Background-We previously demonstrated that conditional overexpression of neuronal nitric oxide synthase (nNOS) inhibited L-type Ca 2ϩ channels and decreased myocardial contractility. However, nNOS has multiple targets within the cardiac myocyte. We now hypothesize that nNOS overexpression is cardioprotective after ischemia/reperfusion because of inhibition of mitochondrial function and a reduction in reactive oxygen species generation. Methods and Results-Ischemia/reperfusion injury in wild-type mice resulted in nNOS accumulation in the mitochondria.Similarly, transgenic nNOS overexpression caused nNOS abundance in mitochondria. nNOS translocation into the mitochondria was dependent on heat shock protein 90. Ischemia/reperfusion experiments in isolated hearts showed a cardioprotective effect of nNOS overexpression. Infarct size in vivo was also significantly reduced. nNOS overexpression also caused a significant increase in mitochondrial nitrite levels accompanied by a decrease of cytochrome c oxidase activity. Accordingly, O 2 consumption in isolated heart muscle strips was decreased in nNOS-overexpressing nNOS ϩ /␣MHC-tTA ϩ mice already under resting conditions. Additionally, we found that the reactive oxygen species concentration was significantly decreased in hearts of nNOS-overexpressing nNOS ϩ /␣MHC-tTA ϩ mice compared with noninduced nNOS ϩ /␣MHC-tTA ϩ animals. Conclusion-We demonstrated that conditional transgenic overexpression of nNOS resulted in myocardial protection after ischemia/reperfusion injury. Besides a reduction in reactive oxygen species generation, this might be caused by nitrite-mediated inhibition of mitochondrial function, which reduced myocardial oxygen consumption already under baseline conditions. (Circulation. 2010;122:1588-1603.)
Nephronophthisis (NPHP), an autosomal-recessive cystic kidney disease, is the most frequent genetic cause of end-stage renal failure in children. NPHP types 1 and 4 are caused by mutations in NPHP1 and NPHP4, encoding the proteins nephrocystin-1 and nephrocystin-4, respectively. Nephrocystin-1 and nephrocystin-4 are expressed in primary cilia of renal epithelial cells. NPHP1 and NPHP4 are highly conserved in Caenorhabditis elegans. However, this species does not have a kidney but an excretory system that consists of an excretory cell, an excretory gland cell, a duct cell, and a pore cell. Therefore, cell type-specific expression pattern and function of the nephrocystin homologs in C. elegans were of interest. Expression of green fluorescence protein fusion constructs that contain the C. elegans promoter regions for nph-1 and nph-4 was not found in the excretory system but in ciliated sensory neurons of the head (amphid neurons) and the tail in hermaphrodites (phasmid neurons) and males (sensory ray neurons). As the knockout phenotype for the PKD homologs lov-1 and pkd-2 shows impaired male mating behavior, RNAi knockdown animals were analyzed for this phenotype. A similar phenotype was found in the nph-1 and nph-4 RNAi knockdown animals compared with the lov-1 and pkd-2 knockout phenotype. Thus, it is suggested that renal cyst-causing genes may be part of a shared functional module, highly conserved in evolution. The NPHP homologs may be necessary for initial assembly of the cilium, whereas the polycystic kidney disease homologs may function as sensory transducers.
We here identified a new suggestive gene locus (NPL1) for autosomal dominant nephrolithiasis. It is localized on chromosome 9q33.2-q34.2. The identification of the responsible gene will provide new insights into the molecular basis of nephrolithiasis.
We confirm the MCKD1 locus on chromosome 1q21 and show further refinement of the MCKD1 locus to 2.1 Mb. This allowed us to exclude another 17 genes as positional candidate genes.
Medullary cystic kidney disease type 1 (MCKD1) is an autosomal dominant, tubulo-interstitial nephropathy that causes renal salt wasting and end-stage renal failure in the fourth to seventh decade of life. MCKD1 was localized to chromosome 1q21. We demonstrated haplotype sharing and confirmed the telomeric border by a recombination of D1S2624 in a Belgian kindred. Since the causative gene has been elusive, high resolution haplotype analysis was performed in 16 kindreds. Clinical data and blood samples of 257 individuals (including 75 affected individuals) from 26 different kindreds were collected. Within the defined critical region mutational analysis of 37 genes (374 exons) in 23 MCKD1 patients was performed. In addition, for nine kindreds RT-PCR analysis for the sequenced genes was done to screen for mutations activating cryptic splice sites. We found consistency with the haplotype sharing hypothesis in an additional nine kindreds, detecting three different haplotype subsets shared within a region of 1.19 Mb. Mutational analysis of all 37 positional candidate genes revealed sequence variations in 3 different genes, AK000210, CCT3, and SCAMP3, that were segregating in each affected kindred and were not found in 96 healthy individuals, indicating, that a single responsible gene causing MCKD1 remains elusive. This may point to involvement of different genes within the MCKD1 critical region.
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