Fibroblast Growth Factor 21 Is Not Required for the Reductions in Circulating Insulin-Like Growth Factor-1 or Global Cell Proliferation Rates in Response to Moderate Calorie Restriction in Adult Mice

<div><p>Calorie restriction (CR) delays aging and extends lifespan in numerous organisms, including mice. Down-regulation of the somatotropic axis, including a reduction in insulin-like growth factor-1 (IGF-1), likely plays an important role in CR-induced lifespan extension, possibly by reducing cell proliferation rates, thereby delaying replicative senescence and inhibiting tumor promotion. Accordingly, elucidating the mechanism(s) by which IGF-1 is reduced in response to CR holds therapeutic potential in the fight against age-related diseases. Up-regulation of fibroblast growth factor 21 (FGF21) is one possible mechanism given that FGF21 expression is induced in response to nutritional deprivation and has been implicated as a negative regulator of IGF-1 expression. Here we investigated alterations in hepatic growth hormone (GH)-mediated IGF-1 production and signaling as well as the role of FGF21 in the regulation of IGF-1 levels and cell proliferation rates in response to moderate CR in adult mice. We found that in response to moderate CR, circulating GH and hepatic janus kinase 2 (JAK2) phosphorylation levels are unchanged but that hepatic signal transducer and activator of transcription 5 (STAT5) phosphorylation levels are reduced, identifying STAT5 phosphorylation as a potential key site of CR action within the somatotropic axis. Circadian measurements revealed that the relative level of FGF21 expression is both higher and lower in CR vs. ad libitum (AL)-fed mice, depending on the time of measurement. Employing FGF21-knockout mice, we determined that FGF21 is not required for the reduction in IGF-1 levels or cell proliferation rates in response to moderate CR. However, compared to AL-fed WT mice, AL-fed FGF21-knockout mice exhibited higher basal rates of cell proliferation, suggesting anti-mitotic effects of FGF21. This work provides insights into both GH-mediated IGF-1 production in the context of CR and the complex network that regulates FGF21 and IGF-1 expression and cell proliferation rates in response to nutritional status.</p></div

Circulating and hepatic mRNA FGF21, IGF-1 and IGFBP-1 levels in WT/AL, WT/CR, FGF21-KO/AL and FGF21-KO/CR mice.

<p>Plasma levels of A) FGF21, C) IGF-1 and E) IGFBP-1 and relative hepatic mRNA expression levels of B) FGF21, D) IGF-1 and F) IGFBP-1 at 1500 h and 1900 h in WT and FGF21-KO mice fed AL or CR (n = 3–12 per genotype per diet per time point). Hepatic mRNA expression levels were normalized to 18S rRNA and then normalized to WT/AL mice at 1500 h. All between-group analyses were performed using a two-way ANOVA with a Bonferroni <i>post hoc</i> test at each time point (* p<0.05, ** p<0.01, *** p<0.001).</p

GH secretory dynamics and hepatic GH signaling in CR mice.

<p>Plasma GH levels described as A) the integrated hourly concentration or B) the maximum pulse amplitude of GH over the 5.25 h sampling period in AL and CR mice (n = 8 per diet). Densitometric analysis and western blot images of hepatic protein levels of phosphorylated to total C) JAK2 and D) STAT5 in AL and CR mice at 2100 h (n = 5–7 per diet. Data normalized to AL). Student's unpaired two-tailed <i>t</i>-tests were used for all between-group analyses (** p<0.0071).</p

Circulating and hepatic mRNA levels of IGF-1 and FGF21 in CR mice.

<p>Plasma A) IGF-1 and B) FGF21 levels in AL and CR mice at 2100 h or 2200 h (n = 16–21 per diet). Relative hepatic mRNA expression levels of C) IGF-1 and D) FGF21 in AL and CR mice at 2100 h or 2200 h (n = 29–31 per diet). Hepatic mRNA expression levels were normalized to 18S rRNA and then normalized to the AL group. Student's unpaired two-tailed <i>t</i>-tests were used for all between-group analyses (* p<0.05, ** p<0.0043, *** p<0.0001).</p