Abstract-The identification of any quantitative trait locus (QTL) via a genome scan is only the first step toward the ultimate goal of gene identification. The next step is the production of congenic strains by which the existence of a QTL may be verified and the implicated chromosomal region be reduced to a size applicable to positional cloning of the causal gene. We used a speed congenic breeding protocol previously verified in mice for 2 blood pressure QTLs on rat chromosome 2. Four congenic strains were produced through introgression of various segments of chromosome 2 from Wistar-Kyoto rats from Glasgow colonies [WKY (Gla) rats] into the recipient stroke-prone spontaneously hypertensive rats from Glasgow colonies [SHRSP (Gla) ], and vice versa. The number of backcross generations required for each strain to achieve complete homozygosity at 83 background genetic markers in a "best" male varied between 3 and 4. Transfer of the region of rat chromosome 2 containing both QTLs from WKY (Gla) into an SHRSP (Gla) genetic background lowered both baseline and salt-loaded systolic blood pressure by Ϸ20 and Ϸ40 mm Hg in male congenic rats compared with the SHRSP parental strain (Fϭ53.4, PϽ0.005; Fϭ28.0, PϽ 0.0005, respectively). In contrast, control animals for stowaway heterozygosity presented no deviation from the blood pressure values recorded for the SHRSP (Gla) , indicating that if such heterozygosity exists, its effect on blood pressure is negligible. A reciprocal strategy in which 1 or both QTLs on rat chromosome 2 were transferred from SHRSP (Gla) into a WKY (Gla) genetic background resulted in statistically significant but smaller blood pressure increases for 1 of these QTLs. These results confirm the existence of blood pressure QTLs on rat chromosome 2 and demonstrate the applicability of a speed congenic strategy in the rat and emphasize the important role of the genetic background. Key Words: hypertension, genetic Ⅲ genes Ⅲ rats, inbred strokeprone SHR G enome wide linkage analysis has proved to be successful during the past decade in the localization of large chromosomal regions containing putative quantitative trait loci (QTLs) for high blood pressure, 1-3 left ventricular hypertrophy, 3 and ischemic stroke 4 in F2 crosses derived from the stroke-prone spontaneously hypertensive rat (SHRSP) and its normotensive reference strain, the Wistar-Kyoto (WKY) rat. In particular, we previously identified 2 separate blood pressure QTLs on rat chromosome 2. 3 The most significant of these had its peak (logarithm of the odds [LOD] 3.6) close to the microsatellite marker D2Mit6 and was determined for both baseline and salt-loaded systolic and diastolic blood pressures in male and female F2 cohorts. The second QTL (LOD 3.1) was localized 73 cM from D2Mit6 and contributed to salt-loaded blood pressure in the male F2 cohort only.The identification of such a blood pressure QTL is only the first step toward the ultimate goal of gene identification, which can be achieved through genetic and physiological analyses of c...
Abstract-We have previously demonstrated that the SHRSP Y chromosome contains a locus that contributes to hypertension in SHRSP/WKY F2 hybrids and that SHRSP exhibit an increased vulnerability to focal cerebral ischemia after permanent middle cerebral artery occlusion (MCAO). This increased vulnerability is inherited as a codominant trait, and a putative role for the Y chromosome has been suggested in F1 hybrids. The objective of this study was to investigate further the role of Y chromosome in blood pressure (BP) regulation and in the vulnerability to cerebral ischemia. We have constructed consomic strains by selectively replacing the Y chromosome from WKY rats with that of SHRSP, and vice versa, by using a marker-assisted breeding strategy. Permanent MCAO was carried out by electrocoagulation, with infarct volume expressed as a percentage of the ipsilateral hemisphere. Systolic blood pressure was measured by radiotelemetry during a baseline period of 5 weeks followed by a 3-week period of salt loading. We observed that the transfer of the Y chromosome from WKY onto SHRSP background significantly reduced systolic BP in consomic strains, SP.WKYGlaY w (nϭ6) versus SHRSP (nϭ6) Key Words: hypertension Ⅲ stroke Ⅲ genetics Ⅲ SHRSP Ⅲ consomics Ⅲ focal cerebral ischemia Ⅲ middle cerebral artery occlusion T he stroke-prone spontaneously hypertensive rat (SHRSP) is generally regarded as a good experimental model of cerebrovascular disease and human essential hypertension. 1 Spontaneous strokes and the increased vulnerability to cerebral ischemia have been well documented in the SHRSP, and several quantitative trait loci (QTLs) for these phenotypes were published. 2,3 Ely and Turner 4 found that the blood pressure of F2 offspring depended on the strain of the male progenitor in a WKY x SHR cross. Male offspring with an SHR male progenitor had significantly higher pressures than male offspring with a WKY progenitor. The blood pressure of F2 males was compatible with a Y-linked effect on blood pressure. Reciprocal Y-consomic strains (SHR Y chromosome on WKY background and WKY Y chromosome on SHR background) were constructed and confirmed the Y chromosome effect on blood pressure. 5 Previous data from our laboratory described 143 F2 rats obtained by crossing SHRSP and WKY, which were phenotyped using a radiotelemetry system. In this study, male F2 hybrids with an SHRSP grandfather had significantly higher blood pressures compared with male F2 hybrids with the WKY grandfather, suggesting that the Y chromosome effect was also present in the SHRSP. 6 The SHRSP strain exhibits an increased frequency of spontaneous strokes and an increased volume of infarction after experimentally induced focal cerebral ischemia compared with the WKY reference strain. 2,3 To investigate the role of the SHRSP Y chromosome in stroke, we used permanent middle cerebral artery occlusion (MCAO) in an F1 reciprocal cross. 7,8 We found that F1 males with an SHRSP male progenitor had smaller infarct than those with a WKY
-Previous studies suggested that atrial natriuretic peptide gene (Anp) and brain natriuretic peptide gene (Bnp) are plausible candidate genes for susceptibility to stroke and for sensitivity to brain ischemia in the stroke-prone spontaneously hypertensive rat (SHRSP). We performed structural and functional analyses of these 2 genes in SHRSP from Glasgow colonies (SHRSPGla) and Wistar-Kyoto rats from Glasgow colonies (WKYGla) and developed a radiation hybrid map of the relevant region of rat chromosome 5. Sequencing of the coding regions of the Anp and Bnp genes revealed no difference between the 2 strains. Expression studies in brain tissue showed no differences at baseline and at 24 hours after middle cerebral artery occlusion. Plasma concentrations of atrial natriuretic peptide (ANP) did not differ between the SHRSPGla and WKYGla, whereas concentrations of brain natriuretic peptide were significantly higher in the SHRSPGla as compared with the WKYGla (n=11 to 14; 163+/-21 pg/mL and 78+/-14 pg/mL; 95% confidence interval 31 to 138, P=0.003). We did not detect any attenuation of endothelium-dependent relaxations to bradykinin or ANP in middle cerebral arteries from the SHRSPGla; indeed the sensitivity to ANP was significantly increased in arteries harvested from this strain (WKYGla: n=8; pD2=7. 3+/-0.2 and SHRSPGla: n=8; pD2=8.2+/-0.15; P<0.01). Moreover, radiation hybrid mapping and fluorescence in situ hybridization allowed us to map the Anf marker in the telomeric position of rat chromosome 5 in close proximity to D5Rat48, D5Rat47, D5Mgh15, and D5Mgh16. These results exclude Anp and Bnp as candidate genes for the sensitivity to brain ischemia and pave the way to further congenic and physical mapping strategies.
Experimental models of genetic hypertension are used to develop paradigms to study human essential hypertension while removing some of the complexity inherent in the study of human subjects. Since 1991 several quantitative trait loci responsible for blood pressure regulation have been identified in various rat crosses. More recently, a series of interesting quantitative trait loci influencing cardiac hypertrophy, stroke, metabolic syndrome and renal damage has also been described. It is recognized that the identification of large chromosomal regions containing a quantitative trait locus is only a first step towards gene identification. The next step is the production of congenic strains and substrains to confirm the existence of the quantitative trait locus and to narrow down the chromosomal region of interest. Several congenic strains have already been produced, with further refinement of the methodology currently in progress. The ultimate goal is to achieve positional cloning of the causal gene, a task which has so far been elusive. There are several areas of cross-fertilization between experimental and human genetics of hypertension, with a successful transfer of two loci directly from rats to humans and with new pharmacogenetic approaches which may be utilized in both experimental and clinical settings.
A genetic variant of the gene for the alpha(1)-isoform of Na(+)-K(+)-ATPase (Atp1a1) was suggested to be involved in the pathogenesis of salt hypertension in Dahl rats through altered Na(+):K(+) coupling ratio. We studied Na(+)-K(+) pump activity in erythrocytes of Dahl salt-sensitive (SS/Jr) rats in relation to plasma lipids and blood pressure (BP) and the linkage of polymorphic microsatellite marker D2Arb18 (located within intron 1 and exon 2 of Atp1a1 gene) with various phenotypes in 130 SS/Jr x SR/Jr F(2) rats. Salt-hypertensive SS/Jr rats had higher erythrocyte Na(+) content, enhanced ouabain-sensitive (OS) Na(+) and Rb(+) transport, and higher Na(+):Rb(+) coupling ratio of the Na(+)-K(+) pump. BP of F(2) hybrids correlated with erythrocyte Na(+) content, OS Na(+) extrusion, and OS Na(+):Rb(+) coupling ratio, but not with OS Rb(+) uptake. In F(2) hybrids there was a significant association indicating suggestive linkage (P < 0.005, LOD score 2.5) of an intragenic marker D2Arb18 with pulse pressure but not with mean arterial pressure or any parameter of Na(+)-K(+) pump activity (including its Na(+):Rb(+) coupling ratio). In contrast, plasma cholesterol, which was elevated in salt-hypertensive Dahl rats and which correlated with BP in F(2) hybrids, was also positively associated with OS Na(+) extrusion. The abnormal Na(+):K(+) stoichiometry of the Na(+)-K(+) pump is a consequence of elevated erythrocyte Na(+) content and suppressed OS Rb(+):K(+) exchange. In conclusion, abnormal cholesterol metabolism but not the Atp1a1 gene locus might represent an important factor for both high BP and altered Na(+)-K(+) pump function in salt-hypertensive Dahl rats.
The localisation of quantitative trait loci (QTL) is the first step towards gene identification. This is then verified by the construction of reciprocal congenic strains. The hypertensive SHRSP and normotensive WKY strains were used in a speed congenic approach to confirm the existence of a QTL on rat chromosome 2. Systolic baseline and salt loaded blood pressures were measured by radiotelemetry. Transfer of the chromosome 2 blood pressure QTL region from WKY into an SHRSP background significantly reduced blood pressure, with the increased significance at the salt loaded period, compared to the SHRSP. The reciprocal congenic blood pressure showed a significantly increased baseline systolic pressure compared to the WKY, with no change in significance at the salt loaded period. Thus we have successfully captured a gene(s) which contribute to blood pressure regulation in both congenic strains. This will facilitate further positional cloning of the causative genes first in this model and then in human essential hypertension.
54 Background Localisation of QTL using a genome wide scan strategy is the first step towards gene identification. This is followed by the construction of congenics by which the existence of the QTL can be verified. Congenic strains for chromosome 2 using the Dahl S as the recipient and WKY and MNS as donors, showed variable blood pressure (BP) depending on the donor alleles 1 . Methods We used a speed congenic approach to produce several congenic strains of chromosome 2 using the SHRSP Gla and the WKY Gla as recipients to test the effect of the genetic background of a given congenic on BP. In some strains, the region introgressed was identical to avoid confounding effects of additional QTLs. Two reciprocal control congenic strains were also produced to determine the presence of any passenger loci. Baseline systolic BP was measured by radiotelemetry. Results Four congenic and 2 parental strains were phenotyped (n=3-6). Transfer of the region of rat chromosome 2 from the WKY into a SHRSP background significantly lowered both day and night-time systolic BP by approximately 16 and 20 mm Hg respectively in male congenic rats compared to the SHRSP parental strains (p<0.005, 95%CI: 8.6-21.0 mm Hg; and p<0.005, 95%CI: 16.7-24.7 mm Hg). In contrast, transfer of the same region from the SHRSP into a WKY background significantly increased both day and night-time systolic BP by approximately 25 and 28 mm Hg, respectively in male congenic rats compared to the WKY parental strains (p<0.05, 95%CI: -49.2-2.0 mmHg; and p<0.05%, 95%CI: -61.2-6.1 mm Hg, respectively). Reciprocal control congenic strain showed no deviation from the BP recorded in the parental strain (Strain WKY.SPGla2b vs. WKY parental strain, 128.8±5.2 mm Hg vs. 139.7±5.7 mm Hg, p= ns; and strain SP.WKYGla2b vs. SHRSP parental strain, 178.0±5.0 mm Hg vs. 179.9±2.1 mm Hg, p= ns). Conclusions The results show the effect of a permissive background on BP and confirm that the genetic background chosen for a congenic strain has a significant effect on the BP phenotype. Moreover, this work sheds new light on the different epistatic or pleiotropic effects according to the genetic background chosen. 1. Deng AY et al . Hypertension 1997;30:199-202
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.