Mechanistic insights into super-enhancer-driven genes as prognostic signatures in patients with glioblastoma.

BACKGROUND: Glioblastoma (GBM) is one of the most common malignant brain tumors in adults and is characterized by high aggressiveness and rapid progression, poor treatment, high recurrence rate, and poor prognosis. Although super-enhancer (SE)-driven genes haven been recognized as prognostic markers for several cancers, whether it can be served as effective prognostic markers for patients with GBM has not been evaluated. METHODS: We first combined histone modification data with transcriptome data to identify SE-driven genes associated with prognosis in patients with GBM. Second, we developed a SE-driven differentially expressed genes (SEDEGs) risk score prognostic model by univariate Cox analysis, KM survival analysis, multivariate Cox analysis and least absolute shrinkage and selection operator (LASSO) regression. Its reliability in predicting was verified by two external data sets. Third, through mutation analysis, immune infiltration, we explored the molecular mechanisms of prognostic genes. Next, Genomics of Drug Sensitivity in Cancer (GDSC) and the Connectivity Map (cMap) database were employed to assess different sensitivities to chemotherapeutic agents and small-molecule drug candidates between high- and low-risk patients. Finally, SEanalysis database was chosen to identify SE-driven transcription factors (TFs) regulating prognostic markers which will reveal a potential SE-driven transcriptional regulatory network. RESULTS: First, we developed a 11-gene risk score prognostic model (NCF2, MTHFS, DUSP6, G6PC3, HOXB2, EN2, DLEU1, LBH, ZEB1-AS1, LINC01265, and AGAP2-AS1) selected from 1,154 SEDEGs, which is not only an independent prognostic factor for patients, but also can effectively predict the survival rate of patients. The model can effectively predict 1-, 2- and 3-year survival of patients and was validated in external Chinese Glioma Genome Atlas (CGGA) and Gene Expression Omnibus (GEO) datasets. Second, the risk score was positively correlated with the infiltration of regulatory T cell, CD4 memory activated T cell, activated NK cell, neutrophil, resting mast cell, M0 macrophage, and memory B cell. Third, we found that high-risk patients showed higher sensitivity than low-risk patients to both 27 chemotherapeutic agents and 4 small-molecule drug candidates which might benefit further precision therapy for GBM patients. Finally, 13 potential SE-driven TFs imply how SE regulates GBM patient's prognosis. CONCLUSION: The SEDEG risk model not only helps to elucidate the impact of SEs on the course of GBM, but also provides a bright future for prognosis determination and choice of treatment for GBM patients.

Regulatory Effect of JAK2/STAT3 on the Immune Function of Endotoxin-tolerant Dendritic Cells and its Involvement in Acute Liver Failure.

Background and Aims: Acute liver failure (ALF) is a potentially fatal clinical syndrome with no effective treatment. This study aimed to explore the role of Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway in modulating the phenotype and immune function of endotoxin-tolerant dendritic cells (ETDCs). In addition, we explored the use of EDTCs in an experimental model of ALF and investigated the associated mechanisms. Methods: In the in vitro experiment, ETDCs were transfected with adenovirus to induce SOCS1+/+ETDCs and SOCS1-/-ETDCs. Thereafter, costimulatory molecules and mixed lymphocyte reaction were assessed. Experimental mice were randomly divided into normal control, ALF, ALF+mock-ETDCs, ALF+SOCS1+/+ETDCs, ALF+AG490, and ALF+AG490+SOCS1+/+ETDCs groups. We examined the therapeutic effect of adoptive cellular immunotherapy by tail-vein injection of target ETDCs 12 h before ALF modeling. AG490, a JAK2/STAT3 inhibitor, was used in the in vivo experiment to further explore the protective mechanism of SOCS1+/+ETDCs. Results: Compared with control ETDCs, SOCS1+/+ETDCs had lower expression of costimulatory molecules, weaker allostimulatory ability, lower levels of IL-6 and TNF-α expression and higher IL-10 secretion. SOCS1-/-ETDCs showed the opposite results. In the in vivo experiments, the ALF+SOCS1+/+ETDCs and ALF+AG490+SOCS1+/+ETDCs groups showed less pathological damage and suppressed activation of JAK2/STAT3 pathway. The changes were more pronounced in the ALF+AG490+SOCS1+/+ETDCs group. Infusion of SOCS1+/+ETDCs had a protective effect against ALF possibly via inhibition of JAK2 and STAT3 phosphorylation. Conclusions: The SOCS1 gene had an important role in induction of endotoxin tolerance. SOCS1+/+ETDCs alleviated lipopolysaccharide/D-galactosamine-induced ALF by downregulating the JAK2/STAT3 signaling pathway.

Post-interventional Evaluation and Follow-Up in Children With Patent Ductus Arteriosus Complicated With Moderate to Severe Pulmonary Arterial Hypertension: A Retrospective Study.

Background: Transcatheter closure is an important treatment for patent ductus arteriosus (PDA) complicated with moderate and severe pulmonary arterial hypertension (PAH). This report presents our experience with transcatheter closure of PDA complicated with moderate and severe PAH. Methods: The 49 cases of PDA complicated with moderate and severe PAH were collected in the Second Affiliated Hospital and Yuying Children's Hospital from January 2014 to December 2019 with transcatheter closure of PDA and follow-up. All patients were invited for transthoracic echocardiography, electrocardiogram, and thoracic radiography check-up. Results: Device implantation was successful in 48 of 49 patients (98.0%). Among them, 30 cases were in the PAH after defect correction (CD) group, and 19 examples were in the Non-PAH after defect correction (NCD) group. Pulmonary systolic pressure, left atrial diameter, and left ventricular end-diastolic diameter immediately after interventional therapy and 6 months later were lower than the pre-operative levels (p < 0.05). The incidence of the immediate residual shunt (RS) in this study was 34.9%, most of which were minimal amount shunt. RS disappeared in all patients within 1 year of therapy. Four patients had thrombocytopenia and one patient had left pulmonary artery stenosis. No other serious adverse event occurred during the follow-up period. The pressure gradient tricuspid valve regurgitation (PGTI) and the right heart catheterization (RHC) consistency points were 93.75% (15/16) and were within the 95% consistency limit by the Bland-Altman method. The Logistic regression analysis concluded that the pre-operative Pp/Ps and the narrowest diameter of PDA are risk factors for post-operative PAH (p < 0.05). The cut-off point of the pre-operative Pp/Ps and the narrowest diameter of PDA were calculated to be 0.595 and 4.75 mm, respectively. Conclusion: Interventional occlusion in children with PDA complicated with moderate and severe PAH is safe, effective, and has few complications. Targeted drug therapy has a good clinical effect. The narrowest diameter of PDA and the pre-operative Pp/Ps may be one of the risk factors of residual PAH after interventional therapy.

Changes and Clinical Significance of PIVKA-II in Hepatitis E Patients.

Increased protein induced by vitamin K absence or antagonist-II (PIVKA-II) levels had been widely reported in patients with hepatocellular carcinoma (HCC) and chronic hepatitis. However, the role of PIVKA-II in hepatitis E is unclear. The aim of this study was to clarify the changes related with PIVKA-II and its clinical significance in hepatitis E. We enrolled 84 patients with hepatitis E hospitalized in two hospitals from December 2019 to June 2021. The levels of serum PIVKA-II and related serological indicators in the patients were determined to elucidate the role of PIVKA-II in hepatitis E. We observed that 59.51% (50/84) of patients showed an increase in PIVKA-II levels. Compared with the normal PIVKA-II group (<32 mAU/L), patients in the elevated PIVKA-II group (>32 mAU/L) had much higher serum total bilirubin (TBIL), direct bilirubin (DBIL), indirect bilirubin (IBIL), and total bile acid (TBA) levels (p < 0.05 for each). Compared with the slightly elevated PIVKA-II group (32-125 mAU/L), patients in the significantly elevated PIVKA-II group (>125 mAU/L) had much lower serum albumin, alanine aminotransferase (ALT), aspartate transaminase (AST) levels, and longer days for the hospital stay (p < 0.05 for each). The association of PIVKA-II with TBIL and DBIL was an inverted U-shaped curve with an inflection point at 199.1 mAU/L). The association of PIVKA-II with IBIL was a U-shaped curve with an inflection point at 18.6 mAU/L while the association of PIVKA-II with albumin was an inverted U-shaped curve with an inflection point at 18.6 mAU/L. With the improvement of the disease, PIVKA-II levels were gradually decreased and finally returned to normal. This trend was consistent with that of bilirubin, and a peak appeared in the third week. Therefore, findings from our study show that the increase in PIVKA-II levels can be related to the degree of hepatic insufficiency in patients with hepatitis E, wherein PIVKA-II levels are transiently increased, and the trend of change can be related to the disease course.

STAT3 Suppresses Cardiomyocytes Apoptosis in CVB3-Induced Myocarditis Via Survivin.

Background: Viral myocarditis (VMC) is a common inflammatory cardiovascular disease with unclear mechanisms, which mainly affects children and adolescents. Apoptosis is the key to CVB3-induced myocarditis, and blocking this process may be beneficial to the therapy of VMC. Hence, this study aimed to explore the protective function of STAT3 on cardiomyocyte apoptosis of VMC and its underlying mechanisms. Methods and Results: In this research, we confirmed that STAT3 was significantly activated in both animal and cell models of VMC. To further clarify what role did STAT3 play in VMC, AG490, an inhibitor of STAT3, was used to suppress p-STAT3. Our results demonstrated that decreased expression of p-STAT3 caused by AG490 significantly aggravated severity of VMC with elevated myocardial inflammation, deteriorative ventricular systolic function and increased mortality. It suggested that STAT3 plays a protective role in VMC. To further identify the anti-apoptosis impact that activated STAT3 made, we constructed lentivirus to regulate the expression of STAT3 in NMCs. We found that up-regulated activated STAT3 attenuated cardiomyocyte apoptosis, but down-regulated one aggravated that, which verified activated STAT3 played an anti-apoptosis role in VMC. Following that, we explored what elements are involved in the anti-apoptotic mechanism of activated STAT3 by using survivin inhibitor YM155. The result showed the anti-apoptotic effect of activated STAT3 does not work in the case of survivin inhibition. Conclusion: Our findings demonstrated STAT3 by targeting survivin alleviated cardiomyocyte apoptosis in CVB3-induced myocarditis.

Synergic effects of artemisinin and resveratrol in cancer cells.

PURPOSE: The aim of this study was to investigate whether resveratrol (Res) combined with artemisinin (ART) possess synergistic effect on different cancer cells. MATERIALS AND METHODS: The viability of HepG2 and HeLa cells treated with ART and Res was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Combination index (CI) analysis and isobologram were used to assess the synergistic effect of ART and Res in different ratios. Wound-healing assay was used to investigate the migration rate. AO staining and fluorescent microscopy measurements were performed to detect the cell apoptosis. Reactive oxygen species (ROS) was measured with 2'7'-dichlorofluorescein diacetate (DCFH-DA). RESULTS: MTT assay indicated that ART and Res inhibited the growth of HeLa and HepG2 cells in a dose-dependent manner. The combination of ART and Res exhibited the strongest anticancer effect at the ratio of 1:2 (ART to Res). The combination of the two drugs also markedly reduced the ability of cell migration. Apoptosis analysis showed that combination of ART and Res significantly increased the apoptosis and necrosis rather than use singly. Additionally, ROS levels were elevated by combining ART with Res. CONCLUSIONS: Taken together, the present study suggested that ART and Res possessed the synergistic anti-tumor effect. ART in combination with Res could be an effective therapeutic strategy for cancer.

Resveratrol and arsenic trioxide act synergistically to kill tumor cells in vitro and in vivo.

BACKGROUND AND AIMS: Arsenic trioxide (As2O3), which used as an effective agent in the treatment of leukaemia and other solid tumors, is largely limited by its toxicity. QT prolongation, torsades de pointes and sudden heart death have been implicated in the cardiotoxicity of As2O3. The present study was designed to explore whether the combination of As2O3 and resveratrol could generate a more powerful anti-cancer effect both in vitro and in vivo. MATERIALS AND METHODS: MTT assay was performed to assess the proliferation of Hela, MCF-7 and NB4 cells. Isobolographic analysis was used to evaluate combination index values from cell viability data. The apoptosis and the cellular reactive oxygen species (ROS) level were assessed by fluorescent microscopy and flow cytometry separately in vitro. The effect of As2O3, alone and in combination with resveratrol on Hela tumor growth in an orthotopic nude mouse model was also investigated. The tumor volume and the immunohistochemical analysis of CD31, CD34 and VEGF were determined. RESULTS: Resveratrol dramatically enhanced the anti-cancer effect induced by As2O3 in vitro. In addition, isobolographic analysis further demonstrated that As2O3 and resveratrol generated a synergistic action. More apoptosis and ROS generation were observed in the combination treatment group. Similar synergistic effects were found in nude mice in vivo. The combination of As2O3 and resveratrol dramatically suppressed both tumor growth and angiogenesis in nude mice. CONCLUSIONS: Combining As2O3 with resveratrol would be a novel strategy to treat cancer in clinical practice.

Tetrandrine enhances the anticancer effects of arsenic trioxide in vitro.

Arsenic trioxide (As2O3), an effective agent to treat leukemia and other solid tumors, is largely limited by its toxicity. QT prolongation, torsades de pointes and sudden death have been implicated in the cardiotoxicity of As2O3. The present study was designed to assess whether the combination of As2O3 and tetrandrine could generate a more powerful anti-cancer effect. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was performed for detecting the proliferation of HepG2 and A549 cells treated with tetrandrine and As2O3. Fluorescent microscopy measurements and flow cytometry were carried out to evaluate the apoptosis in HepG2 cells. The cell cycle arrest of HepG2 cells was also determined by flow cytometry. The cell proliferation assay in HepG2 and A549 cells indicated that tetrandrine significantly enhanced the inhibit effect of As2O3. In addition, the following Isobolograms further demonstrated that combining As2O3 with tetrandrine generated synergism action. Tetrandrine also enhanced the apoptosis, necrosis and cell cycle arrest in As2O3-treated HepG2 cells. Our present study showed that tetrandrine can dramatically enhance the anti- cancer effect induced by As2O3. Combining As2O3 with tetrandrine would be a novel strategy to treat cancer in clinical practice.

Resveratrol protects against vinorelbine-induced vascular endothelial cell injury.

Vinorebine (VNR), a second-generation vinca alkaloid antitumor drug, caused serious local venous toxicities, such as venous irritation, phlebitis and necrotizing vasculitis. This study investigated whether resveratrol (RES), a naturally occurring polyphenol compound, could reduce the vascular injury induced by VNR. Human vascular endothelial cell line ECV-304 cells were exposed to VNR for 10 min and then cells were cultured with serum-free medium with or without RES for 24 h. MTT (3-[4, 5-dimethyl-2-thiazolyl]-2, 5-diphe-nyl-2-tetrazolium bromide) assay was used to detect the cell viability. Reactive oxygen species (ROS) was measured with 2', 7'-dichlorofluorescein diacetate (DCFH-DA). The activity of superoxide dismutase (SOD) was assessed by SOD detection kit. VNR decreased the viability of ECV-304 cells in a dose-dependent manner. Moreover, VNR caused cell apoptosis, the generation of intracellular ROS and the reduction of intracellular SOD. Interestingly, pretreatment with RES for 2 h increased the cell viability in a dose-dependent manner. Cell apoptosis, the intracellular ROS generation and the reduction of intracellular SOD induced by VNR were also attenuated when cells were pretreated with RES for 2 h. These results demonstrated that RES would protect against vascular endothelial cell injury induced by VNR. So the use of RES together with VNR may be a possible therapeutic approach to avoid the vascular injury induced by VNR.

Berberine sensitizes mutliple human cancer cells to the anticancer effects of doxorubicin in vitro.

The clinical use of doxorubicin (DOX), a potent antineoplastic agent, is limited by its serious side-effects, which include acute and chronic cumulative dose-related cardiotoxicity. Berberine (BER), a botanical alkaloid, has been reported to possess cardioprotective and antitumor effects. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-tetrazolium bromide (MTT) assay was used to detect the cell viability of A549, HeLa and HepG2 cells after each cell line was treated with DOX, BER or a combination of DOX and BER for 24 h. Apoptosis was evaluated by acridine orange staining. The results showed that BER and DOX exhibited dose-dependent inhibitory effects on A549 and HeLa cells which were likely mediated by inducing apoptosis. The same result was found in the combination group. Isobologram illustration and combination index (CI) analyses revealed that the combination of DOX and BER generates synergistic effects in A549 (CI=0.61) and HeLa (CI=0.73) cells. These findings indicate that BER sensitizes cells to the anticancer effects of DOX.

Protective effects of berberine on doxorubicin-induced hepatotoxicity in mice.

Doxorubicin, a very potent and often used anti-cancer drug, is largely limited due to the dose-related toxic effects. The present study investigated whether berberine, a natural product alkaloid, can reduce the liver injury induced by doxorubicin. Mice of either gender were randomly divided into four groups: the control group, doxorubicin group, berberine group, and berberine+doxorubicin group. In the tests, body weight, general condition and mortality of the mice were observed, and serum alanine aminotransferase and aspartate transaminase levels were determined to evaluate liver function. Furthermore, the liver was excised for determination of the weight changes, as well as histopathological analysis in the tissues. Mortality rate and significant decline in body weight, and increased plasma alanine aminotransferase and aspartate transaminase activities were observed in doxorubicin-treated mice. These changes were significantly prevented by pretreatment with berberine. Histopathological studies showed that doxorubicin caused structural injuries, such as vascular congestion, inflammatory cell infiltration, hepatocellular degeneration and necrosis, fibrosis in the liver. These histopathological changes were largely attenuated by berberine pretreatment. These findings indicate that berberine has the hepatoprotective effect on doxorubicin-induced liver injury in mice.