PMCCPMCCPMCC

Search tips
Search criteria 

Advanced

 
Logo of diabetesSubscribeSearchDiabetes JournalAmerican Diabetes Association
 
Diabetes. 2009 April; 58(4): 1007–1017.
Published online 2009 January 23. doi:  10.2337/db08-1215
PMCID: PMC2661594

Type 1 Diabetes in the BB Rat: A Polygenic Disease

Abstract

OBJECTIVE

Two type 1 diabetes susceptibility genes have been identified in the spontaneously diabetic biobreeding diabetes-prone (BBDP) rat, the major histocompatibility complex (MHC) (RT1) class II u haplotype (Iddm1) and Gimap5 (Iddm2). The strong effects of these have impeded previous efforts to map additional loci. We tested the hypothesis that type 1 diabetes is a polygenic disease in the BBDP rat.

RESEARCH DESIGN AND METHODS

We performed the most comprehensive genome-wide linkage analysis for type 1 diabetes, age of disease onset (AOO), and insulitis subphenotypes in 574 F2 animals from a cross-intercross between BBDP and type 1 diabetes–resistant, double congenic ACI.BBDP-RT1u,Gimap5 (ACI.BB1u.lyp) rats, where both Iddm1 and Iddm2 were fixed as BBDP.

RESULTS

A total of 19% of these F2 animals developed type 1 diabetes, and eight type 1 diabetes susceptibility loci were mapped, six showing significant linkage (chromosomes 1, 3, 6 [two loci], 12, and 14) and two (chromosomes 2 and 17) suggestive linkage. The chromosomes 6, 12, and 14 intervals were also linked to the severity of islet infiltration by immunocytes, while those on chromosomes 1, 6 (two loci), 14, 17, and a type 1 diabetes–unlinked chromosome 8 interval showed significant linkage to the degree of islet atrophy. Four loci exhibited suggestive linkage to AOO on chromosomes 2 (two loci), 7, and 18 but were unlinked to type 1 diabetes. INS, PTPN22, IL2/IL21, C1QTNF6, and C12orf30, associated with human type 1 diabetes, are contained within the chromosomes 1, 2, 7, and 12 loci.

CONCLUSIONS

This study demonstrates that the BBDP diabetic syndrome is a complex, polygenic disease that may share additional susceptibility genes besides MHC class II with human type 1 diabetes.

The biobreeding diabetes-prone (BBDP) rat spontaneously develops type 1 diabetes with high incidence around puberty through a T-cell–mediated autoimmune destruction of pancreatic β-cells (1). Two genes that contribute to disease pathogenesis in this animal have been identified, the RT1u allele (Iddm1) and the GTPase immunity-associated protein family member 5 (Gimap5, Iddm2) (24). Genetic variation in Gimap5 has been recently associated with humoral antipancreatic autoimmunity in human type 1 diabetes as well as with human systemic lupus erythematosus (5,6). Crosses between BBDP rats and type 1 diabetes–resistant strains have shown that homozygosity for the BBDP Gimap5 allele and at least one BBDP allele of Iddm1 are required for type 1 diabetes development (4,7). These requirements have impeded previous efforts to map additional Iddm loci. However, in an attempt to circumvent these difficulties, two experimental approaches have been used.

First, linkage analyses have been performed in progeny derived from backcrosses (F1N2 or F2N3) between BBDP and type 1 diabetes–resistant strains, specifically designed to enrich for BBDP homozygosity at both Iddm1 and Iddm2. This approach led to the identification of Iddm3 (chromosome 2) in one study, while in others more than one locus was mapped, but they met only suggestive genome-wide statistical criteria (812). However, the design of these crosses had two disadvantages. They could not be used to investigate the effects of homozygosity for non-BBDP alleles since they were backcrosses to BBDP. Furthermore, as Iddm1 and Iddm2 were selected to be homozygous, loci in close proximity to them could be missed, as was likely the case for a locus close to Iddm2 (8,1315).

The second approach examined the genetics of experimentally induced (as opposed to spontaneous) diabetic syndromes following the demonstration that many rat strains carrying the BBDP allele at Iddm1 were highly susceptible to these syndromes (16). This approach had the advantage of eliminating the requirement for BBDP homozygosity at Iddm2, thus increasing the number of informative progeny in crosses. However, only one locus (other than Iddm1) identified by this approach, Iddm14, has been shown to also control spontaneous type 1 diabetes (15). The results of these two approaches strongly suggested that contrary to spontaneous diabetes in humans and nonobese diabetic (NOD) mice, which are both under the control of multiple genes (1719), type 1 diabetes in the BBDP rat was oligogenic with one or two susceptibility genes other than Iddm1 and 2 depending on the resistant strain used (8,9,12,13).

To determine whether this is the case, we have taken a different approach that overcomes some of the limitations encountered in previous studies. First, we introgressed BBDP-derived Iddm1 and Iddm2 onto the genetic background of the diabetes-resistant August Copenhagen Irish (ACI) inbred strain. The resulting double congenic ACI.BBDP-RT1u,Gimap5 (ACI.BB1u.lyp) inbred strain exhibits complete resistance to both spontaneous type 1 diabetes and islet inflammation (20), establishing the requirement for at least one other susceptibility locus and allowing us to test the hypothesis that the BBDP diabetic syndrome is also under complex, polygenic control. Specifically, this congenic line allowed us to fix both Iddm1 and Iddm2 loci as BBDP in a cross/intercross with the BBDP strain. Here, using this F2 cohort, we report the results of the most comprehensive genome-wide linkage analysis performed in the BBDP rat.

RESEARCH DESIGN AND METHODS

The genetic analysis consisted of a three-step approach. In the first step, diabetic animals were genotyped across the genome using 229 microsatellite markers at an average of one marker every 12.5 Mb (maximal distance between two markers was 28.7 Mb). Information on the primers and their physical location was obtained from the Rat Genome Database (available at http://rgd.mcw.edu/) and is listed in supplementary Table 1 (available at http://dx.doi.org/10.2337/db08-1215). This was followed by segregation analysis of type 1 diabetes based on the comparison of the observed to the expected genotype distribution at each marker. In the second step, at each peak marker linked to diabetes with a log10 likelihood ratio (logarithm of odds [LOD]) score >1 in the first step, we genotyped all nondiabetic animals. Single-marker analysis was then performed to check that the linkage was type 1 diabetes specific as opposed to the consequence of an overall bias in Mendelian inheritance. In the third step, markers flanking those peaks linked to type 1 diabetes with a LOD >1.5 in step 2 were also genotyped in all nondiabetic subjects to refine each peak through interval mapping analysis (21). Further refinement of linkage peaks was obtained through genotyping of the whole F2 cohort with additional and newly designed markers.

Before linkage analysis, R/QTL (22) was used to identify genotyping errors and to check the marker order by re-estimating the genetic map for each order through calculating the LOD scores relative to the initial order. For QTL mapping, single-marker and QTL × sex interaction analyses were performed using R/QTL with a binary model used for type 1 diabetes and a normal model for other traits. Segregation analysis, interval mapping (21), and composite interval mapping were conducted using Windows QTL Cartographer, version 2.5 (23). For linkage analysis, we calculated LOD scores in both R/QTL and QTL Cartographer using a full model, allowing for both additive and dominance effects. LOD thresholds were determined by permutation testing (n = 1,000 permutations) (24). Suggestive loci were defined as those that exceeded the 37th percentile (P < 0.63) of permutation distribution, while significant loci exceeded the 95th percentile (P < 0.05). Power calculations showed that for type 1 diabetes there was good power (1-β = 0.8) to detect a relative risk of 2.0 with α = 1 × 10−4, while for quantitative traits we could detect a locus that contributes to 4% of the trait variance at α = 1 × 10−4. The loci identified through the above analyses were designated Iddm25 to Iddm36 by the Rat Genome Database and have been added to the list of previously mapped Iddm loci at http://rgd.mcw.edu/, while details about these 12 loci are available at http://www.t1dbase.org/page/welcome/display/?species = rat. Details of the animals used and phenotypic analysis of the pancreas are provided in the online appendix.

RESULTS

Of 470 F2 animals that remained negative for glycosuria until 165 days, 23 were found postmortem to have glycemia >16.7 mmol/l hence within the diabetic range on the day of killing (25). However, they were excluded from our analysis of cumulative incidence of type 1 diabetes and age of onset (AOO) since we did not have glycemic levels on 2 consecutive days for them. Their pancreata were nevertheless included in the linkage analysis for insulitis. Of 551 F2 animals included for cumulative incidence of type 1 diabetes, 18.9% (n = 104) became diabetic with an AOO of (means ± SD) 120 ± 24 days compared with 84% at 76 ± 8 days in BBDP rats and 20.8% at 180 ± 44 days in the F1 generation, respectively (Fig. 1A). There was a small, but significantly higher, cumulative incidence of type 1 diabetes (P = 0.048) in female F2 animals compared with males (Fig. 1B). This led us to perform linkage analyses for type 1 diabetes, insulitis, and AOO separately in male and female F2 animals. Based on permutation analyses using R/QTL, significant sex difference in linkage (LOD threshold for QTL × sex interaction at 5% level >2.1) was found only for type 1 diabetes at the telomeric locus of chromosome 6 (LOD = 2.3). However, at this locus, although female heterozygotes and ACI homozygotes had higher type 1 diabetes incidence than males, BB homozygote males had a significantly higher incidence than BB females.

FIG. 1.
Analysis of type 1 diabetes–free survival in BBDP, F1 and F2 animals (A) and per gender in the F2 cohort (B). The percent survival of each cohort is labeled on the vertical axis, and the number of days to onset of diabetes is labeled on the horizontal ...

Linkage analysis of type 1 diabetes.

Through genome-wide segregation analysis performed in diabetic animals only, we identified six loci significantly linked to type 1 diabetes on chromosomes 1, 3, 6 (two loci), 12, and 14 and another suggestive linkage peak on chromosome 2 (Fig. 2). Four of these regions are novel, on chromosomes 3, 6 (centromeric), 12, and 14, while three others overlap with regions previously identified using alternative type 1 diabetes–resistant strains (8,9). When the genetic analysis was extended to the whole F2 cohort, these peaks were all confirmed and an additional chromosome 17 locus reaching suggestive linkage was found. Of note, some regions of linkage identified at intermediate stages of our study were lost when the whole cohort was genotyped.

FIG. 2.
Mapping of type 1 diabetes and type 1 diabetes subphenotypes in the F2 cohort along chromosomes 1–20. The Y-axis indicates the LOD score and the X-axis the chromosome number and genetic position. Horizontal lines represent the thresholds for suggestive ...

BBDP type 1 diabetes susceptibility loci.

At five of the eight type 1 diabetes loci (chromosomes 2, 14, 17, and both on 6), the BBDP allele conferred increased susceptibility to the disease (Table 1). The peak of linkage (LOD = 2.90; Iddm26) on chromosome 2 is located at marker D2Arb16 (Table 1), and the 1-LOD interval spans 16 cM (supplementary Table 1 and supplementary Fig. 3A). The incidence of type 1 diabetes in F2 animals that were BBDP homozygous and heterozygous at the peak marker was 24 and 22%, respectively, compared with 8% in ACI homozygous animals (Table 1). This locus overlaps with Iddm3, previously identified in a cross between DRlyp/lyp and F344 (8). Interestingly, the peak marker D2Arb16 is only 0.74 Mb away from Ptpn22 (199.05 Mb), a gene strongly associated with both type 1 diabetes and other autoimmune diseases in humans (2630) (Table 2).

TABLE 1
Peak markers at each locus linked to type 1 diabetes and related traits
TABLE 2
Murine and human chromosomal regions syntenic to Iddm loci

Two type 1 diabetes loci were located close to each other toward the telomere of chromosome 6 (Table 1, supplementary Table 1, and supplementary Fig. 3B). The more proximal locus (LOD = 5.21; Iddm28) had a 1-LOD interval of 20 cM between markers D6Rat23 and D6Got169. The more distal locus is the most significant linkage in this cross (LOD = 7.66; Iddm29), with a 1-LOD interval spanning 10 cM between markers D6Mgh3 and D6Got167. These two loci account for 4.5 and 6.3%, respectively, of the trait variance in this cross. The distal locus overlaps with Iddm8, previously identified in a cross-backcross between the BB/OK and SHR/Mol strains, subsequently confirmed through the development of congenic strains (9). Using composite interval mapping, we confirmed that these two chromosome 6 QTLs are distinct and we better defined their peak location 30 cM apart (supplementary Fig. 4).

Two other susceptibility loci on chromosomes 14 and 17 are also novel (supplementary Table 1 and supplementary Figs. 3C and D). The former (LOD = 4.7; Iddm31) has a 1-LOD interval spanning 12 cM between markers D14Rat1 and D14Mit6; however, this is possibly an underestimation of the true locus size since the peak is at the most centromeric marker along this chromosome. The latter (LOD = 1.98; Iddm36) has a 1-LOD interval of 30 cM.

BBDP type 1 diabetes resistance loci.

There are three loci where the BBDP allele confers resistance (or the ACI allele confers susceptibility) to type 1 diabetes in this cross (supplementary Table 1 and supplementary Figs. 5A–C). The strongest BBDP resistance locus (LOD = 4.18; Iddm25) maps to the telomere of chromosome 1 with a 1-LOD interval of 20 cM between markers D1Mit13 and D1Rat76. This locus overlaps with Iddm9/10 previously identified in crosses between the BB/OK and either DA or SHR/Mol strains (9). Strikingly, this locus contains the Ins-2 gene (202.93 Mb), one of two homologues of the human INS gene that is associated with human type 1 diabetes with the second highest odds ratio (31,32) (Table 2).

Two other resistance loci on chromosomes 3 and 12 are novel. Although the former showed significant linkage when diabetic F2 animals were analyzed, linkage was only suggestive (LOD = 2.68; Iddm27) when the whole cohort was included, with a 1-LOD interval spanning 19 cM between D3Rat24 and D3Rat114. The latter locus is the second most significant resistance locus (LOD = 3.82; Iddm30) and has a 1-LOD interval of 24 cM between markers D12Got11 and D12Rat15. C12orf30, one of the loci associated with human type 1 diabetes (36.23 Mb), is located within this locus—though outside the 1-LOD interval (18) (Table 2).

Linkage analysis of insulitis.

Two subphenotypes of insulitis were assessed in the diabetic and nondiabetic F2 progeny: the intensity of islet infiltration by inflammatory cells and the level of islet integrity. The distribution of these two subphenotypes was very different between the diabetic and nondiabetic F2 animals (Fig. 3). Specifically, while the vast majority of nondiabetic animals had intact islets and no cellular infiltration, all diabetic animals exhibited a mixture of islet atrophy and islet inflammation. Importantly, in both cohorts of diabetic and nondiabetic animals, there was a strong correlation (P < 0.0001, Spearman's rank correlation) between islet integrity and inflammation. However, while this correlation was negative in the nondiabetic F2 animals, where islets exhibiting some degree of inflammation tended to have decreased integrity, it was positive in diabetic F2 animals. This observation in diabetic F2 animals most likely reflects the progressive disappearance of inflammatory cells from islets as their β-cells are destroyed (Fig. 3). Of note, though comparison of pancreatic lesions in nondiabetic male and female F2 animals revealed significantly (P = 0.0013) more severe islet atrophy in females (data not shown), no evidence for significant QTL × sex interaction was found for either insulitis trait.

FIG. 3.
Distribution of insulitis subphenotype scores in diabetic and nondiabetic F2 animals. A, C, and E: Data from diabetic animals. B, D, and F: Data from nondiabetic animals. In graphs A–D, the vertical axis indicates the number of animals, while ...

It is important to note that while the diabetic F2 animals were genotyped across the whole genome, nondiabetic animals were only genotyped at regions that at some point during the study showed evidence for linkage to type 1 diabetes. Consequently, nondiabetic animals only contributed to linkage analysis at these regions. In most cases, the loci controlling the two insulitis subphenotypes overlap with those described above that influence type 1 diabetes susceptibility.

Cellular infiltration.

Three of the loci found to significantly influence cellular infiltration of pancreatic islets overlap with type 1 diabetes loci on chromosomes 6 (LOD = 6.78), 12 (LOD = 6.57), and 14 (LOD = 3.79), as illustrated in supplementary Figs. 3B, 5C, and 3C, respectively, while one novel locus on chromosome 8 (LOD = 2.63) is unlinked to type 1 diabetes (Table 1) and, importantly, regulates cellular infiltration of islets in diabetic F2 animals only (supplementary Table 1, Fig. 2, and supplementary Fig. 6). At the peak of those loci regulating both cellular infiltration and type 1 diabetes, the BBDP allele has consistent susceptibility (chromosomes 6 and 14) or resistance (chromosome 12) effects on the two traits (Table 1). At the novel chromosome 8 locus, found in diabetic F2 animals only, the BBDP allele is associated with a decreased cellular infiltration (Table 1), possibly due to its concomitant deleterious effect on islet integrity hence on the disappearance of β-cells (see below and Fig. 3E).

Islet integrity.

The vast majority of the loci linked to islet integrity are also linked to type 1 diabetes in this cross (Fig. 2 and Table 1). The seven loci showing significant linkage (supplementary Figs. 3–6) map to chromosomes 1 (LOD = 4.19), 6 (LOD = 4.55 and 7.54 for the centromeric and telomeric loci, respectively), 8 (LOD = 3.75), 12 (LOD = 4.56), 14 (LOD = 3.74), and 17 (LOD = 3.28). Other loci on chromosomes 2, 3, and 16 are suggestively linked to islet integrity in this cross (Table 1 and Fig. 2). Again, at all of these loci, the protective or deleterious effect of the BBDP allele was consistent with its effect on type 1 diabetes, conferring resistance to type 1 diabetes and relative protection from islet atrophy on chromosomes 1, 3, and 12 as opposed to increased susceptibility to type 1 diabetes and islet atrophy on chromosomes 2, 6, 8, 14, and 17 (Table 1 and supplementary Table 1).

Linkage analysis of AOO of diabetes.

Four chromosomal regions on chromosomes 2, 7, and 18 were found to influence the age at which type 1 diabetes develops in F2 animals (Fig. 2 and supplementary Fig. 7). Both chromosome 2 AOO loci (LOD = 2.36, Iddm32; and LOD = 2.98, Iddm33 for the centromeric and telomeric loci, respectively) overlap with two suggestive loci controlling islet integrity (Table 1 and supplementary Table 1). As expected, based on the deleterious effect of the BBDP alleles on islet integrity at both loci, these alleles are also associated with early onset of type 1 diabetes (Table 1). Importantly, Il-2 and Il-21, which are associated with type 1 diabetes in both humans and NOD mice, are located within the Iddm32 locus (Table 2) (18,33). The two AOO loci on chromosomes 7 (LOD = 2.71; Iddm34) and 18 (LOD = 2.79; Iddm35) did not overlap with any type 1 diabetes or insulitis loci (supplementary Table 1). However, the former locus contains C1qtnf6, recently found to be associated with human type 1 diabetes by meta-analysis of genome-wide association studies (34). While BBDP homozygosity at the peak of the chromosome 7 locus was associated with early type 1 diabetes development, at the peak of the chromosome 18 locus it delayed onset of the disease (Table 1). Of note, although a type 1 diabetes chromosome 18 locus overlapping with Iddm35 was identified in a cross between BB/OK and SHR/Mol rats (9), it was not linked to AOO, and the BB/OK allele conferred increased susceptibility to type 1 diabetes (35).

DISCUSSION

This study has identified no less than 12 loci designated Iddm25 to Iddm36, influencing the diabetogenic process in a single cross, thus establishing the polygenic basis of the disease in the BBDP rat. Six of these loci on chromosomes 1, 6 (two loci), 12, 14, and 17 showed significant linkage to type 1 diabetes and/or insulitis, while six on chromosomes 2 (three loci), 3, 7, and 18 reached suggestive linkage to type 1 diabetes or AOO. The BBDP, KDP, and LEW.1AR1 rat strains spontaneously develop type 1 diabetes, and it initially appeared that these syndromes had a relatively simple genetic basis with perhaps few susceptibility genes (36,37). However, the current study demonstrates that this does not apply to an intercross between the BBDP and ACI.BB1u.lyp strains used here. Strikingly, although the type 1 diabetes loci mapped here are numerous, together they only account for <30% of the trait variance observed in the F2 cohort. From the study of Klaff et al. (8), we calculated that Iddm1 and -2 jointly contributed 9.4% of the variance of type 1 diabetes in a cross between the type 1 diabetes–susceptible DRlyp/lyp and type 1 diabetes–resistant F344 strains. Assuming that the contribution of these two genes to type 1 diabetes variance would be similar in a cross-intercross between BBDP and noncongenic ACI rats, our results strongly suggest a genetic complexity for the BBDP diabetic syndrome that is reminiscent of the human disease.

Genetic analyses using the BBDP strain previously performed by others made us aware of the very low number (<1%) of diabetic (hence informative) animals that could be expected in such a classical F2 cohort. Therefore, we reasoned that a large “conditioned” F2 cross with the two major Iddm loci fixed as BBDP would produce more informative animals. Consequently, several loci, including some contributing small effects to the disease, could be mapped. Furthermore, this would allow us to assess the contribution of all three genotypes to the diabetogenic process. However, a limitation of using a conditioned cross is that it could not detect genetic interaction between Gimap5 or RT1u and any of the other loci detected in our linkage analysis.

Having found multiple type 1 diabetes loci, the question follows as to why only three of these on chromosomes 1, 2, and 6 were identified in previous studies (8,9). The first obvious explanation is the extent of genetic polymorphism between BB rats and the type 1 diabetes–resistant strains used. In this regard, using recent information on genome-wide single nucleotide polymorphisms (SNPs) available from the rat genome database (http://rgd.mcw.edu/), we found that the BB genotype differed from the ACI, BN, F344, SHR, and WF strain genotypes at 30, 53, 27, 36, and 23% of the SNPs, respectively. Since the genetic polymorphism distinguishing the ACI from the BB strain is not the most extensive, it is unlikely to be the main explanation for the detection of many type 1 diabetes loci in this cross. Rather, this suggests that the type 1 diabetes–resistant strains used in previous studies had distinct combinations of type 1 diabetes resistance and susceptibility alleles compared with the ACI.BB1u.lyp strain. In support of this, the current F2 cohort could not map two type 1 diabetes loci on chromosomes 4 and 18 that were identified independently using distinct type 1 diabetes–resistant strains, though these loci could segregate in our cross (9,13). Further, it is perhaps surprising that the chromosome 2 (Iddm3) locus was previously identified with a much higher LOD score in a relatively small backcross using the type 1 diabetes–resistant F344 strain (8). It is possible that this high LOD score was due to allelic heterogeneity resulting in a differential effect depending on the resistant strain used and/or the close proximity of two susceptibility loci, only one of which segregates in our cross. This is supported by the slightly more centromeric (~20 cM) location of the peak of linkage to type 1 diabetes on chromosome 2 compared with Iddm3 (8).

A second likely explanation for the detection of multiple type 1 diabetes loci in this cross is its size and its “conditioned” nature, both resulting in a number of diabetic, hence informative, animals significantly larger than in previous studies (811). This size-related increase in the power of our study may also have benefited from the use of an intercross as opposed to a backcross, since it allowed the identification of three ACI-susceptible (or BBDP resistance) loci.

The observation of a higher cumulative incidence of type 1 diabetes in female F2 animals was unexpected since contrary to the NOD mouse, the BBDP rat does not exhibit a sex bias in disease susceptibility. This differential, sex-related type 1 diabetes risk in the F2 cohort was, however, minimal, and this could explain our inability to detect loci responsible for the higher disease incidence in females. This raises the possibility of an environmental risk factor for females. Paradoxically, the BBDP susceptibility allele at the only locus showing interaction with sex (chromosome 6) confers higher risk for the disease in males.

A successful approach to map type 1 diabetes loci has taken advantage of the widespread susceptibility to experimentally induced diabetic syndromes among nonlymphopenic, RT1u strains of rats (1316). Linkage analyses of crosses between these and RT1u, but type 1 diabetes–resistant, strains identified some type 1 diabetes loci including Iddm14 on chromosome 4 that influences both spontaneous and experimentally induced type 1 diabetes (14,15). Of two other loci on chromosomes 13 and 17 previously shown to control experimentally induced type 1 diabetes, only the latter segregates with spontaneous type 1 diabetes in our cross (13,38). Again, this could reflect genetic polymorphism between the type 1 diabetes–resistant WF and ACI.BB1u.lyp strains used in these studies. Alternatively, it could indicate that some of the loci influencing spontaneous and experimentally induced diabetic syndromes (hence, some of the pathogenic mechanisms involved in these syndromes) are distinct.

Recent genome-wide association studies have successfully identified multiple novel, small-effect genes associated with type 1 diabetes in humans (1719,34). This identification should facilitate our understanding of antipancreatic autoimmunity at the molecular level and, subsequently, the design of therapeutic approaches interfering with these mechanisms. However, the small effect of these genes has highlighted the importance of type 1 diabetes–prone animals sharing disease susceptibility genes or signaling pathways with their human counterparts. Specifically, they could identify additional genetic risk factors, reveal pathways of previously unknown relevance to the disease, and provide insights into pathogenic mechanisms controlled by shared susceptibility genes. The relevance of type 1 diabetes–prone animals to the human disease has been illustrated by the NOD mouse, which shares several type 1 diabetes genes and signaling pathways with type 1 diabetes in humans (33,39,40). Similarly, the presence of five genes associated with the human disease within the Iddm loci identified in this cross (Table 2) may facilitate a better understanding of their contribution to antipancreatic autoimmunity.

Another contribution of experimental models is that they are accessible to the dissection of disease-related subphenotypes that are difficult to assess in humans. For example, the linkage analysis of insulitis subphenotypes leads to both expected and unexpected results. It is generally accepted that the outcome of chronic islet inflammation is the destruction of β-cells and the subsequent development of type 1 diabetes. It is therefore not surprising that insulitis and type 1 diabetes loci overlap and that genotypes increasing type 1 diabetes susceptibility are associated with both intense inflammatory infiltrates in islets and severe islet atrophy (e.g., chromosomes 6 [telomere], 12, and 14 [Table 1]). More surprising is the observation that some type 1 diabetes loci (e.g., on chromosomes 1, 2, 6 [centromere], and 17) influence islet integrity but do not seem to control islet infiltration by immune cells. At the moment, we can speculate that these loci may control very early steps of the autoimmune response (e.g., they may influence the immunogenicity of β-cells). Alternatively, they may regulate an immune mechanism that destroys islets independently of the intensity of the local cellular inflammation or the intrinsic ability of β-cells to succumb to the inflammatory response.

The incidence of type 1 diabetes is increasing in the Western world, most rapidly in individuals with onset before the age of 5 years (41). Importantly, early-onset type 1 diabetes is associated with an aggressive course characterized by rapid disappearance of residual β-cell function and a higher risk for preclinical (as assessed by autoantibodies) and overt disease in siblings and parents (42,43). There is strong evidence that AOO is under genetic control (44). Some of the genes associated with AOO have been identified in humans, which are all MHC class I and II genes (17,42,4547). However, genome-wide association studies have not identified novel AOO genes, possibly due to the fact that many cohorts restricted inclusion to pediatric onset (18,19,48). Our study identifies four loci that appear to influence AOO (though unlinked to type 1 diabetes itself) suggesting that some of these allelic variations may act upstream of those affecting insulitis and type 1 diabetes susceptibility, possibly at the level of the differentiation of the target tissue and/or components of the immune system.

Supplementary Material

Online-Only Appendix:

Acknowledgments

This work was supported by grants from Genome Canada through the Ontario Genomics Institute, the Canadian Institutes of Health Research (MOP77713), and the Ontario Research Foundation. R.H.W. is supported by a postdoctoral fellowship award from the Canadian Diabetes Association. J.S. is supported by a scholarship award from the Canadian Institutes of Health Research. G.C. holds a studentship award from the Banting and Best Diabetes Centre in Toronto. A.D.P. holds a Canada Research Chair in the Genetics of Complex Diseases.

No potential conflicts of interest relevant to this article were reported.

Footnotes

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

See accompanying original article, p. 796.

REFERENCES

1. Mordes JP, Poussier P, Blankenhorn EP, Greiner DL.: Rat Models of Type 1 Diabetes: Genetics, Environment and Autoimmunity Boca Raton, FL, CRC Press, 2007 [PubMed]
2. MacMurray AJ, Moralejo DH, Kwitek AE, Rutledge EA, Van Yserloo B, Gohlke P, Speros SJ, Snyder B, Schaefer J, Bieg S, Jiang J, Ettinger RA, Fuller J, Daniels TL, Pettersson A, Orlebeke K, Birren B, Jacob HJ, Lander ES, Lernmark A.: Lymphopenia in the BB rat model of type 1 diabetes is due to a mutation in a novel immune-associated nucleotide (Ian)-related gene. Genome Res 12: 1029– 1039, 2002. [PubMed]
3. Hornum L, Romer J, Markholst H.: The diabetes-prone BB rat carries a frameshift mutation in Ian4, a positional candidate of Iddm1. Diabetes 51: 1972– 1979, 2002. [PubMed]
4. Colle E, Guttmann RD, Seemayer T.: Spontaneous diabetes mellitus syndrome in the rat: I. association with the major histocompatibility complex. J Exp Med 154: 1237– 1242, 1981. [PMC free article] [PubMed]
5. Hellquist A, Zucchelli M, Kivinen K, Saarialho-Kere U, Koskenmies S, Widen E, Julkunen H, Wong A, Karjalainen-Lindsberg ML, Skoog T, Vendelin J, Cunninghame-Graham DS, Vyse TJ, Kere J, Lindgren CM.: The human GIMAP5 gene has a common polyadenylation polymorphism increasing risk to systemic lupus erythematosus. J Med Genet 44: 314– 321, 2007. [PMC free article] [PubMed]
6. Shin JH, Janer M, McNeney B, Blay S, Deutsch K, Sanjeevi CB, Kockum I, Lernmark A, Graham J, Arnqvist H, Bjorck E, Eriksson J, Nystrom L, Ohlson LO, Schersten B, Ostman J, Aili M, Baath LE, Carlsson E, Edenwall H, Forsander G, Granstrom BW, Gustavsson I, Hanas R, Hellenberg L, Hellgren H, Holmberg E, Hornell H, Ivarsson SA, Johansson C, Jonsell G, Kockum K, Lindblad B, Lindh A, Ludvigsson J, Myrdal U, Neiderud J, Segnestam K, Sjoblad S, Skogsberg L, Stromberg L, Stahle U, Thalme B, Tullus K, Tuvemo T, Wallensteen M, Westphal O, Aman J.: IA-2 autoantibodies in incident type I diabetes patients are associated with a polyadenylation signal polymorphism in GIMAP5. Genes Immun 8: 503– 512, 2007. [PubMed]
7. Guttmann RD, Colle E, Michel F, Seemayer T.: Spontaneous diabetes mellitus syndrome in the rat: II. T lymphopenia and its association with clinical disease and pancreatic lymphocytic infiltration. J Immunol 130: 1732– 1735, 1983. [PubMed]
8. Klaff LS, Koike G, Jiang J, Wang Y, Bieg S, Pettersson A, Lander E, Jacob H, Lernmark A.: BB rat diabetes susceptibility and body weight regulation genes colocalize on chromosome 2. Mamm Genome 10: 883– 887, 1999. [PubMed]
9. Kloting I, Schmidt S, Kovacs P.: Mapping of novel genes predisposing or protecting diabetes development in the BB/OK rat. Biochem Biophys Res Commun 245: 483– 486, 1998. [PubMed]
10. Kloting I, van den Brandt J, Kloting N, Radovic B.: Alleles of diabetes-resistant BN rats contribute to insulin-dependent type 1 diabetes mellitus. J Autoimmun 20: 119– 123, 2003. [PubMed]
11. Kloting I, Vogt L, Serikawa T.: Locus on chromosome 18 cosegregates with diabetes in the BB/OK rat subline. Diabetes Metab 21: 338– 344, 1995 [PubMed]
12. Jackerott M, Hornum L, Andreasen BE, Markholst H.: Segregation of autoimmune type 1 diabetes in a cross between diabetic BB and brown Norway rats. J Autoimmun 10: 35– 41, 1997. [PubMed]
13. Martin AM, Blankenhorn EP, Maxson MN, Zhao M, Leif J, Mordes JP, Greiner DL.: Non-major histocompatibility complex-linked diabetes susceptibility loci on chromosomes 4 and 13 in a backcross of the DP-BB/Wor rat to the WF rat. Diabetes 48: 50– 58, 1999. [PubMed]
14. Martin AM, Maxson MN, Leif J, Mordes JP, Greiner DL, Blankenhorn EP.: Diabetes-prone and diabetes-resistant BB rats share a common major diabetes susceptibility locus, iddm4: additional evidence for a “universal autoimmunity locus” on rat chromosome 4. Diabetes 48: 2138– 2144, 1999. [PubMed]
15. Fuller JM, Kwitek AE, Hawkins TJ, Moralejo DH, Lu W, Tupling TD, Macmurray AJ, Borchardt G, Hasinoff M, Lernmark A.: Introgression of F344 rat genomic DNA on BB rat chromosome 4 generates diabetes-resistant lymphopenic BB rats. Diabetes 55: 3351– 3357, 2006. [PubMed]
16. Ellerman KE, Like AA.: Susceptibility to diabetes is widely distributed in normal class IIu haplotype rats. Diabetologia 43: 890– 898, 2000. [PubMed]
17. Nejentsev S, Howson JM, Walker NM, Szeszko J, Field SF, Stevens HE, Reynolds P, Hardy M, King E, Masters J, Hulme J, Maier LM, Smyth D, Bailey R, Cooper JD, Ribas G, Campbell RD, Clayton DG, Todd JA.: Localization of type 1 diabetes susceptibility to the MHC class I genes HLA-B and HLA-A. Nature 450: 887– 892, 2007. [PMC free article] [PubMed]
18. Todd JA, Walker NM, Cooper JD, Smyth DJ, Downes K, Plagnol V, Bailey R, Nejentsev S, Field SF, Payne F, Lowe CE, Szeszko JS, Hafler JP, Zeitels L, Yang JH, Vella A, Nutland S, Stevens HE, Schuilenburg H, Coleman G, Maisuria M, Meadows W, Smink LJ, Healy B, Burren OS, Lam AA, Ovington NR, Allen J, Adlem E, Leung HT, Wallace C, Howson JM, Guja C, Ionescu-Tirgoviste C, Simmonds MJ, Heward JM, Gough SC, Dunger DB, Wicker LS, Clayton DG.: Robust associations of four new chromosome regions from genome-wide analyses of type 1 diabetes. Nat Genet 39: 857– 864, 2007. [PMC free article] [PubMed]
19. WTCCC: Genome-wide association study of 14,000 cases of seven common diseases and 3,000 shared controls. Nature 447: 661– 678, 2007. [PMC free article] [PubMed]
20. Colle E, Fuks A, Poussier P, Edouard P, Guttmann RD.: Polygenic nature of spontaneous diabetes in the rat. Permissive MHC haplotype and presence of the lymphopenic trait of the BB rat are not sufficient to produce susceptibility. Diabetes 41: 1617– 1623, 1992. [PubMed]
21. Lander ES, Botstein D.: Mapping mendelian factors underlying quantitative traits using RFLP linkage maps. Genetics 121: 185– 199, 1989. [PubMed]
22. Broman KW, Wu H, Sen S, Churchill GA.: R/qtl: QTL mapping in experimental crosses. Bioinformatics 19: 889– 890, 2003. [PubMed]
23. Wang S, Basten CJ, Zeng Z-B.: Windows QTL Cartographer 2.5 Raleigh, NC, Department of Statistics, North Carolina State University, 2005
24. Churchill GA, Doerge RW.: Empirical threshold values for quantitative trait mapping. Genetics 138: 963– 971, 1994. [PubMed]
25. Ramanathan S, Bihoreau MT, Paterson AD, Marandi L, Gauguier D, Poussier P.: Thymectomy and radiation-induced type 1 diabetes in nonlymphopenic BB rats. Diabetes 51: 2975– 2981, 2002. [PubMed]
26. Chelala C, Duchatelet S, Joffret ML, Bergholdt R, Dubois-Laforgue D, Ghandil P, Pociot F, Caillat-Zucman S, Timsit J, Julier C.: PTPN22 R620W functional variant in type 1 diabetes and autoimmunity related traits. Diabetes 56: 522– 526, 2007. [PubMed]
27. Kyogoku C, Langefeld CD, Ortmann WA, Lee A, Selby S, Carlton VE, Chang M, Ramos P, Baechler EC, Batliwalla FM, Novitzke J, Williams AH, Gillett C, Rodine P, Graham RR, Ardlie KG, Gaffney PM, Moser KL, Petri M, Begovich AB, Gregersen PK, Behrens TW.: Genetic association of the R620W polymorphism of protein tyrosine phosphatase PTPN22 with human SLE. Am J Hum Genet 75: 504– 507, 2004. [PubMed]
28. Begovich AB, Carlton VE, Honigberg LA, Schrodi SJ, Chokkalingam AP, Alexander HC, Ardlie KG, Huang Q, Smith AM, Spoerke JM, Conn MT, Chang M, Chang SY, Saiki RK, Catanese JJ, Leong DU, Garcia VE, McAllister LB, Jeffery DA, Lee AT, Batliwalla F, Remmers E, Criswell LA, Seldin MF, Kastner DL, Amos CI, Sninsky JJ, Gregersen PK.: A missense single-nucleotide polymorphism in a gene encoding a protein tyrosine phosphatase (PTPN22) is associated with rheumatoid arthritis. Am J Hum Genet 75: 330– 337, 2004. [PubMed]
29. Bottini N, Musumeci L, Alonso A, Rahmouni S, Nika K, Rostamkhani M, MacMurray J, Meloni GF, Lucarelli P, Pellecchia M, Eisenbarth GS, Comings D, Mustelin T.: A functional variant of lymphoid tyrosine phosphatase is associated with type I diabetes. Nat Genet 36: 337– 338, 2004. [PubMed]
30. Vang T, Congia M, Macis MD, Musumeci L, Orru V, Zavattari P, Nika K, Tautz L, Tasken K, Cucca F, Mustelin T, Bottini N.: Autoimmune-associated lymphoid tyrosine phosphatase is a gain-of-function variant. Nat Genet 37: 1317– 1319, 2005. [PubMed]
31. Bell GI, Horita S, Karam JH.: A polymorphic locus near the human insulin gene is associated with insulin-dependent diabetes mellitus. Diabetes 33: 176– 183, 1984. [PubMed]
32. Lucassen AM, Julier C, Beressi JP, Boitard C, Froguel P, Lathrop M, Bell JI.: Susceptibility to insulin dependent diabetes mellitus maps to a 4.1 kb segment of DNA spanning the insulin gene and associated VNTR. Nat Genet 4: 305– 310, 1993. [PubMed]
33. Yamanouchi J, Rainbow D, Serra P, Howlett S, Hunter K, Garner VE, Gonzalez-Munoz A, Clark J, Veijola R, Cubbon R, Chen SL, Rosa R, Cumiskey AM, Serreze DV, Gregory S, Rogers J, Lyons PA, Healy B, Smink LJ, Todd JA, Peterson LB, Wicker LS, Santamaria P.: Interleukin-2 gene variation impairs regulatory T cell function and causes autoimmunity. Nat Genet 39: 329– 337, 2007. [PMC free article] [PubMed]
34. Cooper JD, Smyth DJ, Smiles AM, Plagnol V, Walker NM, Allen JE, Downes K, Barrett JC, Healy BC, Mychaleckyj JC, Warram JH, Todd JA.: Meta-analysis of genome-wide association study data identifies additional type 1 diabetes risk loci. Nat Genet 2008 [PMC free article] [PubMed]
35. Kloting I, van den Brandt J, Kuttler B.: Genes of SHR rats protect spontaneously diabetic BB/OK rats from diabetes: lessons from congenic BB.SHR rat strains. Biochem Biophys Res Commun 283: 399– 405, 2001. [PubMed]
36. Yokoi N, Komeda K, Wang HY, Yano H, Kitada K, Saitoh Y, Seino Y, Yasuda K, Serikawa T, Seino S.: Cblb is a major susceptibility gene for rat type 1 diabetes mellitus. Nat Genet 31: 391– 394, 2002. [PubMed]
37. Weiss H, Bleich A, Hedrich HJ, Kolsch B, Elsner M, Jorns A, Lenzen S, Tiedge M, Wedekind D.: Genetic analysis of the LEW. 1AR1-iddm rat: an animal model for spontaneous diabetes mellitus. Mamm Genome 16: 432– 441, 2005. [PubMed]
38. Blankenhorn EP, Rodemich L, Martin-Fernandez C, Leif J, Greiner DL, Mordes JP.: The rat diabetes susceptibility locus Iddm4 and at least one additional gene are required for autoimmune diabetes induced by viral infection. Diabetes 54: 1233– 1237, 2005. [PMC free article] [PubMed]
39. Ueda H, Howson JM, Esposito L, Heward J, Snook H, Chamberlain G, Rainbow DB, Hunter KM, Smith AN, Di Genova G, Herr MH, Dahlman I, Payne F, Smyth D, Lowe C, Twells RC, Howlett S, Healy B, Nutland S, Rance HE, Everett V, Smink LJ, Lam AC, Cordell HJ, Walker NM, Bordin C, Hulme J, Motzo C, Cucca F, Hess JF, Metzker ML, Rogers J, Gregory S, Allahabadia A, Nithiyananthan R, Tuomilehto-Wolf E, Tuomilehto J, Bingley P, Gillespie KM, Undlien DE, Ronningen KS, Guja C, Ionescu-Tirgoviste C, Savage DA, Maxwell AP, Carson DJ, Patterson CC, Franklyn JA, Clayton DG, Peterson LB, Wicker LS, Todd JA, Gough SC.: Association of the T-cell regulatory gene CTLA4 with susceptibility to autoimmune disease. Nature 423: 506– 511, 2003. [PubMed]
40. Lund T, O'Reilly L, Hutchings P, Kanagawa O, Simpson E, Gravely R, Chandler P, Dyson J, Picard JK, Edwards A, et al. : Prevention of insulin-dependent diabetes mellitus in non-obese diabetic mice by transgenes encoding modified I-A beta-chain or normal I-E alpha-chain. Nature 345: 727– 729, 1990. [PubMed]
41. EURODIAB ACE Study Group: Variation and trends in incidence of childhood diabetes in Europe. Lancet 355: 873– 876, 2000. [PubMed]
42. Gillespie KM, Gale EA, Bingley PJ.: High familial risk and genetic susceptibility in early-onset childhood diabetes. Diabetes 51: 210– 214, 2002. [PubMed]
43. Harjutsalo V, Podar T, Tuomilehto J.: Cumulative incidence of type 1 diabetes in 10,168 siblings of Finnish young-onset type 1 diabetic patients. Diabetes 54: 563– 569, 2005. [PubMed]
44. Fava D, Gardner S, Pyke D, Leslie RD.: Evidence that the age at diagnosis of IDDM is genetically determined. Diabetes Care 21: 925– 929, 1998. [PubMed]
45. Valdes AM, Erlich HA, Noble JA.: Human leukocyte antigen class I B and C loci contribute to type 1 diabetes (T1D) susceptibility and age at T1D onset. Hum Immunol 66: 301– 313, 2005. [PMC free article] [PubMed]
46. Valdes AM, Thomson G, Erlich HA, Noble JA.: Association between type 1 diabetes age of onset and HLA among sibling pairs. Diabetes 48: 1658– 1661, 1999. [PubMed]
47. Caillat-Zucman S, Garchon HJ, Timsit J, Assan R, Boitard C, Djilali-Saiah I, Bougneres P, Bach JF.: Age-dependent HLA genetic heterogeneity of type 1 insulin-dependent diabetes mellitus. J Clin Invest 90: 2242– 2250, 1992. [PMC free article] [PubMed]
48. Hakonarson H, Grant SF, Bradfield JP, Marchand L, Kim CE, Glessner JT, Grabs R, Casalunovo T, Taback SP, Frackelton EC, Lawson ML, Robinson LJ, Skraban R, Lu Y, Chiavacci RM, Stanley CA, Kirsch SE, Rappaport EF, Orange JS, Monos DS, Devoto M, Qu HQ, Polychronakos C.: A genome-wide association study identifies KIAA0350 as a type 1 diabetes gene. Nature 448: 591– 594, 2007. [PubMed]

Articles from Diabetes are provided here courtesy of American Diabetes Association