An F344 rat congenic for BB/DP rat-derived diabetes susceptibility loci Iddm1 and Iddm2.

Abstract

The diabetes-prone BB rat (BB/DP) spontaneously develops an insulin-dependent diabetes resembling human Type 1 diabetes (previously named IDDM). As in Type 1 diabetes, the insulinproducing b-cells of the islets of Langerhans are targets for autoimmune destruction (Crisa et al. 1992). Two major diabetes susceptibility genes have been genetically positioned from this model: Iddm1 (Lyp) on rat chromosome (Chr) 4 and Iddm2 (RT1) on rat Chr 20 (Colle et al. 1981; Jacob et al. 1992), but several studies have defined additional loci involved in diabetes development (Kloting et al. 1995, 1998; Kloting and Kovacs 1998; Martin et al. 1999; Klaff et al. 1999). In the latter reference, a locus on Chr 2 (Iddm3), where the F344 allele protects against diabetes, was detected. Because the F344 rat also carries protective alleles of Iddm1 and -2, a precise positioning of this locus was not performed. We set out to produce an F344 rat congenic for the Iddm1 and -2 of BB/DP rat origin (F344.lypRT1). Such a strain will make precise positioning of Iddm3 (and other genes affecting diabetes) more feasible, since fewer animals need to be bred. The F344.LypRT1 strain originated in F2(F344 × BB/DP) rats that were backcrossed to F344. These animals were then intercrossed, and offspring were selected for homozygocity for the BB/DP alleles of Lyp and RT1 by FACS (described in Jackerott et al. 1997). At this point, marker-assisted selection was started (Wakeland et al. 1997) in addition to monitoring the two Iddmloci. Four rounds of backcross to F344 followed, with selection of heterozygotes for RT1 by FACS and for Lyp by PCR [via flanking markers D4Rat75 and D4Mit7 (Npy) (described in Hornum and Markholst 1999)]. These rats were designated “N6”F344.LypRT1, since six rounds of introgression had been performed. For marker-assisted selection, 86 microsatellites evenly distributed throughout the genome were typed in each generation, and animals with the highest number of markers of F344 origin were selected for further breeding. The longest distance between markers (or markers and chromosome ends) is 46 cM. “N6”F344.LypRT1 rats, with all 86 microsatellites of F344 origin, were finally intercrossed, and offspring homozygous for the BB/ DP-derived Iddm1and Iddm2-alleles form the basis of the new F344.LypRT1 congenic strain. The segment of BB/DP origin around Iddm1 on Chr 4 is delimited by D4Rat15 and D4Rat37 with map positions 29.4 cM and 46.5 cM on the SHRSP × BN rat genetic map from the Whitehead Institute (www.genome.wi.mit.edu/rat/public), making the maximal size of the BB/DP-derived segment 17.1 cM. The segment of BB/DP origin around Iddm2 on Chr 20 is delimited by the centromere end and D20Rat43 and -70, both of which map to position 15.7 cM on the FHH × ACI rat genetic map from the Whitehead Institute, making the maximal size of the BB/DP derived segment 15.7 cM. While the diabetes incidence in our BB/DP line is >90% with more than 99% of these becoming diabetic before the age of 120 days, none of 22 (0%) F344.LypRT1 congenic rats became diabetic at this age (p << 0.001), although kept under the same specific pathogen-free conditions. Insulitis was not detected among four F344.LypRT1 congenic rats at the age of 120 days. This clearly demonstrates that loci outside Iddm1 and -2 protect