Journal: PLoS ONE

Article Title: Hepatic Fgf21 Expression Is Repressed after Simvastatin Treatment in Mice

doi: 10.1371/journal.pone.0162024

Figure Lengend Snippet: A. Immunoblotting analysis of Fgf21 in the livers of 3-month old male mice treated with simvastatin (0.1% w/w in chow) for 1 week. Immunoblots for Ampkα and elf2a and Coomassie blue staining of the gel were used as loading controls. B. Relative Fgf21 protein levels as assessed by immunoblotting after normalization to elf2a levels. Data are presented as the mean ± SEM. n = 5 per treatment. *P<0.05. C, D, E, F, G, H . mRNA levels of Fgf21(C), Hmgcr (D), Pcsk9 (E), Acox1 (F), Cyp4a10 (G), Cyp7a1 (H) in the livers of 3-month old male mice treated with simvastatin (0.1% w/w in chow) for 1 week. Relative mRNA levels were assessed by qRT-PCR. Data are presented as the mean ±SEM. n = 7–8 per treatment. *P<0.05. I. Assessment of serum Fgf21 levels by ELISA in 3-month old male mice treated with vehicle or simvastatin. Data are presented as the mean ±SEM. n = 7–8 per treatment. *P<0.05.

Article Snippet: Serum Fgf21 levels were measured using an ELISA kit (MF2100) from R&D Biosystems (Minneapolis, MN) with the serum of mice that had been starved overnight.

Techniques: Western Blot, Staining, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: Hepatic Fgf21 Expression Is Repressed after Simvastatin Treatment in Mice

doi: 10.1371/journal.pone.0162024

Figure Lengend Snippet: A. Body weights of mice before and after exposure to simvastatin expressed as the % of initial weight. *P<0.05 compared with baseline weight. B. Cumulative food intake for the 1-week treatment with simvastatin. *P<0.05 compared with vehicle treatment (0% simvastatin w/w). C. Fgf21 hepatic mRNA levels as assessed by qRT-PCR. *P<0.05 compared with the 0% dose. a, b, c, d, e denote 0%, 0.01%, 0.05%, 0.1% and 0.5% w/w simvastatin in chow, respectively. For panels A, B, C the data are presented as the mean ± SEM. n = 6 per treatment with the exception of the 0.5% dose (n = 3; 5 mice received the treatment and 2 died after the treatment).

Article Snippet: Serum Fgf21 levels were measured using an ELISA kit (MF2100) from R&D Biosystems (Minneapolis, MN) with the serum of mice that had been starved overnight.

Techniques: Quantitative RT-PCR

Journal: PLoS ONE

Article Title: Hepatic Fgf21 Expression Is Repressed after Simvastatin Treatment in Mice

doi: 10.1371/journal.pone.0162024

Figure Lengend Snippet: Fgf21 ( A ) and Pcsk9 ( B ) mRNA levels in the liver of mice administered vehicle (control) or simvastatin intraperitoneally twice (20 and 12 hours before sacrifice). Data are presented as the mean ± SEM, n = 10 per treatment. *P<0.05 compared with control.

Article Snippet: Serum Fgf21 levels were measured using an ELISA kit (MF2100) from R&D Biosystems (Minneapolis, MN) with the serum of mice that had been starved overnight.

Techniques:

Journal: PLoS ONE

Article Title: Hepatic Fgf21 Expression Is Repressed after Simvastatin Treatment in Mice

doi: 10.1371/journal.pone.0162024

Figure Lengend Snippet: mRNA levels of Fgf21 ( A ), Pcsk9 ( B ), Acox1 ( C ), Cyp4a10 ( D ) in primary hepatocytes treated with vehicle or simvastatin (3 different doses) for 12 hours. Data are presented as the means ±SEM from 3 individual experiments, each of which included 3 technical replicates. *P<0.05 (compared with vehicle treatment).

Article Snippet: Serum Fgf21 levels were measured using an ELISA kit (MF2100) from R&D Biosystems (Minneapolis, MN) with the serum of mice that had been starved overnight.

Techniques:

Journal: PLoS ONE

Article Title: Hepatic Fgf21 Expression Is Repressed after Simvastatin Treatment in Mice

doi: 10.1371/journal.pone.0162024

Figure Lengend Snippet: Srebp-2 ( A ) and Fgf21 ( B ) mRNA levels in primary hepatocytes after overexpression of Srebp-2. Data are presented as the means ±SEM from 3 individual experiments, each of which included 3 technical replicates. *P<0.05 (compared with empty vector transfection). LDLR ( C ) and FGF21 ( D ) mRNA levels in HepG2 cells after overexpression of Srebp-2. Data are presented as the means ±SEM from 6 individual experiments, each of which included 3 technical replicates. *P<0.05 (compared with empty vector transfection). ABCA1 ( E ) and FGF21 ( F ) mRNA levels in HepG2 cells after overexpression of miR-33. Data are presented as the means ±SEM from 3 individual experiments, each of which included 3 technical replicates. *P<0.05 (compared with empty vector transfection).

Article Snippet: Serum Fgf21 levels were measured using an ELISA kit (MF2100) from R&D Biosystems (Minneapolis, MN) with the serum of mice that had been starved overnight.

Techniques: Over Expression, Plasmid Preparation, Transfection

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: Cell Metabolism

Article Title: GDF15 Provides an Endocrine Signal of Nutritional Stress in Mice and Humans

doi: 10.1016/j.cmet.2018.12.016

Figure Lengend Snippet: GDF15 Expression Is Regulated by the Cellular ISR Pathway (A and C) GDF15 mRNA expression (A) and immunoblot analysis (C) of ISR components in wild-type (WT) mouse embryonic fibroblasts (MEFs) treated with vehicle control (Con), cobalt chloride (CoCl2, 625 μM), thapsigargin (Tg, 1 μM), tunicamycin (Tn 5 μg/mL), or L-Histidinol (His, 1 mM) for 6 h. (B) GDF15 mRNA expression in human cell lines (HeLA, HuH7, and A549) treated with Tn (5 μg/mL) for 6 h. (D) GDF15 mRNA expression in WT MEFs pre-treated for 1 h either with the PERK inhibitor GSK2606414 (GSK, 200 nM) or eIF2α inhibitor ISRIB (ISR, 100 nM), then co-treated with Tn (5 μg/mL) for a further 6 h. (E–G) GDF15 mRNA expression (E) in EIF2α Ser51 (SS) or phospho mutant (AA) MEFs or (F) in ATF4 wild-type (WT) or ATF4 knockout (KO) MEFs and (G) in control siRNA and CHOP siRNA transfected WT MEFs treated with Tn (5 μg/mL) for 6 h. (H) Diagram outlining pathway by which GDF15 and FGF21 expression is regulated by TN. mRNA expression is presented as fold expression relative to its respective control treatment for each cell type (set at 1) or TN-treated samples (set as 100) with normalization to HPRT gene expression in MEFs and GAPDH in human cells. Data are expressed as mean ± SD from at least three independent experiments. ∗∗∗ p < 0.001 versus control (con) for (A) and (B), and versus TN stimulated for (D)–(G) by two-tailed Student’s t test. Blots shown are representative of three independent experiments with Calnexin used as a loading control. See also Figures S3 and .

Article Snippet: Mouse FGF21 was analyzed by FGF21 Quantakine ELISA (R&D Systems) following the manufacturer’s instructions.

Techniques: Expressing, Western Blot, Control, Mutagenesis, Knock-Out, Transfection, Gene Expression, Two Tailed Test

Journal: Cell Metabolism

Article Title: GDF15 Provides an Endocrine Signal of Nutritional Stress in Mice and Humans

doi: 10.1016/j.cmet.2018.12.016

Figure Lengend Snippet:

Article Snippet: Mouse FGF21 was analyzed by FGF21 Quantakine ELISA (R&D Systems) following the manufacturer’s instructions.

Techniques: Recombinant, SYBR Green Assay, Protease Inhibitor, Reverse Transcription, Western Blot, Injection, Enzyme-linked Immunosorbent Assay, Control, TaqMan Assay, Software, Sterility, Electrophoresis, Real-time Polymerase Chain Reaction

Journal: PLoS ONE

Article Title: Fibroblast Growth Factor 21 (FGF21) Protects against High Fat Diet Induced Inflammation and Islet Hyperplasia in Pancreas

doi: 10.1371/journal.pone.0148252

Figure Lengend Snippet: FGF21 is regulated by the nutritional status in fed, fasted and diet-induced obesity (DIO). Fasting reduced FGF21 mRNA in whole pancreas (A). β-klotho, the co-receptor of FGF21, is expressed in the pancreas and is inversely regulated by fasting (B). FGF21 protein was only detected in pancreas compared to liver and inguinal white adipose tissue (IWAT) (C). Pancreas FGF21 protein level was reduced by fasting in both chow fed and high fat diet fed conditions (D, E). Separation of pancreatic fractions to evaluate the individual contribution of islets and acinar tissue on FGF21 expression show that acinar pancreas contributes to the majority of FGF21 levels while islet cells have very low FGF21 levels and are not nutritionally regulated (F). Acinar expression of FGF21 is regulated with fasting (F, G). n = 6 per group. Each western blot lane represents an individual animal.

Article Snippet: Antibodies were purchased from commercial vendors: FGF21 (AF3057; R&D systems, Minneapolis, MN); Insulin (ab7842; Abcam, Cambridge, MA); β-actin (ab8226; Abcam); α -tubulin (ab7291; Abcam); pERK1/2 (4370; Cell Signaling Technology, Denver, MA); ERK1/2 (9107; Cell Signaling Technology); glucagon (2654; Sigma Aldrich, St. Louis, MO); somatostatin (sc7819; Santa Cruz, Dallas, TX); CD3 (MA1-90582; Thermo Scientific, Waltham, MA).

Techniques: Expressing, Western Blot

Journal: PLoS ONE

Article Title: Fibroblast Growth Factor 21 (FGF21) Protects against High Fat Diet Induced Inflammation and Islet Hyperplasia in Pancreas

doi: 10.1371/journal.pone.0148252

Figure Lengend Snippet: Acute FGF21 exposure induces ERK1/2 signaling events therefore phosphorylation of ERK1/2 (pERK1/2) has been chosen as a marker of FGF21 signaling. Up to 50% of acinar cells show strong nuclear pERK1/2 staining (brown) after FGF21 administration (A i, iii). Islets represent a very unique pattern with pERK1/2 only on the peripheral cells (A ii, iv). Percentage of pERK1/2-labeled acinar cells with saline and FGF21 administration (B). Western blot of pERK1/2 and total ERK1/2 from the same pancreas samples (C). Epididymal white adipose tissue (EWAT) is a positive control for FGF21 signaling (C). S = saline and F = FGF21. Confocal photomicrographs show FGF21-induced pERK1/2 immunoreactivity in islets (D). Absence of pERK1/2 staining (green) after saline treatment (Di; Inset , islet outline is indicated by insulin-immunoreactivity shown in red). FGF21 did not elicit pERK1/2 in insulin (red)-producing β-cells (D ii). Merged images show pERK1/2 co-localization in glucagon-positive α cells (D iii) and somatostatin-positive δ cell (D iv). Split panels show glucagon (iii a ) and pERK (iii b ) labeling or somatostatin (iv a ) and pERK (iv b ) labeling in single cells. A robust and specific pERK1/2 staining was observed in almost 10% of total islet nuclei mostly at the periphery. Outlined region (dashed line) in the inset indicates the region represented in each panel. Representative pERK1/2 labelled cells are indicated by white arrows. n = 4 per group. Scale: 200 μm, A; 20 μm, D.

Article Snippet: Antibodies were purchased from commercial vendors: FGF21 (AF3057; R&D systems, Minneapolis, MN); Insulin (ab7842; Abcam, Cambridge, MA); β-actin (ab8226; Abcam); α -tubulin (ab7291; Abcam); pERK1/2 (4370; Cell Signaling Technology, Denver, MA); ERK1/2 (9107; Cell Signaling Technology); glucagon (2654; Sigma Aldrich, St. Louis, MO); somatostatin (sc7819; Santa Cruz, Dallas, TX); CD3 (MA1-90582; Thermo Scientific, Waltham, MA).

Techniques: Phospho-proteomics, Marker, Staining, Labeling, Saline, Western Blot, Positive Control

Journal: PLoS ONE

Article Title: Fibroblast Growth Factor 21 (FGF21) Protects against High Fat Diet Induced Inflammation and Islet Hyperplasia in Pancreas

doi: 10.1371/journal.pone.0148252

Figure Lengend Snippet: Chronic FGF21 treatment for 3 days by mini osmotic pumps led to the reduction in FGF21 expression in pancreas (A). FGF21 infusion down-regulated insulin but glucagon, somatostatin or amylase expression. While FGF21 infusion did not alter serum glucose levels (B), it reduced the serum levels of pancreatic hormones, insulin (C) and glucagon (D) without affecting the amylase activity (E). n = 10 per group.

Article Snippet: Antibodies were purchased from commercial vendors: FGF21 (AF3057; R&D systems, Minneapolis, MN); Insulin (ab7842; Abcam, Cambridge, MA); β-actin (ab8226; Abcam); α -tubulin (ab7291; Abcam); pERK1/2 (4370; Cell Signaling Technology, Denver, MA); ERK1/2 (9107; Cell Signaling Technology); glucagon (2654; Sigma Aldrich, St. Louis, MO); somatostatin (sc7819; Santa Cruz, Dallas, TX); CD3 (MA1-90582; Thermo Scientific, Waltham, MA).

Techniques: Expressing, Activity Assay

Journal: PLoS ONE

Article Title: Fibroblast Growth Factor 21 (FGF21) Protects against High Fat Diet Induced Inflammation and Islet Hyperplasia in Pancreas

doi: 10.1371/journal.pone.0148252

Figure Lengend Snippet: WT and FGF21 KO mice on chow diet have similar body weight (A), glucose tolerance (B) and insulin secretion in response to glucose (C), despite notable differences in their pancreatic morphology (D i, iii). After consuming high fat diet for 16 weeks, FGF21 KO mice (D iv) had increased islet hyperplasia compared with WT mice (D ii). Islet cell surface area was measured and represented (E). Islet cell proliferation markers were substantially over expressed in obese FGF21 KO animals (F). Islet hyperplasia leads to increased serum insulin (G) in FGF21 KO animals with a slight reduction in blood glucose (H) (n = 7 per group). Scale: 1 mm.

Article Snippet: Antibodies were purchased from commercial vendors: FGF21 (AF3057; R&D systems, Minneapolis, MN); Insulin (ab7842; Abcam, Cambridge, MA); β-actin (ab8226; Abcam); α -tubulin (ab7291; Abcam); pERK1/2 (4370; Cell Signaling Technology, Denver, MA); ERK1/2 (9107; Cell Signaling Technology); glucagon (2654; Sigma Aldrich, St. Louis, MO); somatostatin (sc7819; Santa Cruz, Dallas, TX); CD3 (MA1-90582; Thermo Scientific, Waltham, MA).

Techniques:

Journal: PLoS ONE

Article Title: Fibroblast Growth Factor 21 (FGF21) Protects against High Fat Diet Induced Inflammation and Islet Hyperplasia in Pancreas

doi: 10.1371/journal.pone.0148252

Figure Lengend Snippet: HFD consumption caused severe pancreatic inflammation in FGF21 KO mice (A). H&E staining showing, that compared to WT animals (i), FGF21 KO mice developed severe pancreatic periductal inflammation (outlined area in ii-iii) when consuming a HFD diet for 16 weeks. Lymphocytic nature of inflammatory cells is shown in a higher magnification image (iii). (B) Immunohistochemical analysis for the lymphocytic marker CD3 on the pancreas of WT (i) and FGF21 KO animals (ii-iii) is represented. FGF21 KO showed higher number of CD3, T cell receptor antigen. n = 7 per group. Scale: 1 mm, Ai-ii; 2 mm, Bi-ii; 200 μm, iii.

Article Snippet: Antibodies were purchased from commercial vendors: FGF21 (AF3057; R&D systems, Minneapolis, MN); Insulin (ab7842; Abcam, Cambridge, MA); β-actin (ab8226; Abcam); α -tubulin (ab7291; Abcam); pERK1/2 (4370; Cell Signaling Technology, Denver, MA); ERK1/2 (9107; Cell Signaling Technology); glucagon (2654; Sigma Aldrich, St. Louis, MO); somatostatin (sc7819; Santa Cruz, Dallas, TX); CD3 (MA1-90582; Thermo Scientific, Waltham, MA).

Techniques: Staining, Immunohistochemical staining, Marker

Journal: PLoS ONE

Article Title: Fibroblast Growth Factor 21 (FGF21) Protects against High Fat Diet Induced Inflammation and Islet Hyperplasia in Pancreas

doi: 10.1371/journal.pone.0148252

Figure Lengend Snippet: (A) Representative cytofluorometric dot plots of isolated lymphocytes from WT and FGF21 KO animals consuming high fat diet for 16 weeks. FGF21 KO mice show elevated CD45+ (i), TCRb+ and Thy1+ T lymphocytes (ii) and Foxp3+ Treg cells. CD19+ B lymphocytes were not significantly altered (iii). Corresponding summary data is shown in the right panel. The experiment was repeated twice (n = 4 per group). (B) Gene expression analysis of cytokines is presented. n = 7 per group.

Article Snippet: Antibodies were purchased from commercial vendors: FGF21 (AF3057; R&D systems, Minneapolis, MN); Insulin (ab7842; Abcam, Cambridge, MA); β-actin (ab8226; Abcam); α -tubulin (ab7291; Abcam); pERK1/2 (4370; Cell Signaling Technology, Denver, MA); ERK1/2 (9107; Cell Signaling Technology); glucagon (2654; Sigma Aldrich, St. Louis, MO); somatostatin (sc7819; Santa Cruz, Dallas, TX); CD3 (MA1-90582; Thermo Scientific, Waltham, MA).

Techniques: Isolation, Gene Expression

Journal: Journal of Clinical Investigation

Article Title: Peripherally derived FGF21 promotes remyelination in the central nervous system

doi: 10.1172/jci94337

Figure Lengend Snippet: Figure 3. FGF21 secreted from pancreas induces OPC proliferation. (A) Quantitations of Fgf21 mRNA (left) and FGF21 protein (right) in individual organs of intact mice (n = 9); **P < 0.01. (B) Representative images of FGF21-immunolabeled pancreas of intact mice (n = 3). (C) Double IHC staining for FGF21 with the indicated cell markers in the pancreas of adult mice (n = 3). (D) BrdU incorporation in mouse OPCs after stimulation with serum from mice with FGF21 knockdown in the pancreas (n = 4); *P < 0.05, **P < 0.01, as determined by ANOVA with Tukey’s post hoc test. Error bars represent SEM. Scale bars: 50 μm (B); 10 μm (C).

Article Snippet: To assess the role of FGFs, we used recombinant mouse FGF15 (Abcam), recombinant mouse FGF21 (R&D Systems), and recombinant mouse FGF23 (R&D Systems).

Techniques: Immunolabeling, Immunohistochemistry, BrdU Incorporation Assay, Knockdown

Journal: Journal of Clinical Investigation

Article Title: Peripherally derived FGF21 promotes remyelination in the central nervous system

doi: 10.1172/jci94337

Figure Lengend Snippet: Figure 6. FGF21 promotes human OPC proliferation. (A) Representative image of β-klotho expression in an autopsied sample from healthy patient and a patient with multiple sclerosis. Graphs show quantitations as indicated in the images (n = 4 for healthy patients, 3 for multiple sclerosis patients); **P < 0.01. (B) BrdU incorporation in human OPCs after stimulation with recombinant FGF21 (n = 6 for control, 4 for FGF21); *P < 0.05 as determined by Student’s t test. Error bars represent SEM. Scale bar: 20 μm.

Article Snippet: To assess the role of FGFs, we used recombinant mouse FGF15 (Abcam), recombinant mouse FGF21 (R&D Systems), and recombinant mouse FGF23 (R&D Systems).

Techniques: Expressing, BrdU Incorporation Assay, Recombinant, Control

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