The role of ferroptosis mediated by Bmal1/Nrf2 in nicotine -induce injury of BTB integrity.

Nicotine has shown the toxic effects on male reproductive system, and testicular damage is associated with ferroptosis, which is a non-apoptotic regulated cell death driven by iron-dependent lipid peroxidation. However, the role of nicotine on ferroptosis of testicular cells is largely elusive. In the present study, we showed that nicotine destroyed blood-testis barrier (BTB) by interfering with the circadian rhythm of BTB-related factors (ZO-1, N-Cad, Occludin and CX-43) and induced ferroptosis, as reflected via increased clock-control levels of lipid peroxide and decreased ferritin and GPX4, which involved in the circadian. Inhibition of ferroptosis with Fer-1 alleviated nicotine-induced injury of BTB and impaired sperm in vivo. Mechanically, we uncover that the core molecular clock protein, Bmal1, regulates the expression of Nrf2 via direct E-box binding to its promoter to regulate its activity, and nicotine decreases the transcription of Nrf2 through Bmal1 and inactivates Nrf2 pathway and its downstream antioxidant gene, which leads to the imbalance of redox state and ROS accumulation. Intriguingly, nicotine induced lipid peroxidation and subsequent ferroptosis by Bmal1-mediated Nrf2. In conclusion, our study reveals a clear role for the molecular clock in controlling Nrf2 in testis to mediate the ferroptosis induced by nicotine. These findings provide a potential mechanism to prevent smoking and/or cigarette smoke-induced male reproductive injury.

Solid-Liquid Phase Equilibrium of Ammonium Dihydrogen Phosphate and Agricultural Grade Ammonium Polyphosphate (Degree of Polymerization Ranging from 1 to 8) for Mixed Irrigation Strategy.

Water-soluble ammonium polyphosphate (APP) has the advantages of good solubility and slow-release characteristics and has the potential to be used in combination with monoammonium phosphate (MAP) as a high phosphorus content slow-release fertilizer to improve the utilization rate of phosphorus during irrigation. Herein, the effects of the APP1 concentration and temperature (278.2-313.2 K) on the solubility of MAP, solution density, and pH value in the ternary equilibrium system (APP1-MAP-water) were measured. The simplified Apelblat model, two empirical polynomials, and rational two-dimensional functions can describe the experimental solubility data, solution density, and pH value well, respectively, with reliable modeling parameters (R 2 > 0.99). In the OptiMax1001 reactor, the focused beam reflectance measurement (FBRM), the particle-view measurement (PVM), and the ReactIR 15 probes were used to observe and reverse verify that they can be synergistically codissolved to achieve economic efficiency. Basic thermodynamic data and models can guide their collaborative application in irrigation to improve the phosphorus utilization rate.

Liver Receptor homolog-1 Regulates Apoptosis of Bovine Ovarian Granulosa Cells by Progestogen Receptor Signaling Pathway.

The purpose of the present investigation was to assess the function of LRH-1 on GCs and the mechanisms involved. Here, LRH- was highly expressed in the bovine GCs of atretic follicles. Treatment with 50 µM of LRH-1 agonist (DLPC) significantly induced the expression of LRH-1 (p < 0.05). In particular, LRH-1 activation blocked the progestogen receptor signaling pathway via downregulating progesterone production and progestogen receptor levels (p < 0.05), but had no effect on 17 beta-estradiol synthesis. Meanwhile, LRH-1 activation promoted the apoptosis of GCs and increased the activity of caspase 3 (p < 0.05). Importantly, upregulating the progestogen receptor signaling pathway with progestogen could attenuate the LRH-1-induced proapoptotic effect. Moreover, treatment with progestogen decreased the activity of the proapoptotic gene caspase 3 and increased the expression of antiapoptotic gene Bcl2 in LRH-1 activated GCs (p < 0.05). Taken together, these results demonstrate that LRH-1 might be dependent on the progestogen receptor signaling pathway to modulate bovine follicular atresia.

17ß-Estradiol Enhances Porcine Meiosis Resumption from Autophagy-Induced Gap Junction Intercellular Communications and Connexin 43 Phosphorylation via the MEK/ERK Signaling Pathway.

Estrogen and its analogues are ubiquitous in agricultural environments, with large biological functions of oocyte development. Gap junction intercellular communications (GJICs) are the structural basis in cumulus-oocyte complexes (COCs) and regulate oocyte maturation and developmental material transport through a number of pathways. This study mainly determines the effect and potential mechanism of estrogen (17ß-estradiol) in regulating GJICs in porcine COCs. In our study, 17ß-estradiol increased porcine nuclear maturation in a time-dependent manner. The analysis revealed that 17ß-estradiol upregulated the autophagy in COCs during in vitro maturation. In contrast with the control, 17ß-estradiol decreased GJICs in a time-dependent manner between cumulus cells and oocytes, while it was consistent with the control group at 24 h. Carbenoxolone (CBX) blocks GJICs as a negative control group used in our system. Autophagy inhibitor autophinib decreased oocyte maturation, and the reduced nuclear maturation treated with autophinib was abolished by 17ß-estradiol. Besides, the upregulation effect of autophinib on GJICs and transzonal projections (TZPs) was decreased by 17ß-estradiol. 17ß-Estradiol could reduce serine 368 phosphorylation of connexin 43 (Cx43) protein by autophinib in porcine COCs. These results were dependent upon the MEK/ERK signaling pathway. Furthermore, 17ß-estradiol-induced GJICs and Cx43 phosphorylation were inhibited by autophinib or the MEK/ERK pathway inhibitors (Trametinib and FR 180204), indicating that 17ß-estradiol regulated GJICs through the MEK/ERK signaling pathway. In conclusion, 17ß-estradiol improves the autophagy-mediated nuclear maturation with downregulating GJICs and TZPs in porcine COCs. Such an effect occurs by phosphorylation of Cx43, which was regulated via the MEK/ERK signaling pathway.

Isorhamnetin Promotes Estrogen Biosynthesis and Proliferation in Porcine Granulosa Cells via the PI3K/Akt Signaling Pathway.

Isorhamnetin is a natural flavonoid widely distributed in fruits and vegetables. However, the roles of isorhamnetin involved in steroidogenesis, proliferation, and apoptosis in ovarian granulosa cells (GCs) are poorly understood. We found that isorhamnetin promoted the secretion of estrogen and inhibited the secretion of progesterone and testosterone by modulating steroidogenesis-associated proteins and mRNA such as CYP19A1, StAR, and 3ß-HSD in ovarian GCs. Mechanistically, isorhamnetin stimulated the expression of the proliferating cell nuclear antigen and C-myc and promoted the proliferation of GCs via the PI3K/Akt signaling pathway. Furthermore, isorhamnetin increased the protein expression of CyclinB, CyclinD, CyclinE, and CyclinA, thereby raising the ratio of S-phase cells in response to GC proliferation. Changes in the expression of apoptosis-associated proteins (Bcl2, Bax, and cytochrome c) and intracellular reactive oxygen species levels showed that isorhamnetin inhibited GC apoptosis. Collectively, these findings indicate that isorhamnetin regulates steroidogenesis through the activation of PI3K/Akt, which promotes proliferation, inhibits apoptosis, and alleviates oxidative stress.

17ß-estradiol improves the developmental ability, inhibits reactive oxygen species levels and apoptosis of porcine oocytes by regulating autophagy events.

OBJECTIVES: Estrogen plays a critical role in the development and apoptosis of oocytes. Autophagy is an evolutionarily conserved and exquisitely regulated self-eating cellular process with important biological functions including the regulation of reproduction. This study aimed to determine the effect of autophagy regulated by the biologically active form of estrogen (17ß-estradiol) in porcine oocyte maturation in vitro. MATERIALS AND METHODS: We measured the effects of oocyte developmental competencies and autophagic activity in the porcine oocyte regulated by 17ß-estradiol using autophagic inhibitor (Autophinib). In addition, we studied the role of autophagy in reactive oxygen species (ROS) levels, mitochondrial distribution, Ca2+ production, mitochondrial membrane potential (ΔΨm), and early apoptosis by caspase-3, -8 activity in the mature oocytes. RESULTS: The results showed that the oocyte meiotic progression and early embryonic development were gradually decreased with Autophinib treatment, which was improved by 17ß-estradiol. Immunofluorescence experiments revealed that 17ß-estradiol primarily could promote the autophagy in the mature oocytes, and block the reduced-autophagic events by Autophinib. Moreover, 17ß-estradiol improved the Autophinib induced high ROS levels, abnormal mitochondrial distribution and low Ca2+ production in mature oocytes. Analyses of early apoptosis and ΔΨm showed that autophagy inhibition was accompanied by increased cellular apoptosis, and 17ß-estradiol reduced apoptosis rates of mature oocytes. Importantly, autophagy was downregulated by treatment with Autophinib, an activation of caspase-8 and cleaved caspase-3 increased. Those effects were abolished by 17ß-estradiol, which could upregulate autophagy. CONCLUSIONS: Our study have showed important implications that 17ß-estradiol could promote efficacy of the development of porcine oocytes, enhance the autophagy, reduce ROS levels and apoptosis activity in vitro maturation.

Development and characterization of in vitro self-assembled recombinant human follicle stimulating hormone originated from goat mammary epithelial cells.

Follicle stimulating hormone (FSH), composed of FSHα and FSHß subunits, is essential for female follicle development and male spermatogenesis. The recombinant human FSH (rhFSH) products on the market are mainly generated from mammalian cells and are expensive. Large animal mammary gland bioreactors are urgently needed to produce large amounts of rhFSH. However, there are currently no effective methods to prepare rhFSH by large animals mainly due to the fact that excessive accumulation of FSH might cause many adverse effects in animals. We herein report the development and characterization of functional self-assembled rhFSH produced in goat mammary epithelial cells (GMECs). FSHα and FSHß stably expressed in Chinese hamster ovary (CHO) cell lines were secreted into culture medium and well glycosylated. Importantly, FSHα and FSHß expressed apart were able to assemble into functional FSH. We next inserted human FSHα or FSHß gene separately into goat ß-Lactoglobulin locus in GMECs by CRISPR/Cas9. Inactive FSHα and FSHß subunits expressed from GMECs assembled into rhFSH as analyzed by His-tag pull down assay. Functional assessment of rhFSH by cAMP induction assay, mouse ovulation induction and rat ovarian weight gain experiments showed that the bioactivity of self-assembled rhFSH expressed by GMECs was comparable to that of Gonal-F both in vitro and in vivo. Our study demonstrated that FSHα and FSHß can be separately expressed and assembled into functional rhFSH, and provided the basis for future preparing FSH by goat mammary gland bioreactor with less health problems on the producing animals.

WITHDRAWN: Transcriptional regulation of miR-483-3p mediated by IL-6/STAT3 axis promoted epithelial-mesenchymal transition and tumor stemness in glioma.

This article was originally published in Aging Advance Online Publications on November 4, 2020. Concerns were raised at that time about overlap between data in Figures 1D, 8A and 8B of this article and Figures in an unrelated retracted article (https://www.dovepress.com/cr_data/article_fulltext/s268000/268222/img/CMAR_A_268222_O_F0007g.jpg, (https://doi.org/10.2147/CMAR.S268222). The article was investigated, and the editorial decision was made to withdraw the article entirely in compliance with Aging's withdrawal policy. The institution affiliated with the published Aging article was notified about this action. This article will not appear internally in Aging or any external indexes or archives.

Electric field assisted benzene oxidation over Pt-Ce-Zr nano-catalysts at low temperature.

A novel catalytic system for benzene oxidation at low temperature is constructed by combining electric field with Pt-Ce-Zr nano-catalyst. The 1 wt% Pt/Ce0.75Zr0.25O2 catalyst assisted by electric field shows the best catalytic performance with 90% benzene conversion at 96.5 °C and excellent water resistance. The effect of electric field on catalysts and catalytic process is comprehensively investigated. The results of XRD, TEM, XPS and H2-TPR reveal that the electric field show negligible influence on the crystal structure and surface morphology of the catalyst, but it can lead to more oxygen vacancies. Therefore, more adsorbed oxygen with higher activity will be produced on the catalyst surface. The redox performance is improved due to the fact that valence distribution of Pt is changed in forms of more active sites composed of high valence oxides (PtO) generated in electric field. In situ DRIFTS is used to investigate the oxidation process of benzene and the results prove that electric field could accelerate the production and consumption of intermediate products, and produce new intermediate products such as carboxylic acid species, indicating that the introduction of electric field may open up a new rapid reaction path and promote the activation of benzene at low temperature.

Fibrinogen, Neutrophil-to-Lymphocyte Rate and Platelet-to-Neutrophil Rate as Novel Acute Phase Indicators in Patients with Thromboangiitis Obliterans.

BACKGROUND/OBJECTIVES: The acute exacerbations and progressive deterioration seen in thromboangiitis obliterans (TAO) have been related to poor clinical outcomes. Here, we have studied the association of laboratory biomarkers with the acute phase of TAO (AP-TAO). METHODS/RESULTS: We conducted a retrospective case-control study on 112 patients with TAO and 98 healthy controls; comparing the neutrophil-to-lymphocyte rate (NLR), lymphocyte-to-monocyte rate (LMR), platelet-to-neutrophil rate (PNR), fibrinogen (FIB), and apolipoprotein A-I (ApoA-I). Significantly higher NLR level, as well as lower LMR, PNR, and ApoA-I levels were observed in patients with TAO, particularly the acute phase. Significantly increased FIB was only observed in AP-TAO. A positive correlation was found between NLR and with C-reactive protein (CRP) in the acute phase (r = 0.817, P < 0.001). Moreover, NLR, PNR, and FIB levels of 3.38, 45.12, and 3.69 were shown to be the predictive cut-off values for the AP-TAO (sensitivity 72.5, 82,4, and 66,7%, specificity 92.2, 78.4, and 96.1%; area under the curve [AUC] 0.875, 0.855, and 0.872), respectively. The FIB level was independently associated with the AP-TAO (OR = 11.420, P = 0.007). CONCLUSIONS: NLR, PNR, and FIB may be useful markers for the identification of inflammation and the AP-TAO. FIB may be an independent risk factor for the acute phase.

Phospholipase C inhibits apoptosis of porcine primary granulosa cells cultured in vitro.

Phospholipase C (PLC) can participate in cell proliferation, differentiation and aging. However, whether it has a function in apoptosis in porcine primary granulosa cells is largely uncertain. The objective of this study was to examine the effects of PLC on apoptosis of porcine primary granulosa cells cultured in vitro. The mRNA expression of BAK, BAX and CASP3, were upregulated in the cells treated with U73122 (the PLC inhibitor). The abundance of BCL2 mRNA, was upregulated, while BAX and CASP3 mRNA expression was decreased after treatment with m-3M3FBS (the PLC activator). Both the early and late apoptosis rate were maximized with 0.5 µM U73122 for 4 h. The rate of early apoptosis was the highest at 4 h and the rate of late apoptosis was the highest at 12 h in the m-3M3FBS group. The protein abundance of PLCß1, protein kinase C ß (PKCß), calmodulin-dependent protein kinaseII α (CAMKIIα) and calcineurinA (CalnA) were decreased by U73122, and CAMKIIα protein abundance was increased by m-3M3FBS. The mRNA expression of several downstream genes (CDC42, NFATc1, and NFκB) was upregulated by PLC. Our results demonstrated that apoptosis can be inhibited by altering PLC signaling in porcine primary granulosa cells cultured in vitro, and several calcium-sensitive targets and several downstream genes might take part in the processes.

Raf-ERK1/2 signalling pathways mediate steroid hormone synthesis in bovine ovarian granulosa cells.

Steroid hormones are required for normal reproductive function of female. The aim of this study was to investigate the role of Raf-ERK1/2 on steroid hormone synthesis in bovine ovarian granulosa cells. Immunohistochemistry assay showed that both B-Raf and C-Raf were expressed in granulosa cells, theca cells and Sertoli cells. The protein expression of Raf or ERK1/2 was clearly decreased by Raf inhibitor GSK2118436 or ERK1/2 inhibitor SCH772984, respectively (p < 0.05). In addition, western blotting was performed for investigating the crosstalk between Raf and ERK1/2, the data showed that Raf positively regulated ERK1/2, whereas ERK1/2 had a negative feedback effect on Raf. The biosynthesis of oestradiol or testosterone was significantly decreased by treatment with GSK2118436 or SCH772984 (p < 0.05). Conversely, the progesterone biosynthesis was clearly increased by treatment with those inhibitors (p < 0.05). Furthermore, the mRNA expression of STAR, aromatase and CYP17 was blocked by Raf-ERK1/2 signalling inhibition, which oppositely induced the mRNA expression of CYP11. Together, these findings suggested that Raf-ERK1/2 signalling pathways mediate steroid hormone synthesis via affecting the expression of steroidogenic enzymes.

SIRT2 plays a novel role on progesterone, estradiol and testosterone synthesis via PPARs/LXRα pathways in bovine ovarian granular cells.

SIRT2 has been shown to possess NAD+-dependent deacetylase and desuccinylase enzymatic activities, it also regulates metabolism homeostasis in mammals. Previous data has suggested that resveratrol, a potential activator of Sirtuins, played a stimulation role in steroidogenesis. Unfortunately, to date, the physiological roles of SIRT2 in ovarian granular cells (GCs) are largely unknown. Here, we studied the function and molecular mechanisms of SIRT2 on steroid hormone synthesis in GCs from Qinchuan cattle. Immunohistochemistry and western blotting showed that SIRT2 was expressed not only in GCs and cumulus cells, but also in oocytes and theca cells. We found that the secretion of progesterone was induced, whereas that of estrogen and testosterone secretion was suppressed by treatment with the SIRT2 inhibitor (Thiomyristoyl or SirReal2) or siRNA. Additionally, the PPARs/LXRα signaling pathways were suppressed by SIRT2 siRNA or inhibitors. The mRNA expression of CYP17, aromatase and StAR was suppressed, but the abundance of CYP11A1 mRNA was induced by SIRT2 inhibition. Furthermore, the PPARα agonist or PPARγ antagonist could mimic the effects of SIRT2 inhibition on hormones levels and gene expression associated with steroid hormone biosynthesis. In turn, those effects were abolished by the LXRα agonist (LXR-623). Together, these data support the hypothesis that SIRT2 regulates steroid hormone synthesis via the PPARs/LXRα pathways in GCs.

Comparison of Pathogenic Mechanisms Underlying Single and Recurrent Venous Thromboembolism Based on Gene Expression Profiling.

BACKGROUND: Unprovoked venous thromboembolism (VTE), generally divided into single and recurrent categories, is a common leading cause of morbidity and mortality in a real-world population. This study was aimed to explore the similarities and differences in the mechanisms of single and recurrent VTE. METHODS: Gene expression data (GSE19151) generated from 63 healthy controls, 32 single, and 38 recurrent VTE patients were analyzed. Differentially expressed genes (DEGs) were screened by Affy package and Kyoto Encyclopedia of Genes and Genomes and Gene Ontology enrichment analysis were performed using database for annotation, visualization, and integrated discovery. Based on the Search Tool for the Retrieval of Interacting Genes/Proteins database, protein-protein interaction network was visualized by Cytoscape, and modules were identified by CFinder. Finally, transcription factor regulatory networks were constructed. RESULTS: Totally, 559 and 294 DEGs were obtained from recurrent and single VTE, respectively. There were 202 upregulated and 58 downregulated genes overlapped between them. Terms of regulation of actin cytoskeleton enriched by downregulated genes and oxidative phosphorylation enriched by upregulated genes were found in 2 types of VTE. Leukocyte transendothelial migration and Jak-STAT signaling pathway were found related with recurrent VTE. In addition, genes including signal transducer and activator of transcription 3 (STAT3) involving in the Jak-STAT signaling pathway were highly connected nodes. CONCLUSIONS: Actin cytoskeleton and oxidative phosphorylation may be involved in the common mechanisms of recurrent VTE and single VTE. Leukocyte migration and Jak-STAT signaling pathway and their related genes may be important for the development and recurrence of VTE.

[The prediction and validation of liver fibrosis by a noninvasive model and validation in patients with chronic hepatitis B].

OBJECTIVE: To develop a simple model for the noninvasive diagnosis of liver fibrosis in patients with chronic hepatitis B and to testify its diagnostic value. METHODS: One hundred and ninety patients with chronic hepatitis B who had undergone liver biopsy were divided into 2 groups: one for developing the model (n = 110) and one for validation (n= 80). Histological staging of liver fibrosis, assessed blindly and independently by 2 pathologists, was determined according to Scheuer fibrosis score. Twenty markers involved in the study were analyzed initially in the estimation group to derive a predictive model to discriminate the stages of fibrosis. The model created was then assessed with receiver operating characteristic curve (ROC) analysis. It was also applied to the validation group to test its accuracy. RESULTS: Haptoglobin (HPT), gamma-glutamyl transpeptidase (GGT) and platelet were identified by logistic regression analysis as independent factors of fibrosis. A model developed from the above three markers was established to predict the stage of fibrosis(S). In ROC analysis, the area under curve (AUC) for identifying S > or =1, S > or = 2, S > or = 3 and S =4 was 0.832, 0.835, 0.820 and 0.843 respectively. The model had a similar AUC in the validation group without statistically significant difference. Using a cut-off of <0. 18, significant fibrosis (S > or = 2) could be excluded in 27 patients of the total patient population (negative predictive value 90%). Similarly, applying a cut-off > or = 0.70, significant fibrosis could be identified correctly in 67 patients of the total patient population (positive predictive value 82.7%). The model had a high level of diagnostic value in patients with HBeAg-positive chronic hepatitis B as well as in patients with HBeAg-negative chronic hepatitis B (AUC for identifying S > or = 2, 0.857 vs 0.802). Restricting biopsy to patients with intermediate scores ( > or = 0.70 and <0.18) may prevent liver biopsies in 58.4% of the patients while maintaining 84.7% accuracy. CONCLUSIONS: A model including HPT, GGT and platelet is a simple and reliable index for predicting significant fibrosis in patients with HBeAg-positive chronic hepatitis B as well as in patients with HBeAg-negative chronic hepatitis B.