Candidate genes that determine response to salt in the stroke-prone spontaneously hypertensive rat: congenic analysis

The existence of blood pressure quantitative trait loci exaggerated by salt on rat chromosome 2 has been confirmed previously using congenic strains derived from stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar-Kyoto (WKY) rats. This study aimed to dissect the implicated chromosome 2 region and to identify candidate genes based on microarray expression profiling and real-time PCR. A marker-assisted breeding strategy generated congenic strains SP.WKYGla2a (D2Rat13-D2Rat157), SP.WKYGla2c* (D2Wox9-D2Mgh12), and SP.WKYGla2k (D2Mit21-D2Rat157) using SHRSP as the recipient and WKY as the donor strain. The SP.WKYGla2k strain contains a 10-cM congenic interval, which is encompassed within the larger (64-cM) SP.WKYGla2a congenic region. Salt-loaded systolic blood pressure, measured by radiotelemetry, was significantly lower in the SP.WKYGla2a and SP.WKYGla2k strains compared with SHRSP. Salt sensitivity in SP.WKYGla2c* was not significantly different from SHRSP. Exclusion mapping identified a 6-Mbp region harboring genes responsible for salt-sensitive blood pressure regulation. Microarray expression profiling was carried out in whole homogenized kidneys from parental and SP.WKYGla2a strains. Examination of expression data within the minimal congenic interval identified the positional candidates Edg1 and Vcam1, demonstrating significantly elevated renal RNA expression levels in the SHRSP compared with WKY and SP.WKYGla2a congenic strains. These results were confirmed by quantitative real-time PCR. DNA sequencing identified SNPs in both Edg1 and Vcam1 between SHRSP and WKY rats. In conclusion, we have identified a suggestive minimal interval encompassing a 6-Mbp region on rat chromosome 2. This region contains several physiological candidate genes for salt-sensitive hypertension in the SHRSP, including Edg1 and Vcam1, which are differentially expressed and lie on common and functionally important pathways.