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BackgroundMany patients remain without a diagnosis despite extensive medical evaluation. The Undiagnosed Diseases Network (UDN) was established to apply a multidisciplinary model in the evaluation of the most challenging cases and to identify the biologic characteristics of newly discovered diseases. The UDN, which is funded by the National Institutes of Health, was formed in 2014 as a network of seven clinical sites, two sequencing cores, and a coordinating center. Later, a central biorepository, a metabolomics core, and a model organisms screening center were added.MethodsWe evaluated patients who were referred to the UDN over a period of 20 months. The patients were required to have an undiagnosed condition despite thorough evaluation by a health care provider. We determined the rate of diagnosis among patients who subsequently had a complete evaluation, and we observed the effect of diagnosis on medical care.ResultsA total of 1519 patients (53% female) were referred to the UDN, of whom 601 (40%) were accepted for evaluation. Of the accepted patients, 192 (32%) had previously undergone exome sequencing. Symptoms were neurologic in 40% of the applicants, musculoskeletal in 10%, immunologic in 7%, gastrointestinal in 7%, and rheumatologic in 6%. Of the 382 patients who had a complete evaluation, 132 received a diagnosis, yielding a rate of diagnosis of 35%. A total of 15 diagnoses (11%) were made by clinical review alone, and 98 (74%) were made by exome or genome sequencing. Of the diagnoses, 21% led to recommendations regarding changes in therapy, 37% led to changes in diagnostic testing, and 36% led to variant-specific genetic counseling. We defined 31 new syndromes.ConclusionsThe UDN established a diagnosis in 132 of the 382 patients who had a complete evaluation, yielding a rate of diagnosis of 35%. (Funded by the National Institutes of Health Common Fund.).

Precision or personalized medicine through clinical genome and exome sequencing has been described by some as a revolution that could transform healthcare delivery, yet it is currently used in only a small fraction of patients, principally for the diagnosis of suspected Mendelian conditions and for targeting cancer treatments. Given the burden of illness in our society, it is of interest to ask how clinical genome and exome sequencing can be constructively integrated more broadly into the routine practice of medicine for the betterment of public health. In November 2014, 46 experts from academia, industry, policy and patient advocacy gathered in a conference sponsored by Illumina, Inc. to discuss this question, share viewpoints and propose recommendations. This perspective summarizes that work and identifies some of the obstacles and opportunities that must be considered in translating advances in genomics more widely into the practice of medicine.

Renal preglomerular arterioles regulate vascular tone to ensure a large pressure gradient over short distances, a function that is extremely important for maintaining renal microcirculation. Regulation of renal microvascular tone is impaired in salt-sensitive (SS) hypertension-induced nephropathy, but the molecular mechanisms contributing to this impairment remain elusive. Here, we assessed the contribution of the SH2 adaptor protein p66Shc (encoded by Shc1) in regulating renal vascular tone and the development of renal vascular dysfunction associated with hypertension-induced nephropathy. We generated a panel of mutant rat strains in which specific modifications of Shc1 were introduced into the Dahl SS rats. In SS rats, overexpression of p66Shc was linked to increased renal damage. Conversely, deletion of p66Shc from these rats restored the myogenic responsiveness of renal preglomerular arterioles ex vivo and promoted cellular contraction in primary vascular smooth muscle cells (SMCs) that were isolated from renal vessels. In primary SMCs, p66Shc restricted the activation of transient receptor potential cation channels to attenuate cytosolic [Ca.sup.2+] influx, implicating a mechanism by which overexpression of p66Shc impairs renal vascular reactivity. These results establish the adaptor protein p66Shc as a regulator of renal vascular tone and a driver of impaired renal vascular function in hypertension-induced nephropathy., Introduction One of the major complications of hypertension is nephropathy, which is associated with impaired renal vascular responsiveness and structural changes (1). The microvascular injuries are detected in the majority [...]

Hypertension and its complications represent leading causes of morbidity and mortality. Although the cause of hypertension is unknown in most patients, genetic factors are recognized as contributing significantly to an individual's lifetime risk of developing the condition. Here, we investigated the role of the G protein regulator phosducin (Pdc) in hypertension. Mice with a targeted deletion of the gene encoding Pdc ([Pdc.sup.[-/-]] mice) had increased blood pressure despite normal cardiac function and vascular reactivity, and displayed elevated catecholamine turnover in the peripheral sympathetic system. Isolated postganglionic sympathetic neurons from [Pdc.sup.[-/-]] mice showed prolonged action potential firing after stimulation with acetylcholine and increased firing frequencies during membrane depolarization. Furthermore, [Pdc.sup.[-/-]] mice displayed exaggerated increases in blood pressure in response to post-operative stress. Candidate gene-based association studies in 2 different human populations revealed several SNPs in the PDC gene to be associated with stress-dependent blood pressure phenotypes. Individuals homozygous for the G allele of an intronic PDC SNP (rs12402521) had 12-15 mmHg higher blood pressure than those carrying the A allele. These findings demonstrate that PDC is an important modulator of sympathetic activity and blood pressure and may thus represent a promising target for treatment of stress-dependent hypertension., Introduction Hypertension and its cardiovascular consequences contribute to the leading causes of morbidity and mortality worldwide (1). In over 90% of hypertensive patients, the causes of increased blood pressure are [...]

This study examined the effect of transfer of overlapping regions of chromosome 5 that includes ([4A.sup.+]) or excludes ([4A.sup.-]) the cytochrome P-450 4A (CYP4A) genes from the Lewis rat on the renal production of 20-hydroxyeicosatetraenoic acid (20-HETE) and the development of hypertension-induced renal disease in congenic strains of Dahl salt-sensitive (Dahl S) rats. The production of 20-HETE was higher in the outer medulla of [4A.sup.+] than in Dahl S or [4A.sup.-] rats. Mean arterial pressure (MAP) rose to 190 [+ or -] 7 and 185 [+ or -] 3 mmHg in Dahl S and [4A.sup.-] rats fed a high-salt (HS) diet for 21 days but only to 150 [+ or -] 5 mmHg in the [4A.sup.+] strain. Protein excretion increased to 423 [+ or -] 40 and 481 [+ or -] 37 mg/day in Dahl S and [4A.sup.-] rats vs. 125 [+ or -] 15 mg/day in the [4A.sup.+] strain. Baseline glomerular capillary pressure (Pgc) was lower in [4A.sup.+] rats (38 [+ or -] 1 mmHg) than in Dahl S rats (42 [+ or -] 1 mmHg). Pgc increased to 50 [+ or -] 1 mmHg in Dahl S rats fed a HS diet, whereas it remained unaltered in [4A.sup.+] rats (39 [+ or -] 1 mmHg). Baseline glomerular permeability to albumin ([P.sub.alb]) was lower in [4A.sup.+] rats (0.19 [+ or -] 0.05) than in Dahl S or [4A.sup.-] rats (0.39 [+ or -] 0.02). [P.sub.alb] rose to ~0.61 [+ or -] 0.03 in [4A.sup.-] and Dahl S rats fed a HS diet for 7 days, but it remained unaltered in the [4A.sup.+] rats. The expression of transforming growth factor-[beta]2 was higher in glomeruli of Dahl S rats than in [4A.sup.+] rats fed either a low-salt (LS) or HS diet. Chronic administration of a 20-HETE synthesis inhibitor (HET0016; 10 mg x [kg.sup.-1] x [day.sup.-1] sc) reversed the fall in MAP and renoprotection seen in [4A.sup.+] rats. These results indicate that the introgression of the CYP4A genes from Lewis rats into the Dahl S rats increases the renal formation of 20-HETE and attenuates the development of hypertension and renal disease. hypertension; glomerulosclerosis; 20-hydroxyeicosatetraenoic acid; transforming growth factor-[beta]; Dahl salt-sensitive rats; Rattus norvegicus 5; epoxyeicosatrienoic acids; dihydroxyeicosatrienoic acids; glomerulus; renal hemodynamics; kidney

This study examined the genetic basis of hypertension and renal disease in Dahl SS/Mcwi (Dahl Salt-Sensitive) rats using a complete chromosome substitution panel of consomic rats in which each of the 20 autosomes and the X and Y chromosomes were individually transferred from the Brown Norway (BN) rat onto the Dahl SS/Mcwi genetic background. Male and female rats of each of the two parental and 22 consomic strains (10-12 rats/group) were fed a high-salt (8.0% NaCl) diet for 3 wk. Mean arterial blood pressure rose by 60 mmHg and urinary protein and albumin excretion increased 3- and 20-fold, respectively, in male SS/Mcwi rats compared with BN controls. Substitution of chromosomes 1, 5, 7, 8, 13, or 18 from the BN onto the SS/Mcwi background attenuated the development of hypertension, proteinuria, and albuminuria in male rats. In female rats, substitution of chromosomes 1 and 5 also decreased blood pressure, protein excretion, and albumin excretion. These studies also identified several chromosomes in male (6, 11, Y) and female (4, 6, 11, 19, 20) rats that reduced albuminuria without altering blood pressure. These data indicate that genes contributing to salt-sensitive hypertension are found on multiple chromosomes of the Dahl SS/Mcwi rat. Furthermore, this consomic rat panel provides a stable genetic platform that can facilitate further gene mapping by either linkage studies or the breeding of congenic and subcongenic rats. hypertension; kidney disease; rats; consomic

This study examined the genetic basis for hypertension and renal disease phenotypes in Fawn Hooded hypertensive (FHH) rats using chromosome substitution strains (consomic rats) in which each of the 20 autosomes as well as the X and Y chromosomes were transferred from the normal Brown Norway (BN) rat onto the FHH genetic background. Male and female rats of each of the parental and consomic strains were maintained for 2 wk on high-salt (8.0% NaCl) chow with [N.sup.G]-nitro-L-arginine methyl ester (L-NAME) in the drinking water (12.5 mg/l) to induce hypertension and renal disease. Mean arterial blood pressure (MAP) was significantly higher (by over 60 mmHg) in the male FHH compared with BN rats. Urinary protein and albumin excretion rates were increased by 15- and 40-fold, respectively, in the male FHH compared with the BN. Plasma renin activity was 10-fold higher in the FHH than the BN. Similar significant differences were observed between the female FHH and BN, but the degree of hypertension and proteinuria was of a lesser magnitude. Substitution of chromosome 20 from the BN to the FHH attenuated the development of L-NAME-induced hypertension, normalized plasma renin activity, and decreased plasma creatinine in male rats. In female rats, substitution of chromosome 15 decreased MAP and urinary protein excretion. Urinary excretion of albumin in males was decreased by substitution of chromosomes 1, 15, 16, and 18 from the BN into the FHH genetic background. The present data indicate that genes that can modify L-NAME-induced hypertension and proteinuria are on chromosomes 1, 15, 16, 18, and 20. kidney disease; L-NAME; consomic

A lentiviral construct for an enhanced green fluorescent protein (eGFP) driven by a chicken [beta]-actin promoter, cytomegalovirus enhancer, and intronic sequences from rabbit [beta]-globin (CAG) was used to produce transgenic lines of rats for evaluation of the usefulness of this approach in gene function studies. Fertilized eggs were collected from inbred Dahl S and outbred Sprague-Dawley rats, and ~100 pl of concentrated virus were micro-injected into the perivitrelline space of one-cell embryos. Of 121 embryos injected, 60 pups (49.6%) were born. Transgenic rates averaged 22% in Dahl S and 14% in Sprague-Dawley rats. Copy number ranged from one to four in the founders, and the inheritance of the transgene in a subsequent [F.sub.1] population was 48.2%. The small number of insertion sites enabled us to derive inbred transgenic lines with a single copy of the transgene within one generation. Sequencing of each transgene insertion site revealed that they inserted as single copies with a preference for the introns of genes. The CAG promoter drove high levels of eGFP expression in brain, kidney, heart, and vasculature, making it very suitable for exploring the cardiovascular function of newly discovered genes. The pattern of eGFP expression was similar across five different [F.sub.1] transgenic lines, indicating that the expression of the transgene was independent of its chromosomal position. Thus lentiviral transgenesis provides a powerful tool for the production of transgenic inbred rats and will enhance the usefulness of this species in gene discovery and target validation studies. gene targeting; transgenesis; lentiviral gene transfer; gene expression doi:10.1152/ajpregu.00060.2007

Rat remains a major biomedical model system for common, complex diseases. The rat continues to gain importance as a model system with the completion of its full genomic sequence. Although the genomic sequence has generated much interest, only three complete sequences of the rat mitochondria exist. Therefore, to increase the knowledge of the rat genome, the entire mitochondrial genomes (16,307-16,315 bp) from 10 inbred rat strains (that are standard laboratory models around the world) and 2 wild rat strains were sequenced. We observed a total of 195 polymorphisms, 32 of which created an amino acid change (nonsynonymous substitutions) in 12 of the 13 protein coding genes within the mitochondrial genome. There were 11 single nucleotide polymorphisms within the tRNA genes, six in the 12S rRNA, and 12 in the 16S rRNA including 3 insertions/deletions. We found 14 single nucleotide polymorphisms and 2 insertion/deletion polymorphisms in the D-loop. The inbred rat strains cluster phylogenetically into three distinct groups. The wild rat from Tokyo grouped closely with five inbred strains in the phylogeny, whereas the wild rat from Milwaukee was not closely related to any inbred strain. These data will enable investigators to rapidly assess the potential impact of the mitochondria in these rats on the physiology and the pathophysiology of phenotypes studied in these strains. Moreover, these data provide information that may be useful as new animal models, which result in novel combinations of nuclear and mitochondrial genomes, are developed. genome; mitochondria

The present study examined whether transfer of overlapping regions of chromosome 5 that include (4[A.sup.+]) or exclude the cytochrome P-450 (CYP) 4A genes from the Lewis rat alters the renal production of 20-hydroxyeicosatetraenoic acid (20-HETE) and/or the development of hypertension in congenic strains of Dahl salt-sensitive (S) rats. The expression of CYP4A protein and the production of 20-HETE in the renal outer medulla was greater in the 4[A.sup.+] congenic strain than the levels seen in S rats or in overlapping control congenic strains that exclude the CYP4A region. Mean arterial pressure (MAP) rose from 122 [+ or -] 2 to 190 [+ or -] 7 mmHg in S rats and from 119 [+ or -] 2 and 123 [+ or -] 2 to 189 [+ or -] 7 and 187 [+ or -] 3 mmHg in the two control congenic strains fed an 8.0% NaCl diet for 3 wk. In contrast, MAP only increased from 112 [+ or -] 2 to 150 [+ or -] 5 mmHg in the 4[A.sup.+] congenic strain. Chronic blockade of the formation of 20-HETE with N-(3-chloro-4-morpholin-4-yl) phenyl-N'-hydroxyimido formamide (TS-011; 1 mg/kg bid) restored the salt-sensitive phenotype in the 4[A.sup.+] congenic strain and MAP rose to 181 [+ or -] 6 mmHg after an 8.0% NaCl dietary challenge. TS-011 had no effect on the development of hypertension in S rats or the two control congenic strains. The pressure-natriuretic and diuretic responses were fivefold greater in the 4[A.sup.+] congenic strain than in S rats. These results indicate that transfer of the region of chromosome 5 between markers D5Rat108 to D5Rat31 from the Lewis rat into the Dahl S genetic background increases the renal production of 20-HETE, improves pressure-natriuresis and opposes the development of salt-induced hypertension. cytochrome P-450; kidney; 20-HETE

The present study evaluated whether the impairment in autoregulation of renal blood flow (RBF) in the fawn-hooded Hypertensive (FHH) rat colocalizes with the Rf-1 region on chromosome 1 that has been previously linked to the development of proteinuria in this strain. Autoregulation of RBF was measured in FHH and a consomic strain (FHH.[1.sup.BN]) in which chromosome 1 from the Brown-Norway (BN) rat was introgressed into the FHH genetic background. The autoregulation indexes (AI) averaged 0.80 [+ or -] 0.08 in the FHH and 0.19 [+ or -] 0.05 in the FHH.[1.sup.BN] rats. We next performed a genetic linkage analysis for autoregulation of RBF in 85 F2 rats generated from a backcross of FHH.[1.sup.BN] consomic and FHH rats. The results revealed a significant quantitative trait locus (QTL) with a peak logarithm of the odds score of 6.3 near marker DlRat376. To confirm the existence of this QTL, five overlapping congenic strains were created that spanned the region from markers DlRat234 to D1Mit14. Transfer of a region of BN chromosome 1 from markers D1Mgh13 to DlRat89 into the FHH genetic background improved autoregulation of RBF (AI = 0.23 [+ or -] 0.04) and reduced protein excretion. In contrast, RBF was poorly autoregulated and the rats were not protected from proteinuria in congenic strains in which other regions of chromosome 1 that exclude the D1Rat376 marker were transferred. These results indicate that there is a gene(s) that influences autoregulation of RBF and proteinuria between markers DlMghl3 and D1Rat89 on chromosome 1 that lies within the confidence interval of the Rf-1 QTL previously linked to the development of proteinuria in FHH rats. renal disease; Rf-l; consomic and congenic rats; quantitative trait locus

Linkage analysis studies previously identified genetic loci associated with proteinuria and hypertension on chromosome 1 of fawn-hooded hypertensive (FHH) rats. The present studies were performed on conscious male and female rats to evaluate the influence of transfer of chromosome 1 from the Brown Norway (BN) rat to the FHH genetic background (FHH-[1.sup.BN]). Rats were maintained for 2 wk on 8.0% NaCl chow with [N.sup.G]-nitro-L-arginine methyl ester (L-NAME) in the drinking water (12.5 mg/1) to induce hypertension and accelerate the onset of renal disease. Mean arterial blood pressure (MAP) was significantly higher in the male FHH (188 [+ or -] 3 mmHg, n = 13) compared with the BN (121 [+ or -] 3 mmHg, n = 8); MAP in the FHH-[1.sup.BN] was midway between the two parental strains (167 [+ or -] 5 mmHg, n = 9). Urinary protein and albumin excretion rates in the male FHH-[1.sup.BN] (Uprot = 189 [+ or -] 36 mg/day, Ualb = 69 [+ or -] 16 mg/day, n = 10) were also midway between levels observed in the FHH (Uprot = 485 [+ or -] 54 mg/day; Ualb = 206 [+ or -] 25 mg/day, n = 13) and the BN (Uprot = 32 [+ or -] 5 mg/day, Ualblb = 5 [+ or -] 1 mg/day, n = 8). Creatinine clearance was elevated, and the degree of glomerular damage was significantly reduced in the FHH-[1.sup.BN] compared with the FHH. Qualitatively similar results were obtained from female FHH, FHH-[1.sup.BN], and BN rats. The present results indicate that genes contributing to L-NAME-induced hypertension and renal disease are found on chromosome 1 of the FHH rat. kidney disease; consomic