced at 7 and 14 days after surgery, accompanied by significantly increased numbers of apoptotic germ cells. Although the NDRG2 expression was detected in Leydig cells and spontaneous apoptotic germ cells in the scrotal testes, it was significantly upregulated predominately in apoptotic spermatocytes and round spermatids in the abdominal testes. However, the NDRG2 expression was downregulated in Leydig cells in the abdominal testes. Western blot analysis revealed that the expression of NDRG2 and p53 was upregulated significantly in purified germ cells 7 days after surgery, whereas NDRG2 was downregulated in the purified Leydig cells from the abdominal testes 7 and 14 days after surgery. Conclusions: The upregulated expression of NDRG2 was associated with germ cell apoptosis in the cryptorchid testes, suggesting that the NDRG2 may be a positive regulator for germ cell apoptosis in rat cryptorchidism. [Copyright &y& Elsevier]