Mouse models for autosomal-dominant polycystic kidney disease (ADPKD), derived from homozygous targeted disruption of Pkd1 gene, generally die in utero or perinatally because of systemic defects. We introduced a loxP site and a loxP-flanked mc1-neo cassette into introns 30 and 34, respectively, of the Pkd1 locus to generate a conditional, targeted mutation. Significantly, before excision of the floxed exons and mc1-neo from the targeted locus by Cre recombinase, mice homozygous for the targeted allele appeared normal at birth but developed polycystic kidney disease with a slower progression than that of Pkd-null mice. Further, the homozygotes continued to produce low levels of full-length Pkd1-encoded protein, suggesting that slight Pkd1 expression is sufficient for renal cyst formation in ADPKD. In this viable model, up-regulation of heparin-binding epidermal growth factor-like growth factor accompanied increased epidermal growth factor receptor signaling, which may be involved in abnormal proliferation of the cyst-lining epithelia. Increased apoptosis in cyst epithelia was only observed in the later period that correlated with the cyst regression. Abnormalities in Na ؉ /K ؉ -ATPase, aquaporin-2, and vasopressin V2 receptor expression were also identified. This mouse model may be suitable for further studies of progression and therapeutic interventions of ADPKD.

Juvenile nephronophthisis type I is the most common genetic disorder causing end-stage renal failure in children and young adults. The defective gene responsible has been identified as NPHP1. Its gene product, nephrocystin-1, is a novel protein of uncertain function that is widely expressed in many tissues and not just confined to the kidney. To gain insight into the physiological function of nephrocystin, Nphp1-targeted mutant mice were generated by homologous recombination. Interestingly, homozygous Nphp1 mutant mice were viable without renal manifestations of nephronophthisis. They appeared normal, but males were infertile with oligoteratozoospermia. Histological analysis of the seminiferous tubules showed that spermatogenesis was blocked at the early stages of spermatid elongation, with degenerating spermatids sloughing off into the lumen. Electron microscopic analysis revealed detachment of early elongating spermatids from Sertoli cells, and a failure of sperm head and tail morphogenesis. However, a few mature spermatozoa were still deposited in the epididymis, though they were frequently dead, immotile, or malformed. These novel findings indicate that nephrocystin is critically required for the differentiation of early elongating spermatids into spermatozoa in mice. The possible roles of nephrocystin in the formation and maintenance of Sertoli-spermatid junctions are still under investigation.

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Nephrocystin mutations account for the vast majority of juvenile nephronophthisis, the most common inherited cause of renal failure in children. Nephrocystin has been localized to the ciliary transition zone of epithelial cells or its analogous structure, connecting cilium of retinal photoreceptors. Thus, the retinal degeneration associated with nephronophthisis may be explained by a functional ciliary defect. However, the function of nephrocystin in cilium assembly and maintenance of common epithelial cells and photoreceptors is still obscure. Here, we used Nphp1-targeted mutant mice and transgenic mice expressing EmGFP-tagged nephrocystin to demonstrate that nephrocystin located at connecting cilium axoneme can affect the sorting mechanism and transportation efficiency of the traffic machinery between inner and outer segments of photoreceptors. This traffic machinery is now recognized as intraflagellar transport (IFT); a microtubule-based transport system consisting of motors, IFT particles and associated cargo molecules. Nephrocystin seems to control some of the IFT particle components moving along the connecting cilia so as to regulate this inter-segmental traffic. Our novel findings provide a clue to unraveling the regulatory mechanism of nephrocystin in IFT machinery.

Acute tubular necrosis is followed by regeneration of damaged renal tubular epithelial cells, and renal stem cells are supposed to contribute to this process. The purpose of our study is to test the hypothesis that renal stem cells isolated from adult mouse kidney accelerate renal regeneration via participation in the repair process. A unique population of cells exhibiting characteristics consistent with renal stem cells, mouse kidney progenitor cells (MKPC), was isolated from Myh9 targeted mutant mice. Features of these cells include (1) spindle-shaped morphology, (2) self-renewal of more than 100 passages without evidence of senescence, and (3) expression of Oct-4, Pax-2, Wnt-4, WT-1, vimentin, a-smooth muscle actin, CD29, and S100A4 but no SSEA-1, c-kit, or other markers of more differentiated cells. MKPC exhibit plasticity as demonstrated by the ability to differentiate into endothelial cells and osteoblasts in vitro and endothelial cells and tubular epithelial cells in vivo. The origin of the isolated MKPC was from the interstitium of medulla and papilla. Importantly, intrarenal injection of MKPC in mice with ischemic injury rescued renal damage, as manifested by decreases in peak serum urea nitrogen, the infarct zone, and the necrotic injury. Seven days after the injury, some MKPC formed vessels with red blood cells inside and some incorporated into renal tubules. In addition, MKPC treatment reduces the mortality in mice after ischemic injury. Our results indicate that MKPC represent a multipotent adult stem cell population, which may contribute to the renal repair and prolong survival after ischemic injury. STEM CELLS

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Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common life-threatening inherited diseases, and the PKD1 gene is responsible for most cases of this disease. Previous efforts to establish a mouse model that recapitulates the phenotypic characteristics of ADPKD, which have used conventional or conditional knockout of the mouse orthologue Pkd1, have been unsuccessful or unreliable. In a previous study, we described the generation of a novel Pkd1 hypomorphic allele, in which Pkd1 expression was significantly reduced but not totally blocked. These Pkd1 homozygous mutant mice rapidly developed renal cystic disease, supporting the hypothesis that 'haploinsufficiency' explains development of the ADPKD phenotype. In the present study, we further investigated the Pkd1 haploinsufficiency effect by generating Pkd1 knockdown transgenic mice with co-cistronic expression of two miRNA hairpins specific to Pkd1 transcript and an Emerald GFP reporter driven by a human ubiquitin B promoter. Two transgenic lines which had ∼60-70% reduction of Pkd1 expression developed severe renal cystic disease at a rate similar to that of human ADPKD. These results further support the haploinsufficiency hypothesis, and suggest that the onset and progression of the renal cystic diseases are correlated with the level of Pkd1 expression. The two novel mutant lines of mice appear to be ideal models for the study of ADPKD.

Invasive pneumococcal infection is the most common cause of HUS in Taiwan. Positive T-Ag activation and a direct Coombs' test are rapid predictors of SP-HUS in children with invasive pneumonia.