Diabetes Metab J.  2020 Aug;44(4):566-580. 10.4093/dmj.2019.0089.

Therapeutic Effects of Fibroblast Growth Factor-21 on Diabetic Nephropathy and the Possible Mechanism in Type 1 Diabetes Mellitus Mice

• Weng W
• Ge T
• Wang Y
• He L
• Liu T
• Wang W
• Zheng Z
• Yu L
• Zhang C
• Lu X

• KMID: 2513016
• DOI: http://doi.org/10.4093/dmj.2019.0089

Abstract

Fibroblast growth factor 21 (FGF21) has been only reported to prevent type 1 diabetic nephropathy (DN) in the streptozotocin-induced type 1 diabetes mellitus (T1DM) mouse model. However, the FVB (Cg)-Tg (Cryaa-Tag, Ins2-CALM1) 26OVE/PneJ (OVE26) transgenic mouse is a widely recommended mouse model to recapture the most important features of T1DM nephropathy that often occurs in diabetic patients. In addition, most previous studies focused on exploring the preventive effect of FGF21 on the development of DN. However, in clinic, development of therapeutic strategy has much more realistic value compared with preventive strategy since the onset time of DN is difficult to be accurately predicted. Therefore, in the present study OVE26 mice were used to investigate the potential therapeutic effects of FGF21 on DN.

Four-month-old female OVE26 mice were intraperitoneally treated with recombinant FGF21 at a dose of 100 µg/kg/day for 3 months. The diabetic and non-diabetic control mice were treated with phosphate-buffered saline at the same volume. Renal functions, pathological changes, inflammation, apoptosis, oxidative stress and fibrosis were examined in mice of all groups.

The results showed that severe renal dysfunction, morphological changes, inflammation, apoptosis, and fibrosis were observed in OVE26 mice. However, all the renal abnormalities above in OVE26 mice were significantly attenuated by 3-month FGF21 treatment associated with improvement of renal adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK) activity and sirtuin 1 (SIRT1) expression.

Therefore, this study demonstrated that FGF21 might exert therapeutic effects on DN through AMPK-SIRT1 pathway.

Keyword

AMP-activated protein kinases; Diabetes mellitus, type 1; Diabetic nephropathies; Fibroblast growth factor 21; Fibrosis; Inflammation; Sirtuin 1

Figure

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  Fig. 1 Impact of fibroblast growth factor 21 (FGF21) on diabetes-induced general changes and on renal function. Four-month-old female transgenic type 1 diabetes mellitus (DM) (FVB [Cg]-Tg [Cryaa-Tag, Ins2-CALM1] 26OVE/PneJ [OVE26]) mice and Friend Virus B NIH Jackson (FVB/NJ) control mice were given FGF21 (100 µg/kg) or an equal volume of phosphate-buffered saline (PBS) daily for 3 months (A). Weekly body weight before sacrifice (B), weekly blood glucose before sacrifice (C), urinary albumin (D), glomerular filtration rate (GFR) (E), survival rate (F), kidney weight to tibia length ratio (G), and plasma FGF21 (H) were examined. Data are presented as mean±standard deviation (Friend virus B NIH Jackson [FVB], n=9; diabetes mellitus [DM], n=10; FGF21-treated diabetic mice [FGF21], n=6). i.p., intraperitoneally; UA, urine albumin. aP≤0.05 for each DM vs. FVB/NJ groups, bP≤0.05 for DM/FGF21 vs. DM groups.

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  Fig. 2 Impact of fibroblast growth factor 21 (FGF21) on diabetes-induced pathological changes. Kidney pathology was examined with hematoxylin & eosin staining (H&E) (A, B), periodic acid-Schiff (PAS) staining (C, D), and picrosirius red staining (PRS) (E, F) (×400). Renal expression of fibronectin (FN) (G, H), connective tissue growth factor (CTGF) (G, I), and α-smooth muscle actin (α-SMA) (G, J) were examined by Western blotting assay. Expression of Wilms' tumor 1 gene (WT1) was tested by immunohistochemical staining (K , L) (×400). Black arrows: Wilms tumor positive cells. For PRS, semi-quantitative analysis was conducted by computer imaging analysis. Data are presented as mean±standard deviation (Friend virus B NIH Jackson [FVB], n=9; diabetes mellitus [DM], n=10; FGF21-treated diabetic mice [FGF21], n=6). aP≤0.05 for each DM vs. FVB groups, bP≤0.05 for DM/FGF21 vs. DM groups.

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  Fig. 3 Impact of fibroblast growth factor 21 (FGF21) on diabetes-induced renal inflammation. Renal expression of tumor necrosis factor α (TNF-α) (A, B), CD68 (A, C), and intercellular adhesion molecule-1 (ICAM) (A, D) was tested by Western blotting assay (A). Renal expression of CD68 and CD3 was examined by immunohistochemical staining (E–H) (×400). Data are presented as mean±standard deviation (Friend virus B NIH Jackson [FVB], n=9; diabetes mellitus [DM], n=10; FGF21-treated diabetic mice [FGF21], n=6). aP≤0.05 for each DM vs. FVB groups, bP≤0.05 for DM/FGF21 vs. DM groups.

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  Fig. 4 Impact of fibroblast growth factor 21 (FGF21) on diabetes-induced renal apoptosis. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining (green), nuclei staining with 4′,6-diamidino-2-phenylindole (DAPI) (blue), and merge (A, B) (×400). Red arrows: apoptosis in tubule; purple arrows: apoptosis in glomeruli. BCL-2-associated X protein (BAX) and BCL-2 were examined by Western blotting assay (C, D). Data are presented as mean±standard deviation (Friend virus B NIH Jackson [FVB], n=9; diabetes mellitus [DM], n=10; FGF21-treated diabetic mice [FGF21], n=6). aP≤0.05 for each DM vs. FVB groups, bP≤0.05 for DM/FGF21 vs. DM groups.

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  Fig. 5 Impact of fibroblast growth factor 21 (FGF21) on diabetes-induced renal oxidative stress. The lipid peroxide concentrations of plasma (A) and kidney (B) were measured by detecting thiobarbituric acid (TBA) reactivity reflected by the amount of malondialdehyde (MDA) formed during acid hydrolysis of the lipid peroxide compound. Renal expression of 3-nitrotyrosine (3-NT) (D), 4 hydroxynonenal (4-HNE) (E), heme oxygenase-1 (HO-1) (F), NAD(P)H dehydrogenase [quinone]-1 (NQO-1) (G), catalse (CAT) (H) and superoxide dismutase 2 (SOD2) (I) were tested by Western blotting assay (C). Data are presented as mean±standard deviation (Friend virus B NIH Jackson [FVB], n=9; diabetes mellitus [DM], n=10; FGF21-treated diabetic mice [FGF21], n=6). OVE, FVB (Cg)-Tg (Cryaa-Tag, Ins2-CALM1) 26OVE/PneJ (OVE26). aP≤0.05 for each DM vs. FVB groups, bP≤0.05 for DM/FGF21 vs. DM groups.

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  Fig. 6 Fibroblast growth factor 21 (FGF21) protects diabetic nephropathy through the AMP-activated protein kinase (AMPK)-sirtuin 1 (SIRT1) pathway. Renal expression of ERK1/2 (B), AMPK (C), P38 (D), SIRT1 (F) and signal transducer and activator of transcription 3 (STAT3) (G) was examined by Western blotting assay (A), and renal SIRT1 mRNA expression was examined by reverse transcription polymerase chain reaction (E). Data are presented as mean±standard deviation (Friend virus B NIH Jackson [FVB], n=9; diabetes mellitus [DM], n=10; FGF21-treated diabetic mice [FGF21], n=6). p-, phosphor; T-, total. aP≤0.05 for each DM vs. FVB groups, bP≤0.05 for DM/FGF21 vs. DM groups.

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