Bifidobacterium bifidum BGN4 fractions ameliorate palmitic acid-induced hepatocyte ferroptosis by inhibiting SREBP1-CYP2E1 pathway.

Metabolic dysfunction-associated fatty liver disease (MAFLD) is strongly associated with disturbances in the intestinal microbiota. Herein, the biological effects and mechanism of Bifidobacterium bifidum BGN4 fractions in regulating hepatocyte ferroptosis during MAFLD progression were investigated. To establish an in vitro model of MAFLD, LO2 cells were subjected to palmitic acid (PA). The mRNA and protein expressions were assessed using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. LO2 cell proliferation was examined using 5-diphenyltetrazolium bromide (MTT) and ethynyl-2'-deoxyuridine (EdU) assays, whereas its apoptosis was evaluated by flow cytometry. Furthermore, level of reactive oxygen species (ROS) was measured using 2', 7,-Dichlorodihydrofluorescein diacetate (DCFH-DA) staining. Additionally, the levels of Fe2+, malondialdehyde (MDA), and glutathione (GSH), as well as the activities of superoxide dismutase (SOD) and glutathione peroxidase (GPX) were detected using corresponding kits. Chromatin immunoprecipitation and dual-luciferase reporter gene assays were performed to analyze the interaction between sterol-regulatory element binding protein 1 (SREBP1) and cytochrome P450-2E1 (CYP2E1) promoter. Our results revealed that Bifidobacterium bifidum BGN4 fractions effectively ameliorated PA-induced hepatocyte injury, oxidative stress, and ferroptosis. However, these beneficial effects of BGN4 fractions on PA-induced hepatocyte were dramatically reversed by SREBP1 overexpression, suggesting that BGN4 attenuated MAFLD by acting on SREBP1. Moreover, we observed that BGN4 fractions inhibited CYP2E1 transcription by suppressing SREBP1 nuclear translocation. In addition, CYP2E1 overexpression eliminated the inhibitory effect of BGN4 fractions on PA-induced hepatocyte oxidative stress and ferroptosis. These findings collectively indicated that BGN4 fractions reduced CYP2E1 expression by inhibiting SREBP1 nuclear translocation, thereby suppressing hepatocyte oxidative stress and ferroptosis during the development of MAFLD.

LINC01232 Promotes Metastasis and EMT by Regulating miR-506-5p/PAK1 Axis in Gastric Cancer.

Background: Long non-coding RNA LINC01232 plays an important role in the progression of metastasis in several cancers. However, the function of LINC01232 in gastric cancer is limited. Authors aimed to investigate the role and mechanism of LINC01232 in the metastasis of gastric cancer. Methods: The expression levels and correlation of LINC01232, miR-506-5p, and PAK1 were analyzed by GEPIA or ENCORI, and the abundance of LINC01232 and miR-506-5p was measured in tissues and cells via qRT-PCR, the location of LINC01232 in gastric cells was analyzed by nuclear and cytoplasmic fractionation, while the protein levels of PAK1, E-cadherin and vimentin were additionally quantified by Western blotting. Interactions between LINC01232, miR-506-5p, and PAK1 were detected through luciferase reporter assays, qRT-PCR and Western blotting. Cellular viability was evaluated through CCK8 assays, migration ability was measured by transwell assays, invasion ability was tested by wound healing experiment. Results: LINC01232 was overexpressed in gastric cancer tissues and cells, and mainly located in nucleus. The inhibition of LINC01232 could suppress migration, invasion and EMT of gastric cancer cells. MiR-506-5p was downregulated in gastric cancer tissues and cells. LINC01232 sponged miR-506-5p to accelerate migration and EMT. PAK1 was certified to be a target of miR-506-5p, inhibition of PAK1 could interrupt LINC01232 overexpression-induced migration of gastric cancer cells. Conclusion: The LINC01232/miR-506-5p/PAK1 axis promotes metastasis of gastric cancer cells.

Association between Virulence Factors and TRAF1/4-1BB/Bcl-xL Expression in Gastric Mucosa Infected with Helicobacter pylori.

Objective. CagA+/vacAs1+/vacAm1+ Helicobacter pylori upregulates the expression of tumor necrosis factor receptor-associated factor 1 (TRAF1), tumor necrosis factor receptor superfamily member 9 (4-1BB), and B-cell lymphoma-extra large (Bcl-xL) in human gastric epithelial cells. We investigated the correlation between cagA/vacAs1/vacAm1 and TRAF1/4-1BB/Bcl-xL expression in gastric mucosal tissue of patients with gastric disorders. Methods. We collected gastric mucosa samples from 35 chronic, nonatrophic gastritis (CG) patients, 41 atrophic gastritis patients, 44 intestinal metaplasia with atypical hyperplasia (IM) patients, and 28 gastric carcinoma (Ca) patients. The expression of TRAF1, 4-1BB, and Bcl-xL was determined using western blotting. The expression of cagA, vacAs1, and vacAm1 in H. pylori was examined with polymerase chain reaction. Results. The expression of TRAF1, 4-1BB, and Bcl-xL was significantly upregulated in IM and Ca patients (P < 0.05 compared with CG). There were more cases of cagA+/vacAs1+/vacAm1+ H. pylori infection in samples with elevated TRAF1, 4-1BB, or Bcl-xL expression (P < 0.05). Additionally, there were a remarkably large number of samples with upregulated TRAF1/4-1BB/Bcl-xL expression in cases of cagA+/vacAs1+/vacAm1+ H. pylori infection (44 cases, 67.7%; P < 0.05). Conclusions. The pathogenesis of IM and Ca may be promoted by cagA+/vacAs1+/vacAm1+ H. pylori, possibly via upregulated TRAF1, 4-1BB, and Bcl-xL in gastric mucosal tissue.

The expression level of TRAF1 in human gastric mucosa is related to virulence genotypes of Helicobacter pylori.

OBJECTIVE: To investigate the expression level of tumor necrosis factor receptor-associated factor 1 (TRAF1) in gastric mucosa tissue in patients infected with Helicobacter pylori (H. pylori) and to analyze the relationship between TRAF1 expression and H. pylori virulence. METHODS: Gastric tissue samples were collected from patients with gastritis, atrophic gastritis, intestinal metaplasia with atypical hyperplasia, and gastric cancer. The expression level of TRAF1 in each group was analyzed by real-time polymerase chain reaction (PCR) and Western blot analysis. Virulence genotypes of H. pylori were determined by PCR. RESULTS: Significant differences in TRAF1 mRNA levels were observed between the gastritis and gastric cancer groups, and the atrophic gastritis and gastric cancer groups (p < 0.05). Moreover, significant differences in TRAF1 protein levels were observed between the gastritis and intestinal metaplasia with atypical hyperplasia groups, between the gastritis and gastric cancer groups, and between the atrophic gastritis and gastric cancer groups (all p < 0.05). The virulence genotypes of cytotoxin-associated gene A (cagA), vacAs1, and vacAm1 were more frequent in the TRAF1 high-level group than in the TRAF1 low-level group (p < 0.05). CONCLUSION: Higher TARF1 expression level is associated with infection by CagA(+)/vacAs1(+)/m1(+) virulent H. pylori strains and may promote the proliferation of gastric mucosal cells and induce gastric cancer.

[Influence of endoscopic variceal ligation on liver function and risk factors of rebleeding].

OBJECTIVE: To investigate the influence of endoscopic variceal ligation (EVL) on liver function and analyze the risk factors of rebleeding after EVL. METHODS: A total of 137 cirrhotic patients with esophageal varices who received EVL were retrospectively analyzed, and divided into group A, B, and C according to the Child-Pugh scores of liver function. We compared the liver function 1 week preoperatively and postoperatively. The patients were further divided into a rebleeding group and a non-rebleeding group after the EVL, and risk factors about rebleeding were analyzed. RESULTS: There was no significant difference on ALT, AST, T-Bil, and D-Bil either preoperatively or postoperatively in group A, B, and C (P>0.05). Thirteen patients (9.49%) rebled after the EVL. The course of disease, liver function, prothrombin time, and mass ascites were the risk factors of rebleeding. CONCLUSION: EVL has no obvious effect on liver function, and the course of disease, liver function, prothrombin time and mass ascites are risk factors of rebleeding after EVL.

[Construction of RNAi targeting TRAF1 gene and effect of TRAF1 on gastric cancer cells].

OBJECTIVE: To construct the RNAi targeting tumor necrosis factor receptor associated factor (TRAF1) gene, and to explore the effect of interference targeting TRAF1 on the biological behavior of gastric cancer cells. METHODS: We detected the expression of TRAF1 in BGC823, SGC7901, and MGC803 gastric cancer cell lines through the real-time PCR and Western blot; then we constructed three pLVXshRNA- TRAF1-shRNAs expression vector targeting TRAF1. When TRAF1 was interfered successfully, we selected the strongest interference efficiency ShRNA by real-time PCR and Western blot. Based on interference targeting TRAF1 on gastric cancer, we tested the cell proliferation activity and apoptosis through MTT assay and flow cytometry, and the cell migration by transwell migration assay. RESULTS: The expression of TRAF1 was increased in BGC823, SGC7901, and MGC803 gastric cancer cell lines compared with gastric epithelial cells (P<0.05), and the highest expression was in BGC823 gastric cell line. In the three TRAF1 shRNAs, the strongest interference efficiency shRNA was pLVX-shRNA-TRAF1-shRNA2. When the gene TRAF1 of BGC823 was interfered, the cell growing power was weakened and the apoptosis rate increased, and the cell migration had no difference. CONCLUSION: The expression of TRAF1 is up-regulated in gastric cancer cell lines BGC823, SGC7901, and MGC803, and the most obvious one is BGC823. The interference targeting TRAF1 can successfully inhibit the expression of TRAF1 in gastric cancer cell line BGC823. TRAF1 can inhibit the apoptosis of BGC823 cells.