hNAG-1 increases lifespan by regulating energy metabolism and insulin/IGF-1/mTOR signaling

Abstract

Nonsteroidal anti-inflammatory drug-activated gene (NAG-1) or GDF15 is a divergent member of the transforming growth factor beta (TGF-β) superfamily and mice expressing hNAG-1/hGDF15 have been shown to be resistant to HFD-induced obesity and inflammation. This study investigated if hNAG-1 increases lifespan in mice and its potential mechanisms. Here we report that female hNAG-1 mice had significantly increased both mean and median life spans in two transgenic lines, with a larger difference in life spans in mice on a HFD than on low fat diet. hNAG-1 mice displayed significantly reduced body and adipose tissue weight, lowered serum IGF-1, insulin and glucose levels, improved insulin sensitivity, and increased oxygen utilization, oxidative metabolism and energy expenditure. Gene expression analysis revealed significant differences in conserved gene pathways that are important regulators of longevity, including IGF-1, p70S6K, and PI3K/Akt signaling cascades. Phosphorylation of major components of IGF-1/mTOR signaling pathway was significantly lower in hNAG-1mice. Collectively, hNAG-1 is an important regulator of mammalian longevity and may act as a survival factor. Our study suggests that hNAG-1 has potential therapeutic uses in obesity-related diseases where life span is frequently shorter.

Figure 1. Increased lifespan of female hNAG-1 transgenic mice

(a-b), Kaplan-Meier survival curves for female Wt and hNAG-1 mice from two transgenic lines, line 1377 and line 1398 (a) LFD (b) HFD. (c), Terminal mean body weights of hNAG-1 and Wt mice at 95±5 wk old in two lines (g). (d), Mean abdominal white adipose tissue (WAT) weights of the mice (g). n=9~18. Data are presented as mean ± SE. a, p<0.05, b, p<0.01 and c, p<0.001 as determined by Student's t-test.

Figure 2. hNAG-1 transgenic mice have improved insulin sensitivity

(a), Mean body length of female hNAG-1 and Wt mice in both lines (n=6~8). (b-d), Mean serum levels of growth hormone (b), IGF-1 (c), insulin and leptin (d) in terminal blood of hNAG-1 and Wt mice on LFD and HFD (n=10). e, Insulin tolerance test on old (>95 wk) 1398 female hNAG-1 Tg mice and Wt mice (n=8~9). Data are presented as mean ±SE. a, p<0.05, b, p<0.01 and c, p<0.001 as determined by Student's t-test.

Figure 3. hNAG-1 transgenic mice have increased metabolism

(a), Heat production (W/kg) in female hNAG-1 and Wt mice as determined indirect colorimeter (n=5). (b-c), Consumption of O₂ and VO₂ (ml/h/kg) in female hNAG-1 and Wt mice during day and night time (n=5). (d), Respiratory quotient (RER) as calculated from VO₂ and VCO₂ in 1398 hNAG-1 female mice. Data are presented as mean ± SE. a, p<0.05, b, p<0.01 and c, p<0.001 as determined by Student's t-test.

Figure 4. Gene validation from microarray results

Validation was analyzed by qRT-PCR. (a-b), Validation of the expression of down-regulated genes from microarray study in young (30 wk, a) and old (95 wk, b) abdominal white adipose tissue (WAT) of female mice (n=6). Data are presented as mean ± SE. a, p<0.05, b, p<0.01 and c, p<0.001 as determined by Student's t-test.

Figure 5. hNAG-1 transgenic mice have reduced IGF-1/Insulin/mTOR signaling

Phosphorylation of proteins of IGF-1/insulin/mTOR signaling pathway was analyzed by Western blot (n=3) in abdominal WAT. Both samples from 30 wk and 95 wk old animals were analyzed.

Figure 6. Schematic model for increased survival and lifespan in hNAG-1 mice

Overexpression of hNAG-1 in female mice lowers serum levels of IGF-1 and insulin and thus reduces IGF-1/insulin (IIS)/mTOR signaling. Circulating hNAG-1 also increases lipolysis, thermogenesis, and metabolism in hNAG-1 mice [14], and reduces macrophage infiltration into WAT and reduces inflammation in hNAG-1 mice [15, 16]. hNAG-1 mice are thus resistant to obesity, have increased energy metabolism and increased insulin sensitivity, which leads to increased lifespan, and reduced age- or dietary-induced pathological lesions.