Two genetic loci regulate T cell-dependent islet inflammation and drive autoimmune diabetes pathogenesis

Abstract

Insulin-dependent diabetes mellitus (IDDM) is a polygenic disease caused by progressive autoimmune infiltration (insulitis) of the pancreatic islets of Langerhan, culminating in the destruction of insulin-producing beta cells. Genome scans of families with diabetes suggest that multiple loci make incremental contributions to disease susceptibility. However, only the IDDM1 locus is well characterized, at a molecular and functional level, as alleleic variants of the major histocompatibility complex (MHC) class II HLA-DQB1, DRB1, and DPB1 genes that mediate antigen presentation to T cells. In the nonobese diabetic (NOD) mouse model, the Idd1 locus was shown to be the orthologous MHC gene I-Ab. Inheritance of susceptibility alleles at IDDM1/Idd1 is insufficient for disease development in humans and NOD mice. However, the identities and functions of the remaining diabetes loci (Idd2-Idd19 in NOD mice) are largely undefined. A crucial limitation in previous genetic linkage studies of this disease has been reliance on a single complex phenotype-diabetes that displays low penetrance and is of limited utility for high-resolution genetic mapping. Using the NOD model, we have identified an early step in diabetes pathogenesis that behaves as a highly penetrant trait. We report that NOD-derived alleles at both the Idd5 and Idd13 loci regulate a T lymphocyte-dependent progression from a benign to a destructive stage of insulitis. Human chromosomal regions orthologous to the Idd5 and -13 intervals are also linked to diabetes risk, suggesting that conserved genes encoded at these loci are central regulators of disease pathogenesis. These data are the first to reveal a role for individual non-MHC Idd loci in a specific, critical step in diabetes pathogenesis-T cell recruitment to islet lesions driving destructive inflammation. Importantly, identification of intermediate phenotypes in complex disease pathogenesis provides the tools required to progress toward gene identification at these loci.

Figure 1

Invasive insulitis is T cell dependent. A, Serial pancreatic cross-sections from 80-d-old female NOD, (NOD×NOR) F₁, and NOR mice were stained with H+E, to visualize infiltrating cells (panels a–c); with the anti-MHC class II antibody 10.2-16, to visualize APC (panels d–f); and with the anti-CD3e antibody 145-2C11, to visualize T cells (panels g–i). Panels d–i were counterstained in Mayer's hematoxylin. B, Serial sections from two representative (NOD×NOR) F₂ intercross progeny were stained with H+E (panels a and b), anti-MHC class II (panels c and d), and anti-CD3 (panels e and f). Positive protein expression is indicated by dark brown coloration. Original magnification, ×250.

Figure 2

Insulitis severity in (NOD×NOR) F₂ intercross progeny. The average insulitis severity was calculated in 82 80-d-old (NOD×NOR) F₂ intercross mice by the scoring of H+E–stained pancreatic cross-sections. As shown, a scatter plot comparing individual insulitis severity in male (blackened circles) and female (unblackened circles) animals clearly indicates that insulitis severity behaves as a bimodal trait.

Figure 3

NOD-derived alleles at Idd5 and Idd13 confer invasive insulitis. The LOD scores for the linkage of invasive insulitis in females, males, and all animals are shown vs. the distance in centimorgans from the centromeres of A, chromosome 1 and B, chromosome 2. The relative order of the microsatellite markers along each chromosome (according to Whitehead Institute for Biomedical Research/MIT Center for Genome Research) is shown beneath the x axis of each graph. C57BL/6J-derived regions (checkered boxes) are shown on NOR chromosomes 1 and 2. The remaining regions (unblackened boxes) are NOD-derived. *(NOD×NOR) F₂ intercross animals could not be scored for linkage telomeric to D1Mit305 or centromeric to D2Mit395 since these markers very close to the boundaries of these C57BL/6J-derived chromosome segments in NOR mice.

Figure 4

Histogram scatter plots of the insulitis severity in male (blackened circles) and female (unblackened circles) (NOD×NOR) F₂ intercross progeny distributed by combination of NOD (N) and C57BKS/J (B) alleles at Idd5 (D1Mit305) and Idd13 (D2Mit395).

Figure 5

Chromosomes 1 and 2 in NOR mice. NOR genomic DNA was typed for NOD- (unblackened boxes) or C57BL/6J-derived (checkered boxes) alleles at each microsatellite marker shown on chromosomes 1 (A) and 2 (B). The distance in centimorgans between markers is shown to the far left. Several candidate genes that map to each region are also shown (in italics). * = 1-LOD confidence intervals were calculated with MAPMAKER QTL. The gene locations and distances in centimorgans are adapted from published studies (Seldin , ; Siracusa et al. 1996, 1997), the Whitehead Institute for Biomedical Research/MIT Center for Genome Research, and the Jackson Laboratory. The original location of Idd5 encompasses an interval 20–60 cM from the centromere of chromosome 1 (Cornall et al. ; Garchon et al. 1991). Idd13 was originally mapped to a 4-cM interval of chromosome 2 that spans the β₂m, Il1, and Pcna genes (Serreze et al. 1994), within which the 1-LOD confidence interval for insulitis progression resides.