Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: Construction, purification, and characterization of FGF21-164. ( A ) FGF21-164 sequence and FGF21 sequence. In the FGF21-164 sequence, red marked the mutated amino acid and blue represents the TR acid sequence from hCD164. In the FGF21 sequence, the red-labeled amino acid represents the amino acid sequence whose n-terminal is deleted in FGF21-164. ( B ) Subcloned cell culture supernatants expressing FGF21-164 and FGF21 were analyzed using Western blotting (Lane M: molecular weight marker. Lane 1~7: FGF21-164 clones 1, 2, 3, A4, B2, F3, and H4. Line 8: FGF21 clone2). ( C ) The purified FGF21-164 and FGF21 were assayed using SDS-PAGE and Western blotting (Lane M: molecular weight marker. Lane 1: purified FGF21-164. Line 2: purified FGF21). ( D ) Purity and hydrodynamic radius of FGF21-164 were analyzed using SEC-HPLC. ( E ) MALDI-TOF mass-spectrometry analysis of FGF21-164. ([M + H] + denotes the singly charged ionic forms, and [M + 2H] 2+ denotes doubly charged ionic forms). ( F ) Circular Dichroism (CD) spectra of FGF21-164 and FGF21.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques: Purification, Sequencing, Labeling, Cell Culture, Expressing, Western Blot, Molecular Weight, Marker, Clone Assay, SDS Page, Mass Spectrometry, Circular Dichroism

Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: Glycan forms analysis of the protein FGF21-164. ( A ) O-linked glycan forms of FGF21-164. ( B ) N-linked glycan forms of FGF21-164.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: Effect of FGF21-164 in adipocytes induced by 3T3-L1. ( A ) The process of inducing 3T3-L1’s differentiation into adipocytes. ( B ) Reduction in glucose in medium stimulated by FGF21-164 in adipocytes induced by 3T3-L1. Data are means ± SEM (n = 6). * p < 0.05, *** p < 0.001, and **** p < 0.0001 compared to 0 μg/mL FGF21-164. ( C ) Phospho-Erk1/2-specific bands (Thr202/Tyr204; 44–42 kDa) were detected in 3T3-L1-derived adipocytes after FGF21-164 stimulation. ( D ) Evaluation of lipid droplet accumulation in adipocytes induced by 3T3-L1 with or without FGF21-164. The scale bar represents 50 μm.Data are the means ± SEM (n = 6). * p < 0.05, **** p < 0.0001.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques: Derivative Assay

Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: Pharmacokinetics of FGF21-164. FGF21-164 plasma levels in C57BL/6 mice as determined via targeted LC-MS after i.v. (tail vein) bolus injection of FGF21-164 protein.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques: Liquid Chromatography with Mass Spectroscopy, Injection

Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: The results of an oral glucose tolerance test applied to ob / ob mice after a single FGF21-164 treatment. ( A ) Blood glucose levels were measured at 0, 2, 3, 4, 5, 6, 7, and 8 h after the administration of FGF21-164. Data are the means ± SEM (n = 4–5). * p < 0.05 and ** p < 0.01 compared with the model. ( B ) The decrease in blood glucose levels post-administration relative to the baseline measurement at 0 h. Data are the means ± SEM (n = 4–5). * p < 0.05 compared with the model.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: Effect of a single administration of FGF21-164 on ob / ob mice. ( A , D , G ) Fasting glucose levels. Blood glucose levels were measured at 7, 21, and 28 days after a single dose of FGF21-164 was administered. Data are the means ± SEM (n = 4–5). ns: no significance. * p < 0.05, ** p < 0.01, and **** p < 0.0001. ( B , E , H ) The OGTT at 7, 21, and 28 days. After oral glucose administration, blood glucose levels were measured. Data are the means ± SEM (n = 4–5). * p < 0.05, *** p < 0.001, and **** p < 0.0001 at 6 mg·kg −1 vs. model. # p < 0.05, ## p < 0.01 and #### p < 0.0001 at 12 mg·kg −1 vs. model. ( C , F , I ) The glucose AUC during the OGTT process. Data are the means ± SEM (n = 4–5). ns: no significance. * p < 0.05, *** p < 0.001. ( J ) Body weight measured at 0, 7, 21, and 28 days. Data are the means ± SEM (n = 4–5). ns: no significance.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: A Novel Recombinant Human FGF21 Analog with High Glycosylation Has a Prolonged Half-Life and Affects Glycemic and Body Weight Control

doi: 10.3390/ijms26062672

Figure Lengend Snippet: Effect of repeated administration of FGF21-164. ( A ) Body weight changes. The DIO mice were treated with PBS or FGF21-164 once every 2 days. Data are the means ± SEM (n = 5–7). **** p < 0.0001 vs. Ctrl. #### p < 0.0001 for FGF21-164 vs. the model. ( B ) The liver sections were stained with Oil-red-O. The scale bar represents 100 μm. DIO mice showed many red lipid droplets in hepatic tissue (indicated by the black arrow), while lipid droplet levels were lower after the FGF21-164 treatment.

Article Snippet: Here, we describe a novel human FGF21 analog, designated as FGF21-164, which was generated through fusion protein technology and co-encoded by a mutated FGF21 gene and the TR gene of hCD164.

Techniques: Staining

Journal: Redox Biology

Article Title: Exercise and dietary intervention ameliorate high-fat diet-induced NAFLD and liver aging by inducing lipophagy

doi: 10.1016/j.redox.2020.101635

Figure Lengend Snippet: Exercise increased the FGF21 levels in serum and muscle. (A) FGF21 level in serum by ELISA. (B) FGF21 protein levels in liver and muscle by Western Blot. (C) FGF21 content in liver and muscle by ELISA. (D) FGF21 mRNA levels in liver and muscle by qPCR. Data are shown as the means ± SEM, n = 8 per group. Significance was designated by asterisks with *p < 0.05.

Article Snippet: Rat FGF21 enzyme-linked immunosorbent assay (ELISA) kit was obtained from Cusabio Biotech (Cat. No. CSB-EL008627RA, Cusabio Biotech, Wuhan, China).

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Redox Biology

Article Title: Exercise and dietary intervention ameliorate high-fat diet-induced NAFLD and liver aging by inducing lipophagy

doi: 10.1016/j.redox.2020.101635

Figure Lengend Snippet: FGF21 attenuated LDs accumulation in FFA-treated HepG2 cells through an autophagy dependent way. HepG2 cells were treated with FFA (400 μM). (A) Representative confocal images of LDs by BODIPY 493/503 staining. Scale: 50 μm. (B) TG level in HepG2 cells. (C) Representative immunoblotting of PLIN2, LC3II/LC3I and p62 in HepG2 cells. HepG2 cells were pre-transfected with Atg5 shRNA or empty vector by lenti-virus, then treated with FFA (400 μM) with or without FGF21 (1 μg/ml). (D) Representative confocal images of LDs by BODIPY 493/503 staining. Scale: 50 μm. (E) TG level in HepG2 cells. Experiments were repeated three times. Data are shown as the means ± SEM. Significance was designated by asterisks with *p < 0.05.

Article Snippet: Rat FGF21 enzyme-linked immunosorbent assay (ELISA) kit was obtained from Cusabio Biotech (Cat. No. CSB-EL008627RA, Cusabio Biotech, Wuhan, China).

Techniques: Staining, Western Blot, Transfection, shRNA, Plasmid Preparation, Virus

Journal: Redox Biology

Article Title: Exercise and dietary intervention ameliorate high-fat diet-induced NAFLD and liver aging by inducing lipophagy

doi: 10.1016/j.redox.2020.101635

Figure Lengend Snippet: FGF21 increased lipophagy through AMPK-dependent pathway in FFA-treated HepG2 cells. HepG2 cells were treated with FFA (400 μM), FGF21 (1 μg/ml) as well as AMPK inhibitor-Dorsomorphin (1 μmol/L). (A) Representative of p -AMPK (Thr172)/AMPK total, p -ULK1 (Ser555)/ULK1 total and LC3-II/LC3-I immunoblotting in HepG2 cells treated with FFA, FGF21 and Dorsomorphin, and the quantified data. (B) Representative confocal images of HepG2 cells expressing GFP-RFP-LC3 and quantitation of early autophagosome puncta and autolysosome puncta following FFA, FGF21 and Dorsomorphin treatment. Yellow showed co-localization of GFP and RFP, indicating early autophagosomes. Red only showed autolysosomes, Scale: 20 μm. Experiments were repeated three times. Data are shown as the means ± SEM. Significance was designated by asterisks with *p < 0.05. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Rat FGF21 enzyme-linked immunosorbent assay (ELISA) kit was obtained from Cusabio Biotech (Cat. No. CSB-EL008627RA, Cusabio Biotech, Wuhan, China).

Techniques: Western Blot, Expressing, Quantitation Assay

Journal: Redox Biology

Article Title: Exercise and dietary intervention ameliorate high-fat diet-induced NAFLD and liver aging by inducing lipophagy

doi: 10.1016/j.redox.2020.101635

Figure Lengend Snippet: FGF21 increased the lipophagy and reduced the lipid accumulation in FFA-treated HepG2 cells. HepG2 cells were treated with FFA (400 μM), FGF21 (1 μg/ml) as well as AMPK inhibitor-Dorsomorphin (1 μmol/L). (A) Representative confocal images of colocalization of LDs and autophagosomes by BODIPY 493/503 staining in GFP-LC3 protein expressed HepG2 cells. Scale: 50 μm. (B) TG level in HepG2 cells treated with FFA, FGF21 and Dorsomorphin. Experiments were repeated three times. Data are shown as the means ± SEM. Significance was designated by asterisks with *p < 0.05.

Article Snippet: Rat FGF21 enzyme-linked immunosorbent assay (ELISA) kit was obtained from Cusabio Biotech (Cat. No. CSB-EL008627RA, Cusabio Biotech, Wuhan, China).

Techniques: Staining

Journal: Redox Biology

Article Title: Exercise and dietary intervention ameliorate high-fat diet-induced NAFLD and liver aging by inducing lipophagy

doi: 10.1016/j.redox.2020.101635

Figure Lengend Snippet: FGF21 ameliorated the cell senescence through AMPK dependent autophagic pathway in FFA-treated WI-38 cells. WI-38 cells were treated with FFA (400 μM), FGF21 (1 μg/ml) as well as AMPK inhibitor-Dorsomorphin (1 μmol/L). (A) Representative of p -AMPK (Thr172)/AMPK total, p -ULK1 (Ser555)/ULK1 total and LC3-II/LC3-I immunoblotting in WI-38 cells and quantified data. (B) TG level in WI-38 cells. (C) Representative SA-β-gal staining (Scale: 50 μm), (D) SA-β-gal activity, (E) lipofuscin and (F) MDA contents in WI-38 cells. (G) p16 and (H) p27 mRNA levels in WI-38 cells. Experiments were repeated three times. Data are shown as the means ± SEM. Significance was designated by asterisks with *p < 0.05.

Article Snippet: Rat FGF21 enzyme-linked immunosorbent assay (ELISA) kit was obtained from Cusabio Biotech (Cat. No. CSB-EL008627RA, Cusabio Biotech, Wuhan, China).

Techniques: Western Blot, Staining, Activity Assay

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: High UA levels stimulated HDAC6 and inhibited FGF21 expression in HUVECs. A, NO production detected by NO assay kit (n = 5). B, HDAC6 mRNA expression determined by RT-qPCR (n = 5). C, HDAC6 protein expression determined by WB analysis (n = 4). D, FGF21 mRNA level measured by RT-qPCR (n = 5). E, FGF21 protein level measured by WB analysis (n = 4). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; # P < 0.05, ## P < 0.01, ### P < 0.001 versus 6 mg/dL UA.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Expressing, Quantitative RT-PCR

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: TSA attenuated UA-stimulated endothelial dysfunction in HUVECs. A, Representative cell morphology and ROS generation captured by a microscope (n = 3). B, TNF-α, IL-1β, and IL-6 mRNA levels determined by RT-qPCR (n = 5). C, NO production (n = 5). D, RT-qPCR analysis of CD31 and α-SMA (n = 5). E, WB analysis of CD31 and α-SMA (n = 4). F, α-SMA expression analyzed by immunofluorescence (n = 3). G, WB analysis of p-AKT/AKT and p-eNOS/eNOS (n = 4). H, RT-qPCR analysis of FGF21 (n = 5). I, WB analysis of FGF21 (n = 4). J, Level of histone H3 acetylation on the FGF21 promoter region, as analyzed by ChIP (n = 4). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; # P < 0.05, ## P < 0.01, ### P < 0.001 versus 12 mg/dL UA; & P < 0.05, && P < 0.01 versus 12 mg/dL UA+TSA low.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Microscopy, Quantitative RT-PCR, Expressing, Immunofluorescence

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: FGF21 knockdown abrogated the effects of TSA on UA-induced inflammation, oxidative stress, endothelial dysfunction, and endothelial–interstitial transformation in HUVECs. A, RT-qPCR analysis of FGF21 (n = 3). B, RT-qPCR analysis of TNF-α, IL-1β, and IL-6 (n = 4). C, ROS generation (n = 3). D, NO production (n = 5). E, Levels of p-AKT/AKT and p-eNOS/eNOS measured by WB (n = 4). F, RT-qPCR analysis of CD31 and α-SMA (n = 4). G, WB analysis of CD31 and α-SMA (n = 4). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; ### P < 0.001 versus 12 mg/dL UA; $$ P < 0.01, $$$ P < 0.001 versus 12 mg/dL UA+TSA high; & P < 0.05, && P < 0.01, &&& P < 0.001 versus 12 mg/dL UA+TSA high+NC.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Knockdown, Transformation Assay, Quantitative RT-PCR

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: LY294002 abrogated the effects of TSA on UA-induced vascular endothelial cell injury in HUVECs. A, NO production (n = 5). B, Levels of TNF-α, IL-1β, and IL-6 detected by RT-qPCR (n = 5). C, WB analysis of p-AKT/AKT, p-eNOS/eNOS, and p-FoxO3a/FoxO3a (n = 4). D, WB analysis of FGF21, CD31, and α-SMA (n = 4). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; ## P < 0.01, ### P < 0.001 versus 12 mg/dL UA; $ P < 0.05, $$ P < 0.01, $$$ P < 0.001 versus 12 mg/dL UA+TSA high; &&& P < 0.001 versus 12 mg/dL UA+TSA high+ LY294002.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Quantitative RT-PCR

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: Different from TSA, Mocetinostat, a class-Ⅰ selective HDAC inhibitor did not attenuate UA-stimulated endothelial dysfunction in HUVECs. A, HDAC1,2,3,6 activity detected by HDAC kits (n = 5). B, TNF-α, IL-1β, and IL-6 mRNA level detected by RT-qPCR (n = 5). C, NO production (n = 5). D, RT-qPCR analysis of FGF21 (n = 5). E, WB analysis of FGF21 (n = 4). F, Level of histone H3 acetylation on the FGF21 promoter region, as analyzed by ChIP (n = 4). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; # P < 0.05, ## P < 0.01, ### P < 0.001 versus 12 mg/dL UA; $ P < 0.05, $$ P < 0.01, $$$ P < 0.001 versus 12 mg/dL UA+TSA high.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Activity Assay, Quantitative RT-PCR

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: HDAC6 knockdown alleviated UA-induced inflammation, oxidative stress, endothelial dysfunction, and endothelial–interstitial transformation in HUVECs. A, HDAC6 mRNA level measured by RT-qPCR (n = 3). B, Levels of TNF-α, IL-1β, and IL-6 detected by RT-qPCR (n = 5). C, HDAC6 activity (n = 5). D, NO production (n = 5). E, WB analysis of FGF21 (n = 4). F, WB analysis of p-AKT/AKT, p-eNOS/eNOS, and p-FoxO3a/FoxO3a (n = 4). G, WB analysis of CD31 and α-SMA (n = 4). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; ## P < 0.01, ### P < 0.001 versus 12 mg/dL UA; $$ P < 0.01, $$$ P < 0.001 versus 12 mg/dL UA+NC.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Knockdown, Transformation Assay, Quantitative RT-PCR, Activity Assay

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: TSA or TubA alleviated aortic inflammation and dysfunction in hyperuricemic mice. A, Blood pressure between the groups (n = 15). B, Serum UA (n = 7–8). C, Serum NO (n = 8). D, Serum FGF21 (n = 7–8). E, Serum TNF-α and IL-6 (n = 7–8). F, Levels of TNF-α and IL-6 mRNA in aorta tissue were determined by RT-qPCR (n = 5). G, Vasorelaxant responses to Ach and SNP in mice aortic rings precontracted with phenylephrine (n = 5). H, Representative images of mice aorta histopathology through H&E staining (magnification × 200) and used for analysis of media thickness. I, WB analysis of HDAC1, 2, 3, and 6 in aorta tissue (n = 5). J, WB analysis of p-AKT/AKT, p-eNOS/eNOS, and p-FoxO3a/FoxO3a in aorta tissue (n = 5). K, WB analysis of FGF21, CD31, and α-SMA in aorta tissue (n = 5). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.01, *** P < 0.001 versus CON; # P < 0.05, ## P < 0.01, ### P < 0.001 versus model; $ P < 0.05, $$ P < 0.01 versus TSA.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Quantitative RT-PCR, Histopathology, Staining

Journal: Journal of Cardiovascular Pharmacology

Article Title: HDAC Inhibitors Alleviate Uric Acid–Induced Vascular Endothelial Cell Injury by Way of the HDAC6/FGF21/PI3K/AKT Pathway

doi: 10.1097/FJC.0000000000001372

Figure Lengend Snippet: TSA or TubA alleviated renal inflammation and dysfunction in hyperuricemic mice. A, Serum BUN (n = 7–8). B, Serum CRE (n = 8). C, RT-qPCR analysis of TNF-α and IL-6 in renal tissues (n = 5). D, Representative images of mice renal tissue stained by H&E, Masson, and PAS (magnification × 200), and percentage of fibrotic area was analyzed. E, WB analysis HDAC1, 2, 3, and 6 (n = 5). F, WB analysis of p-AKT/AKT, p-eNOS/eNOS, and p-FoxO3a/FoxO3a (n = 5). G, WB analysis of FGF21, CD31, and α-SMA (n = 5). One-way ANOVA followed by Tukey's post hoc multiple-group comparisons. * P < 0.05, ** P < 0.001, *** P < 0.001 versus CON; # P < 0.05, ## P < 0.01, ### P < 0.001 versus model; $ P < 0.05, $$ P < 0.01 versus TSA.

Article Snippet: The serum levels of FGF21 (CSB-EL008627MO), TNF-α (CSB-E04741m), and IL-6 (CSB-E04639m) were determined using the enzyme-linked immunosorbent assay (ELISA) kits (CUSABIO life Sciences, Wuhan, China) according to the kit's protocols.

Techniques: Quantitative RT-PCR, Staining

Journal: Cell Death & Disease

Article Title: FGF21 augments autophagy in random-pattern skin flaps via AMPK signaling pathways and improves tissue survival

doi: 10.1038/s41419-019-2105-0

Figure Lengend Snippet: FGF21 level was evaluated after the establishment of rat random skin flap model without drug administration. On Day 7, animals were euthanized, and then the flap survival area and subcutaneous microcirculation were evaluated. a Western blot showing FGF21 levels at several time points after the operation of random skin flap. b Histogram showing quantification of the FGF21 expression assessed by western blotting. c Digital photograph of flap survival/necrosis area on the 3rd and 7th day after surgery. d Histogram showing the percentage of survival area of flap on the 7th day. e Digital photograph exhibiting tissue necrosis, edema and the subcutaneous vascular network on the inner surface of the flap. f Histogram showing the percentage of tissue water content of flap. g Subcutaneous vascular network detected by LDBF, a stronger signal intensity representing greater blood flow. h Histogram showing signal intensity of blood flow in flap. i H&E staining exhibiting subcutaneous histology of the flap, showing subcutaneous blood vessels and inflammation (original magnification × 200; scan bar, 50 μm). j Histogram showing mean vessel density (MVD) in the flap (/mm 2 ). k IHC of CD34 which are mainly expressed in vascular endothelial cells; l Histogram showing the density of CD34-positive blood vessels (/mm 2 ). Significance: * p < 0.05 and ** p < 0.01 vs. Control group or Sham group; # p < 0.05 and ## p < 0.01 vs. random skin flap Day 3 group. Data were expressed as means ± SEM ( n = 6 per group).

Article Snippet: Primary antibodies were purchased from companies as following: FGF21 (26272-1-AP), VEGF (19003-1), SOD1 (10269-1), VPS34 (12452-1), MMP9 (10375-2), HO1 (10701-1), cathepsin D (CTSD, 21327-1), and Caspase 3 (CAPS3, 19677-1) from Proteintech Group (Chicago, IL, USA); cytochrome c (CYC, 14796), Bax (32027), endothelial nitric oxide synthase (eNOS, 11940S), AMPK (2532), p-AMPK (2537), mTOR (2983), p-mTOR (2971), FOXO3a (12829), p-FOXO3a (9466), and CARM1 (3379) from Cell Signaling Technology (Beverly, MA, USA); P-TFEB (Ser221) (AF3708) from Affinity Biosciences (OH, USA), SQSTM1/p62 (ab56416) from Abcam (Cambridge, UK); LC3B (L7543) from Sigma-Aldrich Chemical Company (Milwaukee, WI, USA); Cadherin5 (A02632-2) from Boster Biological Technology (Wuhan, China); GAPDH (AP0063) from Biogot Technology (Shanghai, China); Histone-H3 (17168-1-AP) and SKP2 (15010-1-AP) from Proteintech Group (Chicago, IL, USA).

Techniques: Western Blot, Expressing, Staining, Control

Journal: Cell Death & Disease

Article Title: FGF21 augments autophagy in random-pattern skin flaps via AMPK signaling pathways and improves tissue survival

doi: 10.1038/s41419-019-2105-0

Figure Lengend Snippet: On the 7th day after operation of random flap, the rats were sacrificed in the FGF21 group and FGF21 + 3MA group, and then autophagy level, survival area and subcutaneous microcirculation in the flap were evaluated. a Immunofluorescence for the evaluation of LC3II in the ischemic skin flap (scan bar, 15 μm). b Quantification of percentage of positive cells with LC3II labeled autophagosomes in dermal layer. c , d Western blotting performed for levels of autophagic proteins (Belin1, LC3II, CTSD, VPS34, and p62), which was corrected by GAPDH as internal control. a Histogram showing the quantification of autophagic proteins (Belin1, LC3II, CTSD, VPS34, and p62) detected by western blotting. e Digital photograph of flap survival/necrosis area on the 3rd and 7th day after surgery. f Histogram showing the percentage of survival area of flap on the 7th day. g Digital photograph exhibiting tissue necrosis, edema and the subcutaneous vascular network on the inner surface of the flap. h Histogram showing the percentage of tissue water content of flap. i The subcutaneous vascular network detected by LDBF, with stronger signal intensity representing greater blood flow. j Histogram showing signal intensity of blood flow in flap. k H&E staining exhibiting subcutaneous histology of the flap, showing subcutaneous blood vessels and inflammation (original magnification × 200; scan bar, 50 μm). l Histogram showing mean vessel density (MVD) in the flap (/mm 2 ). m IHC of CD34 which mainly were expressed in vascular endothelial cells. n Histogram showing the density of CD34-positive blood vessels (/mm 2 ). Significance: * p < 0.05 and ** p < 0.01 vs the FGF21 group. Data were expressed as means ± SEM ( n = 6 per group).

Article Snippet: Primary antibodies were purchased from companies as following: FGF21 (26272-1-AP), VEGF (19003-1), SOD1 (10269-1), VPS34 (12452-1), MMP9 (10375-2), HO1 (10701-1), cathepsin D (CTSD, 21327-1), and Caspase 3 (CAPS3, 19677-1) from Proteintech Group (Chicago, IL, USA); cytochrome c (CYC, 14796), Bax (32027), endothelial nitric oxide synthase (eNOS, 11940S), AMPK (2532), p-AMPK (2537), mTOR (2983), p-mTOR (2971), FOXO3a (12829), p-FOXO3a (9466), and CARM1 (3379) from Cell Signaling Technology (Beverly, MA, USA); P-TFEB (Ser221) (AF3708) from Affinity Biosciences (OH, USA), SQSTM1/p62 (ab56416) from Abcam (Cambridge, UK); LC3B (L7543) from Sigma-Aldrich Chemical Company (Milwaukee, WI, USA); Cadherin5 (A02632-2) from Boster Biological Technology (Wuhan, China); GAPDH (AP0063) from Biogot Technology (Shanghai, China); Histone-H3 (17168-1-AP) and SKP2 (15010-1-AP) from Proteintech Group (Chicago, IL, USA).

Techniques: Immunofluorescence, Labeling, Western Blot, Control, Staining

Journal: Cell Death & Disease

Article Title: FGF21 augments autophagy in random-pattern skin flaps via AMPK signaling pathways and improves tissue survival

doi: 10.1038/s41419-019-2105-0

Figure Lengend Snippet: On the 7th day after operation, samples were harvested from the Control and FGF21 groups to evaluate the TFEB level. Further, transfection of the AVV TFEB shRNA was performed to inhibit the expression of TFEB in the cells, and then the expression of relative proteins was compared in the FGF21, Scramble and TFEB shRNA group. a Immunofluorescence exhibiting more expression of TFEB in the FGF21 group than that in the Control group (scan bar, 15 μm). b Histogram showing the percentage of TFEB translocation into nucleus in dermal layer. c Western blotting showing levels of P-TFEB (Ser221) and nuclear TFEB. d Histogram showing quantificational comparison of cytoplasmic and nuclear TFEB expressions between the Control and FGF21 groups. e Immunofluorescence exhibiting the expression of LC3II in the FGF21, Scramble and TFEB shRNA groups. f Quantification of percentage of positive cells with LC3II labeled autophagosomes in dermal layer. g Western blotting showing levels of P-TFEB (Ser221) and nuclear TFEB in the FGF21, Scramble and TFEB shRNA groups. h Quantification of cytoplasmic and nuclear TFEB expressions detected by western blotting. i Western blotting showing levels of autophagic proteins (p62, Beclin1, VPS34, CTSD, and LC3II) in the FGF21, Scramble and TFEB shRNA groups, which was corrected by GAPDH as internal control. j Quantification of expression of autophagy-related proteins (p62, Beclin1, VPS34, CTSD, and LC3II) in western blotting. Significance: * p < 0.05 and ** p < 0.01 vs the Control group. # p < 0.05 and ## p < 0 . 01 vs the Scramble group . Data were expressed as means ± SEM ( n = 6 per group).

Article Snippet: Primary antibodies were purchased from companies as following: FGF21 (26272-1-AP), VEGF (19003-1), SOD1 (10269-1), VPS34 (12452-1), MMP9 (10375-2), HO1 (10701-1), cathepsin D (CTSD, 21327-1), and Caspase 3 (CAPS3, 19677-1) from Proteintech Group (Chicago, IL, USA); cytochrome c (CYC, 14796), Bax (32027), endothelial nitric oxide synthase (eNOS, 11940S), AMPK (2532), p-AMPK (2537), mTOR (2983), p-mTOR (2971), FOXO3a (12829), p-FOXO3a (9466), and CARM1 (3379) from Cell Signaling Technology (Beverly, MA, USA); P-TFEB (Ser221) (AF3708) from Affinity Biosciences (OH, USA), SQSTM1/p62 (ab56416) from Abcam (Cambridge, UK); LC3B (L7543) from Sigma-Aldrich Chemical Company (Milwaukee, WI, USA); Cadherin5 (A02632-2) from Boster Biological Technology (Wuhan, China); GAPDH (AP0063) from Biogot Technology (Shanghai, China); Histone-H3 (17168-1-AP) and SKP2 (15010-1-AP) from Proteintech Group (Chicago, IL, USA).

Techniques: Control, Transfection, shRNA, Expressing, Immunofluorescence, Translocation Assay, Western Blot, Comparison, Labeling

Journal: Cell Death & Disease

Article Title: FGF21 augments autophagy in random-pattern skin flaps via AMPK signaling pathways and improves tissue survival

doi: 10.1038/s41419-019-2105-0

Figure Lengend Snippet: On the 7th day after operation, samples were harvested from the Control, FGF21, FGF21 + CC, FGF21 + CC + Torin1 groups for the evaluation. a Western blotting showing the cytoplasmic levels of AMPK, p-AMPK, mTOR, p-mTOR and P-TFEB (Ser221) which were corrected by GAPDH as internal control; and nuclear levels of TFEB which were corrected by Histone-H3 as internal control. b Histogram showing quantificational comparison of AMPK, p-AMPK, mTOR and p-mTOR. c Western blotting showing the nuclear levels of AMPK, p-AMPK, FOXO3a, p-FOXO3a, SKP2, and CARM1 which were corrected by Histone-H3 as internal control. d Histogram showing quantificational comparison of AMPK, p-AMPK, FOXO3a, p-FOXO3a, SKP2, and CARM1 between the Control and FGF21 groups. e Western blotting for Immunoprecipitation of CARM1 and TFEB. f Histogram exhibiting the quantification of CARM1 and TFEB levels under Immunoprecipitation. g Western blotting showing levels of proteins of LC3II, p62, VEGF, SOD1, and CASP3 which were corrected by GAPDH as internal control. h Histogram showing quantificational comparison of LC3II, p62, VEGF, SOD1, and CASP3. Significance: * p < 0.05 and ** p < 0.01 vs the Control group; @ p < 0.05 and @@ p < 0 . 01 vs the FGF21; & p < 0 . 05 and && p < 0.01 vs the FGF21 + CC. Data were expressed as means ± SEM ( n = 6 per group).

Article Snippet: Primary antibodies were purchased from companies as following: FGF21 (26272-1-AP), VEGF (19003-1), SOD1 (10269-1), VPS34 (12452-1), MMP9 (10375-2), HO1 (10701-1), cathepsin D (CTSD, 21327-1), and Caspase 3 (CAPS3, 19677-1) from Proteintech Group (Chicago, IL, USA); cytochrome c (CYC, 14796), Bax (32027), endothelial nitric oxide synthase (eNOS, 11940S), AMPK (2532), p-AMPK (2537), mTOR (2983), p-mTOR (2971), FOXO3a (12829), p-FOXO3a (9466), and CARM1 (3379) from Cell Signaling Technology (Beverly, MA, USA); P-TFEB (Ser221) (AF3708) from Affinity Biosciences (OH, USA), SQSTM1/p62 (ab56416) from Abcam (Cambridge, UK); LC3B (L7543) from Sigma-Aldrich Chemical Company (Milwaukee, WI, USA); Cadherin5 (A02632-2) from Boster Biological Technology (Wuhan, China); GAPDH (AP0063) from Biogot Technology (Shanghai, China); Histone-H3 (17168-1-AP) and SKP2 (15010-1-AP) from Proteintech Group (Chicago, IL, USA).

Techniques: Control, Western Blot, Comparison, Immunoprecipitation

Journal: Molecular Metabolism

Article Title: Genome-wide association study for circulating FGF21 in patients with alcohol use disorder: Molecular links between the SNHG16 locus and catecholamine metabolism

doi: 10.1016/j.molmet.2022.101534

Figure Lengend Snippet: Plasma FGF21 concentrations were associated with recent alcohol use phenotypes. Plasma FGF21 levels were positively correlated with recent alcohol use as determined by TLFB 90 days prior to blood collection i.e. (A) the number of drinking days, (B) the number of heavy drinking days, and (C) total drinks. (D) Plasma FGF21 levels were also positively correlated with blood levels of gamma-glutamyl transferase (GGT) which is commonly used as a marker for heavy alcohol use.

Article Snippet: We measured FGF21 levels in the iPSC-derived brain organoids and HepG2 cell lysates with an FGF21 ELISA kit (EK0994, Boster Biological Technology, Pleasanton, CA) in accordance with the manufacturer's instructions.

Techniques: Clinical Proteomics, Marker

Journal: Molecular Metabolism

Article Title: Genome-wide association study for circulating FGF21 in patients with alcohol use disorder: Molecular links between the SNHG16 locus and catecholamine metabolism

doi: 10.1016/j.molmet.2022.101534

Figure Lengend Snippet: (A) Schematic outline of our proteomics-informed genomics research strategy. (B–C) Q–Q plot and Manhattan plot for GWAS of plasma FGF21 levels. (D) The locus zoom plot displays that the top SNPs on chromosome 17 map within a gene cluster: CYGB , PRCD and SNHG16 . The SNP most highly associated with plasma FGF21 levels in patients with AUD was rs9914222 ( p: 4.6E-09).

Article Snippet: We measured FGF21 levels in the iPSC-derived brain organoids and HepG2 cell lysates with an FGF21 ELISA kit (EK0994, Boster Biological Technology, Pleasanton, CA) in accordance with the manufacturer's instructions.

Techniques: Clinical Proteomics

Journal: Molecular Metabolism

Article Title: Genome-wide association study for circulating FGF21 in patients with alcohol use disorder: Molecular links between the SNHG16 locus and catecholamine metabolism

doi: 10.1016/j.molmet.2022.101534

Figure Lengend Snippet: Biological functions of the rs9914222 SNP. (A) SNP-dependent plasma FGF21 levels in patients with AUD demonstrating that rs9914222 is an eQTL. (B) rs9914222 is associated with SNHG16 mRNA expression in several brain regions. https://gtexportal.org/home/snp/rs9914222 .

Article Snippet: We measured FGF21 levels in the iPSC-derived brain organoids and HepG2 cell lysates with an FGF21 ELISA kit (EK0994, Boster Biological Technology, Pleasanton, CA) in accordance with the manufacturer's instructions.

Techniques: Clinical Proteomics, Expressing

Journal: Molecular Metabolism

Article Title: Genome-wide association study for circulating FGF21 in patients with alcohol use disorder: Molecular links between the SNHG16 locus and catecholamine metabolism

doi: 10.1016/j.molmet.2022.101534

Figure Lengend Snippet: Biological functions of the SNHG16 gene. (A) FGF21 concentration was measured after the knockdown of SNHG16 using siRNA in HepG2 cells. Relative mRNA expression of SNHG16 and COMT was determined after the knockdown of SNHG16. COMT enzyme activity was also measured after the knockdown of SNHG16 in HepG2 cells. At least three independent experiments were performed. ∗ p < 0.05. (B) FGF21 concentration was determined before and after ethanol treatment of HepG2 cells. Relative mRNA expression of SNHG16 and COMT was determined in response to ethanol treatment. COMT enzyme activity was then measured using HepG2 cells treated with ethanol. ∗A p value ≤ 0.05 was considered statistically significant (two tailed paired t test). Three independent experiments were performed. All values are mean ± S.E.M.

Article Snippet: We measured FGF21 levels in the iPSC-derived brain organoids and HepG2 cell lysates with an FGF21 ELISA kit (EK0994, Boster Biological Technology, Pleasanton, CA) in accordance with the manufacturer's instructions.

Techniques: Concentration Assay, Knockdown, Expressing, Activity Assay, Two Tailed Test

Journal: Molecular Metabolism

Article Title: Genome-wide association study for circulating FGF21 in patients with alcohol use disorder: Molecular links between the SNHG16 locus and catecholamine metabolism

doi: 10.1016/j.molmet.2022.101534

Figure Lengend Snippet: Ethanol induced FGF21 in iPSC-derived brain organoids and activated the release of catecholamines. (A) A schematic outline of procedures used during the differentiation of iPSC-derived brain organoids. The panel below the schematic displays representative examples of staining for tyrosin hydroxylase (TH), and Neuron-specific class III beta-tubulin (TUJ1). (B) the effect of ethanol on FGF21 concentration was measure using ELISA. Dopamine, norepinephrine, and epinephrine concentrations were measured using UPLC–Tandem Mass Spectrometry. (C) A schematic outline of the catecholamine biosynthesis pathway. (D) mRNA expression of SNHG16, MAOA, MAOB, COMT, TH, COMT, DDC, DBH, and PNMT in response to ethanol treatment (25 mM) for 7 days. Realtime PCR experiments were performed in iPSC-derived brain organoids (n = 3). The expression of these genes was determined after exposure to drug for 7 days. ∗A p value ≤ 0.05 was considered statistically significant (two tailed paired t test). All values shown are mean ± S.E.M.

Article Snippet: We measured FGF21 levels in the iPSC-derived brain organoids and HepG2 cell lysates with an FGF21 ELISA kit (EK0994, Boster Biological Technology, Pleasanton, CA) in accordance with the manufacturer's instructions.

Techniques: Derivative Assay, Staining, Concentration Assay, Enzyme-linked Immunosorbent Assay, Mass Spectrometry, Expressing, Two Tailed Test

Journal: Molecular Metabolism

Article Title: Genome-wide association study for circulating FGF21 in patients with alcohol use disorder: Molecular links between the SNHG16 locus and catecholamine metabolism

doi: 10.1016/j.molmet.2022.101534

Figure Lengend Snippet: Gene expression of iPSC-derived brain organoids. (A) Schematic diagram illustrating the effects of ethanol on FGF21 which have implications for alcohol use. Specifically, FGF21 can be induced by ethanol. Our GWAS for plasma FGF21 showed that a SNP located 5′ of SNHG16 is associated with AUD. SNHG16 could regulate COMT expression and activity, which play a role in catecholamine metabolism. Finally, we determined that ethanol induced both FGF21 and catecholamines, including dopamine, norepinephrine and epinephrine using iPSC-derived brain organoids. (B – G) mRNA expression of SNHG16, TH, COMT, DDC, DBH, PNMT in response to drug treatment. Realtime PCR experiments were performed in iPSC-derived brain organoids (n = 3). The expression of those genes was determined after exposure to drugs for 7 days. ∗A p value ≤ 0.05 was considered statistically significant (two tailed paired t test). All values are mean ± S.E.M.

Article Snippet: We measured FGF21 levels in the iPSC-derived brain organoids and HepG2 cell lysates with an FGF21 ELISA kit (EK0994, Boster Biological Technology, Pleasanton, CA) in accordance with the manufacturer's instructions.

Techniques: Gene Expression, Derivative Assay, Clinical Proteomics, Expressing, Activity Assay, Two Tailed Test