methionine choline chloride deficient mcd diet  (Valiant Co Ltd)

Journal: Journal of Clinical and Translational Hepatology

Article Title: Investigation of HO-1 Regulation of Liver Fibrosis Related to Nonalcoholic Fatty Liver Disease Through the SIRT1/TGF-ß/Smad3 Pathway

doi: 10.14218/JCTH.2024.00481

Figure Lengend Snippet: (A) liver /body weight. (B) Body weight. Serum (C) ALT and (D) AST in each group. (E) Images of H&E (×200 magnification) staining of representative liver sections and (F) the NAFLD activity score. (G) Images of Masson (×200 magnification) staining of representative liver sections and (H) the Metavir score. (I) Images of Sirius (×200 magnification) red staining of representative liver sections and (J) Relative collagen content (%). Values are the mean ± SD (n = 6 per group). * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. MCD, methionine-and-choline-deficient diet; NAFLD, nonalcoholic fatty liver disease; H&E, hematoxylin-eosin. ALT, alanine aminotransferase; AST, aspartate transaminase; Znpp, zincprotoporphyria.

Article Snippet: The mice were randomly divided into four groups (n = 6) and received group-specific diets for eight weeks: (1) Control group: Mice were fed a normal diet; (2) Methionine- and choline-deficient (MCD) group: Mice were fed an MCD diet (Research Diets, Inc., NJ, New Brunswick, USA); (3) Hemin group: Mice were fed an MCD diet and treated with the HO-1 chemical inducer Hemin three times per week; (4) Zinc protoporphyrin (Znpp) group: Mice were fed an MCD diet and treated with the HO-1 inhibitor Znpp three times per week.

Techniques: Staining, Activity Assay

Journal: Journal of Clinical and Translational Hepatology

Article Title: Investigation of HO-1 Regulation of Liver Fibrosis Related to Nonalcoholic Fatty Liver Disease Through the SIRT1/TGF-ß/Smad3 Pathway

doi: 10.14218/JCTH.2024.00481

Figure Lengend Snippet: (A–F) The expression levels of HO-1, SIRT1, TGF-β, Smad3, Collagen1 and α-SMA were measured by RT-qPCR. (G) Immunofluorescence (×200 magnification) double staining between of HO-1 and SIRT1 in liver tissue. (H–K) Immunofluorescence double staining semi-quantitative analysis. GAPDH served as the loading control. Data are presented as representative results of three independent experiments. Values are the mean ± SD (n = 6 per group). *** p < 0.001, **** p < 0.0001. MCD, methionine-and-choline-deficient diet; HO-1, heme oxygenase 1; SIRT1, Sirtuin 1; TGF-β, transforming growth factor beta; α-SMA, alpha-smooth muscle actin; GAPDH, glyceraldehyde-3-phosphate dehydrogenase. Znpp, zincprotoporphyria.

Article Snippet: The mice were randomly divided into four groups (n = 6) and received group-specific diets for eight weeks: (1) Control group: Mice were fed a normal diet; (2) Methionine- and choline-deficient (MCD) group: Mice were fed an MCD diet (Research Diets, Inc., NJ, New Brunswick, USA); (3) Hemin group: Mice were fed an MCD diet and treated with the HO-1 chemical inducer Hemin three times per week; (4) Zinc protoporphyrin (Znpp) group: Mice were fed an MCD diet and treated with the HO-1 inhibitor Znpp three times per week.

Techniques: Expressing, Quantitative RT-PCR, Immunofluorescence, Double Staining, Control

Journal: Journal of Clinical and Translational Hepatology

Article Title: Investigation of HO-1 Regulation of Liver Fibrosis Related to Nonalcoholic Fatty Liver Disease Through the SIRT1/TGF-ß/Smad3 Pathway

doi: 10.14218/JCTH.2024.00481

Figure Lengend Snippet: (A, C–G) Western blot analysis of HO-1, SIRT1, TGF-β, P-Smad2/3, and Smad2/3 in liver tissue. (B, H, I) Western blot analysis of α-SMA and Collagen1 in liver tissue. Values are the mean ± SD (n = 6 per group). ** p < 0.01, *** p < 0.001, **** p < 0.0001. MCD, methionine-and-choline-deficient diet; HO-1, heme oxygenase 1; SIRT1, Sirtuin 1; TGF-β, transforming growth factor beta; α-SMA, alpha-smooth muscle actin; GAPDH, glyceraldehyde-3-phosphate dehydrogenase. Znpp, zincprotoporphyria.

Article Snippet: The mice were randomly divided into four groups (n = 6) and received group-specific diets for eight weeks: (1) Control group: Mice were fed a normal diet; (2) Methionine- and choline-deficient (MCD) group: Mice were fed an MCD diet (Research Diets, Inc., NJ, New Brunswick, USA); (3) Hemin group: Mice were fed an MCD diet and treated with the HO-1 chemical inducer Hemin three times per week; (4) Zinc protoporphyrin (Znpp) group: Mice were fed an MCD diet and treated with the HO-1 inhibitor Znpp three times per week.

Techniques: Western Blot

Journal: Journal of Clinical and Translational Hepatology

Article Title: Investigation of HO-1 Regulation of Liver Fibrosis Related to Nonalcoholic Fatty Liver Disease Through the SIRT1/TGF-ß/Smad3 Pathway

doi: 10.14218/JCTH.2024.00481

Figure Lengend Snippet: (A, B) Western blot analysis of HO-1 overexpression and silencing in LX2 cells. (C, D) Western blot analysis of LX2 cells treated with SIRT1 activators and inhibitors. (E–I) Western blot analysis of SIRT1, TGF-β, P-Smad2/3, Smad2/3 in LX2 cells. (J–L) Western blot analysis of α-SMA and Collagen I in LX2 cells. Values are the mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. MCD, methionine-and-choline-deficient diet; HO-1, heme oxygenase 1; SIRT1, Sirtuin 1; TGF-β, transforming growth factor beta; α-SMA, alpha-smooth muscle actin; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; OE HO-1, overexpressing HO-1; SiHO-1, small interfering RNA silencing of HO-1.

Article Snippet: The mice were randomly divided into four groups (n = 6) and received group-specific diets for eight weeks: (1) Control group: Mice were fed a normal diet; (2) Methionine- and choline-deficient (MCD) group: Mice were fed an MCD diet (Research Diets, Inc., NJ, New Brunswick, USA); (3) Hemin group: Mice were fed an MCD diet and treated with the HO-1 chemical inducer Hemin three times per week; (4) Zinc protoporphyrin (Znpp) group: Mice were fed an MCD diet and treated with the HO-1 inhibitor Znpp three times per week.

Techniques: Western Blot, Over Expression, Small Interfering RNA

Journal: JHEP Reports

Article Title: Hepatic immune environment differences among common mouse strains in models of MASH and liver cancer

doi: 10.1016/j.jhepr.2025.101380

Figure Lengend Snippet: Changes of liver immune subsets under MASH in BALB/c, C57BL/6, and FVB/N mice fed with MCD diet. Female BALB/c, C57BL/6, and FVB/N mice were fed with MCD diet ( vs. regular diet) to induce MASH. (A) The development of MASH was confirmed by H&E staining. Liver immune cells from MASH mice or control mice were prepared and immune subsets were measured by flow cytometry analysis. The comparison between MCD (green) and control (gray) was performed in each liver immune subsets of the three mouse strains (B–G). The overall changes of various liver immune subsets from three mouse strains are shown (H). The size of circle represents the log 10 transformed fold changes of each immune subset. The color gradient represents the relative frequences of each immune subset. The distribution of fold changes of each type of immune cell is also shown; n = 4 per group, two-way ANOVA with Bonferroni correction, ∗ p <0.05. MASH, metabolic dysfunction-associated steatohepatitis; MCD diet, methionine- and choline-deficient diet; Tregs, regulatory T cells.

Article Snippet: MASH was induced in mice by feeding a methionine- and choline-deficient (MCD) diet (Research diets inc., New Brunswick, NJ, USA Ref. A02082002BR) for a period of 3 weeks, or a Western diet (Envigo, Indianapolis, IN, USA, Ref. TD.120528) with high sugar solution (23.1 g/L d-fructose and 18.9 g/L d-glucose) with weekly intraperitoneal injections of carbon tetrachloride (CCl 4 ) (Sigma, St. Louis, MO, USA, Cat# 289116) at a dose of 0.32 mg/g of body weight for 12 weeks as previously reported.

Techniques: Staining, Control, Flow Cytometry, Comparison, Transformation Assay

Journal: JHEP Reports

Article Title: Hepatic immune environment differences among common mouse strains in models of MASH and liver cancer

doi: 10.1016/j.jhepr.2025.101380

Figure Lengend Snippet: Cross-species comparison of liver immune changes by MASH between mice and humans. The published human scRNA-seq dataset GSE159977 of CD45+ cells from either MASH or healthy livers were processed using Seurat (5.1.0). (A) shows the UMAP split based on healthy or MASH. (B) shows the dot plot of marker genes for each annotated cell clusters. (C) Liver CD45 + cell compositions were measured in naïve BALB/c, C57BL/6, and FVB/N strains by flow cytometry as described in Fig. 1 A. CD45 + cell composition of healthy human liver or human HCC adjacent liver tissues were calculated based on the scRNA-seq datasets of GSE159977 or phs003279.v1.p1, respectively. (D,E) CD45 + or CD4 + T cell compositions of MASH or healthy human livers were calculated from GSE159977 . (F) In total liver CD45 + cells, the frequencies of shared major liver immune subsets between mice and human, including CD8+ T cell, CD4+ T cell, B cells, γδT cells and NK cells, were calculated in MASH or healthy human livers ( GSE159977 ) and MASH or control mice of three strains under either MCD diet or Western + CCl 4 diet MASH model. CCl 4 , carbon tetrachloride; HCC, hepatocellular carcinoma; MASH, metabolic dysfunction-associated steatohepatitis; MAIT cells, mucosal-associated invariant T cells; MCD diet, methionine- and choline-deficient diet; NK, natural killer; UMAP, Uniform Manifold Approximation and Projection.

Article Snippet: MASH was induced in mice by feeding a methionine- and choline-deficient (MCD) diet (Research diets inc., New Brunswick, NJ, USA Ref. A02082002BR) for a period of 3 weeks, or a Western diet (Envigo, Indianapolis, IN, USA, Ref. TD.120528) with high sugar solution (23.1 g/L d-fructose and 18.9 g/L d-glucose) with weekly intraperitoneal injections of carbon tetrachloride (CCl 4 ) (Sigma, St. Louis, MO, USA, Cat# 289116) at a dose of 0.32 mg/g of body weight for 12 weeks as previously reported.

Techniques: Comparison, Marker, Flow Cytometry, Control, Western Blot

Journal: bioRxiv

Article Title: HuR-Driven Reversible Mitochondrial Shuttling Buffers Cytosolic miRNA Levels in Hepatic Cells to Control Apoptosis

doi: 10.1101/2025.04.08.647748

Figure Lengend Snippet: A. Fold change in cellular miRNA content after 16 hours of amino acid starvation of Huh7 cells was analyzed. Relative levels of various miRNAs were quantified using qRT–PCR, and the data were normalized to U6 snRNA (n> 3). B. miRNA-specific Ct values are plotted to illustrate the changes in their relative abundance in purified mitochondria following amino acid starvation of Huh7 cells. The analysis shows a significant decrease in Ct values for the miRNAs let-7b, let-7g, and miR-181a in the mitochondria of starved cells, indicating their enrichment in the mitochondrial fraction during starvation. A bar graph presents the fold enrichment of let-7b and let-7g mito-miRs in purified mitochondria compared to let-7a C. Western blot analysis performed on fed and starved Huh7 extracts showed increased levels of the stress marker protein phospho-eIF2α and cleaved PARP, along with decreased phosphorylation of eIF-4EBP1. β-actin served as the loading control for all samples. D. Cellular miRNA levels in the liver of mice vary under a normal diet (Fed) and after 12 hours of starvation, as estimated by qRT–PCR and normalized against U6 snRNA levels (n=3). E. The fold changes of let-7b and let-7g in the isolated pure mitochondrial fraction obtained from the livers of mice on a normal diet (Fed) or starved for 12 hours. Levels of each miRNA in the Fed condition are considered as units. F-G. The relative enrichment of mito-microRNAs in the outer and intermembrane space (F) as well as in the inner membrane and mitochondrial matrix (G) of mitochondria isolated from control (fed) and 16-hour amino acid-starved Huh7 cells. Relative Ct values are plotted. H. Folds change in mitochondrial miR-122 and let-7g levels in the liver mitochondria of animals exposed to a control and Methionine Choline Deficient (MCD) diet (n=4). I. Western blot analysis (left panel) and relative quantification (right panel) of different proteins in the mitochondrial fractions isolated from control and MCD diet-exposed animal livers J. The effect of Thapsigargin on the expression of stress-responsive proteins in Huh7 cells was determined by western blot analysis, using β-actin as a loading control. K-L. The effect of Thapsigargin treatment on cellular (K) and mitochondrial (L) miRNA content was analyzed. Relative levels of miRNAs were measured and plotted. U6 and let-7a were used for normalization in both cellular and mitochondrial samples. M . Cellular miR-122 levels increased, while mitochondrial levels remained unchanged when cells were treated with a pharmacological blocker of extracellular vesicle production GW4869. Normalization was performed using U6 snRNA and let-7a for cellular and mitochondrial miR-122, respectively. Statistically analyzed data is presented as mean ± SD from three experiments, with significance levels indicated as ns (nonsignificant), ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and p < 0.0001, calculated using the two-tailed Student’s t-test.

Article Snippet: They were randomly divided into two groups and fed either a standard chow diet or an MCD diet (MP Biomedicals; catalog no. 0296043910) for 4 weeks.

Techniques: Quantitative RT-PCR, Purification, Western Blot, Marker, Control, Isolation, Membrane, Expressing, Two Tailed Test