Reduction of Intracellular Chloride Concentration Promotes Foam Cell Formation.

BACKGROUND: Previous work has demonstrated that the volume-regulated chloride channel is activated during foam cell formation, and inhibition of chloride movement prevents intracellular lipid accumulation. However, the mechanism explaining how chloride movement promotes foam cell formation is not clear. METHODS AND RESULTS: Foam cell formation was determined by Oil Red O staining. Western blotting and co-immunoprecipitation were used to examine protein expression and protein-protein interaction. [Cl(-)]iwas measured using 6-methoxy-N-ethylquinolinium iodide dye. The results showed that [Cl(-)]iwas decreased in monocytes/macrophages from patients with hypercholesterolemia and from apoE(-/-)mice fed with a high-fat diet. Lowering [Cl(-)]iupregulated scavenger receptor A (SR-A) expression, increased the binding and uptake of oxLDL, enhanced pro-inflammatory cytokine production and subsequently accelerated foam cell formation in macrophages from humans and mice. In addition, low Cl(-)solution stimulated the activation of JNK and p38 mitogen-activated protein kinases. Inhibition of JNK and p38 blocked Cl(-)reduced medium-induced SR-A expression and lipid accumulation. In contrast, reduction of [Cl(-)]ipromoted the interaction of SR-A with caveolin-1, thus facilitating caveolin-1-dependent SR-A endocytosis. Moreover, disruption of caveolae attenuated SR-A internalization, JNK and p38 activation, and ultimately prevented SR-A expression and foam cell formation stimulated by low Cl(-)medium. CONCLUSIONS: This data provide strong evidence that reduction of [Cl(-)]iis a critical contributor to intracellular lipid accumulation, suggesting that modulation of [Cl(-)]iis a novel avenue to prevent foam cell formation and atherosclerosis.

ClC-3 chloride channel/antiporter defect contributes to inflammatory bowel disease in humans and mice.

BACKGROUND: ClC-3 channel/antiporter plays a critical role in a variety of cellular activities. ClC-3 has been detected in the ileum and colon. OBJECTIVE: To determine the functions of ClC-3 in the gastrointestinal tract. DESIGN: After administration of dextran sulfate sodium (DSS) or 2,4,6-trinitrobenzenesulfonic acid (TNBS), intestines from ClC-3-/- and wild-type mice were examined by histological, cellular, molecular and biochemical approaches. ClC-3 expression was determined by western blot and immunostaining. RESULTS: ClC-3 expression was reduced in intestinal tissues from patients with UC or Crohn's disease and from mice treated with DSS. Genetic deletion of ClC-3 increased the susceptibility of mice to DSS- or TNBS-induced experimental colitis and prevented intestinal recovery. ClC-3 deficiency promoted DSS-induced apoptosis of intestinal epithelial cells through the mitochondria pathway. ClC-3 interacts with voltage-dependent anion channel 1, a key player in regulation of mitochondria cytochrome c release, but DSS treatment decreased this interaction. In addition, lack of ClC-3 reduced the numbers of Paneth cells and impaired the expression of antimicrobial peptides. These alterations led to dysfunction of the epithelial barrier and invasion of commensal bacteria into the mucosa. CONCLUSIONS: A defect in ClC-3 may contribute to the pathogenesis of IBD by promoting intestinal epithelial cell apoptosis and Paneth cell loss, suggesting that modulation of ClC-3 expression might be a new strategy for the treatment of IBD.

CCL18 signaling from tumor-associated macrophages activates fibroblasts to adopt a chemoresistance-inducing phenotype.

The heterogeneity of cancer-associated fibroblasts (CAFs) might be ascribed to differences in origin. CD10 and GPR77 have been reported to identify a chemoresistance-inducing CAF subset in breast cancer. However, the precise mechanism for the formation of the CD10+GPR77+ CAFs remains unknown. In this study, we found that CCL18 expression was positively correlated with the density of CD10+GPR77+ CAFs in breast cancer and associated with a poor response to chemotherapy. Moreover, CCL18 secreted by tumor-associated macrophages (TAMs) activated a CD10+GPR77+ CAF phenotype in normal breast-resident fibroblasts (NBFs), which could then enrich cancer stem cells (CSCs) and induce chemoresistance in breast cancer cells. Mechanistically, CCL18 activated NF-κB signaling via PITPNM3 and thus enhanced the production of IL-6 and IL-8. Furthermore, intratumoral CCL18 injection significantly induced the activation of NBFs and the chemoresistance of xenografts in vivo. In addition, targeting CCL18 by anti-CCL18 antibody could inhibit the formation of CD10+GPR77+ CAFs and recover the chemosensitivity in vivo, leading to effective tumor control. Collectively, these findings reveal that inflammatory signaling crosstalk between TAMs and fibroblasts is responsible for the formation of the CD10+GPR77+ CAFs, suggesting CCL18-PITPNM3 signaling is a potential therapeutic target to block the activation of this specific CAF subtype and tumor chemoresistance.

Mechanisms of Compound Sophora flavescens (Kushen) Decoction for the Treatment of Ulcerative Colitis based on Network Pharmacology and Molecular Docking Technology.

BACKGROUND: The compound Sophora flavescenes (Kushen) decoction was found to reduce the inflammatory symptom of Ulcerative Colitis (UC). However, there exists a very limited understanding of the molecular pharmacological mechanisms. OBJECTIVE: This study aimed to explore the mechanism of compound Sophora flavescens (Kushen) decoction in treating ulcerative colitis from the perspective of network pharmacology. METHODS: Active components and potential targets of compound Sophora flavescens (Kushen) decoction were obtained through the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP) database. GeneCards and other databases were used to predict and screen ulcerative colitis-related genes. Cytoscape software was applied to construct the "drugactive component-disease-target" network. GO function and KEGG pathway enrichment analyses revealed the potential pathway of the compound Sophora flavescenes (Kushen) decoction for UC. RESULTS: After the screening, a total of 124 active ingredients and 163 potential therapeutic targets for UC were obtained from the compound Sophora flavescens (Kushen) decoction. Protein interaction network analysis showed that 15 key targets could be identified for the possible treatment of UC. GO and KEGG analyses showed that the active ingredients in the compound Sophora flavescens (Kushen) decoction were mainly enriched in 2556 biological processes and 172 signaling pathways. CONCLUSION: The study showed that the compound Sophora flavescens (Kushen) decoction has therapeutic effects on UC through multi-component, multi-target, and multi-pathway.

Identification of candidate biomarkers and therapeutic drugs of colorectal cancer by integrated bioinformatics analysis.

Most colorectal cancer (CRC) patients are diagnosed with advanced stages and low prognosis. We aimed to identify potential diagnostic and prognostic biomarkers, as well as active small molecules of CRC. Microarray data (GSE9348, GSE35279, and GSE106582) were obtained from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) were identified by the GEO2R platform. Common DEGs were selected for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. Cytoscape software was used to construct protein-protein interaction networks and identify hub genes. Hub genes were evaluated by Kaplan-Meier survival analysis in the GEPIA database and validated in two independent microarray data (GSE74602 and GSE83889). Common DEGs were used to select active small molecules by the connectivity map database. A total of 166 DEGs were identified as common DEGs. GO analysis demonstrated that common DEGs were significantly enriched in the apoptotic process, cell proliferation, and cell adhesion. KEGG analysis indicated that the most enriched pathways were the PI3K-Akt signaling pathway and extracellular matrix-receptor interaction. COL1A2, THBS2, TIMP1, and CXCL8 significantly upregulated in colorectal tumor. High expressions of COL1A2, THBS2, and TIMP1 were associated with poor survival, while high expressions of CXCL8 were associated with better survival. We selected 11 small molecules for CRC therapy. In conclusion, we found key dysregulated genes associated with CRC and potential small molecules to reverse them. COL1A2, THBS2, TIMP1, and CXCL8 may act as diagnostic and prognostic biomarkers of CRC.

Decrease of intracellular chloride concentration promotes endothelial cell inflammation by activating nuclear factor-κB pathway.

Recent evidence suggested that ClC-3 channel/antiporter is involved in regulation of nuclear factor (NF)-κB activation. However, the mechanism explaining how ClC-3 modulates NF-κB signaling is not well understood. We hypothesized that ClC-3-dependent alteration of intracellular chloride concentration ([Cl(-)](i)) underlies the effect of ClC-3 on NF-κB activity in endothelial cells. Here, we found that reduction of [Cl(-)](i) increased tumor necrosis factor-α (TNFα)-induced expression of intercellular adhesion molecule 1 and vascular cell adhesion molecule 1 and adhesion of monocytes to endothelial cells (P<0.05; n=6). In Cl(-) reduced solutions, TNFα-evoked IκB kinase complex ß and inhibitors of κBα phosphorylation, inhibitors of κBα degradation, and NF-κB nuclear translocation were enhanced. In addition, TNFα and interleukin 1ß could activate an outward rectifying Cl(-) current in human umbilical vein endothelial cells and mouse aortic endothelial cells. Knockdown or genetic deletion of ClC-3 inhibited or abolished this Cl(-) conductance. Moreover, Cl(-) channel blockers, ClC-3 knockdown or knockout remarkably reduced TNFα-induced intercellular adhesion molecule 1 and vascular cell adhesion molecule 1expression, monocytes to endothelial cell adhesion, and NF-κB activation (P<0.01; n=6). Furthermore, TNFα-induced vascular inflammation and neutrophil infiltration into the lung and liver were obviously attenuated in ClC-3 knockout mice (P<0.01; n=7). Our results demonstrated that decrease of [Cl(-)](i) induced by ClC-3-dependent Cl(-) efflux promotes NF-κB activation and thus potentiates TNFα-induced vascular inflammation, suggesting that inhibition of ClC-3-dependent Cl(-) current or modification of intracellular Cl(-) content may be a novel therapeutic approach for inflammatory diseases.