A 3-Mbp fragment on rat chromosome 1 affects susceptibility both to stroke and kidney injury under salt loading in the stroke-prone spontaneously hypertensive rat: a genetic approach using multiple congenic strains

Figures

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  Fig. 1.. Construction of subcongenic strains and evaluation of susceptibility to salt-induced stroke. (A) Genomic structure of the subcongenic strains. Each strain has a chr 1 fragment derived from the donor strain (stroke-resistant SHR/Izm) on the genetic background of the recipient strain (SHRSP/Izm). Congenic regions (homozygous for SHR alleles) are shown as closed columns. Vertical bars indicate regions including a recombination boundary. Pr1.31 and Pr1.10 were constructed in a previous study [4]. Pr1.101, Pr1.102, and Pr1.103 were newly constructed in the present study by backcrossing of Pr1.10 to SHRSP/Izm as described in the Materials and Methods. Numbers in parentheses indicate physical positions (Mbp) of SSR markers based on Rnor_5.0. (B) Drinking water was changed to 1% salt water from 12 weeks of age to evaluate stoke susceptibility under salt loading. The X-axis indicates the period in days from the start of salt loading. Initial sign-free ratios (upper panel) and survival ratios (lower panel) were determined as described in the Materials and Methods. #P<0.01 vs. SHRSP by log-rank test with Bonferroni correction (n=21 to 23; shown in parentheses). $P<0.01 vs. Pr1.31 by…

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  Fig. 2.. Blood pressure and heart rate. Systolic blood pressure (SBP), diastolic BP (DBP), and heart rate (HR) at 12 weeks of age were measured by tail-cuff method before starting urine collection experiments under salt loading (see Materials and Methods). #P<0.05 vs. SHRSP by one-way ANOVA with Dunnett’s post hoc test (n=7–8).

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  Fig. 3.. Gene expression of Kim-1 and Clu in the kidney with or without 1 week of salt loading. The expression of renal tubular injury markers was quantitatively examined in kidneys obtained from (A) non-salt-loaded control rats and (B) salt-loaded rats. The Kim-1 and Clu expression levels were standardized to β-actin expression as described in the Materials and Methods. #P<0.05 vs. SHRSP by one-way ANOVA with Dunnett’s post hoc test (n=5).

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  Fig. 4.. Evaluation for Zfp45L and Ceacam 19. (A) The expression level of Zfp45L relative to β-actin expression was examined by quantitative RT-PCR. cDNA solutions of non-salt-loaded control rats were used as templates for reactions (see Fig. 3). #P<0.05 vs. the other three strains (n=5). (B) DNA sequencing of the chr 1 region including the nonsynonymous variation at Ceacam 19 [4]. G (Ala) to C (Thr) substitution was observed between SHRSP/Izm and SHR/Izm. Note that Pr1.102 and Pr1.103 harbor the SHRSP and SHR alleles at this position. (C) Genomic structures of subcongenic strains based on the results of (A) and (B). Numbers in parentheses indicate the physical positions (Mbp) of six candidate genes [4] and SSR markers based on Rnor_5.0.