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: Scientific Reports

Article Title: Fgf21 regulates T-cell development in the neonatal and juvenile thymus

doi: 10.1038/s41598-017-00349-8

Figure Lengend Snippet: Fgf21 mRNA is enriched in mature mTECs. ( A and B ) Expression levels of Fgf21 mRNA in the various tissues from 4-week-old C57BL/6 mice ( A ) and in the thymus of E15.5 and 0-, 4-, 8- and 12-week-old ( B ). Relative Fgf21 mRNA expression was normalised against 18S rRNA expression. ( C and D ) Thymic cell suspensions from 4-week-old C57BL/6 mice were assessed by flow cytometric analysis using anti-CD45 mAb, anti-EpCAM mAb, anti-MHC II (I-A/I-E) mAb, and UEA-1 lectin, and expression levels of Fgf21 mRNA were determined by RT-realtime PCR in the subpopulations of thymic cells from 4-week-old C57BL/6 mice. ( C ) Anti-CD45 and anti-EpCAM staining discriminated CD45 + , TEC (CD45 − EpCAM + ), and non-TEC stromal cell (CD45 − EpCAM − ) subpopulations. ( D ) Gated TECs were subdivided into cTEC hi (UEA-1 − MHC high ), cTEC lo (UEA-1 − MHC low ), mTEC hi (UEA-1 + MHC high ), and mTEC lo (UEA-1 + MHC low ) subpopulations. ( E ) Thymocytes were removed from embryonic thymi by 1.35 mM dGuo treatment for 6 days, and remaining stromal cells were stimulated with 1 μg/ml RANKL to induce the maturation of TECs. Fgf21 and Aire mRNA were increased along with TEC maturation. Graphs represent the mean ± SD; n ≧5 replicates per group from at least 2 independent experiments.

Article Snippet: Medium containing 500 ng/ml of recombinant human FGF21 (R&D Systems) was replaced at 3-day intervals.

Techniques: Expressing, Staining

Journal: Scientific Reports

Article Title: Fgf21 regulates T-cell development in the neonatal and juvenile thymus

doi: 10.1038/s41598-017-00349-8

Figure Lengend Snippet: CD4SP and CD8SP populations are decreased in Fgf21 KO mice. ( A and B ) Thymi and spleens were isolated from 4-week-old WT and Fgf21 KO mice, and their weights were measured. ( C ) Cell numbers of enzymatically digested suspensions from thymi and spleens were counted. ( D ) Thymocytes and splenocytes from WT and Fgf21 KO mice were stained with anti-CD4 and anti-CD8 mAb. Representative flow cytometry plots show CD4/CD8 analysis of thymocytes and splenocytes from 1-week-old WT and Fgf21 KO mice. ( E ) Charts show the percentage of DN, DP, CD8SP, and CD4SP cells from 1- and 4-week-old WT and Fgf21 KO mice. ( F ) Charts show the percent of CD4SP and CD8SP cells from 1- and 4-week-old WT and Fgf21 KO mice. ( G ) Thymocytes from 1-week-old WT and Fgf21 KO mice were defined by TCRβ and CD69 levels and subdivided into 5 subsets (T1; TCRβ − CD69 − , T2; TCRβ int CD69 − , T3; TCRβ int CD69 + , T4; TCRβ hi CD69 + , and T5; TCRβ hi CD69 − ). ( H ) Charts show the percentage of the 5 subsets from 1-week-old WT and Fgf21 KO mice. ( I ) Gated CD4SP and CD8SP cells from 1-week-old WT and Fgf21 KO mice were defined by CD62L and CD24 levels and subdivided into CD24 + CD62L − immature and CD24 − CD62L + mature SP cells. ( J ) Charts show the percentage of CD24 + CD62L − immature and CD24 − CD62L + mature SP subsets from 1-week-old WT and Fgf21 KO mice. All data shown are the mean ± SD from ≧6 mice per genotype from 2 independent experiments. *P < 0.05, *P < 0.01 versus WT mice.

Article Snippet: Medium containing 500 ng/ml of recombinant human FGF21 (R&D Systems) was replaced at 3-day intervals.

Techniques: Isolation, Staining, Flow Cytometry

Journal: Scientific Reports

Article Title: Fgf21 regulates T-cell development in the neonatal and juvenile thymus

doi: 10.1038/s41598-017-00349-8

Figure Lengend Snippet: Thymic stromal cells are not obviously altered in Fgf21 KO mice. ( A ) Enzymatically digested thymic cell suspensions were stained with anti-CD45 mAb and anti-EpCAM mAb, and the TEC (CD45 − EpCAM + ) number was counted from the thymi of E14.5, E15.5, E18.5, and 4-week-old WT and Fgf21 KO mice. ( B ) Immunohistochemistry of WT and Fgf21 KO thymi with anti-Keratin-5 antibody for a medullary TEC marker (left panels) and anti-Ly-51 antibody for a cortical TEC marker (right panels). ( C ) The ratio of Keratin-5-positive thymic medullary regions was measured from immunostained sections. ( D ) Gated TECs of 1-weeks old were subdivided into mTEC hi (Ly51 − MHC high ), mTEC lo (Ly51 − MHC low ), cTEC hi (Ly51 + MHC high ), and cTEC lo (Ly + MHC low ) subpopulations. Graphs represent the percentage of subpopulations. ( E ) The mean fluorescence intensity (MFI) of surface MHCII on the TEC was measured by flow cytometry with anti-MHCII antibody. ( F ) Gated CD45 − thymic stromal cells of 1-weeks old were stained with anti-CD140a and anti-CD31 antibodies. Graphs represent the cellularity of CD45 − CD140a + fibroblasts and CD45 − CD31a + endothelial cells in the thymus of WT and Fgf21 KO mice. ( G ) The expression levels of thymic factors were measured in the thymus of 1-week-old WT and Fgf21 KO mice. Graphs represent the mean ± SD; n ≧ 5 replicates per group from at least 3 independent experiments.

Article Snippet: Medium containing 500 ng/ml of recombinant human FGF21 (R&D Systems) was replaced at 3-day intervals.

Techniques: Staining, Immunohistochemistry, Marker, Fluorescence, Flow Cytometry, Expressing

Journal: Scientific Reports

Article Title: Fgf21 regulates T-cell development in the neonatal and juvenile thymus

doi: 10.1038/s41598-017-00349-8

Figure Lengend Snippet: FGF21 treatment alters T-cell development in FTOC. WT and Fgf21 KO foetal thymi were cultured with or without recombinant human FGF21 protein (500 ng/ml) for 14 days. ( A ) Total cell numbers of thymocytes in FTOCs are presented. Flow cytometry analysis of CD4/8 ( B ) and TCRβ/CD69 ( C ) distribution and thymocyte subset number ( D and E ). ( F ) Representative flow cytometry data for each FTOC. ( G ) Numbers of mTEC (CD45 − EpCAM + UEA-1 + ), cTEC (CD45 − EpCAM + UEA-1 − ), and non-TEC stromal cells (CD45 − EpCAM − ) are shown. Data represents the mean ± SD of two separate experiments. *,# P < 0.05, **,## P < 0.01, and *** P < 0.001 versus WT without rhFGF21, $ P < 0.05, and $$ P < 0.01 versus Fgf21 KO without rhFGF21.

Article Snippet: Medium containing 500 ng/ml of recombinant human FGF21 (R&D Systems) was replaced at 3-day intervals.

Techniques: Cell Culture, Recombinant, Flow Cytometry

Journal: Scientific Reports

Article Title: Fgf21 regulates T-cell development in the neonatal and juvenile thymus

doi: 10.1038/s41598-017-00349-8

Figure Lengend Snippet: FGF21 treatment results in increased apoptosis of immature thymocytes. FTOC was performed in the presence or absence of recombinant human FGF21 (500 ng/ml) for 14 days. Annexin V staining was performed to detect apoptotic T cells by flow cytometry. ( A ) Representative histograms show the percentage of Annexin V + cells in gated thymocyte subsets. ( B ) Graph showing the analysis of the percentage of Annexin V + cells in thymocytes from FTOCs. All data shown are the mean ± SD. **P < 0.01 versus control.

Article Snippet: Medium containing 500 ng/ml of recombinant human FGF21 (R&D Systems) was replaced at 3-day intervals.

Techniques: Recombinant, Staining, Flow Cytometry, Control

Journal: Scientific Reports

Article Title: Fgf21 regulates T-cell development in the neonatal and juvenile thymus

doi: 10.1038/s41598-017-00349-8

Figure Lengend Snippet: Fgf21 induced by protein-free diet does not affect thymic change by protein malnutrition. ( A ) Serum Fgf21 levels of 4-week-old C57BL/6 mice fed for 1 week with normal chow diet (NC) or protein-free diet (PF) were determined with ELISA assays. Data shown are the mean ± SD from ≧6 mice. ** P < 0.001 versus NC. ( B ) Relative expression levels of Fgf21 mRNA in the liver, thymus, subcutaneous white adipose tissue (sWAT), and skeletal muscle from mice fed with NC or PF. Data shown are the mean ± SD from ≧6 mice. ** P < 0.01, ** P < 0.001 versus NC. ( C–F ) Body weight ( C ), blood glucose ( D ), thymic weight ( E ), and thymic cell number ( F ) of WT and Fgf21 KO mice fed with NC or PF. Data shown are the mean ± SD from ≧5 mice. * P < 0.05, # P < 0.05 versus WT and Fgf21 KO mice fed with NC, respectively. $ P < 0.05 versus WT mice fed with PF. ( G and H ) Thymocytes from WT and Fgf21 KO mice fed with NC or PF were stained with anti-CD4 and anti-CD8 mAb ( G ), or anti-TCRβ and anti-CD69 mAb ( H ). All data shown are the mean ± SD from ≧5 mice. * P < 0.05, # P < 0.05 versus WT and Fgf21 KO mice fed with NC, respectively. $ P < 0.05 versus WT mice fed with PF.

Article Snippet: Medium containing 500 ng/ml of recombinant human FGF21 (R&D Systems) was replaced at 3-day intervals.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing, Staining

Journal: Molecular metabolism

Article Title: Fibroblast growth factor 21 has no direct role in regulating fertility in female mice.

doi: 10.1016/j.molmet.2016.05.010

Figure Lengend Snippet: Figure 3: High fat diet reverses the infertility in FGF21-Tg mice. A) Body weight of WT and FGF21-Tg mice fed either with chow or HFD (n ¼ 4e6). B) Body composition as represented as absolute fat and lean mass (n ¼ 6e7). C) Assessment of estrus cycle in FGF21-Tg mice fed either chow or HFD. (1: leukocytes [diestrus/metestrus], 2: cornified cells [estrus], 3: nucleated cells [proestrus]) D) Representative vaginal cytology from FGF21 Tg mice fed with HFD as (i) diestrus, (ii) proestrus, (iii) estrus, (iv) metestrus. Scale: 200 mm (ieiv). (n ¼ 8). E) Ovarian histology from FGF21-Tg mice; (i) chow, (ii) HFD, (iiieiv) HFD fed pregnant females. Corpus lutea (CL) are encircled with dotted lines. Scale: 1000 mm. Statistical significance was evaluated by Student’s t test; *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Mouse FGF21 and human FGF21 proteins were measured by ELISA (R & D Systems, MN).

Techniques:

Journal: Molecular metabolism

Article Title: Fibroblast growth factor 21 has no direct role in regulating fertility in female mice.

doi: 10.1016/j.molmet.2016.05.010

Figure Lengend Snippet: Figure 5: Infertility of FGF21-Tg mice is independent of FGF21. Serum levels of total FGF21 (A) and active FGF21 (B) in FGF21-Tg mice fed with chow or HFD (n ¼ 9e10). AVP and Kiss-1 gene expression in SCN (C) and AVPV (D), respectively of WT and FGF21-Tg mice fed chow or HFD (n ¼ 8e10). All mice were in the diestrus phase. Statistical significance was evaluated by Student’s t test; *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Mouse FGF21 and human FGF21 proteins were measured by ELISA (R & D Systems, MN).

Techniques: Gene Expression

Journal: Toxicology

Article Title: Alcoholic fatty liver is enhanced in CYP2A5 knockout mice: the role of the PPARα-FGF21 axis

doi: 10.1016/j.tox.2017.01.016

Figure Lengend Snippet: Chronic ethanol induces FGF21 in Cyp2a5+/+ mice but does not in Cyp2a5−/− mice. (A) Serum FGF21 (N=5). (B) Liver expression of FGF21 and PPARα by Western blotting analysis. (C) Quantitation of Western blots (N=4). *P<0.05, compared to Cyp2a5+/+ Control Group. Cont, Control; EtOH, Ethanol.

Article Snippet: Some mice were daily injected mouse recombinant FGF21 (rFGF21, purchased from Creative Biomart, Shirley, NY, USA) subcutaneously at 0.25 mg/kg body weight.

Techniques: Expressing, Western Blot, Quantitation Assay, Control

Journal: Toxicology

Article Title: Alcoholic fatty liver is enhanced in CYP2A5 knockout mice: the role of the PPARα-FGF21 axis

doi: 10.1016/j.tox.2017.01.016

Figure Lengend Snippet: Chronic ethanol induces FGF21 in Nrf2+/+ mice more than in Nrf2−/− mice. (A) Western blots. (B) Quantitation of Western blots (N=3). *P<0.05, compared to Nrf2+/+ Control Group; # P<0.05, compared to Nrf2−/− Control Group; & P<0.05, compared with Nrf2+/+ Ethanol group. Cont, Control; EtOH, Ethanol.

Article Snippet: Some mice were daily injected mouse recombinant FGF21 (rFGF21, purchased from Creative Biomart, Shirley, NY, USA) subcutaneously at 0.25 mg/kg body weight.

Techniques: Western Blot, Quantitation Assay, Control

Journal: Toxicology

Article Title: Alcoholic fatty liver is enhanced in CYP2A5 knockout mice: the role of the PPARα-FGF21 axis

doi: 10.1016/j.tox.2017.01.016

Figure Lengend Snippet: Recombinant FGF21 blunts ethanol-induced hypertriglyceridemia (HTG) in Pparα−/− mice. (A) Ethanol induction of FGF21 was blunted in Pparα−/− mice (N=5). (B) rFGF21 blunted ethanol-induced elevation of liver TG in WT mice but not in Pparα−/− mice (N=5). (C) rFGF21 blunted ethanol-induced HTG in Pparα−/− mice (N=5). (D) Ethanol induction of CYP2E1 and CYP2A5 was not affected in both Pparα+/+ and Pparα−/− mice (N=5). *P<0.05, compared to Control Group. # P<0.05, compared to WT Ethanol Group. $ P<0.05, compared to WT Control Group. ^ P<0.05, compared with PPARα KO Ethanol group. E+F, ethanol plus rFGF21; WT, wild-type; PPARα KO, Pparα−/−.

Article Snippet: Some mice were daily injected mouse recombinant FGF21 (rFGF21, purchased from Creative Biomart, Shirley, NY, USA) subcutaneously at 0.25 mg/kg body weight.

Techniques: Recombinant, Control

Journal: Toxicology

Article Title: Alcoholic fatty liver is enhanced in CYP2A5 knockout mice: the role of the PPARα-FGF21 axis

doi: 10.1016/j.tox.2017.01.016

Figure Lengend Snippet: Alcoholic fatty liver is enhanced in Fgf21alb-cre mice but is not in Fr1alb-cre mice. (A) Ethanol induction of FGF21 was blunted in Fgf21alb-cre mice but was not in Fr1alb-cre mice (N=5). (B) H&E staining in liver sections shows that alcoholic fatty liver was enhanced in Fgf21alb-cre mice but not in Fr1alb-cre mice. (C) Steatosis scores (N=5). (D) Liver TG contents (N=5). *P<0.05, compared to Fr1fl/fl Control Group. # P<0.05, compared to Fgf21fl/fl Ethanol Group. ^ P<0.05, compared with Fgf21fl/fl control group. $ P<0.05, compared to Fr1alb-cre Control Group. @ P<0.05, compared to Fr1alb-cre Control Group. Cont, Control; EtOH, Ethanol.

Article Snippet: Some mice were daily injected mouse recombinant FGF21 (rFGF21, purchased from Creative Biomart, Shirley, NY, USA) subcutaneously at 0.25 mg/kg body weight.

Techniques: Staining, Control

Journal: Toxicology

Article Title: Alcoholic fatty liver is enhanced in CYP2A5 knockout mice: the role of the PPARα-FGF21 axis

doi: 10.1016/j.tox.2017.01.016

Figure Lengend Snippet: Alcoholic fatty liver is more pronounced in Pparα−/−/Cyp2a5−/− mice than in Pparα+/+/Cyp2a5−/− mice. (A) Ethanol induction of FGF21 was blunted in Pparα−/−/Cyp2a5−/− mice (N=4). (B) H&E staining in liver sections shows that alcoholic fatty liver was more pronounced in Pparα−/−/Cyp2a5−/− mice than in Pparα+/+/Cyp2a5−/− mice. (C) Steatosis scores (N=5). (D) Liver TG contents (N=5). *P<0.05, compared to Pparα+/+/Cyp2a5−/− Control Group. # P<0.05, compared to Pparα−/−/Cyp2a5−/− Control Group. & P<0.05, compared to Pparα+/+/Cyp2a5−/− Ethanol Group. Cont, Control; EtOH, Ethanol.

Article Snippet: Some mice were daily injected mouse recombinant FGF21 (rFGF21, purchased from Creative Biomart, Shirley, NY, USA) subcutaneously at 0.25 mg/kg body weight.

Techniques: Staining, Control

Journal: The Journal of Clinical Investigation

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

doi: 10.1172/JCI94337

Figure Lengend Snippet: (A) Concentration-dependence of BrdU incorporation in A2B5+ OPCs cultured with adult mice serum (n = 6). (B) BrdU incorporation in A2B5+ OPCs 1 day after stimulation with adult mouse serum heated or pretreated with the indicated reagents (n = 6). (C) BrdU incorporation in OPCs after serum stimulation with PD173074 (10 nM), an inhibitor of FGFR (n = 4). (D) BrdU incorporation in OPCs after serum stimulation with NF449 (10 μM), an inhibitor of FGFR3 (n = 4). (E) BrdU incorporation in mouse OPCs with FGFR and β-klotho knockdown after serum stimulation (n = 7). (F) BrdU incorporation in OPCs after stimulation with recombinant FGF15, FGF21, and FGF23 (n = 4). (G) BrdU incorporation in OPCs after serum stimulation with neutralizing antibody against FGF21 (n = 5), determined by Student’s t test or by ANOVA with Tukey’s post hoc test or Dunnett’s test. Error bars represent SEM. *P < 0.05, **P < 0.01.

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: Concentration Assay, BrdU Incorporation Assay, Cell Culture, Knockdown, Recombinant

Journal: The Journal of Clinical Investigation

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

doi: 10.1172/JCI94337

Figure Lengend Snippet: (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: The Journal of Clinical Investigation

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

doi: 10.1172/JCI94337

Figure Lengend Snippet: (A) Quantitation of FGF21 protein in the spinal cord 1 day and 3 days after LPC injection (n = 5 for control, 5 for d1, 4 for d3). (B) Representative images of spinal cord sections double-labeled for PDGFRα and Ki67. Sections were obtained from FGF21-KO mice and control littermates 7 days after LPC injection. Graph shows quantitations as indicated in the images (n = 5). Arrows indicate Ki67+ cells colabeled with PDGFRα; arrowheads indicate Ki67+ cells not labeled with PDGFRα. (C) Representative images of spinal cord sections labeled for MBP. Sections were obtained from mouse spinal cord 14 days after LPC injection. Graph shows quantitations as indicated in the images (n = 5). (D) Representative immunoelectron microscopy images of myelin in the spinal cord. Sections were obtained from FGF21-KO mice and control littermates 14 days after LPC injection. Graphs show quantitations of g-ratio indicated in the images (n = 3). (E) Motor function was assessed by ladder-walk test (n = 11 for control littermates, 9 for FGF21-KO mice). (F) Representative images of spinal cord sections labeled for MBP. Graph shows quantitations as indicated in the images (n = 5 for control littermates + vehicle, 5 for FGF21-KO mice + vehicle, 4 for FGF21-KO mice + FGF21). (G) Representative images of spinal cord sections labeled for MBP. Graph shows quantitations as indicated in the images (n = 4); **P < 0.01 as determined by Student’s t test or by ANOVA with Tukey’s post hoc test or Dunnett’s test. *P < 0.05, **P < 0.01. Error bars represent SEM. Scale bars: 50 μm (B); 200 μm (C, F, and G); 2 μm (D).

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: Quantitation Assay, Injection, Control, Labeling, Immuno-Electron Microscopy

Journal: The Journal of Clinical Investigation

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

doi: 10.1172/JCI94337

Figure Lengend Snippet: (A) Representative images of β-klotho expression in the mouse spinal cord 3 days after LPC injection. (B) Representative images of spinal cord sections double-labeled for PDGFRα and Ki67. Graph shows quantitations as indicated in images (n = 3 for control, 3 for CKO); *P < 0.05. Arrows indicate Ki67+ cells colabeled with PDGFRα. (C) Representative images of spinal cord sections labeled for MBP. Graph shows quantitations as indicated in the images (n = 4 each); **P < 0.01. (D) Representative images of brain sections double-labeled for PDGFRα and Ki67 in the mouse cortex, 7 days after traumatic brain injury. FGF21 was administered i.c.v. 24 hours after LPC injection (n = 4); *P < 0.05. Arrows indicate Ki67+ cells colabeled with PDGFRα; arrowheads indicate Ki67+ cells not labeled with PDGFRα. (E) Representative images of brain sections labeled for MBP in the mouse cortex, 14 days after traumatic brain injury. Graph shows quantitations as indicated in the images (n = 6 for vehicle, n = 4 for FGF21); *P < 0.05. (F) Representative immunoelectron microscopy images of myelin in the mouse cortex, 14 days after traumatic brain injury. Graphs show quantitations of g-ratio indicated in the images (n = 4); **P < 0.01 as determined by Student’s t test. Error bars represent SEM. Scale bars: 20 μm (A); 50 μm (B and D); 200 μm (C and E); 2 μm (F).

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, Injection, Labeling, Control, Immuno-Electron Microscopy

Journal: The Journal of Clinical Investigation

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

doi: 10.1172/JCI94337

Figure Lengend Snippet: (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: PLoS ONE

Article Title: Novel Insights into the Cardio-Protective Effects of FGF21 in Lean and Obese Rat Hearts

doi: 10.1371/journal.pone.0087102

Figure Lengend Snippet: FGF21 mRNA [ Fig. 2A1 ] and protein expression [ Fig. 2A2 ]; βKlotho mRNA [ Fig. 2A3 ] and protein expression [ Fig. 2A4 ] in isolated rat cardiomyocytes [A] and rat heart [B]. Fig . 1B:Immunocyto/histochemistry and confocal analysis of FGF21 protein in isolated adult rat cardiomyocytes [ B1 ] and rat heart [ B2 ]. MW.- Molecular Weight Marker for PCR products; BP.- Base Pairs; kDa.-kilo daltons. Fig. 2C -( a ): Trace recordings of left ventricular developed pressure [LVDP (mmHg)] and left ventricular contractility (dp/dt) in control (saline treated) groups; and Fig. 2C -( b ):[LVDP-mmHg] and dp/dt ratio in FGF21 treated groups - following 30 mins of global ischemia and 120 mins reperfusion. Fig. 2D: Rate pressure product [RPP (mmHg/min)] during global ischemia and reperfusion with or without FGF21 treatment [** P <0.01 vs. control]. Fig. 2E : Graphical representation of infarct area (%) in rat hearts treated with or without FGF21. Data shown are means ± SEM (n = 6, in triplicates). ***P<0.001; **P<0.01 vs. control.

Article Snippet: The drugs used for the study were: rat recombinant FGF21 [R&D systems, UK], wortmannin (PI3K inhibitor), U1026 (MAPK inhibitor), Compound C (AMPK inhibitor), TO-901317 [ROR-α (Retinoic acid receptor-related receptor alpha) inhibitor]; TO-901317 (was initially intended as a therapeutic Liver X receptor (LXR) agonist in dementia , however recently, work by Uebanso T et al, have demonstrated its role in the regulation of FGF21 expression ) and collagenase (type I+protease type XIV).

Techniques: Expressing, Isolation, Molecular Weight, Marker, Control, Saline

Journal: PLoS ONE

Article Title: Novel Insights into the Cardio-Protective Effects of FGF21 in Lean and Obese Rat Hearts

doi: 10.1371/journal.pone.0087102

Figure Lengend Snippet: Fig. 3A1: Cardiomyocytes treated with FGF21 (100 nM) for 5–30 minutes. Phosphorylated ERK 1/2 [Fig. 3A1]; Akt [Fig. 3A2] and AMPK [Fig. 3A3] proteins are represented in relation to total proteins, expressed as fold increase over basal. ***P<0.001, **P<0.01, *P<0.05 vs. basal, n = 6 per group. Fig. 3B1: RPP with ischemia/reperfusion and FGF21 treatment following pre-incubation with inhibitors (TO-901317; wortmanin; Compound C or U0126) a P<0.05, b P<0.01 vs. FGF21 only treatment, n = 6 per group. Fig. 3B2: Infarcted area (%) following ischemia/reperfusion and FGF21 treatment following pre-incubation with inhibitors (TO-901317; wortmanin; Compound C or U0126) **P<0.01, *P<0.05 vs. FGF21 treated, n = 6 per group.

Article Snippet: The drugs used for the study were: rat recombinant FGF21 [R&D systems, UK], wortmannin (PI3K inhibitor), U1026 (MAPK inhibitor), Compound C (AMPK inhibitor), TO-901317 [ROR-α (Retinoic acid receptor-related receptor alpha) inhibitor]; TO-901317 (was initially intended as a therapeutic Liver X receptor (LXR) agonist in dementia , however recently, work by Uebanso T et al, have demonstrated its role in the regulation of FGF21 expression ) and collagenase (type I+protease type XIV).

Techniques: Incubation

Journal: PLoS ONE

Article Title: Novel Insights into the Cardio-Protective Effects of FGF21 in Lean and Obese Rat Hearts

doi: 10.1371/journal.pone.0087102

Figure Lengend Snippet: Fig. 4A :FGF21 mRNA expression levels in cardiomyocytes following FGF21 (100 nM) treatment with or without pathway inhibitors [(U0126; wort (wortmanin); Comp C (Compound C) or TO (TO-901317)] (normalised to GAPDH and expressed as fold changes over basal). Fig. 4B : Graphical analysis of FGF21 protein levels following FGF21 treatment. Fig. 4C : Graphical representation of FGF21 ELISA measurements in the conditioned media following FGF21 treatment. Fig. 4D : Graphical representation of FGF21 ELISA measurements of rat heart Langendorff exudates following global ischemia for 5–30 mins and 120 mins of reperfusion; with or without prior FGF21 (100 nM) infusion. Data shown are means ± SEM of triplicates. The values represented are relative to basal. *** P <0.001; ** P <0.01; * P <0.05 vs. FGF21 only treated, a P <0.001 vs. basal, n = 6 per group.

Article Snippet: The drugs used for the study were: rat recombinant FGF21 [R&D systems, UK], wortmannin (PI3K inhibitor), U1026 (MAPK inhibitor), Compound C (AMPK inhibitor), TO-901317 [ROR-α (Retinoic acid receptor-related receptor alpha) inhibitor]; TO-901317 (was initially intended as a therapeutic Liver X receptor (LXR) agonist in dementia , however recently, work by Uebanso T et al, have demonstrated its role in the regulation of FGF21 expression ) and collagenase (type I+protease type XIV).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: Novel Insights into the Cardio-Protective Effects of FGF21 in Lean and Obese Rat Hearts

doi: 10.1371/journal.pone.0087102

Figure Lengend Snippet: Using rat heart Langendorff model and inducing global ischemia for 30[ Fig. 5A1 ], protein [ Fig. 5A2 ], and secretion of FGF21 in Langendorff coronary exudates [ Fig. 5A3 ] were measured. Similarly, changes in FGFR1 mRNA expressions [ Fig. 5A4 ] were measured. Data shown are means ± SEM of triplicates. The values represented are relative to basal. *** P <0.001; ** P <0.01vs. control. n = 6 per group. FGF21 mRNA [ Fig. 5B1 ], protein [ Fig. 5B2 ], FGF21 secretion (in Langendorff coronary exudates) [ Fig. 5B3 ] and FGFR1 mRNA [ Fig. 5B4 ] expressions were measured in obese and lean rat hearts. Graphical representation of a key signalling component of FGF21/FGFR1; βKlotho protein level in obese and lean hearts [ P <0.05 vs.lean; Figure 5C 1 ]. Graphical representation of ERK 1/2 [ Fig. 5C2 ], Akt [ Fig. 5C3 ] and AMPK [ Fig. 5C4 ] phosphorylation levels in lean and obese hearts with FGF21 (100 nM) pre-infusion. Data shown are means ± SEM of triplicates. The values represented are relative to basal. * P <0.05, ** P <0.01 vs. lean control, NS -non-significant; n = 6 per group.

Article Snippet: The drugs used for the study were: rat recombinant FGF21 [R&D systems, UK], wortmannin (PI3K inhibitor), U1026 (MAPK inhibitor), Compound C (AMPK inhibitor), TO-901317 [ROR-α (Retinoic acid receptor-related receptor alpha) inhibitor]; TO-901317 (was initially intended as a therapeutic Liver X receptor (LXR) agonist in dementia , however recently, work by Uebanso T et al, have demonstrated its role in the regulation of FGF21 expression ) and collagenase (type I+protease type XIV).

Techniques: Control, Phospho-proteomics

Journal: PLoS ONE

Article Title: Novel Insights into the Cardio-Protective Effects of FGF21 in Lean and Obese Rat Hearts

doi: 10.1371/journal.pone.0087102

Figure Lengend Snippet: LVDP (mmHg) and left ventricular contractility (dp/dt) in obese control (saline treated) groups [ Fig. 6A1 ] and lean control (saline treated) groups [ Fig. 6A2 ]; following global ischemia and reperfusion. Graphical representation of RPP (mmHg/min)] during global ischemia and reperfusion in obese and lean control rat hearts [ Fig. 6B ]. Graphical representation of the infarcted area (%) in obese and lean control rat hearts following global ischemia and reperfusion [ Fig. 6C ]. Graphical representation of FGF21 levels in obese and lean rat heart coronary effluents following global ischemia and reperfusion [ Fig. 6D ]. Data shown are means ± SEM of triplicates. The values represented are relative to basal. *** P <0.001 vs. t [0] time point, n = 6 per group.

Article Snippet: The drugs used for the study were: rat recombinant FGF21 [R&D systems, UK], wortmannin (PI3K inhibitor), U1026 (MAPK inhibitor), Compound C (AMPK inhibitor), TO-901317 [ROR-α (Retinoic acid receptor-related receptor alpha) inhibitor]; TO-901317 (was initially intended as a therapeutic Liver X receptor (LXR) agonist in dementia , however recently, work by Uebanso T et al, have demonstrated its role in the regulation of FGF21 expression ) and collagenase (type I+protease type XIV).

Techniques: Control, Saline

Journal: PLoS ONE

Article Title: Novel Insights into the Cardio-Protective Effects of FGF21 in Lean and Obese Rat Hearts

doi: 10.1371/journal.pone.0087102

Figure Lengend Snippet: LVDP (mmHg) and left ventricular contractility (dp/dt) in obese control (saline treated) groups [ Fig. 7A1 ], obese (FGF21 treated) groups [ Fig. 7A2 ] and lean (FGF21 treated) groups [ Fig. 7A3 ]; following global ischemia and reperfusion. Graphical representation of the infarcted area (%) in obese control, obese and lean FGF21 (100 nM) treated rat hearts following global ischemia and reperfusion [ Fig. 7B ]. Graphical representation of FGF21 levels in obese and lean FGF21 (100nM) pre-treated rat heart coronary effluents following global ischemia and reperfusion [ Fig 6C ] . Data shown are means ± SEM of triplicates. The values represented are relative to basal. *** P < 0.001 vs. t [0] time point, # P < 0.05 vs. lean at t(5); n = 6 per group.

Article Snippet: The drugs used for the study were: rat recombinant FGF21 [R&D systems, UK], wortmannin (PI3K inhibitor), U1026 (MAPK inhibitor), Compound C (AMPK inhibitor), TO-901317 [ROR-α (Retinoic acid receptor-related receptor alpha) inhibitor]; TO-901317 (was initially intended as a therapeutic Liver X receptor (LXR) agonist in dementia , however recently, work by Uebanso T et al, have demonstrated its role in the regulation of FGF21 expression ) and collagenase (type I+protease type XIV).

Techniques: Control, Saline

Journal: Journal of Diabetes

Article Title: Renoprotective effects of brown adipose tissue activation in diabetic mice

doi: 10.1111/1753-0407.12938

Figure Lengend Snippet: Effects of CL316,243 on serum adipokines (adiponectin and fibroblast growth factor [FGF] 21) and renal expression of Fgf21 components in diabetic mice. A, Serum adiponectin and B, serum Fgf21 concentrations; C, Fgf21 protein and D, relative expression of β‐klotho ( Klb ) and FGF receptor 1c ( Fgfr1c ) mRNA in renal cortical tissues. Data are the mean ± SEM (n = 6 per group). * P < 0.05, ** P < 0.01, *** P < 0.001 compared with control (non‐diabetic) mice; † P < 0.05, †† P < 0.01 compared with the diabetic control (DM‐Con) group. DM + CL, HFD combined with STZ‐induced diabetic mice treated with CL316,243

Article Snippet: Serum creatinine (Cr) was measured using a commercially available kit (Nanjing Jiancheng Bioengineering Institute, Nanjing, China), and FGF21 protein in the renal cortex was determined by ELISA (Elabscience Biotechnology, Wuhan, China).

Techniques: Expressing, Control, Mouse Assay

Journal: Physiological Reports

Article Title: Identification of FGF21 ‐inducing rare sugars that reduces sugar appetite in male BL /6 mice

doi: 10.14814/phy2.70618

Figure Lengend Snippet: Identification of FGF21‐inducing rare sugars and their effects on blood glucose. (a–d) FGF21 level of mouse primary hepatocytes treated for 24 h with vehicle (distilled water‐negative control) or 25 mM D‐glucose, D‐fructose (positive control), or rare sugars (46 samples). n = 3 mice. (e) Plasma FGF21 levels over 24 h after gastric gavage of vehicle (distilled water), D‐glucose, D‐tagatose, D‐allulose, or D‐sorbitol at 5 g/kg in nine‐week‐old male WT mice with ad libitum access to normal chow (NC) diet. n = 4 mice per group. (f) Area under the curve (AUC) over 24 h of data in (e). n = 4 mice per group. (g) Blood glucose levels of the mice receiving the same dose in (e) of either vehicle (distilled water), D‐glucose, D‐tagatose, D‐allulose, or D‐sorbitol after 16 h fasting. n = 4 mice per group. Statistical analyses were done by one‐way ANOVA for (a–d, f) and repeated measures ANOVA for (e, g), followed by Tukey's HSD test. ** p < 0.01; **** p < 0.0001.

Article Snippet: Mouse FGF21 protein levels in each plasma sample were determined using a Mouse/Rat FGF‐21 Quantikine ELISA Kit (R&D Systems) according to the manufacturer's instructions.

Techniques: Negative Control, Positive Control, Clinical Proteomics

Journal: Physiological Reports

Article Title: Identification of FGF21 ‐inducing rare sugars that reduces sugar appetite in male BL /6 mice

doi: 10.14814/phy2.70618

Figure Lengend Snippet: FGF21‐inducing rare sugars activated oxytocin neurons of paraventricular nucleus of hypothalamus in BL/6 mice. (a–e) Immunostaining of mice brain after gastric gavage with either (a) vehicle (distilled water) or 5 g/kg body weight of D‐glucose (b), D‐tagatose (c), D‐allulose (d), or D‐sorbitol (e). Blue arrow ( ) indicates c‐Fos‐positive oxytocin neurons, shown by dark brown spot on the light brown neurons ( ). Magenta arrow ( ) indicates c‐Fos‐negative oxytocin neurons, shown by light brown anti‐oxytocin staining ( ). n = 4 mice per group. Scale bar represents 100 μ m. (f) Percentage of activated oxytocin neurons (c‐Fos positive oxytocin neurons) from the total of PVH oxytocin neurons in the mice receiving either vehicle (distilled water), D‐glucose, or FGF21‐inducing rare sugars in (a–e). Data are presented as box‐whiskers in (f), where the middle line represents median, bottom/ top edges represent 25th/75th percentile of the data, and whiskers represent maximum and minimum value of the data. Statistical analyses were done by one‐way ANOVA, followed by Dunnett's test.

Article Snippet: Mouse FGF21 protein levels in each plasma sample were determined using a Mouse/Rat FGF‐21 Quantikine ELISA Kit (R&D Systems) according to the manufacturer's instructions.

Techniques: Immunostaining, Staining

Journal: Physiological Reports

Article Title: Identification of FGF21 ‐inducing rare sugars that reduces sugar appetite in male BL /6 mice

doi: 10.14814/phy2.70618

Figure Lengend Snippet: Intragastric administration of FGF21‐inducing rare sugars reduced sucrose preference in BL/6 mice. (a–c) 50 mM sucrose intake in mice receiving 5 g/kg body weight of D‐allulose (a), D‐tagatose (b), or D‐sorbitol (c). (d–f) sucrose preference of the same mice in (a–c). n = 10 mice per group for (a, d) and n = 5 mice per group for (b, c, e, f). Data are presented as box‐whiskers, where the middle line represents the median, bottom/top edges represent the 25th/75th percentile of the data, and whiskers represent the maximum and minimum value of the data. Statistical analyses were done by Student's paired t ‐test between “pre” and “post” of each treatment.

Article Snippet: Mouse FGF21 protein levels in each plasma sample were determined using a Mouse/Rat FGF‐21 Quantikine ELISA Kit (R&D Systems) according to the manufacturer's instructions.

Techniques:

Journal: Physiological Reports

Article Title: Identification of FGF21 ‐inducing rare sugars that reduces sugar appetite in male BL /6 mice

doi: 10.14814/phy2.70618

Figure Lengend Snippet: Mixing FGF21‐inducing rare sugars into sucrose solution reduced solution intake and preference in BL/6 mice. (a–c) Intake of 100 mM (final concentration) sucrose solution that was mixed with either vehicle (distilled water), 600 mM (final concentration) of D‐glucose, or 600 mM (final concentration) of D‐allulose (a), D‐tagatose (b), or D‐sorbitol (c). (d–f) Sucrose preference of the same mice in (a–c). n = 6 mice per group. Data are presented as box‐whiskers, where the middle line represents the median, bottom/top edges represent the 25th/75th percentile of the data, and whiskers represent the maximum and minimum value of the data. Statistical analyses were done by repeated measures ANOVA, followed by Tukey's HSD test.

Article Snippet: Mouse FGF21 protein levels in each plasma sample were determined using a Mouse/Rat FGF‐21 Quantikine ELISA Kit (R&D Systems) according to the manufacturer's instructions.

Techniques: Concentration Assay