Polymorphic Variants of Insulin-Like Growth Factor I (IGF-I) Receptor and Phosphoinositide 3-Kinase Genes Affect IGF-I Plasma Levels and Human Longevity: Cues for an Evolutionarily Conserved Mechanism of Life Span Control

` MASSIMILIANO BONAFE, MICHELANGELA BARBIERI, FRANCESCA MARCHEGIANI, FABIOLA OLIVIERI, EMILIA RAGNO, CLAUDIA GIAMPIERI, ELENA MUGIANESI, MATTEO CENTURELLI, CLAUDIO FRANCESCHI, GIUSEPPE PAOLISSO AND

Department of Experimental Pathology, University of Bologna, 40126 Bologna, Italy; Department of Geriatric Medicine and Metabolic Diseases II, University of Naples (B.M., R.E., P.G.), 80138 Naples, Italy; and Italian National Research Centers on Aging (M.F., O.F., G.C., M.E., C.M.), 60100 Ancona, Italy

Current literature indicates that abrogation of the IGF-I re- sponse pathway affects longevity in Caenorhabditis elegans, and that the down-regulation of IGF-I gene expression is as- sociated with an extension of the life span in mice. In this paper we tested the hypothesis that polymorphic variants of IGF-I response pathway genes, namely IGF-IR (IGF-I recep- tor; G/A, codon 1013), PI3KCB (phosphoinositol 3-kinase; T/C, (cid:1)359 bp; A/G, (cid:1)303 bp), IRS-1 (insulin receptor substrate-1; G/A, codon 972), and FOXO1A (T/C, (cid:2)97347 bp), play a role in systemic IGF-I regulation and human longevity. The major findingofthisinvestigationwasthatsubjectscarryingatleast

an A allele at IGF-IR have low levels of free plasma IGF-I and are more represented among long-lived people. Moreover, ge- notype combinations at IGF-IR and PI3KCB genes affect free IGF-I plasma levels and longevity. These findings represent the first indication that free IGF-I plasma levels and human longevity are coregulated by an overlapping set of genes, con- tributingtothehypothesisthattheimpactoftheIGF-I/insulin pathway on longevity is a property that has been evolution- arily conserved throughout the animal kingdom. (J Clin En- docrinol Metab 88: 3299–3304, 2003)

AKT1/AKT2 kinases, affect the life span of the yeast (8). the basis of these data it has been predicted that genes volved in the insulin/IGF-I response pathway play a role longevity throughout evolution (9).

tions in genes that share similarities with the human genes involved in the insulin/IGF-I signal response pathway are responsible for an impairment of IGF-I/insulin signaling and for an extension of the Caenorhabditis elegans life span (1). In particular, the best characterized genes of this ancestral IGF-I/insulin response pathway are: DAF-2, ortholog to the human IGF-I/insulin receptor family; AGE-1, ortholog to human p110 catalytic subunit of phosphoinositide 3-kinase, which is involved in IGF-I/insulin signal transduction; and DAF-16, ortholog to the human FOXO family of Forkhead transcription factors, which regulates IGF-I/insulin-induced changes in gene transcription (2–5).

A major role of IGF-I in mammalian longevity has been suggested by studies on dwarf mice carrying mutations in transcription factors PIT-1 and PROP-1, which are charac- terized by a long life span and reduced IGF-I plasma levels (10–12). Conversely, transgenic animals with elevated plasma IGF-I levels have a short life span (13). In humans there is evidence that long-lived subjects have decreased plasma IGF-I levels (14, 15) and preserved insulin action compared with aged subjects, thus indicating that an efficient insulin response has an impact on human longevity (16–18).

Further recent studies have shown that mutations in the Drosophila melanogaster INR gene (similar to the human in- sulin receptor) and in the CHICO gene, the ortholog to hu- man insulin receptor substrate-1 (IRS-1), the protein associ- ated with the IGF-I/insulin receptor, affect the life span, confirming the hypothesis that the insulin/IGF-I response pathway plays a role in the regulation of invertebrate lon- gevity (6, 7).

On the basis of the above-mentioned literature, we inves- tigated the role of genetic variability at human loci that share similarities with the genes that regulate the insulin/IGF-I response in C. elegans (1–4) and D. melanogaster. In particular, polymorphisms at IGF-IR (IGF-I receptor; 34% protein se- quence identity with the C. elegans DAF-2 gene), PIK3CB (phosphoinositol 3-kinase; 27% protein sequence identity with the C. elegans AGE-1 gene), IRS-1 (30% protein sequence identity with the D. melanogaster CHICO gene), FOXO1A (49% protein sequence identity with the C. elegans DAF-16 gene) were investigated. Their effects on IGF-I plasma levels were assessed. Moreover, the genotype distributions found in long-lived people over 85 yr of age and those found in

Intriguingly, mutations in the Saccaromyces cerevisiae SCH9 gene, which share similarities with insulin/IGF-I-regulated

Abbreviations: BMI, Body mass index; HOMA, homeostatic model IGF-IR, IGF-I receptor; IR, insulin resistance; IRS-1, insulin substrate-1; PI3KCB, phosphoinositol 3-kinase.

CAGAAAT-3(cid:3) . The PCR products were separated by agarose gel electrophoresis, followed by ethidium bromide staining.

people less than 85 yr old were compared. This strategy to select long-lived people from the population was based on three findings: 1) demographic data suggesting that the se- lection for longevity starts at about 85 yr (19); 2) demographic models predicting that changes in the frequency of poly- morphisms associated with different survival capabilities during aging are expected to occur beyond the age of 85 yr (20); and 3) previous data showing that people beyond the age of 85 yr have peculiar metabolic/anthropometric pa- rameters [such as high insulin sensitivity, low body mass index (BMI), and low glycemia] in comparison with the rest of the aging population (18). All of these data suggest that healthy people over 85 yr of age have been highly selected