Biomedical subjects

Howard J Jacob

Publications and source records attributed to Howard J Jacob.

At least 37 records · Page 2Linked to original sources

PubMed2005

Impact of genomics on research in the rat.

The need to translate genes to function has positioned the rat as an invaluable animal model for genomic research. The significant increase in genomic resources in recent years has had an immediate functional application in the rat. Many of the resources for translational research are already in place and are ready to be combined with the years of physiological knowledge accumulated in numerous rat models, which is the subject of this perspective. Based on the successes to date and the research projects under way to further enhance the infrastructure of the rat, we also project where research in the rat will be in the near future. The impact of the rat genome project has just started, but it is an exciting time with tremendous progress.

Animals↗

PubMed2005

Interaction between Rf-1 and Rf-4 quantitative trait loci increases susceptibility to renal damage in double congenic rats.

BACKGROUND: Five quantitative trait loci (QTLs), Rf-1 to Rf-5, were found in Fawn-Hooded hypertensive (FHH) rats influencing susceptibility to renal damage. Previously, we found that single transfer of the Rf-1 QTL from FHH rats onto the renal-resistant August x Copenhagen Irish (ACI) strain caused a small increase in renal susceptibility. To investigate the separate role of the Rf-4 QTL and its interaction with Rf-1, we generated a single congenic strain carrying Rf-4 and a double congenic carrying both Rf-1 and Rf-4. METHODS: Differences in renal susceptibility between ACI, Rf-1A, and Rf-4 single congenics and Rf-1A+4 double congenics were assessed using four different treatments: control (two-kidney), two-kidney with l-arginine analogue N-nitro-l-arginine methyl ester (L-NAME)-induced hypertension, unilateral nephrectomy, and unilateral nephrectomy + L-NAME. In separate experiments, renal blood flow (RBF) autoregulation was compared between two-kidney ACI and congenic rats. RESULTS: Compared to ACI, Rf-1A rats developed more renal damage, while Rf-4 rats did not. The most severe renal damage was found in the Rf-1A+4 double congenic rats. Analysis of variance (ANOVA) demonstrated a significant interaction between the Rf-1A and Rf-4 QTLs. The magnitude of the interaction varied with the type and duration of the treatment. The RBF autoregulation was impaired in Rf-1A single and Rf-1A+4 double congenics, while in Rf-4 single congenics it was similar to that of ACI controls. CONCLUSION: These findings indicate that the Rf-1 QTL directly influences renal susceptibility and autoregulation. In contrast, the Rf-4 QTL shows no direct effects, but significantly increases susceptibility to renal damage via an interaction with Rf-1.

Albuminuria↗

PubMed2005

Genome-wide scanning with SSLPs in the rat.

Genome-wide linkage analysis is a powerful tool for the identification of genes underlying single gene and complex genetic disorders. The most commonly used technique for performing genome wide scans for genetic studies in the rat is by analysis of simple sequence length polymorphism (SSLPs) or microsatellite markers. A sensitive and flexible method for high-throughput genotyping is described. Addition of an M13 tail to the SSLP primer eliminates the need for direct conjugation of the fluorescent dye to the primers, allowing for any combination of primer and fluorophor, and therefore for easy multiplexing of primers in the same reaction. With the use of three different dyes, it is possible to run more than five hundred genotypes in each run of the automatic sequencer. Automation in the fluorescent detection system and data tracking software for processing genotypes, contributes to the ability to genotype large number of samples rapidly and accurately.

Animals↗

PubMed2005

Genetic linkage and association of the growth hormone secretagogue receptor (ghrelin receptor) gene in human obesity.

The growth hormone secretagogue receptor (GHSR) (ghrelin receptor) plays an important role in the regulation of food intake and energy homeostasis. The GHSR gene lies on human chromosome 3q26 within a quantitative trait locus strongly linked to multiple phenotypes related to obesity and the metabolic syndrome. Because the biological function and location of the GHSR gene make it an excellent candidate gene, we tested the relation between common single nucleotide polymorphisms (SNPs) in the GHSR gene and human obesity. We performed a comprehensive analysis of SNPs, linkage disequilibrium (LD), and haplotype structure across the entire GHSR gene region (99.3 kb) in 178 pedigrees with multiple obese members (DNA of 1,095 Caucasians) and in an independent sample of the general population (MONICA Augsburg left ventricular hypertrophy substudy; DNA of 1,418 Caucasians). The LD analysis revealed a disequilibrium block consisting of five SNPs, consistent in both study cohorts. We found linkage among all five SNPs, their haplotypes, and BMI. Further, we found suggestive evidence for transmission disequilibrium for the minor SNP alleles (P < 0.05) and the two most common haplotypes with the obesity affection status ("susceptible" P = 0.025, "nonsusceptible" P = 0.045) in the family cohort using the family-based association test program. Replication of these findings in the general population resulted in stronger evidence for an association of the SNPs (best P = 0.00001) and haplotypes with the disease ("susceptible" P = 0.002, "nonsusceptible" P = 0.002). To our knowledge, these data are the first to demonstrate linkage and association of SNPs and haplotypes within the GHSR gene region and human obesity. This linkage, together with significant transmission disequilibrium in families and replication of this association in an independent population, provides evidence that common SNPs and haplotypes within the GHSR region are involved in the pathogenesis of human obesity.

Body Mass Index↗

PubMed2004

Mapping the genetic determinants of hypertension, metabolic diseases, and related phenotypes in the lyon hypertensive rat.

The complex nature of hypertension makes identifying the pathophysiology and its genetic contributions a challenging task. One powerful approach for the genetic dissection of blood pressure regulation is studying inbred rat models of hypertension, as they provide natural allele variants but reduced heterogeneity (both genetic and etiologic). Furthermore, the detailed physiologic studies to which the rat is amenable allow for the determination of intermediate phenotypes. We have performed a total genome scan in offspring of an F2 intercross between the Lyon hypertensive (LH) and Lyon normotensive rat strains to identify linkage of anthropometric, blood pressure, renal, metabolic, and endocrine phenotypes. Quantitative trait locus (QTL) regions involved in blood pressure regulation, end-stage organ damage, body and organ weight, and lipid metabolism in the LH rat were identified on chromosomes 1, 2, 3, 5, 7, 10, 13, and 17, with 2 phenotypes associated with the metabolic syndrome identified on chromosomes 1 and 17. Regions on chromosomes 2, 13, and 17 were revealed to be important for blood pressure regulation. Regions on chromosome 17 were found to significantly contribute to both metabolic homeostasis and blood pressure regulation; 2 aggregates of a total of 23 QTLs were identified, including several "intermediate phenotypes." These intermediate phenotypes may be used as closer surrogates to the mechanisms leading to hypertension and metabolic dysfunction in the LH rat.

Animals↗

PubMed2004

Transgenic rescue demonstrates involvement of the Ian5 gene in T cell development in the rat.

A single point mutation in a novel immune-associated nucleotide gene 5 (Ian5) coincides with severe T cell lymphopenia in BB rats. We used a transgenic rescue approach in lymphopenic BB-derived congenic F344.lyp/lyp rats to determine whether this mutation is responsible for lymphopenia and to establish the functional importance of this novel gene. A 150-kb P1 artificial chromosome (PAC) transgene harboring a wild-type allele of the rat Ian5 gene restored Ian5 transcript and protein levels, completely rescuing the T cell lymphopenia in the F344.lyp/lyp rats. This successful complementation provides direct functional evidence that the Ian5 gene product is essential for maintaining normal T cell levels. It also demonstrates that transgenic rescue in the rat is a practical and definitive method for revealing the function of a novel gene.

Animals↗

PubMed2004

Rat model of familial combined hyperlipidemia as a result of comparative mapping.

Total genome scan was carried out in 266 F2 intercrosses from the Prague hypertriglyceridemic (HTG) rat that shares several clinical characteristics with human metabolic syndrome. Two loci for plasma triglycerides (TG) were localized on chromosome 2 (Chr 2) (LOD 4.4, 3.2). The first locus overlapped with the rat syntenic region of the human locus for the metabolic syndrome and for small, dense LDL, while the second overlapped with the syntenic region of another locus for small, dense LDL in humans by the comparative mapping approach. Loci for TG on rat Chr 13 (LOD 3.3) and Chr 1 (LOD 2.7) overlapped with the syntenic region of loci for human familial combined hyperlipidemia (FCHL) in Finnish and Dutch populations, respectively. The concordances of loci for TG localized in this study with previously reported loci for FCHL and its related phenotypes are underlying the generalized importance of these loci in dyslipidemia. These data suggest the close relationship between dyslipidemia in HTG rats and human FCHL, establishing a novel animal model for exploration of pathophysiology and therapy based on genomic determinants.

Animals↗

PubMed2004

Influence of diet and genetics on hypertension and renal disease in Dahl salt-sensitive rats.

Experiments examined the influence of diet and genetics on hypertension and renal disease in inbred Dahl salt-sensitive (SS/Mcw) rats and consomic rats in which chromosomes 16 (SS.BN16) or 18 (SS.BN18) of the normotensive Brown Norway rat were inserted into the genetic background of the SS/Mcw. Dahl SS/Mcw breeders and offspring were randomly placed on a purified AIN-76A diet or a grain-based diet, and male offspring were screened for cardiovascular and renal phenotypes following 3 wk on a 4.0% NaCl diet. High-salt arterial blood pressure (162 +/- 5 mmHg, n = 10), urinary protein excretion (147 +/- 16 mg/day, n = 14), and albumin excretion (72 +/- 9 mg/day, n = 14) were significantly elevated in the Dahl SS/Mcw maintained on the purified diet compared with rats fed the grain-based diet. Rats fed the purified diet also exhibited significantly more renal glomerular and tubular damage than rats fed the grain diet. Moreover, feeding the purified diet to the parents led to a significant increase in blood pressure in the offspring, regardless of offspring diet. Similar dietary effects were observed in SS.BN16 and SS.BN18 rats. In rats fed the purified diet, substitution of chromosomes 16 or 18 led to a significant decrease in arterial blood pressure, albumin excretion, and protein excretion compared with the SS/Mcw. Chromosomal substitution did not, however, affect albumin or protein excretion in the consomic rats compared with the SS/Mcw when the rats were maintained on the grain diet. These data demonstrate a significant influence of diet composition on salt-induced hypertension and renal disease in the Dahl SS/Mcw rat.

Animals↗

PubMed2004

Application of chromosomal substitution techniques in gene-function discovery.

A consomic rat strain is one in which an entire chromosome is introgressed into the isogenic background of another inbred strain using marker assisted selection. The development and initial physiologic screening of two inbred consomic rat panels on two genetic backgrounds (44 strains) is well underway. The primary uses of consomic strains are: (1) to assign traits and quantitative trait loci (QTL) to chromosomes by surveying the panel of strains with substituted chromosomes; (2) to rapidly develop congenic strains over a narrow region using several approaches described in this review and perform F2 linkage studies to positionally locate QTL in a fixed genetic background. In addition, consomic strains overcome many of the problems encountered with segregating crosses where, even if linkage is found, each individual in the cross is genetically unique and the combination of genes cannot be reproduced or studied in detail. Consomic strains provide greater statistical power to detect linkage than traditional F2 crosses because of their fixed genetic backgrounds, and can produce sufficient numbers of genetically identical rats to validate the relationship between a trait and a particular chromosome. These strains allow studies to be performed in a replicative or longitudinal manner to elucidate in greater detail the sequential changes responsible for the observed phenotypes of these animals, and they enable one to assess the impact of a causal gene region in a genome by allowing comparisons of the effect of replacement of a specific chromosome upon a disease susceptible or resistant genomic background. Consomics can be used to quickly develop multiple chromosome substitution models to investigate gene-gene interactions of complex traits or diseases. Finally, they often provide the best available inbred control strain for particular physiological comparisons with the inbred parental strains. Consomic rat strains are proving to be a unique scientific resource that greatly extends our understanding of genes and complex normal and pathological function.

Animals↗

PubMed2004

The Collaborative Cross, a community resource for the genetic analysis of complex traits.

The goal of the Complex Trait Consortium is to promote the development of resources that can be used to understand, treat and ultimately prevent pervasive human diseases. Existing and proposed mouse resources that are optimized to study the actions of isolated genetic loci on a fixed background are less effective for studying intact polygenic networks and interactions among genes, environments, pathogens and other factors. The Collaborative Cross will provide a common reference panel specifically designed for the integrative analysis of complex systems and will change the way we approach human health and disease.

Animals↗

PubMed2004

High-density rat radiation hybrid maps containing over 24,000 SSLPs, genes, and ESTs provide a direct link to the rat genome sequence.

The laboratory rat is a major model organism for systems biology. To complement the cornucopia of physiological and pharmacological data generated in the rat, a large genomic toolset has been developed, culminating in the release of the rat draft genome sequence. The rat draft sequence used a variety of assembly packages, as well as data from the Radiation Hybrid (RH) map of the rat as part of their validation. As part of the Rat Genome Project, we have been building a high-density RH map to facilitate data integration from multiple maps and now to help validate the genome assembly. By incorporating vectors from our lab and several other labs, we have doubled the number of simple sequence length polymorphisms (SSLPs), genes, expressed sequence tags (ESTs), and sequence-tagged sites (STSs) compared to any other genome-wide rat map, a total of 24,437 elements. During the process, we also identified a novel approach for integrating the RH placement results from multiple maps. This new integrated RH map contains approximately 10 RH-mapped elements per Mb on the genome assembly, enabling the RH maps to serve as a scaffold for a variety of data visualization tools.

Animals↗

PubMed2004

Comparative recombination rates in the rat, mouse, and human genomes.

Levels of recombination vary among species, among chromosomes within species, and among regions within chromosomes in mammals. This heterogeneity may affect levels of diversity, efficiency of selection, and genome composition, as well as have practical consequences for the genetic mapping of traits. We compared the genetic maps to the genome sequence assemblies of rat, mouse, and human to estimate local recombination rates across these genomes. Humans have greater overall levels of recombination, as well as greater variance. In rat and mouse, the size of the chromosome and proximity to telomere have less effect on local recombination rate than in human. At the chromosome level, rat and mouse X chromosomes have the lowest recombination rates, whereas human chromosome X does not show the same pattern. In all species, local recombination rate is significantly correlated with several sequence variables, including GC%, CpG density, repetitive elements, and the neutral mutation rate, with some pronounced differences between species. Recombination rate in one species is not strongly correlated with the rate in another, when comparing homologous syntenic blocks of the genome. This comparative approach provides additional insight into the causes and consequences of genomic heterogeneity in recombination.

Animals↗

PubMed2004

Integrative genomics: in silico coupling of rat physiology and complex traits with mouse and human data.

Integration of the large variety of genome maps from several organisms provides the mechanism by which physiological knowledge obtained in model systems such as the rat can be projected onto the human genome to further the research on human disease. The release of the rat genome sequence provides new information for studies using the rat model and is a key reference against which existing and new rat physiological results can be aligned. Previously, we described comparative maps of the rat, mouse, and human based on EST sequence comparisons combined with radiation hybrid maps. Here, we use new data and introduce the Integrated Genomics Environment, an extensive database of curated and integrated maps, markers, and physiological results. These results are integrated by using VCMapview, a java-based map integration and visualization tool. This unique environment allows researchers to relate results from cytogenetic, genetic, and radiation hybrid studies to the genome sequence and compare regions of interest between human, mouse, and rat. Integrating rat physiology with mouse genetics and clinical results from human by using the respective genomes provides a novel route to capitalize on comparative genomics and the strengths of model organism biology.

Animals↗

PubMed2004

Initial characterization of a rat model of diabetic nephropathy.

The lack of an appropriate animal model that spontaneously develops diabetic nephropathy has been a significant limitation in the search for genetic factors underlying this disease and the development of new therapeutic strategies to prevent progressive renal disease in diabetes. We introgressed the mitochondria and some passenger loci from the FHH/EurMcwi rat into the genetic background of diabetic GK rats, creating a new rat strain, T2DN (T2DN/Mcwi). Despite the high degree of genetic similarity between T2DN and GK rats (97% at 681 loci), diabetes ensues earlier and progresses more severely in T2DN rats. T2DN rats exhibit proteinuria by 6 months of age, accompanied by renal histologic abnormalities such as focal glomerulosclerosis, mesangial matrix expansion, and thickening of basement membranes. These characteristics progress over time, and nearly all T2DN rats exhibit diffuse global glomerulosclerosis with nodule formation and arteriolar hyalinosis by 18 months of age. The histologic changes in the kidney of T2DN rats closely mimic the changes seen in the kidney of patients with diabetes. These results indicate that the T2DN rat is a suitable model for investigating diabetic nephropathy. Here we report the initial genetic and physiological characterization of this new rat model of diabetic nephropathy.

Animals↗

PubMed2003

Genomic map of cardiovascular phenotypes of hypertension in female Dahl S rats.

Genetic linkage analyses in human populations have traditionally combined male and female progeny for determination of quantitative trait loci (QTL). In contrast, most rodent studies have focused primarily on males. This study represents an extensive female-specific linkage analysis in which 236 neuroendocrine, renal, and cardiovascular traits related to arterial pressure (BP) were determined in 99 female F2 rats derived from a cross of Dahl salt-sensitive SS/JrHsdMcwi (SS) and Brown Norway normotensive BN/SsNHsdMcwi (BN) rats. We identified 126 QTL for 96 traits on 19 of the 20 autosomal chromosomes of the female progeny. Four chromosomes (3, 6, 7, and 11) were identified as especially important in regulation of arterial pressure and renal function, since aggregates of 8-11 QTL mapped together on these chromosomes. BP QTL in this female population differed considerably from those previously found in male, other female, or mixed sex population linkage analysis studies using SS rats. Kidney weight divided by body weight was identified as an intermediate phenotype that mapped to the same region of the genome as resting diastolic blood pressure and was correlated with that same BP phenotype. Seven other phenotypes were considered as "potential intermediate phenotypes, " which mapped to the same region of the genome as a BP QTL but were not correlated with BP. These included renal vascular responses to ANG II and ACh and indices of baroreceptor responsiveness. Secondary traits were also identified that were likely to be consequences of hypertension (correlated with BP but not mapped to a BP QTL). Seven such traits were found, notably heart rate, plasma cholesterol, and renal glomerular injury. The development of a female rat systems biology map of cardiovascular function represents the first attempt to prioritize those regions of the genome important for development of hypertension and end organ damage in female rats.

Animals↗

PubMed2003

Use of a three-color cDNA microarray platform to measure and control support-bound probe for improved data quality and reproducibility.

Construction methodologies for cDNA microarrays lack the ability to determine array integrity prior to hybridization, leaving the array itself a source of uncontrolled experimental variation. We solved this problem through development of a three-color cDNA array platform whereby printed probes are tagged with fluorescein and are compatible with Cy3 and Cy5 target labeling dyes when using confocal laser scanners possessing narrow bandwidths. Here we use this approach to: (i) develop a tracking system to monitor the printing of probe plates at predicted coordinates; (ii) define the quantity of immobilized probe necessary for quality hybridized array data to establish pre-hybridization array selection criteria; (iii) investigate factors that influence probe availability for hybridization; and (iv) explore the feasibility of hybridized data filtering using element fluorescein intensity. A direct and significant relationship (R2 = 0.73, P < 0.001) between pre-hybridization average fluorescein intensity and subsequent hybridized replicate consistency was observed, illustrating that data quality can be improved by selecting arrays that meet defined pre-hybridization criteria. Furthermore, we demonstrate that our three-color approach provides a means to filter spots possessing insufficient bound probe from hybridized data sets to further improve data quality. Collectively, this strategy will improve microarray data and increase its utility as a sensitive screening tool.

Color↗

PubMed2003

Genetic dissection of lymphopenia from autoimmunity by introgression of mutated Ian5 gene onto the F344 rat.

Peripheral T cell lymphopenia (lyp) in the BioBreeding (BB) rat is linked to a frameshift mutation in Ian5, a member of the Immune Associated Nucleotide (Ian) gene family on rat chromosome 4. This lymphopenia leads to type 1 (insulin-dependent) diabetes mellitus (T1DM) at rates up to 100% when combined with the BB rat MHC RT1 u/u genotype. In order, to better study the lymphopenia phenotype without possible confounding effects of diabetes or other autoimmune disease, we generated congenic F344.lyp rats by introgression of lyp on diabetes-resistant MHC RT1 lv1/lv1 F344 rats. Analysis of thymic CD4 and CD8 T lymphocytes revealed no difference in the percentage of CD4(-)CD8(+)and CD4(+)CD8(-)subsets in lyp/lyp compared to +/+ F344 rats. The same subsets was however dramatically reduced in blood (P=0.005), spleen (P=0.019) and mesenteric lymph nodes (MLN) (P<0.0001). Compared to F344 +/+ rats double positive CD4(+)CD8(+)T cells were increased only in lyp/lyp spleen (P=0.034) while double negative CD4(-)CD8(-)were increased in thymus (P=0.033), spleen (P=0.012), MLN (P<0.0001), and peripheral blood (P<0.0001). There were no signs of inflammatory lesions in organs and tissues in F344.lyp/lyp rats examined at 120 days of age or older. We thus conclude that the lymphopenia phenotype was reconstituted by introgression of lyp on to F344 rats without subsequent development of organ-specific autoimmunity. The congenic F344.lyp rat should prove useful to dissect the mechanisms by which the Ian5 frameshift mutation affects T cell selection, differentiation and maturation without organ-specific autoimmunity.

Animals↗

PubMed2003

Expression of G-protein alpha subunit genes in the vestibular periphery of Rattus norvegicus and their chromosomal mapping.

OBJECTIVE: Heterotrimeric G-proteins play an important role in mediating signals transduced across the cell membrane by membrane-bound receptors. The precise role of G-proteins and their coupled receptors in the physiology of the vestibular neuroepithelium is not well understood. The purpose of this study was to better define the role of these proteins by examining their expression in the rat vestibular periphery and characterizing their chromosomal location. MATERIAL AND METHODS: To characterize G-protein alpha subunit gene expression in the target tissue of interest, we performed polymerase chain reaction (PCR) using degenerate G-protein primers corresponding to conserved regions in the G-protein alpha subunit coding sequence on a normalized rat vestibular cDNA library. PCR amplicons were cloned and 50 clones were randomly selected and sequenced. Radiation hybrid (RH) mapping was used to determine the chromosomal location of G alpha(olf) and two previously identified G-protein alpha subunits--G alpha(i2) and G alpha(i2(vest))--in the rat genome. RESULTS: The following G-protein alpha subunits were identified in the normalized cDNA library: G alpha(olf), G alpha(s), G alpha(o) and G alpha(s2). G alpha(olf) maps to chromosome 18 between markers D18Mit17b and D18Mgh2. G alpha(i2) maps to chromosome 8 between markers D8Rat65 and D8Mgh2. G alpha(i2(vest)) maps to chromosome 1 between markers D1Rat132 and D1Rat202. These chromosomal locations in the rat genome are syntenic to chromosomal regions in which the homologous G-protein alpha subunit genes have been localized in the human and mouse genomes, further validating RH mapping as an effective and accurate tool. We were unable to RH map the location of G alpha(o) due to its extensive homology with the hamster gene. CONCLUSION: The characterization of G-protein alpha subunit gene expression in the vestibular periphery and the chromosomal localization of these genes in the rat revealed that a diverse group of these second messengers are expressed.

Amino Acid Sequence↗