OP.2A.02] FURTHER REDUCTION OF THE WIDTH OF STROKE-SUSCEPTIBILITY QUANTITATIVE TRAIT LOCI AND IDENTIFICATION OF CANDIDATE GENES IN THE STROKE-PRONE SPONTANEOUSLY HYPERTENSIVE RAT

Abstract

Objective: In the previous study, we identified two major quantitative trait loci (QTLs) for stroke susceptibility on chr 1 and 18 in the stroke-prone spontaneously hypertensive rat (SHRSP). Most of difference in stroke susceptibility between SHR and SHRSP was explained with these two QTLs. In this study, we attempted to narrow down the QTL regions, and to identify candidate genes in the target regions. Design and method: Sixteen subcongenic and double subcongenic strains were established to cover the QTLs on chr 1 and 18. Stroke susceptibility was evaluated by stroke latency under 1% salt loading. Sequence variations in coding regions of genes were analyzed on whole-genome sequence data of SHRSP and SHR, and confirmed with direct sequencing. Comprehensive gene expression analysis was done on kidney tissue from SHRSP, SHR and two double congenic strains for the QTLs on chr 1 and 18 with Rat GE 44K microarray slides (v.3, Agilent Technologies). The results were confirmed with RT-PCR. Results: Analysis of stroke latency in a double subcongenic strain confirmed that the QTLs for the stroke susceptibility were in a 68 Mbp region on chr 1 and a 20 Mbp region on chr 18. Analysis on subcongenic strains covering these regions suggested that the most probable candidate region on chr 1 was a 240 kbp fragment between D1Got80 and Rat33. Microarray analysis indicated that expression of 3 genes in this region differed significantly between SHRSP and SHR as well as between the two double congenic strains in which the QTLs on chr 1 and 18 were reciprocally exchanged. In addition, analysis on the whole-genome sequence data suggested that 2 genes in this candidate region harbored a missense variant between SHR and SHRSP. In contrast to the QTL on chr 1, it was difficult to narrow down the QTL on chr 18. Conclusions: In conclusion, we successfully narrowed down the QTL on chr 1 and identified several candidate genes in the region. The QTL on chr 18 might harbor multiple causative genes that made the analysis difficult.