Single-cell RNA reveals a tumorigenic microenvironment in the interface zone of human breast tumors.

BACKGROUND: The interface zone, area around invasive carcinoma, can be thought of as the actual tissue of the tumor microenvironment with precedent alterations for tumor invasion. However, the heterogeneity and characteristics of the microenvironment in the interface area have not yet been thoroughly explored. METHODS: For in vitro studies, single-cell RNA sequencing (scRNA-seq) was used to characterize the cells from the tumor zone, the normal zone and the interface zone with 5-mm-wide belts between the tumor invasion front and the normal zone. Through scRNA-seq data analysis, we compared the cell types and their transcriptional characteristics in the different zones. Pseudotime, cell-cell communication and pathway analysis were performed to characterize the zone-specific microenvironment. Cell proliferation, wound healing and clone formation experiments explored the function of differentially expressed gene BMPR1B, which were confirmed by tumor models in vivo. RESULTS: After screening, 88,548 high-quality cells were obtained and identified. Regulatory T cells, M2 macrophages, angiogenesis-related mast cells, stem cells with weak DNA repair ability, endothelial cells with angiogenic activity, fibroblasts with collagen synthesis and epithelial cells with proliferative activity form a unique tumorigenic microenvironment in the interface zone. Cell-cell communication analysis revealed that there are special ligand-receptor pairs between different cell types in the interface zone, which protects endothelial cell apoptosis and promotes epithelial cell proliferation and migration, compared to the normal zone. Compared with the normal zone, the highly expressed BMPR1B gene promotes the tumorigenic ability of cancer cells in the interface zone. CONCLUSIONS: Our work identified a unique tumorigenic microenvironment of the interface zone and allowed for deeper insights into the tumor microenvironment of breast cancer that will serve as a helpful resource for advancing breast cancer diagnosis and therapy.

Omega-3 polyunsaturated fatty acids ameliorated inflammatory response of mammary epithelial cells and mammary gland induced by lipopolysaccharide.

Omega-3 polyunsaturated fatty acids (n-3 PUFAs), essential fatty acids for humans and animals, have been reported to play a beneficial role in a variety of inflammatory diseases. In this study, we investigated the inhibitory effects and potential molecular mechanisms of n-3 PUFAs on the inflammatory response in lipopolysaccharide (LPS)-stimulated mammary alveolar cell line (MAC-T). Results showed that n-3 PUFAs could abate LPS-induced secretions of tumor necrosis factor-α, interleukin (IL)-6 and IL-1ß in MAC-T cells through the nuclear transcription factor kappa B (NF-κB) signal pathway. Meanwhile, n-3 PUFA intervention attenuated histopathologic changes of mammary glands, the white blood cell number decrease, and the alkaline phosphatase level decrease in the serum of mice challenged by LPS. Furthermore, n-3 PUFA intervention improved the ecological structure of the flora in terms of the structural disorder of the non-significant dominant flora induced by LPS in mice. Collectively, both in vitro and in vivo experiments revealed that n-3 PUFAs have a positive effect on LPS-induced inflammatory response, which was possibly mediated by the NF-κB signaling pathway and the intestinal microbiota.

Aplysin Retards Pancreatic Necrosis and Inflammatory Responses in NOD Mice by Stabilizing Intestinal Barriers and Regulating Gut Microbial Composition.

Aplysin is a brominated sesquiterpene with an isoprene skeleton and has biological activities. The purpose of this study is to investigate the inhibitory effect of aplysin on spontaneous pancreatic necrosis in nonobese diabetic (NOD) mice and its potential mechanisms. Results showed that NOD mice at 12 weeks of age showed obvious spontaneous pancreatic necrosis, damaged tight junctions of intestinal epithelia, and widened gaps in tight and adherens junctions. Aplysin intervention was able to alleviate spontaneous pancreatic necrosis in NOD mice, accompanied with decreased serum endotoxin levels and downregulated expressions of Toll-like receptor 4 and its related molecules MyD88, TRAF-6, NF-κB p65, TRIF, TRAM, and IRF-3, as well as protein levels of interleukin-1ß and interferon-ß in pancreatic tissues. In addition, we observed obvious improvements of intestinal mucosal barrier function and changes of gut microbiota in the relative abundance at the phylum level and the genus level in aplysin-treated mice compared with control mice. Together, these data suggested that aplysin could retard spontaneous pancreatic necrosis and inflammatory responses in NOD mice through the stabilization of intestinal barriers and regulation of gut microbial composition.

Blocking C-Raf alleviated high-dose small-volume radiation-induced epithelial mesenchymal transition in mice lung.

The goal of this study was to develop a potential druggable target for lung injury after SABR through the small animal model. Utilising the model, a radiation dose of 70 Gy or 90 Gy was focally (small volume) delivered to the left lung of mice. The highly expressed phosphorylation form of C-Raf was discovered through a protein array experiment, with the protein being extracted from the area of radiated mouse lung tissue, and was confirmed by IHC and western blot. C-Raf activation, along with morphological change and EMT (Epithelial to Mesenchymal Transition) marker expression, was observed after radiation to the mouse type II alveolar cell line MLE-12. C-Raf inhibitor GW5074 was able to reverse the EMT in cells effectively, and was found to be dependent on Twist1 expression. In the animal experiment, pretreatment of GW5074 alleviated EMT and lung injury after 70 Gy radiation was focally delivered to the lung of mice. Conclusively, these results demonstrate that C-Raf inhibitor GW5074 inhibits high-dose small-volume radiation-induced EMT via the C-Raf/Twist1 signalling pathway in mice. Therefore, pharmacological C-Raf inhibitors may be used effectively as inhibitors of SABR-induced lung fibrosis.

LRRC75A antisense lncRNA1 knockout attenuates inflammatory responses of bovine mammary epithelial cells.

Long noncoding RNAs (lncRNAs) play multiple key roles during inflammatory processes. In this study, a novel lncRNA identified by the high-throughput sequencing analysis was found significantly down-regulated in Escherichia coli-introduced cell model of bovine mastitis. Given that this lncRNA consists of the antisense of leucine-rich repeat-containing protein 75A (LRRC75A), it was named LRRC75A antisense lncRNA1 (LRRC75A-AS1). The expression of LRRC75A-AS1 was down-regulated in bovine mammary epithelial cells and mammary tissues under inflammatory condition. Knockout (KO) of LRRC75A-AS1 by CRISPR-Cas9 system in bovine mammary alveolar cell-T (MAC-T) cell line could enhance expressions of tight junction (TJ) proteins Claudin-1, Occludin and ZO-1, reduce cell monolayer permeability, and inhibit Staphylococcus aureus adhesion and invasion. Meanwhile, it also down-regulated expressions of inflammatory factors and attenuated activation of NF-κB pathway. Similarly, knockdown of LRRC75A caused the changes as LRRC75A-AS1 KO did, while overexpression of LRRC75A enabled the opposite effects. TJ of epithelioid cells barriers the pathogenic microorganisms outside during inflammation, in which LRRC75A-AS1 can regulate the expression of TJ proteins through LRRC75A, affecting the development of inflammation.

The lncRNA FAL1 protects against hypoxia-reoxygenation- induced brain endothelial damages through regulating PAK1.

Dysregulation of cerebral microvascular endothelial cells plays an important role in the pathogenesis of stroke. However, the underlying mechanisms still need to be elucidated. In the current study, we found that the long non-coding RNA (lncRNA) FAL1 was significantly reduced in response to oxygen-glucose deprivation and reoxygenation (OGD/R) stimulation in human primary brain microvascular endothelial cells (HBMVECs). Interestingly, overexpression of FAL1 ameliorated OGD/R-induced oxidative stress by reducing the production of reactive oxygen species (ROS) and increasing the level of reduced glutathione (GSH). Also, overexpression of FAL1 suppressed OGD/R-induced secretions of interleukin-6 (IL-6), monocyte chemotactic protein-1 (MCP-1), and high mobility group box-1 (HMGB-1). We then found that OGD/R-induced reduction of cell viability and release of lactate dehydrogenase (LDH) were prevented by overexpression of FAL1. Additionally, exposure to OGD/R significantly reduced the phosphorylated levels of PAK1 and AKT as well as the total level of proliferating cell nuclear antigen (PCNA), which was restored by overexpression of FAL1. Importantly, overexpression of FAL1 restored OGD/R-induced reduction in the expression of endothelial nitric oxide synthase (eNOS) and the subsequent release of nitric oxide (NO). Our results implicate that FAL1 might be involved in the process of brain endothelial cell damage.

Nicotinamide riboside alleviates alcohol-induced depression-like behaviours in C57BL/6J mice by altering the intestinal microbiota associated with microglial activation and BDNF expression.

The gut microbiota play an important role in many central nervous system diseases through the gut microbiota-brain axis. Recent studies suggest that nicotinamide riboside (NR) has neuroprotective properties. However, it is unknown whether NR can prevent or protect against alcohol-induced depression. Furthermore, it is unclear whether its therapeutic action involves changes in the composition of the gut microbiome. Here, we investigated the effects of NR in the mouse model of alcohol-induced depression. Treatment with NR improved the alcohol-induced depressive behaviour in mice. In addition, NR decreased the number of activated microglia in the hippocampus, and it reduced the levels of pro-inflammatory (IL-1ß, IL-6, and TNF-α) and anti-inflammatory (IL-10 and TGF-ß) cytokines in the brain of mice with alcohol-induced depression. Furthermore, NR significantly upregulated BDNF and diminished the inhibition of the AKT/GSK3ß/ß-catenin signalling pathway in the hippocampus of these mice. 16S rRNA sequencing revealed that, compared with control and NR-treated mice, the gut microbiome richness and composition were significantly altered in the depressed mice. Spearman's correlation analysis showed that differential gut bacterial genera correlated with the levels of inflammation-related cytokines and BDNF in the brain. After faecal microbiota transplantation, cognitive behaviours, microglial activity, levels of cytokines and BDNF, and activation state of the AKT/GSK3ß/ß-catenin signalling pathway (which is downstream of the BDNF receptor, TrkB) in recipient mice were similar to those in donor mice. Collectively, our findings show that NR dietary supplementation protects against alcohol-induced depression-like behaviours, possibly by altering the composition of the gut microbiota.

Overexpression of lncRNA H19 changes basic characteristics and affects immune response of bovine mammary epithelial cells.

The function of long non-coding RNA H19 (H19) on cell proliferation has been observed in various cell types, and the increased expression of H19 was also found in the lipopolysaccharide (LPS)-induced inflammatory bovine mammary epithelial cells (MAC-T). However, the roles of H19 in the inflammatory response and physiological functions of bovine mammary epithelial cell are not clear. In the present study, we found that overexpression of H19 in MAC-T cells significantly promoted cell proliferation, increased the protein and mRNA level of ß-casein, and enhanced the expression of tight junction (TJ)-related proteins while inhibited staphylococcus aureus adhesion to cells. In addition, results demonstrated that overexpression of H19 affected the LPS-induced immune response of MAC-T cells by promoting expressions of inflammatory factors, including TNF-α, IL-6, CXCL2 and CCL5, and activating the NF-κB signal pathway. Our findings indicate that H19 is likely to play an important role in maintaining normal functions and regulating immune response of bovine mammary epithelial cells.

NDRG1 negatively regulates proliferation and Milk bio-synthesis of bovine epithelial cells via the mTOR signaling pathway.

The expression of N-myc downstream-regulated gene 1 (NDRG1) was significantly correlated with diverse processes such as cell growth and differentiation, lipid synthesis, stress and immune responses. Here we explored the role of NDRG1 expression in bovine mammary tissue and epithelial cells under an inflammatory condition. Results showed that NDRG1 expression was elevated in bovine mammary tissue with mastitis and mammary epithelial cells treated by heat inactivated Escherichia coli and Staphylococcus aureus compared to normal tissue and untreated cells. Overexpression of NDRG1 significantly inhibited cell proliferation and migration, ß-casein secretion, gene expressions of inflammatory cytokines tumor necrosis factor-α, interleukin (IL)-6 and IL-8, and activation of mTOR signal pathway of mammary epithelial cells, and vice versa by NDRG1 knockdown. These findings suggest that NDRG1 has immense potential in the regulation of properties in bovine mammary epithelial cells under an inflammatory condition.

RNA-seq analysis of different inflammatory reactions induced by lipopolysaccharide and lipoteichoic acid in bovine mammary epithelial cells.

Lipopolysaccharide (LPS) and lipoteichoic acid (LTA) are key virulence factors of Escherichia coli and Staphylococcus aureus respectively, and both of them could cause inflammatory reaction in bovine mammary glands. In this study, we used bovine mammary epithelial cells (BMECs) as pattern recognition receptors and stimulated them with LPS or LTA to investigate the global transcriptional response variations of BMECs to these two different virulent factors through RNA-Seq analysis. We found 100 differentially expressed genes (DEGs) with 95 up-regulated and 5 down-regulated genes in LPS-treated group, whereas 24 DEGs with 12 up-regulated and 12 down-regulated genes in LTA-treated group compared to control. Although the number and expression changes of DEGs are significantly different between LPS vs Control and LTA vs Control, KEGG pathway enrichment analysis showed the majorities of DEGs in each pair were enriched on cytokine-cytokine receptor interaction, NF-κB signaling pathway, and NOD-like receptor signaling pathway, especially cytokines and chemokines. These results provided a comprehensive analysis of gene expression profiles elicited by LPS and LTA in BMECs, contributing to the understanding of early "pathogen-host" interactions during intramammary infections.

A novel long non-coding RNA regulates the immune response in MAC-T cells and contributes to bovine mastitis.

The long non-coding RNAs (lncRNAs) are known to transcriptionally regulate a wide spectrum of diseases. Here, we screened for potentially functional lncRNAs in a mammary epithelial cell model of bovine mastitis by RNA-Seq technology and identified a class of previously undetected mastitis-related lncRNAs. A novel lncRNA was widely expressed in a variety of bovine tissues with diverse relative abundance and had a relatively low expression in mammary tissue. Given its predicted target gene is TUBA1C, we name it lncRNA-TUB. We found a higher expression of lncRNA-TUB in mammary epithelial cells that received a proinflammatory stimulus compared to normal cells. Knockout of lncRNA-TUB by the CRISPR/Cas9 system revealed that it plays crucial roles in the morphological shape, proliferation, migration and ß-casein secretion of mammary epithelial cells. In addition, lncRNA-TUB mediates Escherichia coli-induced inflammatory factor secretion and Staphylococcus aureus adhesion to epithelial cells. Our results suggest that the lncRNAs identified here function in bovine mastitis, and that lncRNA-TUB affects the basic biological characteristics and functions of bovine mammary epithelial cells in inflammatory conditions, providing valuable insights into the mechanisms of bovine mastitis.

LncRNA XIST mediates bovine mammary epithelial cell inflammatory response via NF-κB/NLRP3 inflammasome pathway.

OBJECTIVES: The correlations between long non-coding RNAs (lncRNAs) and diverse mammal diseases have been clarified by many researches, but the cognition about bovine mastitis-related lncRNAs remains limited. This study aimed to investigate the potential role of lncRNA X-inactive specific transcript (XIST) in the inflammatory response of bovine mammary epithelial cells. MATERIALS AND METHODS: Two inflammatory bovine mammary alveolar cell-T (MAC-T) models were established by infecting the cells with Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). The expressions of pro-inflammatory cytokines were measured, and the proliferation, viability and apoptosis of the inflammatory cells were evaluated after XIST was knocked down by an siRNA. The relationship among XIST, NF-κB pathway and NOD-like receptor protein 3 (NLRP3) inflammasome was investigated using an inhibitor of NF-κB signal pathway. RESULTS: The expression of XIST was abnormally increased in bovine mastitic tissues and inflammatory MAC-T cells. Silencing of XIST significantly increased the expression of E. coli or S. aureus-induced pro-inflammatory cytokines. Additionally, knockdown of XIST could inhibit cell proliferation, suppress cell viability and promote cell apoptosis under inflammatory conditions. Furthermore, XIST inhibited E. coli or S. aureus-induced NF-κB phosphorylation and the production of NLRP3 inflammasome. CONCLUSIONS: The expression of XIST was promoted by activated NF-κB pathway and, in turn, XIST generated a negative feedback loop to regulate NF-κB/NLRP3 inflammasome pathway for mediating the process of inflammation.

MCPIP1 mediates inflammatory responses induced by lipopolysaccharide and lipoteichoic acid in bovine mammary epithelial cells.

Monocyte chemoattractant protein-induced protein 1 (MCPIP1) is a kind of zinc finger RNA binding protein, which exerts immune responses in a variety of cell types. However, the role of MCPIP1 in bovine mammary epithelial cells during mastitis has not been studied. In this study, we explored the functions of MCPIP1 in the inflammatory process induced by virulence factors of pathogens in bovine mammary alveolar cell-T (MAC-T) cell line. Our results showed that MCPIP1 was significantly highly expressed both in the mammary tissue of dairy cows with mastitis and in inflammatory MAC-T cells induced by lipopolysaccharide (LPS) or lipoteichoic acid (LTA). Furthermore, we found that overexpression of MCPIP1 in MAC-T cells abated the LPS-induced increase at the gene expression levels of inflammatory mediators tumor necrosis factor-α-α, interleukin (IL)-1ß, IL-6 and IL-8, enhanced the LPS- and LTA-induced inhibition of epithelial proliferation and promoted the LPS- and LTA-induced oxidative and DNA damage. These findings indicated that MCPIP1 has an enormous potential in regulating the inflammatory response of bovine mammary epithelial cells during infection and may provide an effective therapeutic target for bovine mastitis to reduce the damage caused by inflammatory reactions.

IL-1ß induces increased tight junction permeability in bovine mammary epithelial cells via the IL-1ß-ERK1/2-MLCK axis upon blood-milk barrier damage.

Bovine mastitis occurs frequently in dairy cows and is often caused by various aetiological organisms, for example, Escherichia coli. Lipopolysaccharide (LPS) is a key virulence factor of E. coli. In this study, we stimulated bovine mammary epithelial cells (BMECs) with LPS to investigate the global transcriptional response and identify specific proinflammatory factors that play important roles in blood-milk barrier damage during mastitis caused by E. coli. By performing RNA-seq, we identified a large number of significantly differentially expressed genes (DEGs) between the LPS-treated BMECs and the control cells. Among the DEGs, interleukin-1ß (IL-1ß) was selected because its messenger RNA expression was induced by LPS and its enrichment is involved in multiple inflammatory signal pathways, and its roles in blood-milk barrier damage during the process of mastitis were investigated. Exogenous IL-1ß treatment damaged the integrity of the blood-milk barrier, as indicated by the increased BMEC tight junction (TJ) permeability and confirmed by in vitro and in vivo experiments. Furthermore, the IL-1ß-induced increase in the BMEC TJ permeability was mediated by the IL-1ß-ERK1/2-MLCK axis pathway. Our data provide insights into the functions of IL-1ß in blood-milk barrier damage caused by mastitis in dairy cows.

Elevated plasma macrophage migration inhibitor factor as a risk factor for the development of post-stroke depression in ischemic stroke.

BACKGROUND: Macrophage migration inhibitory factor (MIF), a central cytokine of the innate immunity and inflammatory responses, has been reported to link to the pathophysiology of cardiovascular disease and depression. The aim of this study was to test the possible association between plasma MIF and the development of post-stroke depression (PSD) in Chinese patients with acute ischemic stroke (AIS). METHODS: The first-ever AIS patients who were hospitalized at Affiliated Hospital of Weifang Medical College during the period from November 2015 to September 2017 were included. Neurological and neuropsychological evaluations were conducted at the 3-month follow-up. Plasma concentrations of MIF were tested by Quantikine Human MIF Immunoassay. Plasma levels of homocysteine (HCY), C-reactive protein (CRP) and Interleukin 6 (IL-6) were also tested. Results were expressed as percentages for categorical variables and as medians (Interquartile range-IQR) for the continuous variables. RESULTS: Finally, 333 stroke patients were included, and 95 out of those patients (28.5%) were classified as major depression. In the patients with major depression, plasma levels of MIF were higher compared with those in patients free-depression [27.3(IQR, 23.5-34.9) ng/ml vs. 20.9(IQR, 17.0-24.8) ng/ml; Z = 8.369, P < 0.001]. For each 1unit increase of MIF, the unadjusted and adjusted risk of PSD increased by 18% (odds ratios [OR]: 1.18; 95% confidence interval [CI], 1.13-1.23, P < 0.001) and 11% (1.11; 1.02-1.16, P = 0.001), respectively. In a multivariate model using the elevated levels of MIF (≥median) vs. normal (

CYP1A1 Relieves Lipopolysaccharide-Induced Inflammatory Responses in Bovine Mammary Epithelial Cells.

The expression of cytochrome P4501A1 (CYP1A1) enzyme is changed in various organs during the host response to inflammation or infection, leading to alterations in the metabolism of endogenous and exogenous compounds. Results of this study showed that CYP1A1 expression was significantly downregulated in the mammary tissue of bovine with mastitis, in inflammatory epithelial cells (INEs) extracted from the tissue, and in lipopolysaccharide- (LPS-) induced INEs compared with their corresponding counterparts. Overexpression of CYP1A1 in bovine mammary epithelial cells alleviated the LPS-induced inhibition of epithelial proliferation, abated the LPS-induced increase of gene expression and protein secretion of inflammatory cytokine tumor necrosis factor-α and interleukin-6, and attenuated the LPS-induced activation of NF-κB signaling. These findings suggest that CYP1A1 has immense potential in the regulation of inflammatory responses in bovine mammary epithelial cells during mastitis and may serve as a useful therapeutic target in mitigating injuries caused by inflammatory overreaction.

MicroRNA-27b Modulates Inflammatory Response and Apoptosis during Mycobacterium tuberculosis Infection.

Mycobacterium tuberculosis poses a significant global health threat. MicroRNAs play an important role in regulating host anti-mycobacterial defense; however, their role in apoptosis-mediated mycobacterial elimination and inflammatory response remains unclear. In this study, we explored the role of microRNA-27b (miR-27b) in murine macrophage responses to M. tuberculosis infection. We uncovered that the TLR-2/MyD88/NF-κB signaling pathway induced the expression of miR-27b and miR-27b suppressed the production of proinflammatory factors and the activity of NF-κB, thereby avoiding an excessive inflammation during M. tuberculosis infection. Luciferase reporter assay and Western blotting showed that miR-27b directly targeted Bcl-2-associated athanogene 2 (Bag2) in macrophages. Overexpression of Bag2 reversed miR-27b-mediated inhibition of the production of proinflammatory factors. In addition, miR-27b increased p53-dependent cell apoptosis and the production of reactive oxygen species and decreased the bacterial burden. We also showed that Bag2 interacts with p53 and negatively regulates its activity, thereby controlling cell apoptosis and facilitating bacterial survival. In summary, we revealed a novel role of the miR-27b/Bag2 axis in the regulation of inflammatory response and apoptosis and provide a potential molecular host defense mechanism against mycobacteria.

A subchronic feeding safety evaluation of transgenic milk containing human ß-defensin 3 on reproductive system of C57BL/6J mouse.

Bovine mastitis is an infectious disease of the mammary gland which has been generally treated by antibiotic delivery. While the increasing drug-resistant bacteria and the high consumption of the antibiotic had become a noticeable concern. In a previous study, a mammary special vector expressing human ß-defensin 3 (hBD3) was transfected into bovine fetal fibroblasts to produce mastitis-resistant bovine. This investigation focused on potential unintended effects of transgenic milk containing hBD3 produced by these mastitis-resistant bovine on the reproductive system of C57BL/6J mice. Mice were fed with diets containing transgenic milk or conventional milk, nutritionally balanced to an AIN93G diet for 90 days, and non-milk diet was selected as the negative group. The reproductive system was given special attention including reproductive organ/body ratios, necropsy and histopathology, serum sex hormone, sperm parameters, estrus cycle and the expression level of some specific genes which could indicate the development and function of reproductive system. No diet-related significant differences were observed among three groups in this 90-day feeding study. The results indicated that hBD3 milk does not appear to exert any effect on the reproductive system in C57BL/6J rats compared with conventional milk or the control diet.

lncRNA H19 is involved in TGF-ß1-induced epithelial to mesenchymal transition in bovine epithelial cells through PI3K/AKT Signaling Pathway.

Increased levels of long noncoding RNA H19 (H19) have been observed in many inflammatory and organ fibrosis diseases including ulcerative colitis, osteoarthritis, liver fibrosis, renal fibrosis and pulmonary fibrosis. However, the role of H19 in bovine mastitis and mastitis-caused fibrosis is still unclear. In our study, H19 was characterized as a novel regulator of EMT induced by transforming growth factor-ß1 (TGF-ß1) in bovine mammary alveolar cell-T (MAC-T) cell line. We found that H19 was highly expressed in bovine mastitis tissues and inflammatory MAC-T cells induced by virulence factors of pathogens. TGF-ß1 was also highly expressed in inflammatory MAC-T cells, and exogenous TGF-ß1 could induce EMT, enhance extracellular matrix protein expression, and upregulate H19 expression in epithelial cells. Stable expression of H19 significantly promotes EMT progression and expression of ECM protein induced by TGF-ß1 in MAC-T cells. Furthermore, by using a specific inhibitor of the PI3K/AKT pathway, we demonstrated that TGF-ß1 upregulated H19 expression through PI3K/AKT pathway. All these observations imply that the lncRNA H19 modulated TGF-ß1-induced epithelial to mesenchymal transition in bovine epithelial cells through PI3K/AKT signaling pathway, which suggests that mammary epithelial cells might be one source for myofibroblasts in vivo in the mammary glands under an inflammatory condition, thereby contributing to mammary gland fibrosis.

Sp110 enhances macrophage resistance to Mycobacterium tuberculosis via inducing endoplasmic reticulum stress and inhibiting anti-apoptotic factors.

Tuberculosis remains a leading health problem worldwide and still accounts for about 1.3 million deaths annually. Expression of the mouse Sp110 nuclear body protein (Sp110) upregulates the apoptotic pathway, which plays an essential role in enhancing host immunity to Mycobacterium tuberculosis (Mtb). However, the mechanism of this upregulation is unclear. Here, we have identified 253 proteins in mouse macrophages that interact with Sp110, of which 251 proteins were previously uncharacterized. The results showed that Sp110 interacts with heat shock protein 5 (Hspa5) to activate endoplasmic reticulum (ER) stress-induced apoptosis, and that this is essential for Sp110 enhanced macrophage resistance to Mtb. Inhibition of the ER stress pathway abolishing the Sp110-enhanced macrophage apoptosis and resulted in increased intracellular survival of Mtb in macrophages overexpressing Sp110 Further studies revealed that Sp110 also interacts with the RNA binding protein, Ncl to promote its degradation. Consequently, the expression of Bcl2, usually stabilized by Ncl, was downregulated in Sp110 overexpressing macrophages. Moreover, overexpression of Sp110 promotes degradation of ribosomal protein Rps3a, resulting in upregulation of the activity of the pro-apoptotic poly (ADP-ribose) polymerase (PARP). In addition, macrophages from transgenic cattle with increased Sp110 expression confirmed that activation of the ER stress response is the main pathway through which Sp110-enhanced macrophages impart resistance to Mtb. This work has revealed the mechanism of Sp110 enhanced macrophage apoptosis in response to Mtb infection, and provides new insights into the study of host-pathogen interactions.