Use of the polymerase chain reaction to clone and sequence a cDNA encoding the bovine alpha 3 chain of type IV collagen.

A novel type IV collagen, alpha 3(IV), has previously been isolated from a collagenase digest of bovine and human glomerular and lens basement membranes. The cloning and sequencing of a cDNA encoding the alpha 3(IV) chain is described here. Using the polymerase chain reaction, with primers derived from the known 27-residue bovine alpha 3(IV) amino acid sequence, a 68-base pair bovine genomic fragment (KEM68) which encodes the known peptide sequence, was synthesized. KEM68 was then used to screen a bovine lens cDNA library and a 1.5-kilobase partial cDNA clone obtained, encoding 471 residues of the bovine alpha 3(IV) chain: 238 residues from the triple helical collagenous domain and all 233 residues of the noncollagenous domain. The collagenous repeat sequence has three interruptions, coinciding with those in the alpha 1(IV) chain. The noncollagenous domain has 12 cysteine residues in identical positions to those of other type IV collagens and 71, 61, and 70% overall similarity with the human alpha 1(IV), alpha 2(IV), and alpha 5(IV) chains. The noncollagenous domain of alpha 3(IV) is of particular interest as it appears to be the component of glomerular basement membrane that reacts maximally with the Goodpasture antibody. Furthermore, such antigenicity is absent from collagenase digests of the glomerular basement membrane of some patients with Alport syndrome. The alpha 3(IV) cDNA clone described here now permits study of the molecular pathology of COL4A3 in Alport syndrome

Effect of Pioglitazone on Survival and Renal Function in a Mouse Model of Polycystic Kidney Disease

BACKGROUND/AIMS: Cystic epithelia in polycystic kidney disease display features similar to malignant cells. Thiazolidinediones have been shown to have anti-neoplastic properties, therefore we tested the hypothesis that pioglitazone reduces cyst formation, improves renal function, and prolongs survival in a mouse model of polycystic kidney disease. METHODS: PC-Pkd1-KO mice, which have homozygous mutations of the Pkd1 gene in principal cells, were used. On the day after giving birth, mothers were fed standard mouse chow with or without pioglitazone (30 mg/kg chow). After weaning, the assigned diet was continued. At 1 month of age, blood pressure was measured and animals were sacrificed to determine kidney weight, body weight, and serum urea. Kidneys were evaluated for proliferation using Ki-67, apoptosis using TUNEL analysis, and cyst number using MRI. Survival was observed. RESULTS: Pioglitazone did not alter renal function, cell proliferation, apoptosis, or cyst formation in animals with polycystic kidney disease, however it did increase survival. Pioglitazone reduced blood pressure in PC-Pkd1-KO, but not in controls. CONCLUSION: These findings suggest that pioglitazone may have a unique antihypertensive effect in polycystic kidney disease, and that such an effect may promote improved survival