fh1 small molecule  (AbMole Bioscience)

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: The potential target gene of FH1‐induced hepatocyte differentiation. (a) Venn diagram of the intersecting targets of FH1 and hepatocyte differentiation‐related genes. (b) PPI interaction network of FH1 targets and the intersecting targets of hepatocyte differentiation.

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques:

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: HGF/c‐Met signalling is essential for FH1‐induced hepatocyte differentiation. (a–d) GO enrichment analysis results demonstrating that the FH1‐targeted genes involved in hepatocyte differentiation are associated with biological process (BP), cellular component (CC) and molecular function (MF) categories. (e) KEGG pathway analysis results. (f) Venn diagram demonstrating the overlapping Met interactors between the BioGRID and HIPPIE databases and the HIPPIE score for overlapping proteins.

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques:

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: The binding of FH1 to CIN85 and c‐Met. (a) Vina score of FH1 and hub genes according to the CB. Dock 2 database. (b) Predicted binding sites of CIN85 and c‐Met for FH1. (c) Real‐time kinetic binding sensorgrams of different concentrations of FH1 (125–1000 μmol/L) are shown. The response (nm) indicates the optical thickness of the SSA biosensor layer.

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques: Binding Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: FH1 induces MSCs migration. (a) Volcano plot showing differentially expressed genes in FH1‐treated samples. (b) GSEA revealed that the differentially expressed genes were enriched mainly in the HGF/c‐Met signalling pathway. (c) MSCs seeded in six‐well plates and treated with FH1 for 24, 48 and 96 h. At the endpoint of treatment, the cells were stained with Actin‐Tracker Green‐488. The PF + FH1 group was pretreated with PF4217903 for 1 h and then cultured in medium containing 15 μM FH1 for 48 h ( n = 3). (d) FH1 had a chemoattractant effect in hUC‐MSCs. Cells were left untreated or treated with FH1 for 48 h or pretreated with PF4217903 for 1 h before FH1 was added to the Transwell directional migration assay. All the experiments are representative of three replicates ( n = 3).

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques: Migration, Staining, Cell Culture

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: Effect of FH1 on MSCs proliferation. (a, b) Cells were seeded at a density of 5000 cells/cm 2 in 6‐well microplates in EM and treated with or without 15 μM FH1 for the indicated times. The cells were pulsed with EdU for 2 h and then observed via fluorescence microscopy or harvested for flow cytometric analysis ( n = 3). (c) Cells were cultured in EM with or without 15 μM FH1 treatment for 24, 48, or 72 h and then stained with 1× iodide staining solution containing 0.05 mg/mL propidium iodide, 1 mg/mL RNase A and 0.3% Triton X‐100. The cells were incubated in the dark for 30 min. The number of cells in G0/G1 phase was analysed through flow cytometry ( n = 3). (d, e) Western blotting analysis of the protein expression levels of p21, p27 and bcl‐2 in MSCs cultured in EM medium either left untreated or treated with FH1 for the indicated times ( n = 3). Full‐length blots/gels are presented in Figure . Data information: The bars represent the means of three independent experiments ± SEM. Statistics: Two‐tailed unpaired Student's t ‐test. ***p < 0.001; ****p < 0.0001.

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques: Fluorescence, Microscopy, Cell Culture, Staining, Incubation, Flow Cytometry, Western Blot, Expressing, Two Tailed Test

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: Effect of FH1 treatment on the expression of Met in MSCs. (a) Western immunoblotting analysis of c‐Met expression and FH1‐induced activation in MSCs ( n = 3). Full‐length blots/gels are presented in Figures and . (b) Quiescent MSCs were either left unstimulated (−) or stimulated with 15 μM FH1 for the indicated times, and total cell protein was examined through Western blotting and immunoprobed with antibodies against the active phosphorylated forms of ERK1/2, p38 and Akt (pERK1/2, pp38 and pAkt) or against total proteins (ERK1/2, p38 and Akt). Protein phosphorylation induced by 10 min of FH1 stimulation was inhibited by pretreatment for 1 h with the following specific inhibitors: PD98059 (30 M PD), SB230580 (30 M SB) and LY29402 (100 nM WM). All the experiments are representative of three replicates ( n = 3). Full‐length blots/gels of AKT and p‐AKT are presented in Figure ; Full‐length blots/gels of ERK and p‐ERK are presented in Figure ; Full‐length blots/gels of p38 and p‐p38 are presented in Figure . (c) Western blotting analysis of the expression of Met and β‐catenin in MSCs with FH1‐induced activation. Cells treated with FH1 tend to express high levels of the receptors of Met and β‐catenin. Pretreatment with the specific inhibitors PD98059 (30 M PD), SB230580 (30 M SB), or LY29402 (100 nM WM) had no effect on the expression of Met, β‐catenin or the proliferation marker PCNA in MSCs with or without with FH1. Full‐length blots/gels are presented in Figures and . All the experiments are representative of three replicates ( n = 3).

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques: Expressing, Western Blot, Activation Assay, Phospho-proteomics, Marker

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: The RNA profile of FH1‐induced hepatocytes. (a) Hierarchical clustering of PHHs, FH1‐iHeps, HSs and HBs according to differentially expressed genes. (b) Heatmap showing the expression of selected genes in PHHs versus FH1‐iHeps. (c) KEGG pathway analysis results. (d) KOG results showing the differential signalling pathway‐related functions.

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques: Expressing

Journal: Journal of Cellular and Molecular Medicine

Article Title: The Potential Hepatocyte Differentiation Targets and MSC Proliferation by FH1

doi: 10.1111/jcmm.70601

Figure Lengend Snippet: The therapeutic effects of FH1‐ iHeps on ALF. (a) Schematic diagram of cell transplantation into the livers of C57BL/6J mice. Mice were intraperitoneally injected with CCl 4 to induce fulminant hepatic injury. Eight hours after CCl 4 treatment, hUC‐MSCs and iHeps (1 × 10 6 cells/animal, 300 μL) were intravenously injected into the mice, and the control animals received an equal volume of PBS. (b) Kaplan–Meier survival curve of mice with ALF ( n = 6). (c) Serum levels of ALT, AST and human albumin (ALB) in CCl 4 ‐treated mice before (day 0 [d0]) and after (d7) transplantation of Huc‐MSCs or iHeps ( n = 3). (d) Macroscopic images of freshly isolated livers from mice treated with CCl 4 for 8 h; PBS, hADSCs and iHeps were intravenously injected into the mice ( n = 3). (e) H&E staining of liver sections ( n = 3). (f) PAS staining ( n = 3). (g) The integration of iHeps in mouse livers was determined by immunostaining for human ALB in serial sections ( n = 3). Scale bars = 50 μm (e, f) and 100 μm (g). Data information: The bars represent the means of three independent experiments ± SEM. Statistics: Two‐tailed unpaired Student's t ‐test.

Article Snippet: In the progenitor hepatic formation stage, the cells were incubated in serum‐free IMDM supplemented with 100 nM IDE1 (MCE), 10 μM LY294002, 250 nM LDN‐193189 (Abmole Bioscience) and 20 ng/mL basic fibroblast growth factor (bFGF; PeproTech) for 48 h, after which the medium was changed to serum‐free IMDM supplemented with 100 nM IDE1, 10 μM LY294002 and 20 ng/mL bFGF for another 24 h. For the hepatocyte maturation stage, the maturation medium was composed of William's E medium (Gibco) supplemented with 15 μM FH1 (Abmole Bioscience), 20 ng/mL bFGF, 30 ng/mL oncostatin M (OSM; PeproTech), 2 × 10 −5 mol/L dexamethasone (Dex; Abmole Bioscience) and 1% insulin‐transferrin‐selenium supplement‐X (ITS‐X; Sigma Aldrich).

Techniques: Transplantation Assay, Injection, Control, Isolation, Staining, Immunostaining, Two Tailed Test

Journal: Frontiers in Microbiology

Article Title: A Chimeric Humanized Mouse Model by Engrafting the Human Induced Pluripotent Stem Cell-Derived Hepatocyte-Like Cell for the Chronic Hepatitis B Virus Infection

doi: 10.3389/fmicb.2018.00908

Figure Lengend Snippet: Differentiated hepatic cells were long maintained to support a long-term genotype C HBV infection. (A) Fold changes of the mRNA levels of the human hepatic specific genes in hiPSC-HLCs treated with FH1 as compared to that untreated after being normalized to GAPDH ( n = 6). The primers were as shown in Supplementary Table . (B) FACS analysis for the positive ratio of the hepatic specific genes in hiPSC-HLCs treated with FH1 as compared to that untreated ( n = 6). (C) Protein levels of hALB, HBV DNA, HBsAg, and HBeAg in the supernatant, and (D) intracellular HBV RNA and cccDNA of HBV-infected hiPSC-HLCs at different time points (MOI = 200, n = 6). (E) Schematic design of the in vitro ETV, RNAi, PA452, genistein, and Myrcludex B treatments from days 10 to 20 after HBV infection. (F) Detection of relative HBV infection marker suppression of the indicated antivirals at 10 days after treatment ( n = 6). S/CO, signal-to-cutoff. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001.

Article Snippet: From day 4 to 10, the functional components [1% dimethyl sulfoxide (DMSO), 10 -7 M dexamethasone (DEX, LONZA, cat # CC4182-1), 100 ng/mL hepatocyte growth factor (HGF, Sigma-Aldrich, cat # 1404), 5 μg/mL of small molecule FH1 from APExBIO (cat # BRD-K4477), 5 μg/mL insulin (LONZA, cat # CC-4321BB), 500 μM basic fibroblast growth factor (bFGF, LONZA, cat # CC4182-3), epidermal growth factor (EGF) from HCM BulletKit of LONZA (cat # CC4317BB)] were added to the medium.

Techniques: Infection, In Vitro, Marker