Effect of Rab18 on liver injury and lipid accumulation by regulating perilipin 2 and peroxisome proliferator-activated receptor gamma in non-alcoholic fatty liver disease.

BACKGROUND AND AIM: Nonalcoholic fatty liver disease (NAFLD) is currently one of the most common chronic liver diseases worldwide, characterized by the presence of lipid droplets. Rab18 is an important lipid droplet protein; however, its effects and mechanisms of action on NAFLD remain unclear. METHODS: Free fatty acid-stimulated AML-12 cells and high-fat diet (HFD)-fed mice were used as NAFLD models. Lentiviruses overexpressing Rab18 (Rab18-OE) or knockdown (Rab18-KD) were used to generate stable cell lines for genetic analysis. Blood serum levels of alanine aminotransferase, aspartate aminotransferase, total cholesterol, triglycerides, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, glucose, and leptin were measured using a biochemical autoanalyzer. Hematoxylin and eosin staining was performed to detect pathological damage to the liver. Lipid accumulation in the cells was assessed by Oil Red O staining. Target expression was measured using qPCR, western blotting, and immunocytochemistry. RESULTS: Rab18 mRNA and protein expression levels increased in free fatty acid-stimulated AML-12 cells and the livers of HFD-fed mice. Rab18-OE increased lipid accumulation in vitro, which was attenuated by Rab18-KD. In vivo, Rab18-OE augmented liver pathological damage, serum alanine aminotransferase/aspartate aminotransferase activity, and triglyceride, total cholesterol, and low-density lipoprotein levels, whereas Rab18-KD decreased these indicators. Rab18-KD also downregulated blood glucose levels in HFD-fed mice. Mechanistically, Rab18-OE and Rab18-KD regulated the mRNA and protein expression levels of perilipin 2 (PLIN2) and peroxisome proliferator-activated receptor gamma (PPARγ) in vitro and in vivo, respectively. Immunocytochemistry revealed that Rab18 colocalized with PLIN2 and PPARγ in AML-12 cells. CONCLUSION: Rab18 expression was elevated in vitro and in vivo in the NAFLD mouse model. Rab18 regulates PLIN2 and PPARγ expression to exaggerate liver injury and lipid accumulation in patients with NAFLD. Thus, Rab18 may be a crucial protein in this disease and a potential therapeutic target.

Systematic pan-cancer analysis identifies cuproptosis-related gene DLAT as an immunological and prognostic biomarker.

Lipoylated dihydrolipoamide S-acetyltransferase (DLAT), the component E2 of the multi-enzyme pyruvate dehydrogenase complex, is one of the key molecules of cuproptosis. However, the prognostic value and immunological role of DLAT in pan-cancer are still unclear. Using a series of bioinformatics approaches, we studied combined data from different databases, including the Cancer Genome Atlas, Genotype Tissue-Expression, the Cancer Cell Line Encyclopedia, Human Protein Atlas, and cBioPortal to investigate the role of DLAT expression in prognosis and tumor immunity response. We also reveal the potential correlations between DLAT expression and gene alterations, DNA methylation, copy number variation (CNV), tumor mutational burden (TMB), microsatellite instability (MSI), tumor microenvironment (TME), immune infiltration levels, and various immune-related genes across different cancers. The results show that DLAT displays abnormal expression within most malignant tumors. Through gene set enrichment analysis (GSEA), we found that DLAT was significantly associated with immune-related pathways. Further, the expression of DLAT was also confirmed to be correlated with the tumor microenvironment and diverse infiltration of immune cells, especially tumor-associated macrophages (TAMs). In addition, we found that DLAT is co-expressed with genes encoding major histocompatibility complex (MHC), immunostimulators, immune inhibitors, chemokines, and chemokine receptors. Meanwhile, we demonstrate that DLAT expression is correlated with TMB in 10 cancers and MSI in 11 cancers. Our study reveals that DLAT plays an essential role in tumorigenesis and cancer immunity, which may be used to function as a prognostic biomarker and potential target for cancer immunotherapy.

DNMT3a-Mediated Enterocyte Barrier Dysfunction Contributes to Ulcerative Colitis via Facilitating the Interaction of Enterocytes and B Cells.

Objective: Dysfunction of the enterocyte barrier is associated with the development of ulcerative colitis (UC). This study was aimed at exploring the effect of DNMT3a on enterocyte barrier function in the progression of UC and the underlying mechanism. Method: Mice were given 3.5% dextran sodium sulphate (DSS) in drinking water to induce colitis. The primary intestinal epithelial cells (IECs) were isolated and treated with lipopolysaccharide (LPS) to establish an in vitro inflammatory model. We detected mouse clinical symptoms, histopathological damage, enterocyte barrier function, B cell differentiation, DNA methylation level, and cytokine production. Subsequently, the effect of DNMT3a from IECs on B cell differentiation was explored by a cocultural experiment. Result: DSS treatment significantly reduced the body weight and colonic length, increased disease activity index (DAI), and aggravated histopathological damage. In addition, DSS treatment induced downregulation of tight junction (TJ) protein, anti-inflammatory cytokines (IL-10 and TGF-ß), and the number of anti-inflammatory B cells (CD1d+) in intestinal epithelial tissues, while upregulated proinflammatory cytokines (IL-6 and TNF-α), proinflammatory B cells (CD138+), and DNA methylation level. Further in vitro results revealed that DNMT3a silencing or TNFSF13 overexpression in IECs partly abolished the result of LPS-induced epithelial barrier dysfunction, as well as abrogated the effect of IEC-regulated B cell differentiation, while si-TACI transfection reversed these effects. Moreover, DNMT3a silencing decreased TNFSF13 methylation level and induced CD1d+ B cell differentiation, and the si-TNFSF13 transfection reversed the trend of B cell differentiation but did not affect TNFSF13 methylation level. Conclusion: Our study suggests that DNMT3a induces enterocyte barrier dysfunction to aggravate UC progression via TNFSF13-mediated interaction of enterocyte and B cells.

LncRNA LINC00662 promotes colon cancer tumor growth and metastasis by competitively binding with miR-340-5p to regulate CLDN8/IL22 co-expression and activating ERK signaling pathway.

BACKGROUND: LncRNA LINC00662 is closely related to the occurrence and development of cancer. This study aims to explore the effect of LINC00662 on colon cancer tumor growth and metastasis and its molecular mechanism. METHODS: CCK8, colony formation, transwell, scratch wound, TUNEL, flow cytometry, RT-PCR, western blotting and immunohistochemistry assays were used to detect the proliferation, apoptosis, invasion and migration of colon cancer cell and mRNA and protein expressions. Luciferase reporter and RNA pull down assays were used to detect the combination of LINC00662 and miR-340-5p or IL22 and the combination of miR-340-5p and CLDN8/IL22. Co-immunoprecipitation were used to detect the co-expression of CLDN8 and IL22 in colon cell lines. The targets of LINC00662 were predicated by Starbase v2.0. The target genes of miR-340-5p were predicated by miRDB and TargetScan. GO and KEGG enrichment analysis were performed by DAVID website. RESULTS: LINC00662 was up-regulation in colon cancer tissues and cell lines. Univariate Cox regression analysis showed that the LINC00662 expression level was related to the poor prognosis. LINC00662-WT and miR-340-5p mimics co-transfection depressed luciferase activity and IL22/CLDN8-WT and miR-340-5p inhibitors co-transfection memorably motivated luciferase activity. LINC00662 overexpression promoted cell proliferation, invasion and migration, and inhibited cell apoptosis in colon cancer. In vivo xenograft studies in nude mice manifested that LINC00662 overexpression prominently accelerate tumor growth. There was an opposite reaction in the biological functions of colon cells and tumor growth between LINC00662 overexpression and LINC00662 inhibition in vitro and in vivo. The functions of miR-340-5p mimics regulating the biological functions of colon cells and tumor growth were consistent with those of LINC00662 inhibition. CLDN8 and IL22, as target genes of miR-340-5p, reversed the functions of LINC00662 affecting the biological functions of colon cells and the protein levels of Bax, Bcl-2, XIAP, VEGF, MMP-2, E-cadherin and N-cadherin. Co-immunoprecipitation experiments indicated that CLDN8 directly interact with IL22 in colon cell lines. LINC00662 regulated CLDN8 and IL22 expressions and the activation of ERK signaling pathway via targeting miR-340-5p. CONCLUSION: LINC00662 overexpression promoted the occurrence and development of colon cancer by competitively binding with miR-340-5p to regulate CLDN8/IL22 co-expression and activating ERK signaling pathway.

CLDN8 promotes colorectal cancer cell proliferation, migration, and invasion by activating MAPK/ERK signaling.

BACKGROUND: Claudin 8 (CLDN8), an integral membrane protein that constitutes tight junctions in cell membranes, was recently implicated in tumor progression. However, its roles in colorectal cancer (CRC) progression and metastasis remain unknown. METHODS: In this study, we examined the effect of CLDN8 on the progression of CRC, including cell proliferation, migration, and invasion, and determines its underlying molecular mechanism using in vitro CRC cell lines and in vivo mouse xenograft models. RESULTS: We found that CLDN8 expression in human CRC tissues was significantly higher than that in adjacent normal tissues. The knockdown of CLDN8 markedly suppressed the proliferation, migration, and invasion of SW480 and HT-29 CRC cells, whereas the overexpression of CLDN8 notably promoted tumor progression in SW480 and HT-29 CRC cells. Mechanistic studies revealed that CLDN8 upregulated p-ERK (p-PKB/AKT) and MMP9 in CRC cells. Notably, the MAPK/ERK inhibitor PD98095 dramatically attenuated the effects of CLDN8 on p-ERK and MMP9. Moreover, PD98095 remarkably blocked the tumor-promoting activity of CLDN8. The knockdown of CLDN8 also inhibited the in vivo tumor growth in a nude mouse xenograft model. Collectively, CLDN8 promoted CRC cell proliferation, migration, and invasion, at least in part, by activating the MAPK/ERK signaling pathway. CONCLUSION: These findings suggest that CLDN8 exhibits an oncogenic effect in human CRC progression.

Lower gastrointestinal bleeding: role of 64-row computed tomographic angiography in diagnosis and therapeutic planning.

AIM: To determine the value of computed tomographic angiography (CTA) for diagnosis and therapeutic planning in lower gastrointestinal (GI) bleeding. METHODS: Sixty-three consecutive patients with acute lower GI bleeding underwent CTA before endovascular or surgical treatment. CTA was used to determine whether the lower GI bleeding was suitable for endovascular treatment, surgical resection, or conservative treatment in each patient. Treatment planning with CTA was compared with actual treatment decisions or endovascular or surgical treatment that had been carried out in each patient based on CTA findings. RESULTS: 64-row CTA detected active extravasation of contrast material in 57 patients and six patients had no demonstrable active bleeding, resulting in an accuracy of 90.5% in the detection of acute GI bleeding (57 of 63). In three of the six patients with no demonstrable active bleeding, active lower GI bleeding recurred within one week after CTA, and angiography revealed acute bleeding. The overall location-based accuracy, sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) for the detection of GI bleeding by 64-row CTA were 98.8% (249 of 252), 95.0% (57 of 60), 100% (192 of 192), 100% (57 of 57), and 98.5% (192 of 195), respectively. Treatment planning was correctly established on the basis of 64-row CTA with an accuracy, sensitivity, specificity, PPV and NPV of 98.4% (248 of 252), 93.3% (56 of 60), 100% (192 of 192), 100% (56 of 56), and 97.5% (192 of 196), respectively, in a location-based evaluation. CONCLUSION: 64-row CTA is safe and effective in making decisions regarding treatment, without performing digital subtraction angiography or surgery, in the majority of patients with lower GI bleeding.