Citations

Background— Long QT syndrome (LQTS) is the most common cardiac channelopathy with 15 elucidated LQTS-susceptibility genes. Approximately 20% of LQTS cases remain genetically elusive. Methods and Results— We combined whole-exome sequencing and bioinformatic/systems biology to identify the pathogenic substrate responsible for nonsyndromic, genotype-negative, autosomal dominant LQTS in a multigenerational pedigree, and we established the spectrum and prevalence of variants in the elucidated gene among a cohort of 102 unrelated patients with “genotype-negative/phenotype-positive” LQTS. Whole-exome sequencing was used on 3 members within a genotype-negative/phenotype-positive family. Genomic triangulation combined with bioinformatic tools and ranking algorithms led to the identification of a CACNA1C mutation. This mutation, Pro857Arg-CACNA1C, cosegregated with the disease within the pedigree, was ranked by 3 disease-network algorithms as the most probable LQTS-susceptibility gene and involves a conserved residue localizing to the proline, gltamic acid, serine, and threonine (PEST) domain in the II-III linker. Functional studies reveal that Pro857Arg-CACNA1C leads to a gain of function with increased I Ca,L and increased surface membrane expression of the channel compared to wild type. Subsequent mutational analysis identified 3 additional variants within CACNA1C in our cohort of 102 unrelated cases of genotype-negative/phenotype-positive LQTS. Two of these variants also involve conserved residues within Ca v 1.2’s PEST domain. Conclusions— This study provides evidence that coupling whole-exome sequencing and bioinformatic/systems biology is an effective strategy for the identification of potential disease-causing genes/mutations. The identification of a functional CACNA1C mutation cosegregating with disease in a single pedigree suggests that CACNA1C perturbations may underlie autosomal dominant LQTS in the absence of Timothy syndrome.

Heme oxygenase-2 (HO-2), the constitutive isoform of the heme-degrading enzyme heme oxygenase, may serve as an anti-inflammatory vasorelaxant, in part, by generating carbon monoxide. Arteriovenous fistulas (AVFs) are employed as hemodialysis vascular accesses because they provide an accessible, high-blood-flow vascular segment. We examined the role of vascular expression of HO-2 in AVF function. An AVF was created in mice by anastomosing the carotid artery to the jugular vein. HO-2 expression was detected by immunohistochemistry in the intact carotid artery, mainly in endothelial cells and smooth muscle cells; expression of HO-2 protein and mRNA was modestly increased in the artery of the AVF. Creating an AVF in HO-2−/−mice compared with an AVF in HO-2+/+mice led to markedly reduced AVF blood flow and increased numbers of nonfunctioning AVFs. The impairment of AVF function in the setting of HO-2 deficiency could not be ascribed to either preexisting intrinsic abnormalities in endothelium-dependent and endothelium-independent relaxation of the carotid artery in HO-2-deficient mice or to impaired vasorelaxant responses in the intact carotid artery in vivo. HO-1 mRNA was comparably induced in the AVF in HO-2+/+and HO-2−/−mice, whereas the AVF in HO-2−/−mice compared with that in HO-2+/+mice exhibited exaggerated induction of matrix metalloproteinase (MMP)-9 but similar induction of MMP-2. HO-2 deficiency also led to lower AVF blood flow when AVFs were created in uremia, the latter induced by subtotal nephrectomy. We conclude that HO-2 critically contributes to the adequacy of AVF blood flow and function.