Intake of tobacco nitrosamines of smokers in various provinces of China and their cancer risk: A meta-analysis.

Nitrosamines are a class of carcinogens which have been detected widely in food, water, some pharmaceuticals as well as tobacco. The objectives of this paper include reviewing the basic information on tobacco consumption and nitrosamine contents, and assessing the health risks of tobacco nitrosamines exposure to Chinese smokers. We searched the publications in English from "Web of Science" and those in Chinese from the "China National Knowledge Infrastructure" in 2022 and collected 151 literatures with valid information. The content of main nitrosamines in tobacco, including 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK), N-nitrosonornicotine (NNN), N-nitrosoanatabine (NAT), N-nitrosoanabasine (NAB), total tobacco-specific nitrosamines (TSNA), and N-nitrosodimethylamine (NDMA) were summarized. The information of daily tobacco consumption of smokers in 30 provinces of China was also collected. Then, the intakes of NNN, NNK, NAT, NAB, TSNAs, and NDMA via tobacco smoke were estimated as 1534 ng/day, 591 ng/day, 685 ng/day, 81 ng/day, 2543 ng/day, and 484 ng/day by adult smokers in 30 provinces, respectively. The cancer risk (CR) values for NNN and NNK inhalation intake were further calculated as 1.44 × 10-5 and 1.95 × 10-4. The CR value for NDMA intake via tobacco smoke (inhalation: 1.66 × 10-4) indicates that NDMA is similarly dangerous in tobacco smoke when compared with the TSNAs. In China, the CR values caused by average nitrosamines intake via various exposures and their order can be estimated as the following: smoke (3.75 × 10-4) > food (1.74 × 10-4) > drinking water (1.38 × 10-5). Smokers in China averagely suffer 200% of extra cancer risk caused by nitrosamines in tobacco when compared with non-smokers.

Effect of Double-Ovsynch and Presynch-Ovsynch on postpartum ovarian cysts and inactive ovary in high-yielding dairy cows.

Introduction: Optimizing the management of dairy cattle reproduction can reduce postpartum ovarian disease in high-yielding dairy cows and thus enhance ranch economic benefit. The hypothesis of this study was that the Double-Ovsynch (DO) protocol in high-producing dairy cows would result in a lower incidence of follicular cysts but a higher incidence of luteal cysts compared to those undergoing the Presynch-Ovsynch (PS) protocol. Methods: In this experiment, 384 cows (204 primiparous and 180 multiparous) were allocated to the DO group, which followed the protocol: GnRH-7d-PGF2α-3d-GnRH-7d-Ovsynch-56 h (GnRH-7d-PGF2α-56 h-GnRH-16hTAI), starting on 39 ± 3 days in milk (DIM). Additionally, 359 cows (176 primiparous and 183 multiparous) were assigned to the PS group, which followed the protocol: PGF2α-14d-PGF2α-12d-Ovsynch-56 h, starting on 31 ± 3 DIM. In DO, B-mode ultrasound examinations were conducted 1 day after the GnRH-7d-PGF2α-3d-GnRH protocol to diagnose the presence of ovarian diseases followed by reexamination after 7 days of suspected cases. In PS, B-mode ultrasound examinations were conducted 1 day after the PGF2α-14d-PGF2α protocol to diagnose the presence of ovarian diseases followed by reexamination after 7 days. For all cows confirmed to having ovarian diseases, a second B-mode ultrasound examination was conducted at the time of the second GnRH and timed artificial insemination (TAI). If the ovary showed a normal developing follicle in combination with normal ovulation, the ovarian disease was considered to be cured. Results: The current study revealed no significant difference in the overall incidence and cure rate of postpartum ovarian diseases between DO and PS (incidence rate: 3.9% vs. 6.7%, cure rate: 50% vs. 41.7%, DO vs. PS). Also, there was no significant difference in the incidence and cure rate of luteal cysts between DO and PS (incidence rate: 2.9% vs. 2.2%, cure rate: 50.0% vs. 50.0%). The incidence of follicular cysts was significantly lower in the DO group than in the PS group (0.8% vs. 2.8%, DO vs. PS, p = 0.037), but there was no significant difference in the cure rates (66.7% vs. 50%). The occurrence of inactive ovary was lower in DO compared to PS (0.2% vs. 1.7%, p = 0.047). There was no significant difference in the pregnancy rate between the DO and PS groups (48.2% vs. 41.8%), although the DO group had a higher rate. What is different from our assumption is that PS did not effectively reduce the incidence of postpartum luteal cysts.

LncRNA STAT3-AS regulates endometrial receptivity via the STAT3 signaling pathway.

Endometrial receptivity is critical for the successful establishment of pregnancy in ruminants. Interferon tau (IFNT) plays a key role in promoting embryo attachment by activating the Janus kinase/signal transducer and activator of transcription pathway, which induces the expression of a series of interferon-stimulated genes (ISGs). In our previous study, sequencing analysis of goat endometrial epithelial cells (gEECs) treated with 20 ng/mL IFNT revealed a differentially expressed long non-coding RNA located on the STAT3 antisense chain, which we designated STAT3-AS. The aim of this study was to investigate the role and mechanism of STAT3-AS in establishing endometrial receptivity in goats. The results showed that STAT3-AS was expressed in both the nucleus and cytoplasm of gEECs, and its expression increased significantly in the uterus on day 15 of pregnancy. STAT3-AS expression was upregulated in gEECs treated with IFNT alone or in combination with progesterone and estradiol. Knockdown of STAT3-AS using specific short interfering RNA significantly inhibited the expression of classical ISGs such as interferon-stimulated gene 15 and 2',5'-oligodenylate synthetase 2, as well as uterine endometrial receptivity-related genes including homeobox gene A11, integrin beta 3, and vascular endothelial growth factor. Moreover, gEEC proliferation and the STAT3 pathway were suppressed in the absence of STAT3-AS. However, pretreatment with the STAT3 activator RO8191 restored the effect of silencing STAT3-AS on endometrial receptivity. Overall, these results suggest that STAT3-AS is an important regulator of endometrial receptivity in goats and that it regulates endometrial receptivity through the STAT3 pathway.

Reactive Oxygen Species Damage Bovine Endometrial Epithelial Cells via the Cytochrome C-mPTP Pathway.

After parturition, bovine endometrial epithelial cells (BEECs) undergo serious inflammation and imbalance between oxidation and antioxidation, which is widely acknowledged as a primary contributor to the development of endometritis in dairy cows. Nevertheless, the mechanism of oxidative stress-mediated inflammation and damage in bovine endometrial epithelial cells remains inadequately defined, particularly the molecular pathways associated with mitochondria-dependent apoptosis. Hence, the present study was designed to explore the mechanism responsible for mitochondrial dysfunction-induced BEEC damage. In vivo, the expressions of proapoptotic protein caspase 3 and cytochrome C were increased significantly in dairy uteri with endometritis. Similarly, the levels of proapoptotic protein caspase 3, BAX, and cytochrome C were markedly increased in H2O2-treated BEECs. Our findings revealed pronounced BEEC damage in dairy cows with endometritis, accompanied by heightened expression of cyto-C and caspase-3 both in vivo and in vitro. The reduction in apoptosis-related protein of BEECs due to oxidant injury was notably mitigated following N-acetyl-L-cysteine (NAC) treatment. Furthermore, mitochondrial vacuolation was significantly alleviated, and mitochondrial membrane potential returned to normal levels after the removal of ROS. Excessive ROS may be the main cause of mitochondrial dysfunction. Mitochondrial permeability transition pore (mPTP) blockade by cyclophilin D (CypD) knockdown with CSA significantly blocked the flow of cytochrome C (cyto-C) and Ca2+ to the cytoplasm from the mitochondria. Our results indicate that elevated ROS and persistent opening of the mPTP are the main causes of oxidative damage in BEECs. Collectively our results reveal a new mechanism involving ROS-mPTP signaling in oxidative damage to BEECs, which may be a potential avenue for the clinical treatment of bovine endometritis.

Downregulation of XBP1s aggravates lipopolysaccharide-induced inflammation by promoting NF-κB and NLRP3 pathways' activation in goat endometrial epithelial cells.

After delivery, bacterial contamination and uterine tissue degeneration in animals can lead to the development of uterine diseases, such as endometritis, accompanied by endoplasmic reticulum stress (ERS). Increasing evidence suggests that spliced X-box binding protein 1 (XBP1s), a critical component of ERS, is involved in several pathological processes in various organisms. However, the specific molecular mechanisms by which XBP1s mediates the inflammatory response in goat endometrial epithelial cells (gEECs) remain largely unknown. In the present study, XBP1s protein was induced into the nucleus in the lipopolysaccharide (LPS, 5 µg/mL)-induced inflammatory response of gEECs. Lipopolysaccharide-induced expression and nucleation of XBP1s were reduced by the inhibition of Toll-like receptor 4 (TLR4) using TAK-242 (1 µM; a TLR4 inhibitor). Expression and nucleation of XBP1s were similarly reduced when the activity of inositol-requiring enzyme 1α (IRE1α) was inhibited using 4µ8C (10 µM; an IRE1α inhibitor). In addition, inhibition of IRE1a increased IL-1ß, TNF-α, and IL-8 levels and secretion of IL-6 induced by LPS. Notably, phosphorylation of nuclear factor kappa-B (NF-κB) P65 protein and expression of NOD-like receptor thermal protein domain associated protein 3 (NLRP3) were similarly increased. Furthermore, knockdown of XBP1s in gEECs consistently promoted NF-κB P65 protein phosphorylation, NLRP3 protein expression, and inflammatory cytokine secretion. In summary, the current results suggest that in the LPS-induced inflammatory response in gEECs, LPS generates intracellular signaling cascades in gEECs via TLR4, which may promote XBP1s protein expression and nucleation by activating IRE1a. However, downregulation of XBP1s expression exacerbates inflammation by promoting activation of the NF-κB and NLRP3 inflammatory vesicle pathways. These results will potentially contribute to the treatment and prevention of endometritis in ruminants.

Transcription Factor Nrf2 Modulates Lipopolysaccharide-Induced Injury in Bovine Endometrial Epithelial Cells.

Endometritis in high-yield dairy cows adversely affects lactation length, milk quality, and the economics of dairy products. Endoplasmic reticulum stress (ERS) in bovine endometrial epithelial cells (BEECs) occurs as a consequence of diverse post-natal stressors, and plays a key role in a variety of inflammatory diseases. Nuclear-factor-erythroid-2-related factor 2 (Nrf2) is an important protective regulatory factor in numerous inflammatory responses. However, the mechanism by which Nrf2 modulates inflammation by participating in ERS remains unclear. The objective of the present study was to explore the role of Nrf2 in lipopolysaccharide (LPS)-induced injury to BEECs and to decipher the underlying molecular mechanisms of this injury. The expression of Nrf2- and ERS-related genes increased significantly in bovine uteri with endometritis. Isolated BEECs were treated with LPS to stimulate the inflammatory response. The expression of Nrf2 was significantly higher in cells exposed to LPS, which also induced ERS in BEECs. Activation of Nrf2 led to enhanced expression of the genes for the inflammation markers TNF-α, p65, IL-6, and IL-8 in BEECs. Moreover, stimulation of Nrf2 was accompanied by activation of ERS. In contrast, Nrf2 knockdown reduced the expression of TNF-α, p65, IL-6, and IL-8. Additionally, Nrf2 knockdown decreased expression of ERS-related genes for the GRP78, PERK, eIF2α, ATF4, and CHOP proteins. Collectively, our findings demonstrate that Nrf2 and ERS are activated during inflammation in BEECs. Furthermore, Nrf2 promotes the inflammatory response by activating the PERK pathway in ERS and inducing apoptosis in BEECs.

A Comparative Proteomic Analysis to Explore the Influencing Factors on Endometritis Using LC-MS/MS.

The inflammatory system activated by uterine infection is associated with decreased fertility. Diseases can be detected in advance by identifying biomarkers of several uterine diseases. Escherichia coli is one of the most frequent bacteria that is involved in pathogenic processes in dairy goats. The purpose of this study was to investigate the effect of endotoxin on protein expression in goat endometrial epithelial cells. In this study, the LC-MS/MS approach was employed to investigate the proteome profile of goat endometrial epithelial cells. A total of 1180 proteins were identified in the goat Endometrial Epithelial Cells and LPS-treated goat Endometrial Epithelial Cell groups, of which, 313 differentially expressed proteins were accurately screened. The proteomic results were independently verified by WB, TEM and IF techniques, and the same conclusion was obtained. To conclude, this model is suitable for the further study of infertility caused by endometrial damage caused by endotoxin. These findings may provide useful information for the prevention and treatment of endometritis.

USP18 promotes endometrial receptivity via the JAK/STAT1 and the ISGylation pathway.

Interferon-tau (IFNT), a pregnancy recognition signal in ruminants, promotes the establishment of endometrial receptivity by inducing the expression of interferon-stimulated genes (ISGs) via the Janus kinase/signal transducer and activator of transcription (JAK/STATs) signaling pathway. However, the precise mechanisms remain largely unknown. Ubiquitin-specific protease 18 (USP18) acts specifically on the ISGylation modification system to exert deubiquitination and participates in the regulation of the type I IFN signaling pathway. The purpose of this study was to determine the role and mechanism of USP18 on endometrial receptivity in goat. USP18 was mainly localized in the uterine luminal and glandular epithelium, and its expression levels were significantly increased from days 5-18 of early pregnancy. Progesterone (P4), estradiol (E2), and IFNT significantly stimulated USP18 expression in goat endometrial epithelial cells (gEECs) cultured in vitro. Meanwhile, the markers of endometrial receptivity HOXA11, ITGB1, ITGB3, and ITGB5 were significantly upregulated after USP18 overexpression in gEECs. However, USP18 interference significantly inhibited the expression of HOXA10, ITGB1, ITGB3, and ITGB5 in gEECs. In addition, both the phosphorylation levels of STAT1 and the expression of ISGylation-modified proteins were significantly increased after USP18 silencing in gEECs. Furthermore, pretreatment with the STAT1 inhibitor Fludara markedly restored the effect of USP18 interference in gEECs. In summary, USP18 may play an important role in promoting goat endometrial receptivity by regulating the JAK/STAT1 pathway and ISGylation.

Interferon-τ -induced ISG15-AS regulates endometrial receptivity during early goat pregnancy.

Endometrial receptivity is a critical process for the successful establishment of pregnancy in ruminants. However, the biological role of long non-coding RNAs (lncRNAs) in the development of endometrial receptivity is poorly understood. In this study, we performed RNA-seq analysis of immortalised goat endometrial epithelial cells (gEECs) treated with interferon-τ (IFNT). Transcriptome profiles showed that 8069 high-confidence putative lncRNAs, including 6498 intronic lncRNA transcripts, 1078 lincRNAs and 493 antisense lncRNAs were identified in gEECs with or without IFNT treatment. Functional clustering analysis was performed by using cis and trans lncRNAs prediction. GO and KEGG analyses revealed that differentially expressed lncRNAs may regulate tissue remodelling and immune responses. Subsequently, six of the 21 differentially expressed antisense lncRNAs were validated using qRT-PCR. Through functional screening and co-expression analysis of lncRNAs in gEECs, we identified that ISG15-AS was mainly expressed in the luminal and glandular epithelium on days 5 and 15 and was strongly upregulated on day 18 of pregnancy in vivo. Similarly, ISG15-AS was abundant in the nucleus and cytoplasm, and was significantly upregulated after treatment with IFNT in gEECs. In addition, ISG15 is an IFNT-responsive gene, that displayed an evident increase in vivo and in vitro. Moreover, sense ISG15 was significantly upregulated following ISG15-AS silencing. The key genes related to ISGylation and endometrial receptivity in gEECs dramatically increased after ISG15-AS inhibition. Collectively, our results indicate that a novel antisense lncRNA, ISG15-AS, may be important in regulating endometrial receptivity through ISGylation.

MSX1 Regulates Goat Endometrial Function by Altering the Plasma Membrane Transformation of Endometrial Epithelium Cells during Early Pregnancy.

MSX1 is an important member of the muscle segment homeobox gene (Msh) family and acts as a transcription factor to regulate tissue plasticity, yet its role in goat endometrium remodeling remains elusive. In this study, an immunohistochemical analysis showed that MSX1 was mainly expressed in the luminal and glandular epithelium of goat uterus, and the MSX1 expression was upregulated in pregnancy at days 15 and 18 compared with pregnancy at day 5. In order to explore its function, goat endometrial epithelial cells (gEECs) were treated with 17 ß-estrogen (E2), progesterone (P4), and/or interferon-tau (IFNτ), which were used to mimic the physiological environment of early pregnancy. The results showed that MSX1 was significantly upregulated with E2- and P4-alone treatment, or their combined treatment, and IFNτ further enhanced its expression. The spheroid attachment and PGE2/PGF2α ratio were downregulated by the suppression of MSX1. The combination of E2, P4, and IFNτ treatment induced the plasma membrane transformation (PMT) of gEECs, which mainly showed the upregulation of N-cadherin (CDH2) and concomitant downregulation of the polarity-related genes (ZO-1, α-PKC, Par3, Lgl2, and SCRIB). The knockdown of MSX1 partly hindered the PMT induced by E2, P4, and IFNτ treatment, while the upregulation of CDH2 and the downregulation of the partly polarity-related genes were significantly enhanced when MSX1 was overexpressed. Moreover, MSX1 regulated the CDH2 expression by activating the endoplasmic reticulum (ER) stress-mediated unfolded protein response (UPR) pathway. Collectively, these results suggest that MSX1 was involved in the PMT of the gEECs through the ER stress-mediated UPR pathway, which affects endometrial adhesion and secretion function.

Exon skipping caused by splicing mutation in TNNT1 nemaline myopathy.

The TNNT1 gene encoding the slow skeletal muscle TnT has been identified as a causative gene for nemaline myopathy. TNNT1 nemaline myopathy is mainly characterized by neonatal-onset muscle weakness, pectus carinatum and respiratory insufficiency. Herein, we report on a Chinese girl with TNNT1 nemaline myopathy with mild clinical phenotypes without thoracic deformities or decreased respiratory function. Muscle biopsy showed moderate to marked type 1 fiber atrophy and nemaline rods. Next-generation sequencing identified the compound heterozygous c. 587dupA (p. D196Efs*41) and c. 387+5G>A mutations in the TNNT1 gene according to the transcript NM_003283.4. RNA sequencing revealed complete exon 9 skipping caused by the c. 387+5G>A mutation. Through quantitative PCR, we found that both the truncation c. 587dupA (p. D196Efs*41) and the splicing c. 387+5G>A mutations triggered nonsense-mediated mRNA decay (NMD). Western blotting showed the residual amount of the truncated TNNT1 protein by deletion of exon 9, which may ameliorate the disease to some extent.

Staphylococcus aureus-Induced Necroptosis Promotes Mitochondrial Damage in Goat Endometrial Epithelial Cells.

Endometrial cell death is induced by bacterial infection, resulting in damage to the physical barriers and immune function. An in-depth understanding of the mechanisms of endometrial epithelial cell necroptosis might provide new insights into the treatment of uterine diseases. In the present study, we investigated the effect of Staphylococcus aureus on goat endometrial epithelial cell (gEEC) necroptosis, and the underlying molecular mechanism. We found that S. aureus induced significant necroptosis in gEECs by increasing the expression of key proteins of the RIPK1/RIPK3/MLKL axis; importantly, this effect was alleviated by inhibitors of RIPK1, RIPK3, and MLKL. Moreover, we found that the main triggers of gEEC necroptosis induced by S. aureus were not the toll-like receptors (TLRs) and tumor necrosis factor receptor (TNFR), but membrane disruption and ion imbalance. Moreover, we observed a significant decrease in the mitochondrial membrane potential, indicating mitochondrial damage, in addition to increased cytochrome c levels and reactive oxygen species (ROS) generation in S. aureus-infected gEECs; these, effects were also suppressed by the inhibitors of RIPK1, RIPK3, and MLKL. Taken together, these data revealed the molecular mechanism of S. aureus-induced gEEC necroptosis and provided potential new targeted therapies for clinical intervention in bacterial infections.

Review on corrosion and corrosion scale formation upon unlined cast iron pipes in drinking water distribution systems.

The qualified finished water from water treatment plants (WTPs) may become discolored and deteriorated during transportation in drinking water distribution systems (DWDSs), which affected tap water quality seriously. This water stability problem often occurs due to pipe corrosion and the destabilization of corrosion scales. This paper provides a comprehensive review of pipe corrosion in DWDSs, including corrosion process, corrosion scale formation, influencing factors and monitoring technologies utilized in DWDSs. In terms of corrosion process, corrosion occurrence, development mechanisms, currently applied assays, and indices used to determine the corrosion possibility are summarized, as well as the chemical and bacterial influences. In terms of scale formation, explanations for the nature of corrosion and scale formation mechanisms are discussed and its typical multilayered structure is illustrated. Furthermore, the influences of water quality and microbial activity on scale transformation are comprehensively discussed. Corrosion-related bacteria at the genus level and their associated corrosion mechanism are also summarized. This review helps deepen the current understanding of pipe corrosion and scale formation in DWDSs, providing guidance for water supply utilities to ensure effective measures to maintain water quality stability and guarantee drinking water safety.

Novel compound heterozygous mutations in the TTN gene: elongation and truncation variants causing limb-girdle muscular dystrophy type 2J in a Han Chinese family.

INTRODUCTION: Limb-girdle muscular dystrophy (LGMD) is a group of clinically heterogeneous muscle disorders commonly manifesting proximal limb girdle muscle weakness. There have been more than 30 subtypes of LGMD associated with causative genes and limb-girdle muscular dystrophy type 2J (LGMD2J) is caused by mutations in the TTN gene. METHODS: We report a Han Chinese family with LGMD2J. The proband and his sister both presented with weakness in the proximal lower limbs bilaterally. Muscle biopsy and genetic analysis were performed. RESULTS: Muscle biopsy of the proband showed dystrophic changes accompanied by rimmed vacuoles. Whole-exome sequencing identified novel compound heterozygous mutations in the TTN gene, including elongation (c.107962_107963delAT, p.I35988Sfs*26) and truncation (c.99125_99128dupACAG, p.S33043Rfs*9) variants in the proband and his sister. Both two variants have never been reported. Notably, we are the first to identify an elongation mutation in the TTN gene, broadening the genetic mutation spectrum of LGMD2J. DISCUSSION: Several variants in the last exon of the TTN gene have been reported, one of which was associated with LGMD2J. Besides, LGMD2J should be distinguished from other myopathies caused by mutations in the TTN gene. The pathogenesis of and specific curative methods for LGMD2J remain to be further elucidated.

Metabolic Profiling Revealed Prediction Biomarkers for Infantile Hemangioma in Umbilical Cord Blood Sera: A Prospective Study.

Infantile hemangioma (IH), the most common benign tumor in infancy, mostly arises and has rapid growth before 3 months of age. Because irreversible skin changes occur in the early proliferative stage, early medical treatment is essential to reduce the permanent sequelae caused by IH. Yet there are still no early screening biomarkers for IH before its visible emergence. This study aimed to explore prediction biomarkers using noninvasive umbilical cord blood (UCB). A prospective study of the metabolic profiling approach was performed on UCB sera from 28 infants with IH and 132 matched healthy controls from a UCB population comprising over 1500 infants (PeptideAtlas: PASS01675) using liquid chromatography-mass spectrometry. The metabolic profiling results exhibited the characteristic metabolic aberrance of IH. Machine learning suggested a panel of biomarkers to predict the occurrence of IH, with the area under curve (AUC) values in the receiver operating characteristic analysis all >0.943. Phenylacetic acid had potential to predict infants with large IH (diameter >2 cm) from those with small IH (diameter <2 cm), with an AUC of 0.756. The novel biomarkers in noninvasive UCB sera for predicting IH before its emergence might lead to a revolutionary clinical utility.

The endoplasmic reticulum stress-mediated unfolded protein response protects against infection of goat endometrial epithelial cells by Trueperella pyogenes via autophagy.

Trueperella pyogenes is an important bacterial pathogen of a wide range of domestic and wild animals. Autophagy plays a key role in eliminating T. pyogenes in a process that is dependent on mechanistic target of rapamycin (mTOR). The endoplasmic reticulum (ER) stress response also is critical for autophagy regulation. However, the relationship between ER stress and T. pyogenes is uncharacterized and the intracellular survival mechanisms of T. pyogenes have not been investigated adequately. In this study, we show that T. pyogenes invades goat endometrial epithelial cells (gEECs). Meanwhile, we observed that GRP78 was upregulated significantly, and that unfolded protein response (UPR) also were activated after infection. Additionally, treatment with activators and inhibitors of ER stress downregulated and upregulated, respectively, intracellular survival of T. pyogenes. Blocking the three arms of the UPR pathway separately enhanced T. pyogenes survival and inflammatory reaction to different levels. We also show that LC3-labeled autophagosomes formed around the invading T. pyogenes and that autolysosome-like vesicles were visible in gEECs using transmission electron microscopy. Moreover, tunicamycin did not inhibit the intracellular survival of T. pyogenes under conditions in which autophagy was blocked. Finally, severe challenge with T. pyogenes induced host cell apoptosis which also may indicate a role for ER stress in the infection response. In summary, we demonstrate here that ER stress and UPR are novel modulators of autophagy that inhibit T. pyogenes intracellular survival in gEECs, which has the potential to be developed as an effective therapeutic target in T. pyogenes infectious disease.

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

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-ß-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.

Intake of volatile nitrosamines by Chinese residents in different provinces via food and drinking water.

N-nitrosamines are potent carcinogens, particularly N-nitrosodimethylamine (NDMA) and N-nitrosodiethylamine (NDEA), which are commonly found in a variety of foods and drinking water. We calculated the food and drinking water intakes of NDMA, NDEA, and total volatile nitrosamines (TVNA) by Chinese residents in different provinces by multiplying the reported total diet study results by the nitrosamine contents in food and drinking water. The weighted content of nitrosamines in each category of foods and the concentration of nitrosamines in drinking water was obtained through literature review. The exogenous NDMA, NDEA and TVNA intakes of adult residents in 20 provinces ranged from 171 to 425 ng/d, 41 to 140 ng/d and 373 to 1028 ng/d, respectively. The main contributors to NDMA and TVNA intakes were vegetables, cereals, aquatic products, and meats while the main sources of NDEA intake were vegetables and cereals. The average total NDMA intake per capita in China (251 ng/d) was similar to that in Germany in 1991 (231 ng/d) but higher than that in the United States (136 ng/d), Canada (87.6 ng/d) and France (188 ng/d). Large differences in nitrosamine intakes were observed between the coastal provinces and inland provinces. Drinking water was estimated to contribute 13.1%, 1.3% and 10.8% of the exogenous intakes of NDMA, NDEA and TVNA, respectively. Based on our results, we recommend setting the NDMA drinking water criterion of 40 ng/L. Overall, this study presents basic information regarding nitrosamines intake via food and drinking water in China that will facilitate risk assessment, generation of health advisories and policy making.

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

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.

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

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.