Journal: International Journal of Molecular Sciences

Article Title: Associations Between Klotho/FGF-Related Protein Expression in Peripheral Blood Mononuclear Cells, Inflammation, and Muscle Function in Middle-Aged Adults with Obesity: A Pilot Study

doi: 10.3390/ijms27041983

Figure Lengend Snippet: Densitometric quantification and representative Western blot of the Klotho/FGF System Components. Patients with obesity group (PG) n = 30 and control group (CG) n = 15, were analyzed for the Klotho/FGF system (( A ), klotho, KL; ( B ), beta-klotho, KLB; ( C ), fibroblast growth factor 19, FGF19; ( D ), fibroblast growth factor 21, FGF21; and ( E ), fibroblast growth factor 23, FGF23). Mean values ± SEM are represented. Significance levels: * p < 0.05.

Article Snippet: Antibodies against FGF-23 (Cat# MAB2629, RRID: AB_2247024) and KLB (Cat# AF5889) were purchased from R&D Systems (Bio-Techne, Minneapolis, MN, USA).

Techniques: Western Blot, Control

Journal: EBioMedicine

Article Title: A novel GLP-1 and FGF21 dual agonist has therapeutic potential for diabetes and non-alcoholic steatohepatitis

doi: 10.1016/j.ebiom.2020.103202

Figure Lengend Snippet: Summary of the kinetic and affinity constants (KD) for the interaction between FGF21 mutants and β-Klotho using streptavidin (SA) biosensors.

Article Snippet: Rabbit Anti-Human β-Klotho monoclonal antibody (R&D Systems, Cat #MAB58891) or isotype control antibody (R&D Systems, Cat # AB-105-C), and anti-Rabbit IgG (PE) secondary antibody (R&D Systems, Cat # F0110) were used for β-Klotho expression.

Techniques:

Journal: EBioMedicine

Article Title: A novel GLP-1 and FGF21 dual agonist has therapeutic potential for diabetes and non-alcoholic steatohepatitis

doi: 10.1016/j.ebiom.2020.103202

Figure Lengend Snippet: Dual target GLP-1-Fc-FGF21 D1 functions through both GLP-1R and FGF21R/β-Klotho pathways. GLP-1-Fc-FGF21 D1, Fc-FGF21 S1 and Dulaglutide were investigated in cell based assays for their trans-activating activity. (a) ERK phosphorylation was measured in HEK293 cells expressing β-Klotho treated with native FGF21, GLP-1-Fc-FGF21 D1, Fc-FGF21 S1 or Dulaglutide. (b) cAMP secreted from HEK93 cells expressing GLP-1R treated with GLP-1-Fc-FGF21 D1, Fc-FGF21 S1 or dulaglutide. (c) HEK93 cells expressing GLP-1R and β-Klotho were treated with GLP-1-Fc-FGF21 D1, Fc-FGF21 S1, Dulaglutide or a 1:1 mixture of Fc-FGF21 S1 and Dulaglutide and the secreted cAMP were measured. (d) ERK phosphorylation was measured in HEK293 cells expressing GLP-1R and β-Klotho treated with GLP-1-Fc-FGF21 D1, Fc-FGF21 S1, Dulaglutide or a 1:1 mixture of Fc-FGF21 S1 and Dulaglutide. Data are showed as mean +/- standard errors of means, n =3.

Article Snippet: Rabbit Anti-Human β-Klotho monoclonal antibody (R&D Systems, Cat #MAB58891) or isotype control antibody (R&D Systems, Cat # AB-105-C), and anti-Rabbit IgG (PE) secondary antibody (R&D Systems, Cat # F0110) were used for β-Klotho expression.

Techniques: Activity Assay, Phospho-proteomics, Expressing

Journal: Gut and Liver

Article Title: Expression of Fibroblast Growth Factor 21 and β-Klotho Regulates Hepatic Fibrosis through the Nuclear Factor-κB and c-Jun N-Terminal Kinase Pathways

doi: 10.5009/gnl17443

Figure Lengend Snippet: Analysis of FGF19, FGF21, and β-Klotho expression and inflammatory markers in liver tissue. Expression levels of mRNA for FGF19 (A), FGF21 (B), β-Klotho (C), IL-1β (D), IL-6 (E), and TNF-α (F) were determined by real-time polymerase chain reaction. G1 (F0–F1, n=10), G2 (F2–F3, n=10), G3 (F4A–F4C, n=15). FGF, fibroblast growth factor; IL, interleukin; TNF, tumor necrosis factor. *p<0.05, † p<0.01.

Article Snippet: The lysates were subjected to Western blotting analysis under reducing conditions using previously validated human β-Klotho antibodies (R & D Systems), anti-phospho-Erk1/2, total Erk1/2, phospho-AKT, total AKT, phospho-JNK, and total JNK antibodies (Cell Signaling Technology, Danvers, MA, USA), phospho-IκBα, total IκBα, proliferating cell nuclear antigen (PCNA), and GAPDH antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and FGF21 antibody (Boster Biological Technology, Pleasanton, CA, USA).

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: Gut and Liver

Article Title: Expression of Fibroblast Growth Factor 21 and β-Klotho Regulates Hepatic Fibrosis through the Nuclear Factor-κB and c-Jun N-Terminal Kinase Pathways

doi: 10.5009/gnl17443

Figure Lengend Snippet: Correlation between the Fibrosis Stage and mRNA Expression of FGF19/21, β-Klotho, and Inflammatory Markers

Article Snippet: The lysates were subjected to Western blotting analysis under reducing conditions using previously validated human β-Klotho antibodies (R & D Systems), anti-phospho-Erk1/2, total Erk1/2, phospho-AKT, total AKT, phospho-JNK, and total JNK antibodies (Cell Signaling Technology, Danvers, MA, USA), phospho-IκBα, total IκBα, proliferating cell nuclear antigen (PCNA), and GAPDH antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and FGF21 antibody (Boster Biological Technology, Pleasanton, CA, USA).

Techniques: Expressing

Journal: Gut and Liver

Article Title: Expression of Fibroblast Growth Factor 21 and β-Klotho Regulates Hepatic Fibrosis through the Nuclear Factor-κB and c-Jun N-Terminal Kinase Pathways

doi: 10.5009/gnl17443

Figure Lengend Snippet: Inflammatory cytokine effects on β-Klotho expression in Huh-7 cells. Huh-7 cells were incubated with increasing amounts of interleukin-6 (IL-6), IL-1β, and tumor necrosis factor-α (TNF-α) for 6 hours. β-Klotho expression levels were determined by immunoblotting. GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The lysates were subjected to Western blotting analysis under reducing conditions using previously validated human β-Klotho antibodies (R & D Systems), anti-phospho-Erk1/2, total Erk1/2, phospho-AKT, total AKT, phospho-JNK, and total JNK antibodies (Cell Signaling Technology, Danvers, MA, USA), phospho-IκBα, total IκBα, proliferating cell nuclear antigen (PCNA), and GAPDH antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and FGF21 antibody (Boster Biological Technology, Pleasanton, CA, USA).

Techniques: Expressing, Incubation, Western Blot

Journal: Gut and Liver

Article Title: Expression of Fibroblast Growth Factor 21 and β-Klotho Regulates Hepatic Fibrosis through the Nuclear Factor-κB and c-Jun N-Terminal Kinase Pathways

doi: 10.5009/gnl17443

Figure Lengend Snippet: Effects of IL-1β on β-Klotho and FGF21 in Huh-7 cells. Huh-7 cells were incubated with IL-1β for 6 hours. β-Klotho expression levels (A and G), protein kinase B (B), extracelluar signal-regulated kinases (C), JNK (D), IκBα (E), and FGF21 (F and G) were determined by immunoblotting. IL, interleukin; FGF, fibroblast growth factor; JNK, c-Jun N-terminal kinase.

Article Snippet: The lysates were subjected to Western blotting analysis under reducing conditions using previously validated human β-Klotho antibodies (R & D Systems), anti-phospho-Erk1/2, total Erk1/2, phospho-AKT, total AKT, phospho-JNK, and total JNK antibodies (Cell Signaling Technology, Danvers, MA, USA), phospho-IκBα, total IκBα, proliferating cell nuclear antigen (PCNA), and GAPDH antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and FGF21 antibody (Boster Biological Technology, Pleasanton, CA, USA).

Techniques: Incubation, Expressing, Western Blot

Journal: Gut and Liver

Article Title: Expression of Fibroblast Growth Factor 21 and β-Klotho Regulates Hepatic Fibrosis through the Nuclear Factor-κB and c-Jun N-Terminal Kinase Pathways

doi: 10.5009/gnl17443

Figure Lengend Snippet: Signaling pathways that inhibit β-Klotho and induce FGF21 by IL-1β in Huh-7 cells. Huh-7 cells were pretreated with an NF-κB inhibitor (Bay11-7082, A and E), JNK inhibitor (SP600125, B and F), protein kinase B inhibitor (LY294002, C and G), or extracelluar signal-regulated kinases inhibitor (PD98059, D and H) for 20 minutes and then treated with 10 ng/mL of IL-1β for 6 hours to detect β-Klotho and FGF21. (I) Huh-7 cells were pretreated with 1 μM NF-κB inhibitor Bay11-7082, after which β-Klotho and FGF21 levels were determined by immunoblotting. (J) Huh-7 cells were pretreated with 0.5 μM JNK inhibitor SP600125, after which β-Klotho and FGF21 levels were determined by immunoblotting. (K) β-Klotho and FGF21 levels were assessed 6 hours after pretreatment with 1 μM NF-κB inhibitor Bay11-7082 in Huh-7 cells. (L) β-Klotho and FGF21 levels were assessed 6 hours after pretreatment with 0.5 μM JNK inhibitor SP600125 in Huh-7 cells. FGF, fibroblast growth factor; IL, interleukin; NF-κB, nuclear factor-κB; JNK, c-Jun N-terminal kinase; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The lysates were subjected to Western blotting analysis under reducing conditions using previously validated human β-Klotho antibodies (R & D Systems), anti-phospho-Erk1/2, total Erk1/2, phospho-AKT, total AKT, phospho-JNK, and total JNK antibodies (Cell Signaling Technology, Danvers, MA, USA), phospho-IκBα, total IκBα, proliferating cell nuclear antigen (PCNA), and GAPDH antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and FGF21 antibody (Boster Biological Technology, Pleasanton, CA, USA).

Techniques: Protein-Protein interactions, Western Blot

Journal: Gut and Liver

Article Title: Expression of Fibroblast Growth Factor 21 and β-Klotho Regulates Hepatic Fibrosis through the Nuclear Factor-κB and c-Jun N-Terminal Kinase Pathways

doi: 10.5009/gnl17443

Figure Lengend Snippet: FGF21 inhibits IL-1β-induced growth retardation of hepatocytes. Huh-7 cells were incubated with 10 ng/mL of IL-1β (A), 500 ng/mL of FGF21 (B), or IL-1β+FGF21 (C), after which the expression of β-Klotho and PCNA was determined. (D) IL-1β-treated Huh-7 cells were incubated with or without FGF21 in a dose-dependent manner. Cell viability was measured by an methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay. IL, interleukin; FGF, fibroblast growth factor; PCNA, proliferating cell nuclear antigen; GAPDH, glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The lysates were subjected to Western blotting analysis under reducing conditions using previously validated human β-Klotho antibodies (R & D Systems), anti-phospho-Erk1/2, total Erk1/2, phospho-AKT, total AKT, phospho-JNK, and total JNK antibodies (Cell Signaling Technology, Danvers, MA, USA), phospho-IκBα, total IκBα, proliferating cell nuclear antigen (PCNA), and GAPDH antibodies (Santa Cruz Biotechnology, Santa Cruz, CA, USA), and FGF21 antibody (Boster Biological Technology, Pleasanton, CA, USA).

Techniques: Incubation, Expressing, MTT Assay