TMEM88 Modulates Lipid Synthesis and Metabolism Cytokine by Regulating Wnt/β-Catenin Signaling Pathway in Non-Alcoholic Fatty Liver Disease

Objective: To clarify the molecular mechanism of TMEM88 regulating lipid synthesis and metabolism cytokine in NAFLD.Methods:In vivo, NAFLD model mice were fed by a Methionine and Choline-Deficient (MCD) diet. H&E staining and immunohistochemistry experiments were used to analyze the mice liver tissue. RT-qPCR and Western blotting were used to detect the lipid synthesis and metabolism cytokine. In vitro, pEGFP-C1-TMEM88 and TMEM88 siRNA were transfected respectively in free fat acid (FFA) induced AML-12 cells, and the expression level of SREBP-1c, PPAR-α, FASN, and ACOX-1 were evaluated by RT-qPCR and Western blotting.Results: The study found that the secretion of PPAR-α and its downstream target ACOX-1 were upregulated, and the secretion of SREBP-1c and its downstream target FASN were downregulated after transfecting with pEGFP-C1-TMEM88. But when TMEM88 was inhibited, the experimental results were opposite to the aforementioned conclusions. The data suggested that it may be related to the occurrence, development, and end of NAFLD. Additionally, the study proved that TMEM88 can inhibit Wnt/β-catenin signaling pathway. Meanwhile, TMEM88 can accelerate the apoptotic rate of FFA-induced AML-12 cells.Conclusion: Overall, the study proved that TMEM88 takes part in regulating the secretion of lipid synthesis and metabolism cytokine through the Wnt/β-catenin signaling pathway in AML-12 cells. Therefore, TMEM88 may be involved in the progress of NAFLD. Further research will bring new ideas for the study of NAFLD.

Exploration of the Effect on Genome-Wide DNA Methylation by miR-143 Knock-Out in Mice Liver

MiR-143 play an important role in hepatocellular carcinoma and liver fibrosis via inhibiting hepatoma cell proliferation. DNA methyltransferase 3 alpha (DNMT3a), as a target of miR-143, regulates the development of primary organic solid tumors through DNA methylation mechanisms. However, the effect of miR-143 on DNA methylation profiles in liver is unclear. In this study, we used Whole-Genome Bisulfite Sequencing (WGBS) to detect the differentially methylated regions (DMRs), and investigated DMR-related genes and their enriched pathways by miR-143. We found that methylated cytosines increased 0.19% in the miR-143 knock-out (KO) liver fed with high-fat diet (HFD), compared with the wild type (WT). Furthermore, compared with the WT group, the CG methylation patterns of the KO group showed lower CG methylation levels in CG islands (CGIs), promoters and hypermethylation in CGI shores, 5′UTRs, exons, introns, 3′UTRs, and repeat regions. A total of 984 DMRs were identified between the WT and KO groups consisting of 559 hypermethylation and 425 hypomethylation DMRs. Furthermore, DMR-related genes were enriched in metabolism pathways such as carbon metabolism (serine hydroxymethyltransferase 2 (Shmt2), acyl-Coenzyme A dehydrogenase medium chain (Acadm)), arginine and proline metabolism (spermine synthase (Sms), proline dehydrogenase (Prodh2)) and purine metabolism (phosphoribosyl pyrophosphate synthetase 2 (Prps2)). In summary, we are the first to report the change in whole-genome methylation levels by miR-143-null through WGBS in mice liver, and provide an experimental basis for clinical diagnosis and treatment in liver diseases, indicating that miR-143 may be a potential therapeutic target and biomarker for liver damage-associated diseases and hepatocellular carcinoma.

Microscopic Profile of Mice Liver Tissue (Mus musculus) Fixed with Neutral Buffered Formalin (NBF 10%) and Helly Solution

The fixation solution that is widely used in anatomical pathology laboratories is NBF 10%, the excess of NBF 10% because the pH is close to neutral, can be stored in large quantities and a long time. Helly's fixation solution is a good fixation solution for the cytoplasm, and only requires 2-3 hours of fixation. Knowing the microscopic picture of the preparation of hepar mencit tissue (Mus musculus) fixated with NBF 10% and Helly solution. This research is an experimental study with a descriptive analysis approach. picture of hepar mencit tissue preparation (Mus musculus) fixation with NBF 10% obtained as much as 100% good preparation. While the fixated with Helly solution obtained as much as 66% good preparation. Conclusion: Microscopic picture of hepar mencit tissue preparation (Mus musculus) fixated with NBF is 10% better than Helly solution.

Necrosis Description of Mice Liver Induced with Monosodium Glutamate and Methanol Robusta Coffee Bean Extract (Coffea canephora)

This study aimed to analyze the effect of methanol robusta coffee bean extract (Coffea canephora) on liver histopathological description in mice induced with monosodium glutamate (MSG). This study was used 20 male mice for 5 treatment groups: K + (MSG 0.12 mg and Vitamin C 6 mg), K- (MSG 0.12 mg and CMC Na 0.1 ml), P1 (MSG 0.12 mg and RCE 0.1 mg), P2 (MSG 0.12 mg and RCE 0.2 mg), and P3 (MSG 0.12 mg and RCE 0.4 mg) per orally for 42 days. Liver histological scores were analyzed using Kruskal Wallis test. The results of Kruskal Wallis test scored p=0,391 which indicates that the data is not significant (p>0.05). In the control and treatment groups there were no significant. In conclusion, the giving methanol robusta coffee bean extract did not show a decrease in hepatosite damage that was different from the negative control treatment group.

UJI TOKSISITAS AKUT EKSTRAK ETANOL DAUN KAYU MANIS (Cinnamomum burmanii) PADA FUNGSI HATI TIKUS PUTIH (Mus musculus L.) BETINA

Cinnamon leaves (Cinnamomum burmanii) are plants that have many pharmacological effects including lowering blood sugar levels, inhibition of bacterial growth, antioxidants and mymetic insulin activity. However, acute toxicity testing has not been carried out on cinnamon leaves before. This study aims to determine the level of toxicity and effect of extract administration on SGOT, SGPT as well as changes in the histopathological form of the liver of test animals. This study used The Complete Randomized Design method with 5 treatment groups with na-CMC administration 0.5% as a negative control, P1 dose 250 mg/kgBB, P2 Dose 500 mg/kgBB, P3 dose 1000 mg/kgBB and P4 at a dose of 2000 mg/kgBB. Each treatment consists of 5 mice. The parameters observed in this study are SGPT, SGOT and histopathological examination of mice liver organs. The data was analyzed using the One Way ANOVA test with duncan's advanced test. The results showed that the administration of cinnamon leaf ethanol extract did not cause death in test animals so it was practically not toxic. SGPT and SGOT values in female white mice test animals showed statistically meaningful differences in SGPT and SGOT values (p<0.05) compared to negative controls. However, it is still in the normal range. The result of histopathological observations is that there is a change in hepatocytes of liver organs compared to negative control.

Trimethylamine n-Oxide (TMAO) Modulates the Expression of Cardiovascular Disease-Related microRNAs and Their Targets

Diet is a well-known risk factor of cardiovascular diseases (CVDs). Some microRNAs (miRNAs) have been described to regulate molecular pathways related to CVDs. Diet can modulate miRNAs and their target genes. Choline, betaine, and l-carnitine, nutrients found in animal products, are metabolized into trimethylamine n-oxide (TMAO), which has been associated with CVD risk. The aim of this study was to investigate TMAO regulation of CVD-related miRNAs and their target genes in cellular models of liver and macrophages. We treated HEPG-2, THP-1, mouse liver organoids, and primary human macrophages with 6 µM TMAO at different timepoints (4, 8, and 24 h for HEPG-2 and mouse liver organoids, 12 and 24 h for THP-1, and 12 h for primary human macrophages) and analyzed the expression of a selected panel of CVD-related miRNAs and their target genes and proteins by real-time PCR and Western blot, respectively. HEPG-2 cells were transfected with anti-miR-30c and syn-miR-30c. TMAO increased the expression of miR-21-5p and miR-30c-5p. PER2, a target gene of both, decreased its expression with TMAO in HEPG-2 and mice liver organoids but increased its mRNA expression with syn-miR-30c. We concluded that TMAO modulates the expression of miRNAs related to CVDs, and that such modulation affects their target genes.