The intestinal fatty acid binding protein locus (FABP2) was investigated as a possible genetic factor in determining insulin action in the Pima Indian population. A polymorphism at codon 54 of FABP2 was identified that results in an alanineencoding allele (frequency 0.71) and a threonine-encoding allele (frequency 0.29). Pimas who were homozygous or heterozygous for the threonine-encoding allele were found to have a higher mean fasting plasma insulin concentration, a lower mean insulin-stimulated glucose uptake rate, a higher mean insulin response to oral glucose and a mixed meal, and a higher mean fat oxidation rate compared with Pimas who were homozygous for the alanine-encoding allele. Since the FABP2 threonine-encoding allele was found to be associated with insulin resistance and increased fat oxidation in vivo, we further analyzed the FABP2 gene products for potential functional differences. Titration microcalorimetry studies with purified recombinant protein showed that the threonine-containing protein had a twofold greater affinity for long-chain fatty acids than the alanine-containing protein. We conclude that the threonine-containing protein may increase absorption and/or processing of dietary fatty acids by the intestine and thereby increase fat oxidation, which has been shown to reduce insulin action. (J. Clin.

Genetic factors influence the development of type II diabetes mellitus, but genetic loci for the most common forms of diabetes have not been identified. A genomic scan was conducted to identify loci linked to diabetes and body-mass index (BMI) in Pima Indians, a Native American population with a high prevalence of type II diabetes. Among 264 nuclear families containing 966 siblings, 516 autosomal markers with a median distance between adjacent markers of 6.4 cM were genotyped. Variance-components methods were used to test for linkage with an age-adjusted diabetes score and with BMI. In multipoint analyses, the strongest evidence for linkage with age-adjusted diabetes (LOD = 1.7) was on chromosome 11q, in the region that was also linked most strongly with BMI (LOD = 3.6). Bivariate linkage analyses strongly rejected both the null hypothesis of no linkage with either trait and the null hypothesis of no contribution of the locus to the covariation among the two traits. Sib-pair analyses suggest additional potential diabetes-susceptibility loci on chromosomes 1q and 7q.

Homozygosity for the severe combined immunodeficiency (scid) mutation results in a block in T-and B-lymphocyte development. An unusually high incidence of spontaneous thymic lymphoma development was observed after transfer ofthis mutation from the C.B-17 congenic strain background onto the diabetes-susceptible nonobese diabetic (NOD) background. Thymomagenesis in the NOD-scid/scid mouse was associated with expression ofan NOD mouse-unique endogenous ecotropic murine leukemia provirus locus (Ehv-30, mapped to proximal region of chromosome 11) not expressed in the standard substrain NOD/Lt thymus. All tumors exhibited insertions of ecotropic proviruses, whereas only a subset also exhibited proviral integrations of mink cell focusforming retrovirus. Neither class of retrovirus was associated with consistent integration into genes previously associated with activation of oncogenesis. We propose that the unusual features of T-cell ontogeny characteristic of the NOD inbred strain synergize withe the scid-imparted block in thymocyte development, leading to activation of the NOD-unique Emv-30 to initiate thynomagenesis.T lymphocyte-mediated destruction of the insulin-producing pancreatic 8 cells plays a key role in the pathogenesis of spontaneous autoimmune diabetes in the nonobese diabetic (NOD) mouse strain, a model for insulin-dependent diabetes mellitus (1). Diabetes can be adoptively transferred by either T lymphocytes or bone marrow (2, 3). In addition to several abnormal T-lymphocyte functions (4, 5), a dominantly inherited accumulation ofT cells in peripheral lymphoid tissues (6) and a defect in thymic maturation (7) have been associated with diabetogenesis in the NOD/Lt mouse substrain.Studies designed to delineate the separate diabetogenic functions of CD4' and CD8' T-cell subsets in adoptive transfer experiments generally have required irradiation of the recipient to deplete endogenous T-cell immunity. Using a different approach to eliminate endogenous T-cell function, we have developed a NOD strain congenic for the severe combined immunodeficiency (scid) mutation. Homozygotes for this recessive mutation lack functional T and B lymphocytes (8); thus, NOD-scid/scid mice are diabetes-resistant (9). During the establishment of this congenic strain, the diabetes-resistant NOD-scid/scid homozygous mice were distinguished from their nonimmunodeficient sibs (either +/+ or +/scid) by an unusually high frequency of spontaneous thymic lymphoma (thymoma) development in the former. This was especially noteworthy because thymic lymphomas have not been detected in intact NOD/Lt-+/+ mice that do not develop diabetes, although such longer-lived

Type 2 diabetes mellitus is a common chronic disease that is thought to have a substantial genetic basis. Identification of the genes responsible has been hampered by the complex nature of the syndrome. Abnormalities in insulin secretion and insulin action predict the development of type 2 diabetes and are, themselves, highly heritable traits. Since fewer genes may contribute to these precursors of type 2 diabetes than to the overall syndrome, such genes may be easier to identify. We, therefore, undertook an autosomal genomic scan to identify loci linked to prediabetic traits in Pima Indians, a population with a high prevalence of type 2 diabetes.363 nondiabetic Pima Indians were genotyped at 516 polymorphic microsatellite markers on all 22 autosomes. Linkage analyses were performed using three methods (single-marker, nonparametric multipoint [MAPMAKER/SIBS], and variance components multipoint). These analyses provided evidence for linkage at several chromosomal regions, including 3q21-24 linked to fasting plasma insulin concentration and in vivo insulin action, 4p15-q12 linked to fasting plasma insulin concentration, 9q21 linked to 2-h insulin concentration during oral glucose tolerance testing, and 22q12-13 linked to fasting plasma glucose concentration. These results suggest loci that may harbor genes contributing to type 2 diabetes in Pima Indians. None of the linkages exceeded a LOD score of 3.6 (a 5% probability of occurring in a genome-wide scan). These findings must, therefore, be considered tentative until extended in this population or replicated in others. (

Different isoenzymes of pyruvate dehydrogenase kinase (PDK) inhibit the mitochondrial pyruvate dehydrogenase complex by phosphorylation of the E1alpha subunit, thus contributing to the regulation of glucose metabolism. By positional cloning in the 7q21.3-q22.1 region linked with insulin resistance and non-insulin-dependent diabetes mellitus in the Pima Indians, we identified a gene encoding an additional human PDK isoform, as evidenced by its amino acid sequence identity (>65%) with other mammalian PDKs, and confirmed by biochemical analyses of the recombinant protein. We performed detailed comparative analyses of the gene, termed PDK4, in insulin-resistant and insulin-sensitive Pima Indians, and detected five DNA variants with comparable frequencies in both subject groups. Using quantitative reverse transcription polymerase chain reaction, we found that the variants identified in the promoter and 5'-untranslated region did not correlate with differences in mRNA level in skeletal muscle and adipose tissue. We conclude that alterations in PDK4 are unlikely to be the molecular basis underlying the observed linkage at 7q21.3-q22.1 in the Pima Indians. Information about the genomic organization and promoter sequences of PDK4 will be useful in studies of other members of this family of mitochondrial protein kinases that are important for the regulation of glucose metabolism.

A polygenic basis for susceptibility to insulin-dependent diabetes in nonobese diabetic (NOD) mice has been established by outcross to a related inbred strain, nonobese normal (NON). Analysis of first and second backcross progeny has shown that at least three recessive genes are required for development of overt diabetes. One, Idd-1s, is tightly linked to the H-2K locus on chromosome 17; another, Idd-2s, is localized proximal to the Thy-1/Alp-1 cluster on chromosome 9. Segregation of a third, Idd-3s, could be shown in a second backcross. Neither Idd-1s nor Idd-2s could individually be identified as the locus controlling insulitis; leukocytic infiltrates in pancreas were common in most asymptomatic BC1 mice. Both F1 and BC1 mice exhibited the unusually high percentage of splenic T lymphocytes characteristic of NOD, suggesting dominant inheritance of this trait. The polygenic control of diabetogenesis in NOD mice, in which a recessive gene linked to the major histocompatibility complex is but one of several controlling loci, suggests that similar polygenic interactions underlie this type of diabetes in humans.

No abstract

SummaryInsulin-dependent diabetes meUitus (IDDM) in NOD/Lt mice represents a complex polygenic disease. NOR/Lt is a recombinant congenic strain (RCS) in which limited regions of the NOD/Lt genome have been replaced by genome from the C57BL/KsJ strain. NOR. mice are insulitis resistant and diabetes free despite genetic identity with NOD at numerous chromosomal regions containing previously described insulin-dependent diabetes (Idd) genes, including the strongly diabetogenic H2g 7 major histocompatibility complex (MHC) haplotype. The present study revealed BKs-derived genome on segments of chromosomes (Chr) 1, 2, 4, 5, 7, 11, 12, and 18, approximating 11.6% of the total NOR genome analyzed. (NOD x NOR)F2 segregation analysis was employed to identify chromosomal regions in NOR containing Idd resistance alleles. IDDM developed in 33% (10/30) of F1 females, and 29.3% (36/123) of F2 females aged to 1 yr. A previously unrecognized diabetes resistance locus (designated Idcl13") strongly protective in homozygous state was identified on NOR Chr 2 in linkage with the Illcr structural gene. The existence of this locus was confirmed by construction of a NOD stock congenic for NORderived markers on Chr 2. Our analysis shows the utility of RCS and congenic stocks for the identification and isolation of non-MHC genes with strong antidiabetogenic functions.