Impaired blood pressure response in Btg2 mutant female rats

Abstract

Sex‐specific blood pressure loci were previously identified on chromosome 13 using SS‐BN13 congenic rat strains. Previous work identified Btg2 as a candidate gene for salt‐sensitive hypertension in female rats. Two small congenic SS‐BN13 lines were developed that differed by 23 Kb with only Btg2 residing within this region. The strain containing BN Btg2 allele had significantly higher mean arterial pressure (MAP) (186±3.1 mmHg) than SS Btg2allele in females (151±5.8mmHg). Subsequent qRT‐PCR analysis demonstrated significant difference in Btg2 expression between these strains in kidney cortex (Btg2BN: 1.8±0.2 fold change, Btg2SS: 0.4±.01). Based on this previous data we hypothesized that mutating Btg2BN would mitigate the elevated blood pressure response we had seen in rats with the Btg2BN allele. TALEN and CRISPR/Cas9 reagents were used to target exon 1 of Btg2 and generate three mutant lines on the SS‐BN13 congenic with Btg2BN allele: m7 (TALEN) with 44bp deletion, m11 (CRISPR/Cas9) with 3bp deletion and 1bp insertion, and m13 (CRISPR/Cas9) with 16 bp deletion. Following telemetry transmitter implantation, blood pressure was measured in male and female rats on low salt and during 21 days of 8% NaCl diet. On day 17 of high salt, urine was collected for determination of protein and microalbumin. After 21 days on high salt, female homozygous m7 mutant rats MAP was 8 mmHg higher compared to female wild type littermates (167±8mmHg vs. 175±7mmHg). Urinary microalbumin and protein measurement in these female rats demonstrated a similar trend both showing a 1.4 fold increase in homozygous compared to wild type (93±18mg/day vs.131±41mg/day, 135±21mg/day vs.186±54mg/day respectively). Normalized kidney weight showed a 6% increase in female homozygous versus wildtype animals (0.0118±0.0002g/g vs.0.0125±0.0004g/g). Male homozygous and wild type m7 rats were not different after 21 days of high salt for these phenotypes. Preliminary results in the m13 mutant Btg2 rats suggest similar trends in the female vs. male rats. Taken together, our hemodynamic, urinary and morphometric data suggest the m7 mutant Btg2 allele produces an impaired blood pressure and renal response in female rats.Support or Funding InformationFunding: The National Heart, Lung, and Blood Institute of the National Institutes of Health, R24HL114474