Publications by authors named "Yaping Jin"

144 Publications

Glyphosate exposure attenuates testosterone synthesis via NR1D1 inhibition of StAR expression in mouse Leydig cells.

Sci Total Environ 2021 Sep 25;785:147323. Epub 2021 Apr 25.

Northwest A&F University, Yangling 712100, Shaanxi, China; Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling 712100, Shaanxi, China; Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Northwest A&F University, Yangling 712100, Shaanxi, China. Electronic address:

Glyphosate is a broad-spectrum herbicide that impairs testosterone synthesis in mammals. Leydig cells (LCs), the primary producers of testosterone, demonstrate rhythmic expression of circadian clock genes both in vivo and in vitro. The nuclear receptor NR1D1 is an important clock component that constitutes the subsidiary transcriptional/translational loop in the circadian clock system. Nr1d1 deficiency resulted in diminished fertility in both male and female mice. However, whether NR1D1 is involved in the glyphosate-mediated inhibition of testosterone synthesis in LCs remains unclear. Here, the involvement of NR1D1 in glyphosate-mediated inhibition of testosterone synthesis was investigated both in vitro and in vivo. Glyphosate exposure of TM3 cells significantly increased Nr1d1 mRNA levels, but decreased Bmal1, Per2, StAR, Cyp11a1, and Cyp17a1 mRNA levels. Western blotting confirmed elevated NR1D1 and reduced StAR protein levels following glyphosate exposure. Glyphosate exposure also reduced testosterone production in TM3 cells. In primary LCs, glyphosate exposure also upregulated Nr1d1 mRNA levels and downregulated the mRNA levels of other clock genes (Bmal1 and Per2) and steroidogenic genes (StAR, Cyp17a1, Cyp11a1, and Hsd3b2), and inhibited testosterone synthesis. Moreover, glyphosate exposure significantly reduced the amplitude and shortened the period of PER2::LUCIFERASE oscillations in primary LCs isolated from mPer2 knock-in mice. Four weeks of oral glyphosate upregulated NR1D1 at both the mRNA and protein levels in mouse testes, and this was accompanied by a reduction in StAR expression. Notably, serum testosterone levels were also drastically reduced in mice treated with glyphosate. Moreover, dual-luciferase reporter and EMSA assays revealed that in TM3 cells NR1D1 inhibits the expression of StAR by binding to a canonical RORE element present within its promoter. Together, these data demonstrate that glyphosate perturbs testosterone synthesis via NR1D1 mediated inhibition of StAR expression in mouse LCs. These findings extend our understanding of how glyphosate impairs male fertility.
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http://dx.doi.org/10.1016/j.scitotenv.2021.147323DOI Listing
September 2021

Ufmylation regulates granulosa cell apoptosis via ER stress but not oxidative stress during goat follicular atresia.

Theriogenology 2021 Jul 22;169:47-55. Epub 2021 Apr 22.

College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, 712100, China; Key Laboratory of Animal Biotechnology, Ministry of Agriculture and Rural Affairs, Northwest A&F University, Yangling, Shaanxi, 712100, China. Electronic address:

Follicular atresia is primarily caused by granulosa cell (GC) apoptosis, although the mechanisms are largely unknown. Ufmylation is a recently identified ubiquitin-like post-translational modifier that plays an important role in cell proliferation and apoptosis. The purpose of this study was to investigate the effects of Ufmylation on GC apoptosis during goat follicular atresia. Ubiquitin-fold modifier 1 (UFM1) and its target DDRGK domain containing 1 (DDRGK1) proteins were identified in granulosa cells (GCs) isolated from all stages of preantral follicles and from healthy (HF), early atretic (EF) and progressed atretic (PF) antral follicles. The expression levels were higher in GCs derived from antral atretic follicles than healthy follicles. Although the viability of GCs was not affected after overexpression of UFM1, siRNA-mediated UFM1 silencing significantly inhibited GC proliferation and induced apoptosis. Notably, components of the ufmylation pathway were significantly upregulated in GCs induced by the ER stress agent tunicamycin (Tm) and thapsigargin (Tg), but not affected by oxidative stress inducer HO. Furthermore, UFM1 silencing markedly increased the apoptosis of GCs upon Tg treatment by stimulating the ER stress-related gene expression. Our results provide evidence that UFM1 and its target DDRGK1 are expressed in the goat GCs during follicular development and atresia, and ufmylation may play an important role in the prevention of ER stress but not oxidative stress-induced GCs apoptosis.
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http://dx.doi.org/10.1016/j.theriogenology.2021.04.009DOI Listing
July 2021

Low-dose bisphenol A impairs the function of mouse decidual stromal cells by activating LUMAN-mediated unfolded protein response.

Food Chem Toxicol 2021 Apr 27;153:112242. Epub 2021 Apr 27.

College of Food Science and Engineering, Northwest A&F University, Yangling, Shaanxi, China. Electronic address:

The nonsteroidal estrogenic compound bisphenol A (BPA) is widely present in several industrial and medical products including plastic food containers and sealants in dentistry. There are growing concerns on the toxic effects of this compounds since BPA is known to have reproductive toxicity. This study evaluated the effects of low-dose BPA exposure on decidual stromal cells (DSCs) of mice. The results showed that although 10 nM of BPA have no significant effect on the cell viability, it alters the expression of decidualization-related genes including Prl8a2, Prl3c1, Ptgs2, and Mmp2. Moreover, we found that low-dose BPA exposure induces UPR response in DSCs. However, the expression of the three major UPR receptors (Perk, Ire 1, and Xbp1) did not change significantly. Interestingly, the expression of Luman, a novel receptor of UPR, was significantly upregulated in a dose-dependent manner. Lentivirus containing shLuman sequence was used to generate stable Luman silencing DSCs. It's showed that Luman knockdown could affect the expression of decidualization-related genes in decidual cells after BPA treatment. In summary, these results suggest that Luman plays a key role in low dose BPA-induced decidual toxicity of DSCs in mouse.
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http://dx.doi.org/10.1016/j.fct.2021.112242DOI Listing
April 2021

RNA-Seq Analysis Reveals the Role of Omp16 in -Infected RAW264.7 Cells.

Front Vet Sci 2021 4;8:646839. Epub 2021 Mar 4.

College of Veterinary Medicine, Northwest A&F University, Yangling, China.

Brucellosis is an endemic zoonotic infectious disease in the majority of developing countries, which causes huge economic losses. As immunogenic and protective antigens at the surface of spp., outer membrane proteins (Omps) are particularly attractive for developing vaccine and could have more relevant role in host-pathogen interactions. Omp16, a homolog to peptidoglycan-associated lipoproteins (Pals), is essential for survival . At present, the functions of Omp16 have been poorly studied. Here, the gene expression profile of RAW264.7 cells infected with vaccine strain 2 ( S2) and ΔOmp16 was analyzed by RNA-seq to investigate the cellular response immediately after entry. The RNA-sequence analysis revealed that a total of 303 genes were significantly regulated by S2 24 h post-infection. Of these, 273 differentially expressed genes (DEGs) were upregulated, and 30 DEGs were downregulated. These DEGs were mainly involved in innate immune signaling pathways, including pattern recognition receptors (PRRs), proinflammatory cytokines, and chemokines by Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. In ΔOmp16-infected cells, the expression of 52 total cells genes was significantly upregulated and that of 9 total cells genes were downregulated compared to S2-infected RAW264.7 cells. The KEGG pathway analysis showed that several upregulated genes were proinflammatory cytokines and chemokines, such as interleukin (IL)-6, IL-11, IL-12β, C-C motif chemokine (CCL2), and CCL22. All together, we clearly demonstrate that ΔOmp16 can alter macrophage immune-related pathways to increase proinflammatory cytokines and chemokines, which provide insights into illuminating the pathogenic strategies.
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http://dx.doi.org/10.3389/fvets.2021.646839DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7970042PMC
March 2021

FLCN regulates transferrin receptor 1 transport and iron homeostasis.

J Biol Chem 2021 Feb 17:100426. Epub 2021 Feb 17.

College of Veterinary Medicine, Northwest A&F University, Yangling, Shanxi, China, 712100. Electronic address:

Birt-Hogg-Dubé (BHD) syndrome is a multiorgan disorder caused by inactivation of the folliculin (FLCN) protein. Previously, we identified FLCN as a binding protein of Rab11A, a key regulator of the endocytic recycling pathway. This finding implies that the abnormal localization of specific proteins whose transport requires the FLCN-Rab11A complex may contribute to BHD. Here, we used human kidney-derived HEK293 cells as a model, and we report that FLCN promotes the binding of Rab11A with transferrin receptor 1 (TfR1), which is required for iron uptake through continuous trafficking between the cell surface and the cytoplasm. Loss of FLCN attenuated the Rab11A-TfR1 interaction, resulting in delayed recycling transport of TfR1. This delay caused an iron deficiency condition that induced hypoxia-inducible factor (HIF) activity, which was reversed by iron supplementation. In a Drosophila model of BHD syndrome, we further demonstrated that the phenotype of BHD mutant larvae was substantially rescued by an iron-rich diet. These findings reveal a conserved function of FLCN in iron metabolism and may help to elucidate the mechanisms driving BHD syndrome.
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http://dx.doi.org/10.1016/j.jbc.2021.100426DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7995610PMC
February 2021

Circadian clock gene BMAL1 controls testosterone production by regulating steroidogenesis-related gene transcription in goat Leydig cells.

J Cell Physiol 2021 Feb 17. Epub 2021 Feb 17.

Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.

Testosterone is produced by Leydig cells (LCs) and undergoes diurnal changes in serum levels in rats, mice, and humans, but little is known in goats. The present study revealed that goat serum testosterone levels displayed diurnal rhythmic changes (peak time at ZT11.2). Immunohistochemical staining showed that BMAL1, a circadian clock protein, is highly expressed in goat LCs. ELISA revealed that both hCG (0-5 IU/ml) and 22R-OH-cholesterol (0-30 μM) addition stimulated testosterone synthesis in primary goat LCs in a dose-dependent manner. Treating goat LCs with hCG (5 IU/ml) significantly increased intracellular cAMP levels. Additionally, real-time quantitative polymerase chain reaction (PCR) analysis revealed that the circadian clock (BMAL1, PER1, PER2, DBP, and NR1D1) and steroidogenesis-related genes (SF1, NUR77, StAR, HSD3B2, CYP17A1, CYP11A1, and HSD17B3) showed rhythmic expression patterns in goat LCs following dexamethasone synchronization. Several Bmal1-Luc circadian oscillations were clearly observed in dexamethasone-treated goat LCs transfected with the pLV6-Bmal1-Luc plasmid. BMAL1 knockdown significantly downregulated mRNA levels of PER2, NR1D1, DBP, StAR, HSD3B2, SF1, NUR77, and GATA4, and dramatically decreased StAR and HSD3B2 protein levels and testosterone production. In contrast, BMAL1 overexpression significantly increased the mRNA and protein expression levels of StAR and HSD17B3 and enhanced testosterone production. Reporter assays revealed that goat BMAL1, or in combination with mouse CLOCK, activated goat HSD17B3 transcription in vitro. These data indicate that BMAL1 contributes to testosterone production by regulating transcription of steroidogenesis-related genes in goat LCs, providing a basis for further exploring the underlying mechanism by which the circadian clock regulates ruminant reproductive capability.
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http://dx.doi.org/10.1002/jcp.30334DOI Listing
February 2021

Integrated Proteomic and Transcriptomic Analyses Reveal the Roles of Homolog of BAX Inhibitor 1 in Cell Division and Membrane Homeostasis of S2.

Front Microbiol 2021 28;12:632095. Epub 2021 Jan 28.

College of Veterinary Medicine, Northwest A&F University, Yangling, China.

BAX inhibitor 1 (BI-1) is an evolutionarily conserved transmembrane protein first identified in a screening process for human proteins that suppress BAX-induced apoptosis in yeast cells. Eukaryotic BI-1 is a cytoprotective protein that suppresses cell death induced by multiple stimuli in eukaryotes. , the causative agent of brucellosis that threatens public health and animal husbandry, contains a conserved gene that encodes BI-1-like protein. To explore the role of the homolog of BI-1, BrBI, in S2, we constructed the deletion mutant strain and its complemented strain. deletion altered the membrane properties of S2 and decreased its resistance to acidic pH, HO, polymyxin B, and lincomycin. Additionally, deleting led to defective growth, cell division, and viability in S2. We then revealed the effect of deletion on the physiological characteristics of S2 via integrated transcriptomic and proteomic analyses. The integrated analysis showed that deletion significantly affected the expression of multiple genes at the mRNA and/or protein levels. Specifically, the affected divisome proteins, FtsB, FtsI, FtsL, and FtsQ, may be the molecular basis of the impaired cell division of the mutant strain, and the extensively affected membrane proteins and transporter-associated proteins were consistent with the phenotype of the membrane properties' alterations of the mutant strain. In conclusion, our results revealed that BrBI is a bacterial cytoprotective protein involved in membrane homeostasis, cell division, and stress resistance in S2.
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http://dx.doi.org/10.3389/fmicb.2021.632095DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7876416PMC
January 2021

promotes prostaglandin E synthesis by upregulating transcription in response to increasing estradiol levels in pregnant mice.

Am J Physiol Endocrinol Metab 2021 04 8;320(4):E747-E759. Epub 2021 Feb 8.

Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, China.

Prostaglandin G/H synthase 2 (PTGS2) is a rate-limiting enzyme in prostaglandin synthesis. The present study assessed the role of the uterine circadian clock on transcription in response to steroid hormones during early pregnancy. We demonstrated that the core clock genes (, , , and ), , and and their encoded proteins, have rhythmic expression in the mouse uterus from to () of pregnancy. Progesterone (P) treatment of cultured uterus endometrial stromal cells (UESCs) isolated from reporter gene knock-in mice on D4 induced a phase shift in oscillations. This P-induced phase shift of oscillations was significantly attenuated by the P antagonist RU486. Additionally, the amplitude of oscillations was increased by estradiol (E) treatment in the presence of P. Consistently, the mRNA levels of clock genes ( and ), , and were markedly increased by E treatment of UESCs in the presence of P. Treatment with E also promoted prostaglandin E (PGE) synthesis by UESCs. Depletion of in UESCs by small-interfering RNA (siRNA) decreased the transcript levels of clock genes ( and ), , and compared with nonsilencing siRNA treatment. knockdown also inhibited PGE synthesis. Moreover, the mRNA expression levels of clock genes ( and ), , and , and their respective proteins were significantly decreased in the uterus of mice. Thus, these data suggest that in mice promotes PGE synthesis by upregulating in response to increases in E on D4 of pregnancy. Rhythmic expression of Bmal1 and Ptgs2 was observed in the uterus isolated from of pregnant mice. E increased the expression of Bmal1 and Ptg2 in UESCs isolated from mice on D4. The expression of Ptgs2 was significantly decreased in Bmal1-siRNA treated UESCs. knockdown also inhibited PGE synthesis. Thus, these data suggest that Bmal1 in mice promotes PGE synthesis by upregulating Ptgs2 in response to increases in E on D4 of pregnancy.
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http://dx.doi.org/10.1152/ajpendo.00466.2020DOI Listing
April 2021

Trueperella pyogenes pyolysin inhibits lipopolysaccharide-induced inflammatory response in endometrium stromal cells via autophagy- and ATF6-dependent mechanism.

Braz J Microbiol 2021 Jun 16;52(2):939-952. Epub 2021 Jan 16.

Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, College of Veterinary Medicine, Northwest A&F University, Yangling, 712100, Shaanxi, China.

Trueperella pyogenes (T. pyogenes) is a common opportunistic pathogen of many livestock and play an important regulation role during multibacterial infection and interaction with the host by its primary virulence factor pyolysin (PLO). The purpose of this study was to investigate the regulation role of PLO which serve as a combinational pathogen with lipopolysaccharide (LPS) during endometritis. In this study, the expression of bioactive recombinant PLO (rPLO) in a prokaryotic expression system and its purification are described. Moreover, we observed that rPLO inhibited the innate immune response triggered by LPS and that methyl-β-cyclodextrin (MBCD) abrogated this inhibitory effect in goat endometrium stromal cells (gESCs). Additionally, we show from pharmacological and genetic studies that rPLO-induced autophagy represses gene expression by inhibiting NLRP3 inflammasome activation. Importantly, this study reported that ATF6 serves as a primary regulator of the cellular inflammatory reaction to rPLO. Overall, these observations suggest that T. pyogenes PLO could create an immunosuppressive environment for other pathogens invasion by regulating cellular signaling pathways.
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http://dx.doi.org/10.1007/s42770-021-00422-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8105434PMC
June 2021

Luman/CREB3 knock-down inhibit hCG induced MLTC-1 apoptosis.

Theriogenology 2021 Feb 3;161:140-150. Epub 2020 Dec 3.

Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, 712100, China; Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Northwest A&F University, Yangling, Shaanxi, 12100, China. Electronic address:

Luman has been reported to be involved in the formation of COP II-mediated transport vesicles that affect protein transportation and secretion. Western blotting, immunohistochemistry, immunofluorescence, and RT-qPCR indicated that Luman is widely expressed in the male mouse reproductive system. In sperm, Luman was mainly located in the sperm tail, and the expression level increased with sperm maturity. In the testis, Luman was located in Leydig cells. In MLTC-1, a high-concentration hCG treatment significantly increased GRP78, ATF6, p-IRE1, and p-EIF2S1 expression but had no effect on Luman expression. To investigate the role of Luman in hCG-induced ER stress (ERS), experiments were conducted to examine the consequences of short hairpin RNA (shRNA)-mediated Luman knockdown in MLTC-1 cells. Luman knockdown decreased the percentage of S phase cells and up-regulated Cyclin A1, Cyclin B1, and Cyclin D2 expression. ELISA and WB results showed that with Luman knockdown, Cyp11a1, p-IRE1, and p-EIF2S1 expression and testosterone secretion were significantly increased, while GRP78 and CHOP expression were decreased. Flow cytometry results showed that Luman knockdown reduced MLTC-1 cell apoptosis. RT-qPCR and WB results showed that Luman knockdown significantly up-regulated BCL-2 expression and decreased Caspase-3 and BAX expression. These data suggest that Luman is widely expressed in the male mouse reproductive system. In MLTC-1 cells, Luman knockdown up-regulated p-IRE1, p-EIF2S1, and BCL-2 expression and decreased GRP78, CHOP, BAX, and Caspase-3 expression. We propose that Luman knockdown reduces cell apoptosis through the ERS pathway, thereby promoting cell survival and testosterone secretion. These findings provide new insights into the role of Luman in hCG-induced ERS.
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http://dx.doi.org/10.1016/j.theriogenology.2020.11.010DOI Listing
February 2021

Bisphenol A attenuates testosterone production in Leydig cells via the inhibition of NR1D1 signaling.

Chemosphere 2021 Jan 23;263:128020. Epub 2020 Aug 23.

Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, 712100, Shaanxi, China; Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Northwest A&F University, Yangling, 712100, Shaanxi, China. Electronic address:

Bisphenol A (BPA) is an endocrine-disrupting compound that impairs testosterone synthesis in male mammals. A circadian clock gene deficiency leads to diminished fertility and even infertility in male mice. However, whether circadian clock signaling pathways mediate the suppressive effect of BPA on testosterone synthesis in Leydig cells (LCs) remains unknown. The present study aims to detect the effect of BPA on cellular circadian clock and testosterone synthesis in mouse LCs, and examine the mechanisms underlying NR1D1 signaling. BPA treatment significantly attenuated the transcription levels of Nr1d1 and steroidogenic genes (Hsd3b2 and Hsd17b3) in TM3 cells, but increased other circadian clock gene levels (Per2 and Dbp). BPA treatment also significantly downregulated NR1D1 and StAR protein expression, but upregulated BMAL1 protein expression in TM3 cells. Furthermore, there was a marked decline in testosterone production in BPA-treated TM3 cells. Intraperitoneal injection of BPA profoundly reduced NR1D1 and StAR protein levels and steroidogenic gene transcription levels (Cyp11a1, Hsd3b2, and Hsd17b3), while enhancing BMAL1 protein and other circadian clock gene (Per2 and Dbp) levels in mouse testes. Notably, serum testosterone levels were also drastically reduced in BPA-treated mice. Moreover, SR9009, an NR1D1 agonist, augmented testosterone production in TM3 cells via elevated expression of steroidogenic genes (StAR, Cyp11a1 and Hsd17b3). Conversely, Nr1d1 knockdown inhibited testosterone accumulation and attenuated steroidogenic gene expression. Moreover, treatment with SR9009 partially reversed the BPA effect on the circadian clock and testosterone production. Taken together, our study demonstrates that BPA perturbs testosterone production, at least partially, via inhibiting NR1D1 signaling in LCs.
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http://dx.doi.org/10.1016/j.chemosphere.2020.128020DOI Listing
January 2021

Zearalenone perturbs the circadian clock and inhibits testosterone synthesis in mouse Leydig cells.

J Toxicol Environ Health A 2021 Feb 4;84(3):112-124. Epub 2020 Nov 4.

Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University , Yangling, China.

Zearalenone (ZEA), a mycotoxin, is known to impair reproductive capability by disrupting the synthesis and secretion of testosterone by Leydig cells (LCs), although the mechanism is unknown. Robust rhythmicity of circadian clock and steroidogenic genes were identified in LCs. The aim of this study was to examine whether ZEA significantly attenuated the transcription of core clock genes (, and ) as well as steroidogenic genes (, and ) in mouse testis Leydig cell line (TM3). Western blotting confirmed declines in BMAL1, NR1D1, and StAR protein levels. ZEA also suppressed secreted testosterone levels. In primary LCs, isolated from PER2::LUCIFERASE reporter gene knock in mice, ZEA diminished the amplitude of expression, and induced a phase shift and period extension. In primary LCs, ZEA also suppressed the expression levels of core clock and steroidogenic genes, reduced protein levels of BMAL1, and decreased testosterone secretion. expression of core clock and steroidogenic genes were reduced in testes of mice exposed to ZEA for 1 week leading to decreased serum testosterone levels. In summary, data suggest that ZEA may impair testosterone synthesis through attenuation of the circadian clock in LCs culminating in reproductive dysfunction in male mammals .
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http://dx.doi.org/10.1080/15287394.2020.1841699DOI Listing
February 2021

Prostaglandin F2α Induces Goat Corpus Luteum Regression via Endoplasmic Reticulum Stress and Autophagy.

Front Physiol 2020 11;11:868. Epub 2020 Sep 11.

Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, College of Veterinary Medicine, Northwest A&F University, Yangling, China.

Corpus luteum (CL) is a transient endocrine tissue that produces progesterone for maintaining pregnancy in mammals. In addition, the regression of CL is necessary for the initiation of the estrous cycle. Extensive research has shown that the prostaglandin F2α (PGF2α) induces the regression of CL in ruminants. However, the mechanisms of endoplasmic reticulum (ER) stress and autophagy in the regression of goat CL induced by PGF2α are still unclear. In this study, ovaries of dioestrus goats and goats that were 3 months pregnant were collected to detect the location of the ER stress-related protein GRP78. The relationship between the different stages of the luteal phase of goat CL during the estrous cycle and changes in the expression of ER stress-related proteins and autophagy-related proteins was confirmed by western blot analysis. The results showed that both ER stress and autophagy were activated in the late luteal phase of the goat CL. To reveal the function of ER stress and autophagy in the CL regression process induced by PGF2α, we used 4-phenyl butyric acid (4-PBA) and chloroquine (CQ) for inhibiting ER stress and autophagy, respectively. Through the apoptotic rate detected by the flow cytometry and the expression of ER stress- and autophagy-related proteins detected by western blotting, we demonstrated that ER stress promoted goat luteal cell apoptosis and autophagy, and that apoptosis can be enhanced by the inhibition of autophagy. In addition, knockdown of EIF2S1, which blocked the PERK pathway activation, promoted apoptosis by reducing autophagy in goat luteal cells treated with PGF2α. In conclusion, our study indicates that ER stress promotes goat luteal cell apoptosis to regulate the regression of CL and activates autophagy to inhibit the goat luteal cell apoptosis via PERK signaling pathway.
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http://dx.doi.org/10.3389/fphys.2020.00868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7516216PMC
September 2020

UFMylation is associated with LPS-induced inflammatory response in goat endometrial epithelial cells.

Reprod Domest Anim 2020 Dec 27;55(12):1725-1734. Epub 2020 Oct 27.

College of Veterinary Medicine, Northwest A&F University, Yangling, China.

The endometrium plays an important role in the defence against invading pathogens, although the mechanisms are not clear. UFMylation is a recently discovered novel ubiquitination-like modification system that plays a pivotal role in inflammation and the immune response. The purpose of this study was to investigate the effects of UFMylation on lipopolysaccharide (LPS)-induced inflammatory responses in immortalized goat endometrial epithelial cells (gEECs). Ubiquitin-fold modifier conjugating enzyme 1 (UFM1) and DDRGK domain containing 1 (DDRGK1) were mainly localized in the luminal epithelium and glandular epithelium of mouse and goat endometrial tissues. The expression levels of UFM1, ubiquitin-like modifier activating enzyme 5 (UBA5), UFM1 specific ligase 1 (UFL1) and DDRGK1, as key components of the UFMylation system, were significantly activated by 5 μg/mL LPS-induced inflammatory response in gEECs for 6 hr. Meanwhile, the expression levels of interleukin-6 (IL-6) were significantly upregulated, and tumour necrosis factor-α (TNF-α) was significantly down-regulated after overexpression of UFM1 in gEECs. Additionally, we observed UFM1 and DDRGK1 were markedly increased on LPS-stimulated mouse endometritis in vivo. In conclusion, the current study demonstrated that UFMylation was significantly activated by LPS and might be involved in regulating inflammatory response in gEECs.
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http://dx.doi.org/10.1111/rda.13832DOI Listing
December 2020

Bta-miR-34b inhibits proliferation and promotes apoptosis via the MEK/ERK pathway by targeting MAP2K1 in bovine primary Sertoli cells.

J Anim Sci 2020 Oct;98(10)

Department of Clinical Veterinary Medicine, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.

Immature Sertoli cell (SC) proliferation determines the final number of mature SCs and further regulates spermatogenesis. Accumulating evidence demonstrated that microRNAs (miRNAs) play an important role in SC proliferation, differentiation, and apoptosis. However, the effect and molecular mechanism of miRNA on bovine immature SC remain to be poorly understood. In this study, miRNA sequencing of testes collected in mature (24-mo old) and immature (neonatal) bulls was conducted to determine the miRNA expression profiles. MicroRNA-34b was one of the differentially expressed miRNAs and was selected for in-depth functional studies pertaining to SC growth. The results showed that miR-34b mimic transfection in primary Sertoli cells (PSC) inhibited cell proliferation and induced cell cycle arrested at G2 phase and decreased the expression of cell cycle-related genes such as CCNB1, CDK1, CDC25C, and C-MYC. MicroRNA-34b overexpression also leads to increased cell apoptosis, with proapoptotic genes P53 and BAX upregulated, while antiapoptotic gene BCL2 decreased. However, miR-34b knockdown had the opposite effects. Through a combination of transcriptome sequencing, bioinformatics analysis, dual-luciferase reporter assay, and Western blotting, mitogen-activated protein kinase kinase1 (MAP2K1), also known as MEK1, was identified as a target of miR-34b. In addition, PSC proliferation inhibition was mediated by cell cycle arrest and apoptosis with MAP2K1 interference. Overexpression of MAP2K1 effectively reversed the miR-34b-repressed PSC cell growth. Moreover, both miR-34b overexpression and MAP2K1 knockdown decreased the protein levels of P-ERK1/2, while MAP2K1 overexpression showed opposite effects. In summary, data suggest that miR-34b regulates PSC proliferation and apoptosis through the MEK/ERK signaling pathway. These data provide a theoretical and experimental framework for further clarifying the regulation of cell growth in PSC of bovine.
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http://dx.doi.org/10.1093/jas/skaa313DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7598574PMC
October 2020

Omp16, a conserved peptidoglycan-associated lipoprotein, is involved in Brucella virulence in vitro.

J Microbiol 2020 Sep 1;58(9):793-804. Epub 2020 Sep 1.

College of Veterinary Medicine, Northwest A&F University, Yangling, P. R. China.

Brucella, the bacterial agent of common zoonotic brucellosis, primarily infects specific animal species. The Brucella outer membrane proteins (Omps) are particularly attractive for developing vaccine and improving diagnostic tests and are associated with the virulence of smooth Brucella strains. Omp16 is a homologue to peptidoglycan-associated lipoproteins (Pals), and an omp16 mutant has not been generated in any Brucella strain until now. Very little is known about the functions and pathogenic mechanisms of Omp16 in Brucella. Here, we confirmed that Omp16 has a conserved Pal domain and is highly conserved in Brucella. We attempted to delete omp16 in Brucella suis vaccine strain 2 (B. suis S2) without success, which shows that Omp16 is vital for Brucella survival. We acquired a B. suis S2 Omp16 mutant via conditional complementation. Omp16 deficiency impaired Brucella outer membrane integrity and activity in vitro. Moreover, inactivation of Omp16 decreased bacterial intracellular survival in macrophage RAW 264.7 cells. B. suis S2 and its derivatives induced marked expression of IL-1β, IL-6, and TNF-a mRNA in Raw 264.7 cells. Whereas inactivation of Omp16 in Brucella enhanced IL-1β and IL-6 expression in Raw 264.7 cells. Altogether, these findings show that the Brucella Omp16 mutant was obtained via conditional complementation and confirmed that Omp16 can maintain outer membrane integrity and be involved in bacterial virulence in Brucella in vitro and in vivo. These results will be important in uncovering the pathogenic mechanisms of Brucella.
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http://dx.doi.org/10.1007/s12275-020-0144-yDOI Listing
September 2020

Serological survey of SARS-CoV-2 for experimental, domestic, companion and wild animals excludes intermediate hosts of 35 different species of animals.

Transbound Emerg Dis 2020 Jul 7;67(4):1745-1749. Epub 2020 May 7.

National Research Center for Veterinary Medicine, Luoyang, China.

The pandemic SARS-CoV-2 has been reported in 123 countries with more than 5,000 patients died from it. However, the original and intermediate hosts of the virus remain unknown. In this study, 1,914 serum samples from 35 animal species were used for detection of SARS-CoV-2-specific antibodies using double-antigen sandwich ELISA after validating its specificity and sensitivity. The results showed that no SARS-CoV-2-specific antibodies were detected in above samples which excluded the possibility of 35 animal species as intermediate host for SARS-CoV-2. More importantly, companion animals including pet dogs (including one dog the SARS-CoV-2 patient kept and two dogs which had close contact with it) and cats, street dogs and cats also showed serological negative to SARS-CoV-2, which relieved the public concerns for the pets as SARS-CoV-2 carriers.
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http://dx.doi.org/10.1111/tbed.13577DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7264586PMC
July 2020

Comparison of accelerated CXL alone, accelerated CXL-ICRS, and accelerated CXL-TG-PRK in progressive keratoconus and other corneal ectasias.

J Cataract Refract Surg 2020 02;46(2):276-286

From the Kensington Eye Institute (Singal, Ong Tone, Stein, Bujak, Chan, Chew, El-Defrawy, Jin, Kranemann, Rabinovitch, Rootman, Slomovic, Hatch), Department of Ophthalmology and Vision Sciences, University of Toronto (Singal, Ong Tone, Stein, Bujak, Chan, Chew, El-Defrawy, Jin, Kranemann, Rabinovitch, Rootman, Slomovic, Hatch), Kensington Crosslinking Working Group (Singal, Ong Tone, Stein, Bujak, Chan, Chew, El-Defrawy, Jin, Kranemann, Rabinovitch, Rootman, Slomovic, Hatch), and St. Michael's Hospital (Cohen, Dai), Toronto, Ontario, Canada.

Purpose: To compare accelerated corneal crosslinking (CXL) alone, CXL with simultaneous intrastromal corneal ring segments (CXL-ICRS), and CXL with simultaneous topography-guided photorefractive keratectomy (CXL-TG-PRK) in progressive keratoconus, pellucid marginal degeneration (PMD), or laser in situ keratomileusis (LASIK)-induced ectasia.

Setting: The Kensington Eye Institute and Bochner Eye Institute, Toronto, Canada.

Design: Prospective nonrandomized interventional study.

Methods: Visual and topographical outcomes using a comparative analysis adjusting for preoperative maximum keratometry (Kmax) were evaluated 1 year postoperatively.

Results: Four hundred fifty-two eyes from 375 patients with progressive keratoconus, PMD, or LASIK-induced ectasia that underwent accelerated (9 mW/cm, 10 minutes) CXL alone (n = 204), CXL-ICRS (n = 126), or CXL-TG-PRK (n = 122) were included. Change in logarithm of the minimum angle of resolution uncorrected distance visual acuity was significant with CXL-ICRS (-0.31; 95% CI, -0.38 to -0.24) and CXL-TG-PRK (-0.16; 95% CI, -0.24 to -0.09), but not with CXL alone. No significant differences in change were found between the 3 groups. Change in corrected distance visual acuity (CDVA) was significant in all 3 groups: -0.12 (95% CI, -0.15 to -0.10) with CXL alone, -0.23 (95% CI, -0.27 to -0.20) with CXL-ICRS, and -0.17 (95% CI, -0.21 to -0.13) with CXL-TG-PRK. Improvement in CDVA was greater with CXL-ICRS than with CXL alone (-0.08 ± 0.02; P < .0001) and CXL-TG-PRK (-0.05 ± 0.02; P = .005). Change in Kmax was significant with CXL-ICRS [-3.21 diopters (D); 95% CI, -3.98 to -2.45] and CXL-TG-PRK (-3.69 D; 95% CI, -4.49 to -2.90), but not with CXL alone (-0.05 D; 95% CI, -0.66 to 0.55).

Conclusions: CXL alone might be best for keratoconic patients who meet the inclusion criteria. CXL-ICRS might be more effective for eyes with more irregular astigmatism and worse CDVA and CXL-TG-PRK for eyes requiring improvements in irregular astigmatism but still have good CDVA.
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http://dx.doi.org/10.1097/j.jcrs.0000000000000049DOI Listing
February 2020

Exploring the role of Luman/CREB3 in regulating decidualization of mice endometrial stromal cells by comparative transcriptomics.

BMC Genomics 2020 Jan 30;21(1):103. Epub 2020 Jan 30.

College of Veterinary Medicine, Northwest A&F University, Yangling, 712100, Shaanxi, China.

Background: Luman is a member of CREB3 (cAMP responsive element-binding) subfamily of the basic leucine-zipper (bZIP) transcription factors. It may play an important regulatory role during the decidualization process since Luman was highly expressed in the decidual cells. However, the exact molecular mechanisms of how Luman regulating decidualization is unknown.

Results: Using an in vitro model, we prove that Luman knockdown significantly affects the decidualization process of mice endometrial stromal cells (ESCs) as the expression of two decidual markers PRL8a2 and PRL3c1 were repressed. We employed massively parallel RNA sequencing (RNA-Seq) to understand the changes in the transcriptional landscape associated with knockdown of Luman in ESCs during in vitro decidualization. We found significant dysregulation of genes related to protein processing in the endoplasmic reticulum (ER). Several genes involved in decidualization including bone morphogenetic proteins (e.g. BMP1, BMP4, BMP8A, BMP2, and BMP8B), growth factor-related genes (e.g. VEGFB, FGF10, and FGFR2), and transcription factors (IF4E, IF4A2, WNT4, WNT9A, ETS1, NOTCH1, IRX1, IDB1, IDB2, and IDB3), show altered expression. We also found that the knockdown of Luman is associated with increased expression of cell cycle-related genes including cycA1, cycB1, cycB2, CDK1, CDK2, and PLPK1, which resulted in an increased proportion of ESCs in the G1 phase. Differentially expressed genes (DEGs) were highly enriched on ECM-receptor interaction signaling, endoplasmic reticulum protein processing, focal adhesion, and PI3K-Akt signaling pathways.

Conclusions: Luman knockdown results in widespread gene dysregulation during decidualization of ESCs. Genes involved in protein processing in ER, bone morphogenetic protein, growth factor, and cell cycle progression were identified as particularly important for explaining the decidual deficiency observed in this in vitro model. Therefore, this study provides clues as to the underlying mechanisms that may expand our understanding of gene regulation during decidualization.
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http://dx.doi.org/10.1186/s12864-020-6515-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6993373PMC
January 2020

Progesterone-induced RNA Hand2os1 regulates decidualization in mice uteri.

Reproduction 2020 03;159(3):303-314

College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.

Decidualization is a critical process for successful embryo implantation and subsequent placenta formation. The characterization and physiological function of lncRNA during decidualization remain largely unknown. In the present study, we conducted RNA-sequencing analysis to compare gene expression between decidua of days 6 and 8, and normal pregnant endometrium (day 4). A total of 2332 high-confidence putative lncRNA transcripts were expressed. Functional clustering analysis of cis and trans lncRNA targets showed that differentially expressed lncRNAs may regulate multiple gene ontology terms and pathways that have important functions in decidualization. Subsequent analyses using qRT-PCR validated that eight of all lncRNAs were differentially regulated in mice uteri during decidualization, both in vivo and in vitro. Furthermore, we showed that differentially expressed lncRNA of Hand2os1 was specifically detected in stromal cells on days 2 to 5 of pregnancy and was strongly upregulated in decidual cells on days 6-8 of pregnancy. Similarly, Hand2os1 expression was also strongly expressed in decidualized cells following artificial decidualization, both in vivo and in vitro. In uterine stromal cells, P4 was able to significantly upregulate the expression of Hand2os1, but upregulation was impeded by RU486, whereas E2 appeared to have no regulating effect on Hand2os1 expression. Concurrently, Hand2os1 significantly promoted the decidual process in vitro and dramatically increased decidualization markers Prl8a2 and Prl3c1. Our results provide a valuable catalog for better understanding of the functional roles of lncRNAs in pregnant mouse uteri, as it relates to decidualization.
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http://dx.doi.org/10.1530/REP-19-0401DOI Listing
March 2020

Accelerated Corneal Cross-Linking: Efficacy, Risk of Progression, and Characteristics Affecting Outcomes. A Large, Single-Center Prospective Study.

Am J Ophthalmol 2020 05 13;213:76-87. Epub 2020 Jan 13.

Kensington Eye Institute, Toronto, Ontario, Canada; Department of Ophthalmology and Vision Sciences, University of Toronto, Toronto, Ontario, Canada.

Purpose: We examined the efficacy and preoperative characteristics that affect outcomes of accelerated (9 mW/cm for 10 minutes) corneal cross-linking (CXL).

Design: Prospective single-center observational cohort study.

Methods: We enrolled 612 eyes of 391 subjects with progressive keratoconus (n = 589), pellucid marginal degeneration (n = 11), and laser in situ keratomileusis-induced ectasia (n = 12). We evaluated best spectacle-corrected visual acuity (BSCVA), topography, refraction, endothelial cell density, corneal thickness, haze, intraocular pressure, and visual function before and 12 months after the CXL procedure. We tabulated the proportion of those with progression of maximum keratometry (Kmax). We included participant's race, age, sex, and the presence of preoperative apical scarring and environmental allergies in a multivariable linear regression model to determine the effect of these characteristics on outcomes.

Results: At 1 year there was no significant change in mean Kmax (n = 569). Progression of Kmax was higher in subgroups with a baseline Kmax >58 diopters (n = 191) and those 14-18 years of age (n = 53). Preoperative BSCVA, Kmax, refraction, corneal cylinder, coma, central corneal thickness, and vision function were statistically and clinically significant predictors of outcomes (P < .001). Preoperative apical scarring led to worsening haze (P = .0001), more astigmatism (P = .002), more central corneal thinning (P = .002), and was protective to the endothelium (P = .008). Race, age, and sex affected some outcomes.

Conclusion: Mean Kmax was stable at 1 year after accelerated CXL. Younger patients and those with a higher preoperative Kmax need to be monitored closely for progression. Preoperative BSCVA, topography, refraction, CCT, and apical scarring were significant predictors of outcomes.
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http://dx.doi.org/10.1016/j.ajo.2020.01.006DOI Listing
May 2020

Alterations of NMDA and AMPA receptors and their signaling apparatus in the hippocampus of mouse offspring induced by developmental arsenite exposure.

J Toxicol Sci 2019 ;44(11):777-788

Department of Occupational and Environmental Health, School of Public Health, China Medical University, China.

Loss of cognitive function due to arsenic exposure is a serious health concern in many parts of the world, including China. The present study aims to determine the molecular mechanism of arsenic-induced neurotoxicity and its consequent effect on downstream signaling pathways of mouse N-methyl-D-aspartate receptors (NMDARs) and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs). Drinking water containing 0, 25, 50 or 100 mg/L arsenite was provided each day to mother mice throughout gestation period until postnatal day (PND) 35 to expose the newborn mice to arsenite during early developmental period. The effect of arsenite in the expressions of different components of NMDAR (NR1, NR2A, NR2B) and AMPAR (GluR1, GluR2, GluR3), including calcium/calmodulin-dependent protein kinase II (CaMKII) and phosphorylated-CaMKII (p-CaMKII), at PND 7, 14, 21 and 35 was estimated and analyzed from the hippocampus of mice. A significant inhibition in the protein and mRNA expressions of NR1, NR2A, NR2B and GluR1 was observed in mice exposed to 50 mg/L arsenite since PND 7. Down regulation of GluR2 and GluR3 both at mRNA and protein levels was observed in mice exposed to 50 mg/L arsenite till PND 14. Moreover, both CaMKII as well as p-CaMKII expressions were significantly limited since PND 7 in 50 mg/L arsenite exposed mice group. Findings form this study suggested that the previously reported impairment in learning and memorizing abilities in later stage due to early life arsenite exposure is associated with the alterations of NMDARs, AMPARs, CaMKII and p-CaMKII expressions.
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http://dx.doi.org/10.2131/jts.44.777DOI Listing
March 2020

Upregulation of CYP2E1 expression causes oxidative damage induced by 2-chloroethanol in primary cultured rat astrocytes.

Neurotoxicology 2019 12 1;75:233-244. Epub 2019 Oct 1.

Department of Environmental and Occupational Health, School of Public Health, China Medical University, Shenyang, Liaoning, People's Republic of China. Electronic address:

Brain edema caused by subacute poisoning with 1,2-dichloroethane (1,2-DCE) has gained much attention during recent years, but its underlying mechanism is poorly understood. As an intermediate metabolite of 1,2-DCE in vivo, 2-chloroethanol (2-CE) can be transformed into chloroacetaldehyde and reactive oxygen species (ROS) through cytochrome P450 2E1 (CYP2E1) mediated metabolism. In previous studies, it was found that CYP2E1 expression is enhanced in the brain of mice treated with 1,2-DCE. This study was designed to verify the roles of CYP2E1 overexpression in 2-CE induced cytotoxicity in rat astrocytes, and the contribution of specific signaling molecules to the upregulation of CYP2E1 expression caused by 2-CE. The results of this study demonstrate that treatment with 2-CE can enhance CYP2E1 protein and mRNA levels, cause an increase in ROS and MDA levels, and higher percentages of apoptotic cells in rat astrocytes. Pretreatment with either diallyl sulfide or vitamin C, the inhibitor of CYP2E1 or scavenger of ROS, respectively, can suppress the levels of CYP2E1 expression, ROS and MDA, ameliorate cell apoptosis, and attenuate phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) in these cells. Additionally, pretreatment with the inhibitor of either ERK1/2 or transcriptional factor specificity protein 1 (SP1) can suppress the CYP2E1 expression, and alleviate the oxidative damage caused to these cells. In conclusion, our findings demonstrate that CYP2E1 overexpression plays a crucial role in 2-CE induced oxidative damage of rat astrocytes, and that CYP2E1 expression is upregulated partially through the activation of the ERK1/2 and SP1 signaling pathways by ROS generated during CYP2E1-mediated 2-CE metabolism. This study provides novel information that can be used in elucidating the mechanism by which 1,2-DCE induces brain edema.
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http://dx.doi.org/10.1016/j.neuro.2019.09.016DOI Listing
December 2019

Neuroinflammatory Reactions in the Brain of 1,2-DCE-Intoxicated Mice during Brain Edema.

Cells 2019 08 27;8(9). Epub 2019 Aug 27.

Department of Occupational and Environmental Health, School of Public Health, China Medical University, Shenyang 110122, Liaoning, China.

We previously reported that expression of matrix metalloproteinase-9 (MMP-9) mRNA and protein was upregulated during 1,2-dichloroethane (1,2-DCE) induced brain edema in mice. We also found that the p38 mitogen-activated protein kinase (p38 MAPK) signaling pathway resulted in MMP-9 overexpression and nuclear factor-κB (NF-κB) activation in mice treated with 1,2-DCE. In this study, we further hypothesized that inflammatory reactions mediated by the p38 MAPK/ NF-κB signaling pathway might be involved in MMP-9 overexpression, blood-brain barrier (BBB) disruption and edema formation in the brain of 1,2-DCE-intoxicated mice. Our results revealed that subacute poisoning by 1,2-DCE upregulates protein levels of glial fibrillary acidic protein (GFAP), ionized calcium-binding adapter molecule 1 (Iba-1), interleukin-1β (IL-1β), vascular cell adhesion molecule-1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), inducible nitric oxide synthase (iNOS) and p-p65 in mouse brains. Pretreatment with an inhibitor against p38 MAPK attenuates these changes. Moreover, pretreatment with an inhibitor against NF-κB attenuates alterations in brain water content, pathological indications notable in brain edema, as well as mRNA and protein expression on levels of MMP-9, VCAM-1, ICAM-1, iNOS, and IL-1β, tight junction proteins (TJs), GFAP and Iba-1 in the brain of 1,2-DCE-intoxicated mice. Furthermore, pretreatment with an inhibitor against MMP-9 obstructs the decrease of TJs in the brain of 1,2-DCE-intoxicated mice. Lastly, pretreatment with an antagonist against the IL-1β receptor also attenuates changes in protein levels of p-p38 MAPK, p-p65, p-IκB, VCAM -1, ICAM-1, IL-1β, and Iba-1 in the brain of 1,2-DCE-intoxicated-mice. Taken together, findings from the current study indicate that the p38 MAPK/ NF-κB signaling pathway might be involved in the activation of glial cells, and the overproduction of proinflammatory factors, which might induce inflammatory reactions in the brain of 1,2-DCE-intoxicated mice that leads to brain edema.
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http://dx.doi.org/10.3390/cells8090987DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6770564PMC
August 2019

VceC Mediated IRE1 Pathway and Inhibited CHOP-induced Apoptosis to Support Replication in Goat Trophoblast Cells.

Int J Mol Sci 2019 Aug 22;20(17). Epub 2019 Aug 22.

Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, Northwest A&F University, Yangling 712100, China.

The effectors of the type IV secretion system (T4SS) of bacteria play important roles in mediating bacterial intracellular proliferation and manipulating host-related pathway responses to bacterial infection. Spp. inhibit the apoptosis of host cells to benefit their own intracellular proliferation. However, the underlying mechanisms between T4SS effectors and -inhibited apoptosis in goat trophoblast cells remain unclear. Here, based on vaccine strain 2, the VceC was deleted by allelic exchange. We show that ΔVceC was able to infect and proliferate to high titers in goat trophoblast cells (GTCs) and increase C/EBP-homologous protein (CHOP)-mediated apoptosis. GRP78 expression decreased upon ΔVceC infection. In addition, we discovered that the inositolrequiring enzyme 1 (IRE1) pathway was inhibited in this process. Changing endoplasmic reticulum (ER) stress affected intracellular replication in GTCs. The replication of ΔVceC was more sensitive under the different ERstress conditions in the GTC line after treatment with ER stress inhibitors 4 phenyl butyric acid (4-PBA) or ER stress activator Tm. Together, our findings show that VceC has a protective effect on the intracellular persistence of infection, and inhibits ER stress-induced apoptosis in the CHOP pathway. The present work provides new insights for understanding the mechanism of VceC in the establishment of chronic infection.
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http://dx.doi.org/10.3390/ijms20174104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6747397PMC
August 2019

CREBZF regulates testosterone production in mouse Leydig cells.

J Cell Physiol 2019 12 23;234(12):22819-22832. Epub 2019 May 23.

College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.

CREBZF, including the two isoforms SMILE (long isoform of CREBZF) and Zhangfei (short isoform of CREBZF), has been identified as a novel transcriptional coregulator of a variety of nuclear receptors. Our previous studies found that SMILE is expressed in the mouse uterine luminal and glandular epithelium and is upregulated by estrogen. In the present study, CREBZF was age-dependently and -specifically expressed in mouse interstitial Leydig cells during sexual maturation. The expression pattern of CREBZF exhibited an age-related increase, and SMILE was the dominant isoform in the mouse testis. Although hCG did not affect CREBZF expression, CREBZF silencing significantly inhibited hCG-stimulated testosterone production in primary Leydig cells and MLTC-1 cells. Meanwhile, the serum concentration of testosterone was significantly decreased after microinjection of lentiviral-mediated shRNA-CREBZF into the mature mouse testis. In addition, CREBZF silencing markedly decreased P450c17, 17β-HSD, and 3β-HSD expression following hCG stimulation in primary Leydig cells, and this inhibitory effect was obviously reversed by overexpression of CREBZF. Furthermore, CREBZF significantly upregulated the mRNA levels of Nr4a1 and Nr5a1, which are the essential orphan nuclear receptors for steroidogenic gene expression. Together our data indicate that CREBZF promotes hCG-induced testosterone production in mouse Leydig cells by affecting Nr4a1 and Nr5a1 expression levels and subsequently increasing the expression of steroidogenic genes such as 3β-HSD, 17β-HSD, and P450c17, suggesting a potential important role of CREBZF in testicular testosterone synthesis.
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http://dx.doi.org/10.1002/jcp.28846DOI Listing
December 2019

Endoplasmic reticulum stress is involved in lipopolysaccharide-induced inflammatory response and apoptosis in goat endometrial stromal cells.

Mol Reprod Dev 2019 07 1;86(7):908-921. Epub 2019 May 1.

Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.

Endoplasmic reticulum (ER) stress is involved in regulating cell metabolism, apoptosis, autophagy, and survival. However, there is not enough information about the role of ER stress in lipopolysaccharide (LPS)-induced apoptosis and inflammatory cytokine secretion in the uterus. In this study, we found that LPS induced apoptosis and inflammation in goat endometrial stromal cells (ESCs). LPS treatment inhibited cell viability and cell proliferation. In addition, the genes associated with proliferation, such as proliferating cell nuclear antigen and MKI67, were affected by LPS treatment. Moreover, LPS increased the secretion of interleukin (IL)-1β and IL-8, promoting the levels of MYD88, caspase1, and TRL4. The 4-phenylbutyric acid pretreatment inhibited the expression of unfolded protein response proteins and the secretion of inflammatory cytokines in LPS-treated cells. However, blockage of inositol-requiring enzyme 1 and activating transcription factor 6 did not significantly reduce apoptosis and inflammatory cytokine secretion. Collectively, ER stress involved in LPS-induced apoptosis and inflammatory cytokine increased in goat ESCs. This study provides new insight into the function of ER stress in the pathological process.
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http://dx.doi.org/10.1002/mrd.23152DOI Listing
July 2019

Influence of Age, Sex, and Generation on Physician Payments and Clinical Activity in Ontario, Canada: An Age-Period-Cohort Analysis.

Am J Ophthalmol 2019 09 17;205:184-196. Epub 2019 Apr 17.

Department of Ophthalmology and Visual Sciences, University of Toronto, Toronto, Ontario, Canada; Dalla Lana School of Public Health, University of Toronto, Toronto, Ontario, Canada.

Purpose: To compare the effect of age, sex and generation on physician practice patterns in Ontario, Canada.

Design: Retrospective cohort study.

Methods: Physician and patient data from 1992-2013 were used to calculate the yearly number of physicians, distinct patients seen, patient visits, government payments, physician age, sex, specialty, and year of birth. Age-period-cohort models were used for analysis.

Results: There was a negligible change in the number of distinct patients for all physicians and family physicians and a 20.6% decrease for ophthalmologists. There were small declines in yearly visits for all physicians (14.2%) and family physicians (17.3%) and a 10.0% increase for ophthalmologists. There were a lower number of visits (and patients for ophthalmologists) in each succeeding recent birth cohort. For all groups and birth cohorts, male physicians had a significantly greater number of visits and patients. Median payments increased over time in all groups and were less for women with an average women-to-men ratio of 0.64 for all physicians, 0.75 for family physicians, and 0.59 for ophthalmologists. After adjusting for the number of visits and patients, sex differences in payments remained significant for all physicians and ophthalmologists but were no longer significant for family physicians.

Conclusion: Younger cohorts of Ontario physicians have greater yearly payments compared with older cohorts at the same age despite similar or a slightly lower number of visits and patients. The sex gap of payments was mostly explained by differences in the number of patients and visits for family physicians but remained significant for all physicians and ophthalmologists.
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http://dx.doi.org/10.1016/j.ajo.2019.04.006DOI Listing
September 2019

COPS5 negatively regulates goat endometrial function via the ERN1 and mTOR-autophagy pathways during early pregnancy.

J Cell Physiol 2019 08 29;234(10):18666-18678. Epub 2019 Mar 29.

Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China.

In ruminant, adequate endometrial function is a major factor affecting implantation and economic efficiency. However, the precise mechanisms regulating goat endometrial function during the peri-implantation period of pregnancy are still unclear. Here, we investigated the functional role and signal transduction of the fifth component of the constitutive photomorphogenic-9 signalosome (COPS5) in the regulation of endometrial function in endometrial epithelial cells (EECs). Our results showed that hormones decreased COPS5 expression, and COPS5-mediated regulation of endometrial function. We also found that knockdown of COPS5 hindered EECs proliferation by the G1-phase cell cycle arrest. Hormones affected the activity of COPS5 through hormones receptors, while feedback from the expression of COPS5 regulated the transcription of the receptor. Moreover, knockdown of endoplasmic reticulum (ER) to nucleus signaling 1 (ERN1) via si-ERN1 partly inhibited endometrial function in shCOPS5 EECs. In addition, blocking the mTOR pathway by rapamycin promoted endometrial function in si-ERN1-transfected shCOPS5 EECs. Overall, these results suggest that COPS5 negatively regulates goat endometrial function via the ERN1 and mTOR-autophagy pathways and provide new insights into the mechanistic pathways of COPS5 during female reproductive development.
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http://dx.doi.org/10.1002/jcp.28505DOI Listing
August 2019

Coordination between the circadian clock and androgen signaling is required to sustain rhythmic expression of in mouse liver.

J Biol Chem 2019 04 12;294(17):7046-7056. Epub 2019 Mar 12.

From the Departments of Clinical Veterinary Medicine and

ELOVL3 is a very long-chain fatty acid elongase, and its mRNA levels display diurnal rhythmic changes exclusively in adult male mouse livers. This cyclical expression of hepatic is potentially controlled by the circadian clock, related hormones, and transcriptional factors. It remains unknown, however, whether the circadian clock, in conjunction with androgen signaling, functions in maintaining the rhythmic expression of in a sexually dimorphic manner. Under either zeitgeber or circadian time, WT mouse livers exhibited a robust circadian rhythmicity in the expression of circadian clock genes and In contrast, male mice displayed severely weakened expression of hepatic circadian clock genes, resulting in relatively high, but nonrhythmic, expression levels. ChIP assays revealed that NR1D1 binds to the promoter upon circadian change in WT mouse livers , and a diminished binding was observed in male mouse livers. Additionally, female mouse livers exhibited constant low levels of expression. Castration markedly reduced expression levels in male mouse livers but did not disrupt circadian variation of Injection of female mice with 5α-dihydrotestosterone induced rhythmicity in the liver. In AML12 cells, 5α-dihydrotestosterone also elevated expression in a time-dependent manner. In contrast, flutamide efficiently attenuated this induction effect. In conclusion, a lack of either the circadian clock or androgen signaling impairs hepatic expression, highlighting the observation that coordination between the circadian clock and androgen signaling is required to sustain the rhythmic expression of in mouse liver.
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http://dx.doi.org/10.1074/jbc.RA118.005950DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6497949PMC
April 2019