Comprehensive cell surface protein profiling of human mesenchymal stromal cells from peritoneal dialysis effluent and comparison with those from human bone marrow and adipose tissue.

Peritoneal mesenchymal stromal cells (pMSCs) are isolated from peritoneal dialysis (PD) effluent, and treatment with the pMSCs reduces peritoneal membrane injury in rat model of PD. This study was designed to verify the identity of the pMSCs. pMSCs were grown in plastic dishes for 4-7 passages, and their cell surface phenotype was examined by staining with a panel of 242 antibodies. The positive stain of each target protein was determined by an increase in fluorescence intensity as compared with isotype controls in flow cytometrical analysis. Here, we showed that pMSCs predominantly expressed CD9, CD26, CD29, CD42a, CD44, CD46, CD47, CD49b, CD49c, CD49e, CD54, CD55, CD57, CD59, CD63, CD71, CD73, CD81, CD90, CD98, CD147, CD151, CD200, CD201, ß2-micoglobulin, epithelial growth factor receptor, human leukocyte antigen (HLA) class 1, and, to a lesser extent, CD31, CD45RO, CD49a, CD49f, CD50, CD58, CD61, CD105, CD164, and CD166. These cells lacked expression of most hematopoietic markers such as CD11b, CD14, CD19, CD34, CD40, CD80, CD79, CD86, and HLA-DR. There was 38.55% difference in the expression of 83 surface proteins between bone marrow (BM)-derived MSCs and pMSCs, and 14.1% in the expression of 242 proteins between adipose tissue (AT)-derived MSCs and pMSCs. The BM-MSCs but not both AT-MSCs and pMSCs express cytokine receptors (IFNγR, TNFI/IIR, IL-1R, IL-4R, IL-6R, and IL-7R). In conclusion, pMSCs exhibited a typical cell surface phenotype of MSCs, which was not the same as on BM-MSCs or AT-MSCs, suggesting that the pMSCs may represent a different MSC lineage from peritoneal cavity.

Four-Pyroptosis Gene-Based Nomogram as a Novel Strategy for Predicting the Effect of Immunotherapy in Hepatocellular Carcinoma.

Background: Immunotherapy has been considered as a promising cancer treatment for hepatocellular carcinoma (HCC). However, due to the particular immune environment of the liver, identifying patients who could benefit from immunotherapy is critical in clinical practice. Methods: The pyroptosis gene expression database of 54 candidates from The Cancer Genome Atlas (TCGA) were collected to discover the critical prognostic-related pyroptosis genes. A novel pyroptosis gene model was established to calculate the risk score. Kaplan-Meier analysis and receiver operating characteristic curve (ROC) were used to verify its predictive ability. The International Cancer Genome Consortium (ICGC) data was collected as external validation data to verify the model's accuracy. We employed multiple bioinformatics tools and algorithms to evaluate the tumor immune microenvironment (TIME) and the response to immunotherapy. Results: Our study found that most pyroptosis genes were expressed differently in normal and tumor tissues and that their expression was associated with the prognosis. Then, a precise four-pyroptosis gene model was generated. The one-year area under the curves (AUCs) among the training, internal, and external validation patients were 0.901, 0.727, and 0.671, respectively. An analysis of survival data revealed that individuals had a worse prognosis than patients with low risk. The analysis of TIME revealed that the low-risk group had more antitumor cells, fewer immunosuppressive cells, stronger immune function, less immune checkpoint gene expression, and better immunotherapy response than the high-risk group. Immunophenoscore (IPS) analysis also demonstrated that the low-risk score was related to superior immune checkpoint inhibitors therapy. Conclusion: A nomogram based on the four-pyroptosis gene signature was a novel tool to predict the effectiveness of immunotherapy for HCC. Therefore, individualized treatment targeting the pyroptosis genes may influence TIME and play an essential role in improving the prognosis in HCC patients.

Resveratrol, novel application by preconditioning to attenuate myocardial ischemia/reperfusion injury in mice through regulate AMPK pathway and autophagy level.

Myocardial ischemia/reperfusion injury (MI/RI) is the main cause of deaths in the worldwide, leading to severe cardiac dysfunction. Resveratrol (RSV) is a polyphenol plant-derived compound. Our study aimed to elucidate the underlying molecular mechanism of preconditioning RSV in protecting against MI/RI. Mice were ligated and re-perfused by the left anterior descending branch with or without RSV (30 mg/kg·ip) for 7 days. Firstly, we found that RSV pretreatment significantly alleviated myocardial infarct size, improved cardiac function and decreased oxidative stress. Furthermore, RSV activated p-AMPK and SIRT1, ameliorated inflammation including the level of TNF-α and IL-1ß, and promoting autophagy level. Moreover, neonatal rat ventricular myocytes (NRVMs) and H9c2 cells with knockdown the expression of AMPK, SIRT1 or FOXO1 were used to uncover the underlying molecular mechanism for the cardio-protection of RSV. In NRVMs, RSV increased cellular viability, decreased LDH release and reduced oxidative stress. Importantly, Compound C(CpC) and EX527 reversed the effect of RSV against MI/RI in vivo and in vitro and counteracted the autophagy level induced by RSV. Together, our study indicated that RSV could alleviate oxidative stress in cardiomyocytes through activating AMPK/SIRT1-FOXO1 signallingpathway and enhanced autophagy level, thus presenting high potential protection on MI/RI.

Farnesoid X receptor functions in cervical cancer via the p14ARF-mouse double minute 2-p53 pathway.

BACKGROUND: Cervical cancer is the second most common cancer among women living in developing countries. Farnesoid X receptor (FXR) is a member of the nuclear receptor family, which regulates the development and proliferation of cancer. However, the role of and molecular mechanism by which FXR acts in cervical cancer are still unknown. METHODS AND RESULTS: The relationship between FXR and the proliferation of cervical cancer cell lines was detected by MTT and colony formation assays. Immunohistochemistry was used to detect the expression of FXR in cervical cancer tissue slides. Western blotting was used to detect the expression of p14ARF, mouse double minute 2 (MDM2) and p53 when FXR was overexpressed or siRNA was applied. Western blotting was used to detect the expression of MDM2 and p53 when pifithrin-α (PFT-α) was applied. FXR activation inhibited the proliferation of cervical cancer cell lines. FXR was significantly decreased in cervical squamous cell carcinoma, which was correlated with TNM stage, but not with metastasis. Overexpression of FXR activated the p14ARF-MDM2-p53 pathway. As a p53 inhibitor, PFT-α increased MDM2 in Lenti-vector cells, but had no effect on MDM2 in Lenti-FXR cells. CONCLUSIONS: FXR inhibits cervical cancer by upregulating the p14ARF-MDM2-p53 pathway. Activation of FXR may be a potential strategy for the treatment of cervical cancer.

Nine-Year Median Follow-up of Cardiotoxicity and Efficacy of Trastuzumab Concurrently With Anthracycline-Based and Anthracycline-Free Neoadjuvant Chemotherapy in HER2-Positive Breast Cancer Patients.

BACKGROUND: The combination of trastuzumab with anthracycline chemotherapy drugs is associated with synergistic cardiotoxicity. The aim of this study is to compare the efficacy and late-onset cardiac toxicity of neoadjuvant chemotherapy regimens, trastuzumab plus paclitaxel followed by 5-fluorouracil, epirubicin, and cyclophosphamide (PH-FECH) versus trastuzumab plus docetaxel and carboplatin (TCH), for human epidermal growth factor receptor 2-positive (HER2+) breast cancer (BC). METHODS: Patients with HER2+ BC who received neoadjuvant chemotherapy with PH-FECH or TCH between 2002 and 2009 at MD Anderson Cancer Center were included. The primary endpoint was progression-free survival (PFS). Secondary endpoints included pathological complete response (pCR), overall survival, cardiac events, breast cancer-specific survival, noncardiac toxicities, and chemotherapy interruption. RESULTS: We identified 249 consecutive patients (184 who received PH-FECH and 65 who received TCH). The 10-year PFS was higher in the PH-FECH group than in the TCH group (83.6% vs. 72.2%; P = .044). The pCR rate was significantly higher in the PH-FECH group (58.2% vs. 41.5%; P = .021). The rate of cardiac events was higher in the PH-FECH group, but the difference was not significant (13.0% vs. 7.7%; P = .352). More patients developed late-onset cardiotoxicity in the PH-FECH group (3.8%) than in the TCH group (1.5%). Hypertension (odds ratio, 4.402 [95% confidence interval, 1.020-18.998]; P = .047) was an independent predictor of late-onset cardiotoxicity. CONCLUSIONS: Both neoadjuvant regimens are effective and tolerable in patients with HER2+ BC. The PH-FECH regimen offers a higher pCR rate and higher PFS but no difference in overall survival or breast cancer-specific survival. Higher frequency of cardiac toxicity with PH-FECH was noted.

Sevoflurane Alleviates Myocardial Ischemia Reperfusion Injury by Inhibiting P2X7-NLRP3 Mediated Pyroptosis.

Myocardial ischemia is common in aging population. This study investigates the protective effect of Sevoflurane on myocardial ischemia reperfusion injury (MIRI) and its underlying mechanism. A total of 87 patients with a history of myocardial ischemia who underwent abdominal surgery with Sevoflurane general anesthesia were recruited in the study. The clinical data, blood pressure, heart rate, pressure-rate quotient (PRQ) and rate-pressure product (RPP) were recorded. Serum samples were collected and heart-type fatty acid binding protein (H-FABP), ischemia modified albumin (IMA), interleukin-1ß (IL-1ß), and interleukin-18 (IL-18) were measured to observe whether Sevoflurane anesthesia had protective effect on myocardium. In addition, MIRI rats and hypoxia/reoxygenation (H/R) injury cell model was established using neonatal rat ventricular myocytes (NRVM). Rats or NRVM were pretreated with sevoflurane for 45min before hypoxia. The mRNA expression of purinergic receptor-7 (P2X7) and NLR family pyrin domain containing 3(NLRP3) were examined. The protein expression of P2X7, NLRP3, apoptosis-associated speck-like protein (ASC), cysteine aspartic acid specific protease-1(Caspase-1), Gasdermin-D (GSDMD), Bcl-2 Associated X Protein (Bax), B-cell lymphoma-2 (Bcl-2) in myocardial tissue and cells were evaluated. The serum contents of IL-1ß, IL-18, Malondialdehyde (MDA), Superoxide dismutase (SOD), Lactate dehydrogenase (LDH), Creatine kinase (CK), and Creatine kinase isoenzymes (CK-MB) were measured. The cellular localization and fluorescence intensity of NLRP3 and ASC in cells were detected. It was found that the secretion of IL-1ß and IL-18 decreased in the patients. After I45 min/R3h in SD rats and H3h/R1h in NRVM, the protein expressions of P2X7, NLRP3, ASC, Caspase-1 and GSDMD were increased, the release of IL-1ß, IL-18, CK, CK-MB, LDH and MDA were increased, and SOD activity was decreased. Sevoflurane treatment inhibited the high expression of P2X7, NLRP3, ASC, Caspase-1 and GSDMD, inhibited the release of LDH, CK,CK-MB and MDA in cells, and improved the activity of SOD, indicating that Sevoflurane alleviated the damage of MIRI of rats and H/R of NRVM, and had myocardial protective effect. Taken together, our study suggests that Sevoflurane inhibited the expression of IL-1ß, IL-18 and GSDMD by inhibiting the P2X7-NLRP3 signaling pathway. It reduced the H/R injury of cardiomyocytes and protected the cardiac function by regulating inflammatory reaction and pyroptosis.

Combination of curcumin with N-n-butyl haloperidol iodide inhibits hepatocellular carcinoma malignant proliferation by downregulating enhancer of zeste homolog 2 (EZH2) - lncRNA H19 to silence Wnt/ß-catenin signaling.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common cause of cancer-related death worldwide. Curcumin (C) has been extensively investigated in different types of malignancies, including hepatocellular carcinoma, but its physicochemical properties have significantly influenced its clinical use. Several approaches are being explored to enhance curcumin's therapeutic response, including its combination with various drugs. PURPOSE: This study aimed to evaluate the anti-tumor effect of curcumin (C) in combination with F2 (N-n-butyl haloperidol iodide) on hepatocellular carcinoma and its potential underlying mechanism in vitro and in vivo. METHODS: Cell proliferation was evaluated by CCK-8 and colony formation assays, and apoptosis was measured by flow cytometry. The migratory and invasive abilities of Hep3B and SMMC-7721 cells were measured by wound-healing and matrigel transwell assays. In order to investigate the molecular pathways, various experiments such as western blotting, qPCR, RNA-seq, immunostaining and transfection were performed. To evaluate the anti-HCC effects in vivo, a xenograft tumor model was used. RESULTS: Our findings showed that the combination of curcumin (C) & F2 (F2C) strongly inhibited malignant proliferation and migration in SMMC-7721 and Hep3B cells. The F2C treatment downregulates enhancer of zeste homolog 2 (EZH2) transcription and protein expression, which is key epigenetic regulator responsible for HCC development. Moreover, the inhibition of EZH2 by F2C led to Wnt/ß-catenin signaling inhibition by decreasing tri-methylation of histone H3 at lysine 27 (H3K27me3) and long non-coding RNA H19 expression. The inhibition of F2C was associated with the suppression of tumorigenicity in xenograft HCC models. CONCLUSION: These findings suggested that, F2C inhibited HCC formation, migration and its modulatory mechanism seemed to be associated with downregulation of EZH2, silencing Wnt/ß-catenin signaling by interacting with H19, suggesting that F2C may be a promising drug in the clinical treatment of HCC.

Interaction of Hepatitis B Virus X Protein with the Pregnane X Receptor Enhances the Synergistic Effects of Aflatoxin B1 and Hepatitis B Virus on Promoting Hepatocarcinogenesis.

BACKGROUND AND AIMS: Hepatitis B virus (HBV) infection has been found to increase hepatocellular sensitivity to carcinogenic xenobiotics, by unknown mechanisms, in the generation of hepatocellular carcinoma. The pregnane X receptor (PXR) is a key regulator of the body's defense against xenobiotics, including xenobiotic carcinogens and clinical drugs. In this study, we aimed to investigate the molecular mechanisms of HBV X protein (HBx)-PXR signaling in the synergistic effects of chemical carcinogens in HBV-associated hepatocarcinogenesis. METHODS: The expression profile of PXR-cytochrome p450 3A4 (CYP3A4) signaling was determined by PCR, western blotting, and tissue microarray. Cell viability and aflatoxin B1 (AFB1) cytotoxicity were measured using the cell counting kit-8 assay. Target gene expression was evaluated using transient transfection and real time-PCR. The genotoxicity of AFB1 was assessed in newborn mice with a single dose of AFB1. RESULTS: HBx enhanced the hepatotoxicity of AFB1 by activating CYP3A4 and reducing glutathione S-transferase Mu 1 (GSTM1) in cell lines. Activation of PXR by pregnenolone 16α-carbonitrile increased AFB1-induced liver tumor incidence by up-regulating oncogenic KRAS to enhance interleukin (IL)-11:IL-11 receptor subunit alpha-1 (IL11RA-1)-mediated inflammation in an HBx transgenic model. CONCLUSIONS: Our finding regarding AFB1 toxicity enhancement by an HBx-PXR-CYP3A4/ GSTM1-KRAS-IL11:IL11RA signaling axis provides a rational explanation for the synergistic effects of chemical carcinogens in HBV infection-associated hepatocarcinogenesis.

The Nuclear Farnesoid X Receptor Reduces p53 Ubiquitination and Inhibits Cervical Cancer Cell Proliferation.

The role of farnesoid X receptor (FXR) in cervical cancer and the underlying molecular mechanism remain largely unknown. Therefore, this study aimed to assess the mechanism of FXR in cervical cancer. Western blot, qRT-PCR, and immunohistochemistry demonstrated that FXR was significantly reduced in squamous cell carcinoma tissues, although there were no associations of metastasis and TNM stage with FXR. In Lenti-FXR cells obtained by lentiviral transfection, the overexpression of FXR reduced cell viability and colony formation. Compared with the Lenti-Vector groups, the overexpression of FXR induced early and late apoptosis and promoted G1 arrest. With time, early apoptosis decreased, and late apoptosis increased. In tumor xenograft experiments, overexpression of FXR upregulated small heterodimer partner (SHP), murine double minute-2 (MDM2), and p53 in the nucleus. Co-immunoprecipitation (Co-IP) showed that SHP directly interacted with MDM2, which is important to protect p53 from ubiquitination. Nutlin3a increased MDM2 and p53 amounts in the Lenti-Vector groups, without effects in the Lenti-FXR groups. Silencing SHP reduced MDM2 and p53 levels in the Lenti-FXR groups, and Nutlin3a counteracted these effects. Taken together, these findings suggest that FXR inhibits cervical cancer via upregulation of SHP, MDM2, and p53.

Overexpressed WDR3 induces the activation of Hippo pathway by interacting with GATA4 in pancreatic cancer.

BACKGROUND: WD repeat domain 3 (WDR3) is involved in a variety of cellular processes including gene regulation, cell cycle progression, signal transduction and apoptosis. However, the biological role of WDR3 in pancreatic cancer and the associated mechanism remains unclear. We seek to explore the immune-independent functions and relevant mechanism for WDR3 in pancreatic cancer. METHODS: The GEPIA web tool was searched, and IHC assays were conducted to determine the mRNA and protein expression levels of WDR3 in pancreatic cancer patients. MTS, colony formation, and transwell assays were conducted to determine the biological role of WDR3 in human cancer. Western blot analysis, RT-qPCR, and immunohistochemistry were used to detect the expression of specific genes. An immunoprecipitation assay was used to explore protein-protein interactions. RESULTS: Our study proved that overexpressed WDR3 was correlated with poor survival in pancreatic cancer and that WDR3 silencing significantly inhibited the proliferation, invasion, and tumor growth of pancreatic cancer. Furthermore, WDR3 activated the Hippo signaling pathway by inducing yes association protein 1 (YAP1) expression, and the combination of WDR3 silencing and administration of the YAP1 inhibitor TED-347 had a synergistic inhibitory effect on the progression of pancreatic cancer. Finally, the upregulation of YAP1 expression induced by WDR3 was dependent on an interaction with GATA binding protein 4 (GATA4), the transcription factor of YAP1, which interaction induced the nuclear translocation of GATA4 in pancreatic cancer cells. CONCLUSIONS: We identified a novel mechanism by which WDR3 plays a critical role in promoting pancreatic cancer progression by activating the Hippo signaling pathway through the interaction with GATA4. Therefore, WDR3 is potentially a therapeutic target for pancreatic cancer treatment.

Partial abrogation of FXR-KNG1 signaling by carboxyl-terminal truncated HBx-C30 in hepatitis B virus-associated hepatocellular carcinoma.

Hepatitis B virus (HBV) X protein (HBx) is a key regulator of HBV-associated hepatocarcinogenesis. C-terminal-truncated HBx is frequently detected in hepatocellular carcinoma (HCC). The role of HBx, especially C-terminal-truncated HBx, in HCC pathogenesis has been controversial. To elucidate the biological role of C-terminal-truncated HBx underlying HBV-associated hepato-oncogenesis, we constructed a vector expressing HBx-C30 (deletion of 30 aa from the C terminus of HBx) and functionally analyzed its regulation on farnesoid X receptor (FXR) signaling, which is known to inhibit hepatocarcinogenesis. In the present study, we found full-length HBx and HBx C-terminal truncation coexist in HCC, and both full length HBx and HBx-C30 can activate FXR signaling. Moreover, HBx-C30 weakly coactivates FXR-KNG1 signaling compared to full-length HBx.

Clusterin regulates macrophage expansion, polarization and phagocytic activity in response to inflammation in the kidneys.

Clusterin (CLU) is a multifunctional protein localized extracellularly and intracellularly. Although CLU-knockout (KO) mice are more susceptible to renal ischemia-reperfusion injury (IRI), the mechanisms underlying the actions of CLU in IRI are not fully understood. Macrophages are key regulators of IRI severity and tissue repair. Therefore, we investigated the role of CLU in macrophage polarization and phagocytosis. Renal IRI was induced in wild-type (WT) or CLU-KO C57BL/6 mice by clamping the renal pedicles for 30 min at 32°C. Peritoneal macrophages were activated via an intraperitoneal injection of lipopolysaccharide (LPS). Renal tissue damage was examined using histology, whereas leukocyte phenotypes were assessed using flow cytometry and immunohistochemistry. We found that monocytes/macrophages expressed the CLU protein that was upregulated by hypoxia. The percentages of macrophages (F4/80+ , CD11b+ or MAC3+ ) infiltrating the kidneys of WT mice were significantly less than those in CLU-KO mice after IRI. The M1/M2 phenotype ratio of the macrophages in WT kidneys decreased at day 7 post-IRI when the injury was repaired, whereas that in KO kidneys increased consistently as tissue injury persisted. In response to LPS stimulation, WT mice produced fewer M1 macrophages, but not M2, than the control did. Phagocytosis was stimulated by CLU expression in macrophages compared with the CLU null controls and by the exogenous CLU protein. In conclusion, CLU suppresses macrophage infiltration and proinflammatory M1 polarization during the recovery period following IRI, and enhances phagocytic activity, which may be partly responsible for tissue repair in the kidneys of WT mice after injury.

Transcriptomic identification of HBx-associated hub genes in hepatocellular carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most common malignancies around the world. Among the risk factors involved in liver carcinogenesis, hepatitis B virus (HBV) X protein (HBx) is considered to be a key regulator in hepatocarcinogenesis. Whether HBx promotes or protects against HCC remains controversial, therefore exploring new HBx-associated genes is still important. METHODS: HBx was overexpressed in HepG2, HepG2.2.15 and SMMC-7721 cell lines, primary mouse hepatocytes and livers of C57BL/6N mice. High-throughput RNA sequencing profiling of HepG2 cells with HBx overexpression and related differentially-expressed genes (DEGs), pathway enrichment analysis, protein-protein interaction networks (PPIs), overlapping analysis were conducted. In addition, Gene Expression Omnibus (GEO) and proteomic datasets of HBV-positive HCC datasets were used to verify the expression and prognosis of selected DEGs. Finally, we also evaluated the known oncogenic role of HBx by oncogenic array analysis. RESULTS: A total of 523 DEGs were obtained from HBx-overexpressing HepG2 cells. Twelve DEGs were identified and validated in cells transiently transfected with HBx and three datasets of HBV-positive HCC transcription profiles. In addition, using the Kaplan-Meier plotter database, the expression levels of the twelve different genes were further analyzed to predict patient outcomes. CONCLUSION: Among the 12 identified HBx-associated hub genes, HBV-positive HCC patients expressing ARG1 and TAT showed a good overall survival (OS) and relapse-free survival (RFS). Thus, ARG1 and TAT expression could be potential prognostic markers.

LncRNA HOTAIR-mediated MTHFR methylation inhibits 5-fluorouracil sensitivity in esophageal cancer cells.

BACKGROUND: Esophageal cancer (EC) represents one of the most aggressive digestive neoplasms globally, with marked geographical variations in morbidity and mortality. Chemoprevention is a promising approach for cancer therapy, while acquired chemoresistance is a major obstacle impeding the success of 5-fluorouracil (5-FU)-based chemotherapy in EC, with the mechanisms underlying resistance not well-understood. In the present study, we focus on exploring the role of long non-coding RNA (lncRNA) HOTAIR in EC progression and sensitivity of EC cells to 5-FU. METHODS: Paired cancerous and pre-cancerous tissues surgically resected from EC patients were collected in this study. Promoter methylation of the MTHFR was assessed by methylation-specific PCR. RIP and ChIP assays were adopted to examine the interaction of DNA methyltransferases (DNMTs) with lncRNA HOTAIR and MTHFR, respectively. EC cells resistant to 5-FU were induced by step-wise continuous increasing concentrations of 5-FU. The sensitivity of EC cells to 5-FU in vivo was evaluated in nude mice treated with xenografts of EC cells followed by injection with 5-FU (i.p.). RESULTS: We found reciprocal expression patterns of lncRNA HOTAIR and MTHFR in EC tissues and human EC cells. Interference with lncRNA HOTAIR enhanced 5-FU-induced apoptosis, exhibited anti-proliferative activity, and reduced promoter methylation of the MTHFR in EC cells. Besides, overexpression of MTHFR attenuated the acquired chemoresistance induced by overexpression of lncRNA HOTAIR in EC cells. At last, enhanced chemosensitivity was observed in vivo once nude mice xenografted with lncRNA HOTAIR-depleted EC cells. CONCLUSION: Together, our study proposes that pharmacologic targeting of lncRNA HOTAIR sensitizes EC cells to 5-FU-based chemotherapy by attenuating the promoter hypermethylation of the MTHFR in EC.

The significance of Runx2 mediating alcohol-induced Brf1 expression and RNA Pol III gene transcription.

Runx2 (Runt-related transcription factor 2) is a key transcription factor which is associated with osteoblast differentiation and expressed in ER+ (estrogen receptor positive) human breast cancer cell lines. Runx2 also participates in mammary gland development. Deregulation of RNA Pol III genes (polymerase III-dependent genes) is tightly linked to tumor development, while Brf1 (TFIIB-related factor 1) specifically regulates these gene transcription. However, nothing is known about the effect of Runx2 on Brf1 expression and Pol III gene transcription. Expression of Runx2, Brf1 and Pol III genes from the samples of human breast cancer and cell culture model were determined by the assays of RT-qPCR, immunoblot, luciferase reporter activity, immunohistochemistry, chromatin immunoprecipitation and Immunofluorescence. High expression of Runx2 is observed in the cases of breast cancer. The patients of high Runx2 expression at early stages display longer survival period, whereas the cases of high Runx2 at advanced stages reveal faster recurrence. The identification of signaling pathway indicates that JNK1 and c-Jun mediate Runx2 transcription. Repression of Runx2 reduces Brf1 expression and Pol III gene transcription. Further analysis indicates that Runx2 is colocalized with Brf1 in nucleus of breast cancer tissue. Both Runx2 and Brf1 synergistically modulate Pol III gene transcription. These studies indicate that Brf1 overexpression is able to be used as an early diagnosis biomarker of breast cancer, while high Runx2 expression indicates long survival period and faster recurrence. Runx2 mediates the deregulation of Brf1 and Pol III genes and its abnormal expression predicts the worse prognosis of breast cancer.

SDF-1/CXCR4 axis enhances the immunomodulation of human endometrial regenerative cells in alleviating experimental colitis.

Endometrial regenerative cells (ERCs) are a new type of mesenchymal-like stromal cells, and their therapeutic potential has been tested in a variety of disease models. SDF-1/CXCR4 axis plays a chemotaxis role in stem/stromal cell migration. The aim of the present study was to investigate the role of SDF-1/CXCR4 axis in the immunomodulation of ERCs on the experimental colitis. The immunomodulation of ERCs in the presence or absence of pretreatment of SDF-1 or AMD3100 was examined in both in vitro cell culture system and dextran sulphate sodium-induced colitis in mice. The results showed that SDF-1 increased the expression of CXCR4 on the surface of ERCs. As compared with normal ERCs, the SDF-1-treated, CXCR4 high-expressing ERCs more significantly suppressed dendritic cell population as well as stimulated both type 2 macrophages and regulatory T cells in vitro and in vivo. Meanwhile, SDF-1-pretreated ERCs increased the generation of anti-inflammatory factors (e.g., IL-4, IL-10) and decreased the pro-inflammatory factors (e.g., IL-6, TNF-α). In addition, SDF-1-pretreated CM-Dil-labeled ERCs were found to engraft to injured colon. Our results may suggest that an SDF-1-induced high level of CXCR4 expression enhances the immunomodulation of ERCs in alleviating experimental colitis in mice.

Angiotensin II confers resistance to apoptosis in cardiac myofibroblasts through the AT1/ERK1/2/RSK1 pathway.

Myofibroblast apoptosis is essential for normal resolution of wound repair, including cardiac infarction repair. Impaired cardiac myofibroblast (CMF) apoptosis is associated with excessive extracellular matrix (ECM) deposition, which could be responsible for pathological cardiac fibrosis. Conventionally, angiotensin II (Ang II), a soluble peptide, is implicated in fibrogenesis because it induces cardiac fibroblast (CFb) proliferation, differentiation, and collagen synthesis. However, the role of Ang II in regulation of CMF survival and apoptosis has not been fully clarified. In this report, we cultured neonatal rat CFbs, which transform into CMFs after passage 3 (6-8 days), and investigated the effects of Ang II on CMFs challenged by TNF-α combined with cycloheximide and the underlying mechanisms. Here, we show that Ang II rapidly activates MAPKs but not AKT in CMFs and confers apoptosis resistance, as evidenced by the inhibition of caspase-3 cleavage, early apoptotic cells and late apoptotic cells. This inhibitory effect of Ang II was reversed by blockade of AT1 or inactivation of ERK1/2 or RSK1 but not AT2, indicating that activation of the prosurvival AT1/ERK1/2/RSK1 signaling pathway mediates apoptosis resistance. TGF-ß, a latent fibrotic factor, was found to have no relation to Ang II-induced apoptosis resistance in our study. Furthermore, Ang II-mediated apoptosis resistance, which was conferred by activation of the AT1/ERK1/2/RSK1 signaling pathway, was also confirmed in human adult ventricular cardiac myofibroblasts. Collectively, our findings suggest a novel profibrotic mechanism of Ang II in which it promotes myofibroblast resistance to apoptosis in addition to classical mechanisms, providing a potential novel therapeutic approach by targeting prosurvival signaling pathways. © 2018 IUBMB Life, 71(1):261-276, 2019.

The influences of red wine in phenotypes of human cancer cells.

Alcohol intake increases the risk of cancer development. Approximately 3.6% human cancers worldwide derive from chronic alcohol drinking, including oral, liver, breast and other organs. Our studies in vivo and in vitro have demonstrated that diluted ethanol increase RNA Pol III gene transcription and promotes cell proliferation and transformation, as well as tumor formation. However, it is unclear about the effect of red wines on the human cancer cells. In present study, we investigated the roles of red wine in human cancer cell growth, colony formation and RNA Pol III gene transcription. Low concentration (12.5 mM to 25 mM) of ethanol enhances cell proliferation of breast and esophageal cancer lines, whereas its higher concentration (100 mM to 200 mM) slightly decreases the rates. In contrast, red wines significantly repress cell proliferation of different human cancer lines from low dose to high dose. The results reveal that the red wine also inhibits colony formation of human breast cancer and esophageal carcinoma cells. The effects of repression on different human cancer lines are in a dose-dependent manner. Further analysis indicates that ethanol increases RNA Pol III gene transcription, whereas the red wines significantly reduce transcription of the genes. Interestingly, the effects of mature wine (brick red) on cancer cell phenotypes are much stronger than young wine (intense violet). Together, these new findings suggest that red wines may contain some bioactive components, which are able to inhibit human cancer cell growth and colony formation.

Verteporfin enhances the sensitivity of LOVO/TAX cells to taxol via YAP inhibition.

Studies have reported that taxol (TAX) is an effective drug for the treatment of colorectal cancer; however, its application inevitably results in drug resistance. Overexpression of Yes-associated protein (YAP) is considered one of the factors that cause TAX resistance, which may be inhibited by verteporfin (VP) treatment. The present study aimed to confirm the role of YAP in TAX resistance and to investigate whether the drug sensitivity of the TAX-resistant LOVO/TAX cell line to TAX is affected by VP treatment. The role of YAP in TAX resistance was first determined through vector-mediated overexpression and inhibition of YAP in cells. Reverse-transcription quantitative polymerase chain reaction and western blot analysis were performed for detection of associated mRNA and protein, respectively. An MTT assay was used to detect the drug sensitivity of cells to TAX. The results suggested that compared with that in the native LOVO cell line, YAP expression was significantly increased in LOVO/TAX cells. YAP gene silencing markedly enhanced the drug sensitivity of LOVO/TAX cells to TAX and, on the contrary, the drug sensitivity notably declined when YAP was overexpressed in LOVO cells. The results indicated that YAP gene expression and TAX resistance were correlated. VP treatment suppressed YAP expression and increased the drug sensitivity of LOVO/TAX cells to TAX in a dose-dependent manner. In addition, compared with VP alone, VP and TAX combination therapy had a greater inhibitory effect on YAP expression. VP treatment enhanced the drug sensitivity of LOVO/TAX cells to TAX through inhibiting YAP expression.

Skin Allografting Activates Anti-tumor Immunity and Suppresses Growth of Colon Cancer in Mice.

INTRODUCTION: The tumor cells could escape from the immune elimination through the immunoediting mechanisms including the generation of immunosuppressive or immunoregulative cells. By contrast, allograft transplantation could activate the immune system and induce a strong allogenic response. The aim of this study was to investigate the efficacy of allogenic skin transplantation in the inhibition of tumor growth through the activation of allogenic immune response. METHODS: Full-thickness skin transplantation was performed from C57BL/6 (H-2b) donors to BALB/c (H-2d) recipients that were receiving subcutaneous injection of isogenic CT26 colon cancer cells (2 × 106 cells) at the same time. The tumor size and pathological changes, cell populations and cytokine profiles were evaluated at day 14 post-transplantation. RESULTS: The results showed that as compared to non-transplant group, the allogenic immune response in the skin-grafting group inhibited the growth of tumors, which was significantly associated with increased numbers of intra-tumor infiltrating lymphocytes, increased populations of CD11c+MHC-classII+CD86+ DCs, CD3+CD4+ T cells, CD3+CD8+ T cells, and CD19+ B cells, as well as decreased percentage of CD4+CD25+Foxp3+ T cells in the spleens. In addition, the levels of serum IgM and IgG, tumor necrosis factor (TNF)-α and interferon (IFN)-γ were significantly higher within the tumor in skin transplant groups than that in non-transplant group. CONCLUSIONS: Allogenic skin transplantation suppresses the tumor growth through activating the allogenic immune response, and it may provide a new immunotherapy option for the clinical refractory tumor treatment.