System Review about Function Role of ESCC Driver Gene KDM6A by Network Biology Approach.

Background. KDM6A (Lysine (K)-Specific Demethylase 6A) is the driver gene related to esophageal squamous cell carcinoma (ESCC). In order to provide more biological insights into KDM6A, in this paper, we treat PPI (protein-protein interaction) network derived from KDM6A as a conceptual framework and follow it to review its biological function. Method. We constructed a PPI network with Cytoscape software and performed clustering of network with Clust&See. Then, we evaluate the pathways, which are statistically involved in the network derived from KDM6A. Lastly, gene ontology analysis of clusters of genes in the network was conducted. Result. The network includes three clusters that consist of 74 nodes connected via 453 edges. Fifty-five pathways are statistically involved in the network and most of them are functionally related to the processes of cell cycle, gene expression, and carcinogenesis. The biology themes of clusters 1, 2, and 3 are chromatin modification, regulation of gene expression by transcription factor complex, and control of cell cycle, respectively. Conclusion. The PPI network presents a panoramic view which can facilitate for us to understand the function role of KDM6A. It is a helpful way by network approach to perform system review on a certain gene.

The p38 MAPK inhibitor SB203580 differentially modulates LPS-induced interleukin 6 expression in macrophages.

The p38 mitogen-activated protein kinase (MAPK) plays a key role in lipopolysaccharide (LPS)-induced signal transduction pathways that lead to inflammatory cytokine synthesis in macrophages; however, whether the inhibition of p38 MAPK regulates LPS-induced inflammatory cytokine expression in different types of macrophages remains the subject of debate. Herein, we assessed whether the inhibition of p38 MAPK by SB203580 regulates LPS-induced expression of the inflammatory cytokines tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6) in RAW264.7 and resident peritoneal macrophages. Lipopolysaccharide stimulation of RAW264.7 macrophages or mouse resident peritoneal macrophages significantly increased TNF-α and IL-6 production. The addition of SB203580 to cultures dramatically blocked LPS-induced TNF-α production in RAW264.7 and mouse resident peritoneal macrophages, and dramatically blocked LPS-induced IL-6 production in RAW264.7 macrophages, but not in mouse resident peritoneal macrophages. Additionally, high concentrations of SB203580 resulted in increased IL-6 production. However, LPS-stimulation significantly up-regulated the mRNA transcript levels of TNF-α and IL-6 in RAW264.7 and mouse resident peritoneal macrophages, whereas pretreatment with SB203580 dramatically down-regulated LPS-induced mRNA transcript levels of TNF-α and IL-6 in these cells. Our data show that SB203580 differentially modulates LPS-induced production of the inflammatory cytokine IL-6 in two different sources of macrophages, and that this course of regulation occurs at the IL-6 mRNA post-transcriptional stage.

New insights into COL26A1 in thyroid carcinoma: prognostic prediction, functional characterization, immunological drug target and ceRNA network.

Background: Thyroid carcinoma (THCA) is one of the most commonly diagnosed malignancies. Collagen is the main component in extracellular matrix. Rising studies have determined the oncogenic effect of collagen in cancer progression, which is intriguing to be further explored. Collagen type XXVI alpha 1 chain (COL26A1) is a newly discovered collagen subtype, functions of which still remain poorly demonstrated in THCA. Methods: Based on the transcriptome data from The Cancer Genome Atlas (TCGA) and other public databases, we conducted investigations of COL26A1 in THCA with respects to diagnostic/prognostic prediction, functional characterization, immune infiltration, chemical drug target and non-coding RNA regulatory network. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) and western blot were used to verify the expression of COL26A1 in THCA. Results: COL26A1 was significantly upregulated in THCA, and the high COL26A1 expression inferred poor prognosis [hazard ratio (HR) =4.76; 95% confidence interval (CI): 1.36-16.73; P=0.015]. The diagnostic area under the curve (AUC) of COL26A1 achieved 0.736 (95% CI: 0.669-0.802). COL26A1 was also identified as an independent prognostic predictor for THCA (HR =3.928; 95% CI: 3.716-4.151; P<0.001). Besides, logistic regression analysis indicated that age >45 years [odds ratio (OR) =1.532; 95% CI: 1.081-2.176; P=0.017], pathological stage III (OR =2.055; 95% CI: 1.314-3.184; P=0.001), tall cell subtype (OR =5.533; 95% CI: 2.420-14.957; P<0.001), residual tumor R1 (OR =1.844; 95% CI: 1.035-3.365; P=0.041) and extrathyroidal extension (OR =1.800; 95% CI: 1.225-1.660; P=0.003) were risk factors associated with high COL26A1 expression in THCA. Functional characterizations implied that COL26A1 was associated with immunological processes and oncogenic signaling pathways. High COL26A1 expression was accompanied by more abundant infiltration of immune cells and higher stromal/immune score. In addition, most immune checkpoints were significantly positively co-expressed with COL26A1, including PD-1, PD-L1 and CTLA4. Drugs including trichloroethylene, acetamide and thioacetamide etc. that can decrease the expression of COL26A1 were also identified. The predicted long noncoding RNA (lncRNA)-microRNA (miRNA)-COL26A1 regulatory axes were successfully deciphered. qRT-PCR and western blot verified the upregulation of COL26A1 in THCA. Conclusions: Our work has primarily appraised COL26A1 as a promising biomarker for diagnosis/prognosis and immuno/therapeutic target in THCA.

Huperzine A ameliorates cognitive deficits and oxidative stress in the hippocampus of rats exposed to acute hypobaric hypoxia.

Acute exposure to high altitudes can cause neurological dysfunction due to decreased oxygen availability to the brain. In this study, the protective effects of Huperzine A on cognitive deficits along with oxidative and apoptotic damage, due to acute hypobaric hypoxia, were investigated in male Sprague-Dawley rats. Rats were exposed to simulated hypobaric hypoxia at 6,000 m in a specially fabricated animal decompression chamber while receiving daily Huperzine A orally at the dose of 0.05 or 0.1 mg/kg body weight. After exposure to hypobaric hypoxia for 5 days, rats were trained in a Morris Water Maze for 5 consecutive days. Subsequent trials revealed Huperzine A supplementation at a dose of 0.1 mg/kg body weight restored spatial memory significantly, as evident from decreased escape latency and path length to reach the hidden platform, and the increase in number of times of crossing the former platform location and time spent in the former platform quadrant. In addition, after exposure to hypobaric hypoxia, animals were sacrificed and biomarkers of oxidative damage, such as reactive oxygen species, lipid peroxidation, lactate dehydrogenase activity, reduced glutathione, oxidized glutathione and superoxide dismutase were studied in the hippocampus. Expression levels of pro-apoptotic proteins (Bax, caspase-3) and anti-apoptotic protein (Bcl-2) of hippocampal tissues were evaluated by Western blotting. There was a significant increase in oxidative stress along with increased expression of apoptotic proteins in hypoxia exposed rats, which was significantly improved by oral Huperzine A at 0.1 mg/kg body weight. These results suggest that supplementation with Huperzine A improves cognitive deficits, reduces oxidative stress and inhibits the apoptotic cascade induced by acute hypobaric hypoxia.

Survivin Regulates Bad Gene Expression by Binding to Its Promoter and Modulates Cell Cycle and Apoptosis in Esophageal Carcinoma Cell.

Esophageal cancer (EC) is the eighth most prevalent cancer and the sixth leading cause of cancer-related mortality worldwide. As an antiapoptotic and a proapoptotic protein, respectively, survivin and Bad play an important role in carcinogenesis of the most human cancers including EC. However, the regulatory relationships between them remain unclear. We sought to investigate the effects of survivin knockdown and overexpression on the expression of Bad gene, cell cycle progression, and apoptosis of esophageal carcinoma cell. The mRNA expression levels of survivin and Bad were determined in EC tissue samples. The knockdown and overexpression experiments were performed in ECA109 and KYSE450 cells via transfection with survivin overexpression and shRNA plasmids. A Bad overexpression experiment was conducted to confirm the biological effect on knockdown of survivin via modulating Bad expression. RT-qPCR and Western blot analysis were used to detect mRNA and protein expression, respectively. Cell cycle and apoptosis were analyzed by flow cytometry. The chromatin immunoprecipitation (ChIP) was conducted to determine the binding sites of survivin on the promoter of Bad gene. By analyzing the mRNA expression of survivin and Bad in 40 ESCC patient specimens, we found that the positive expression rate of survivin in tumor tissues (88%, 35/40) was remarkably high, compared with the distal nontumor tissues (48%, 19/40, p < 0.01). On the other hand, the positive expression rate of Bad in tumor tissues (70%, 28/40) was remarkably low, compared with the distal nontumor tissues (95%, 38/40, p < 0.01). Overexpression of survivin decreases Bad mRNA and protein expression and promotes transformation of cell cycle to S phase. Conversely, knockdown of survivin increases Bad mRNA and protein expression and induces cell cycle arrest and apoptosis. Bad overexpression inducing apoptosis of esophageal carcinoma cell shows the similar apoptotic effect with survivin knockdown. ChIP assays indicate that survivin directly binds to the Bad promoter region, diminishing the transcriptional activity of Bad. In conclusion, the result suggested that survivin regulates Bad gene expression by binding to its promoter and modulates cell cycle and apoptosis in esophageal carcinoma cell.

Ginkgolide B Maintains Calcium Homeostasis in Hypoxic Hippocampal Neurons by Inhibiting Calcium Influx and Intracellular Calcium Release.

Ginkgolide B (GB), a terpene lactone and active ingredient of Ginkgo biloba, shows protective effects in neuronal cells subjected to hypoxia. We investigated whether GB might protect neurons from hypoxic injury through regulation of neuronal Ca2+ homeostasis. Primary hippocampal neurons subjected to chemical hypoxia (0.7 mM CoCl2) in vitro exhibited an increase in cytoplasmic Ca2+ (measured from the fluorescence of fluo-4), but this effect was significantly diminished by pre-treatment with 0.4 mM GB. Electrophysiological recordings from the brain slices of rats exposed to hypoxia in vivo revealed increases in spontaneous discharge frequency, action potential frequency and calcium current magnitude, and all these effects of hypoxia were suppressed by pre-treatment with 12 mg/kg GB. Western blot analysis demonstrated that hypoxia was associated with enhanced mRNA and protein expressions of Cav1.2 (a voltage-gated Ca2+ channel), STIM1 (a regulator of store-operated Ca2+ entry) and RyR2 (isoforms of Ryanodine Receptor which mediates sarcoplasmic reticulum Ca2+ release), and these actions of hypoxia were suppressed by GB. Taken together, our in vitro and in vivo data suggest that GB might protect neurons from hypoxia, in part, by regulating Ca2+ influx and intracellular Ca2+ release to maintain Ca2+ homeostasis.

1400W ameliorates acute hypobaric hypoxia/reoxygenation-induced cognitive deficits by suppressing the induction of inducible nitric oxide synthase in rat cerebral cortex microglia.

Nitric oxide (NO) is involved in neuronal modifications, and overproduction of NO contributes to memory deficits after acute hypobaric hypoxia-reoxygenation. This study investigated the ability of the iNOS inhibitor 1400W to counteract spatial memory deficits following acute hypobaric hypoxia-reoxygenation, and to affect expression of NOS, NO, 3-NT and MDA production, and apoptosis in rat cerebral cortex. We also used primary rat microglia to investigate the effect of 1400W on expression of NOS, NO, 3-NT and MDA production, and apoptosis. Acute hypobaric hypoxia-reoxygenation impaired spatial memory, and was accompanied by activated microglia, increased iNOS expression, NO, 3-NT and MDA production, and neuronal cell apoptosis in rat cerebral cortex one day post-reoxygenation. 1400W treatment inhibited iNOS expression without affecting nNOS or eNOS. 1400W also reduced NO, 3-NT and MDA production, and prevented neuronal cell apoptosis in cerebral cortex, in addition to reversing spatial memory impairment after acute hypobaric hypoxia-reoxygenation. Hypoxia-reoxygenation activated primary microglia, and increased iNOS and nNOS expression, NO, 3-NT, and MDA production, and apoptosis. Treatment with 1400W inhibited iNOS expression without affecting nNOS, reduced NO, 3-NT and MDA production, and prevented apoptosis in primary microglia. Based on the above findings, we concluded that the highly selective iNOS inhibitor 1400W inhibited iNOS induction in microglial cells, and reduced generation of NO, thereby mitigating oxidative stress and neuronal cell apoptosis in the rat cerebral cortex, and improving the spatial memory dysfunction caused by acute hypobaric hypoxia-reoxygenation.

Survivin activates NF­κB p65 via the IKKß promoter in esophageal squamous cell carcinoma.

Survivin and transcription factor p65 (NF­κB p65) participate in the progression of esophageal squamous cell carcinoma (ESCC). However, the mechanism of NF­κB p65 activation in ESCC remains to be elucidated. The aim of the present study was to investigate the role of survivin in the activation of NF­κB p65 in ESCC. The expression levels of survivin, NF­κB p65, inhibitor of nuclear factor κB kinase subunit α (IKKα) and inhibitor of nuclear factor κB kinase subunit ß (IKKß) were detected in ESCC tissue samples. Eca109 and KYSE150 cells were cultured and survivin activity was modulated via transfection with an overexpression plasmid, a small hairpin RNA plasmid and a specific inhibitor. Quantitative reverse transcription-polymerase chain reaction and western blotting assays were conducted to assess the effects of survivin on the expression levels of IKKα, IKKß and NF­κB p65. Cell cycle and apoptosis assays were conducted to detect surviving-dependent cellular behavior changes. In addition, the luciferase reporter gene assay and chromatin immunoprecipitation assay were conducted to determine the genomic sites responsible for surviving-induced activation of NF­κB p65. The present study demonstrated that the expression of survivin is positively correlated with IKKα and IKKß in ESCC tissues. Survivin affected the mRNA and protein expression levels of IKKα, IKKß, and NF­κB p65 in Eca109 and KYSE150 cells. Furthermore, survivin increased the transcriptional activity of the IKKß promoter and bound to the IKKß promoter region in the Eca109 cells. Downregulation of survivin arrested the cell cycle at the G2/M phase and induced apoptosis. Results of the present study suggest that survivin activates NF­κB p65 in Eca109 cells via binding to the IKKß promoter region and upregulating IKKß promoter transcriptional activity. Survivin overexpression activates NF­κB p65, which is important in the acquisition and maintenance of the oncogenic characteristics of ESCC.

Geniposide suppresses LPS-induced nitric oxide, PGE2 and inflammatory cytokine by downregulating NF-κB, MAPK and AP-1 signaling pathways in macrophages.

Inflammatory responses are important to host immune reactions, but uncontrolled inflammatory mediators may aid in the pathogenesis of other inflammatory diseases. Geniposide, an iridoid glycoside found in the herb gardenia, is believed to have broad-spectrum anti-inflammatory effects in murine models but its mechanism of action is unclear. We investigated the action of this compound in murine macrophages stimulated by lipopolysaccharide (LPS), as the stimulation of macrophages by LPS is known to induce inflammatory reactions. We determined the effect of geniposide on LPS-induced production of the inflammatory mediators, nitric oxide (NO) and prostaglandin E2 (PGE2), the mRNA and protein expression of the NO and PGE2 synthases, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), respectively, and the mRNA and protein expression of the inflammatory cytokine, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). Furthermore, nuclear factor (NF)-κB, mitogen-activated protein kinase (MAPK) and activator protein (AP)-1 activity were assayed. To understand the action of geniposide on the NF-κB and MAPK pathways, we studied the effect of NF-κB and MAPK inhibitors on the LPS-induced production of NO, PGE2 and TNF-α. Our findings clearly showed that geniposide mainly exerts its anti-inflammatory effects by inhibiting the LPS-induced NF-κB, MAPK and AP-1 signaling pathways in macrophages, which subsequently reduces overexpression of the inducible enzymes iNOS and COX-2 and suppresses the expression and release of the inflammatory factors, TNF-α, IL-6, NO and PGE2. Thus, geniposide shows promise as a therapeutic agent in inflammatory diseases.

Construction and analysis of the protein-protein interaction network related to essential hypertension.

BACKGROUND: Essential hypertension (EH) is a complex disease as a consequence of interaction between environmental factors and genetic background, but the pathogenesis of EH remains elusive. The emerging tools of network medicine offer a platform to explore a complex disease at system level. In this study, we aimed to identify the key proteins and the biological regulatory pathways involving in EH and further to explore the molecular connectivities between these pathways by the topological analysis of the Protein-protein interaction (PPI) network. RESULT: The extended network including one giant network consisted of 535 nodes connected via 2572 edges and two separated small networks. 27 proteins with high BC and 28 proteins with large degree have been identified. NOS3 with highest BC and Closeness centrality located in the centre of the network. The backbone network derived from high BC proteins presents a clear and visual overview which shows all important regulatory pathways for blood pressure (BP) and the crosstalk between them. Finally, the robustness of NOS3 as central protein and accuracy of backbone were validated by 287 test networks. CONCLUSION: Our finding suggests that blood pressure variation is orchestrated by an integrated PPI network centered on NOS3.