Purpose: To investigate the involvement of the inter-alpha inhibitor family of proteins in calcium oxalate stone formation we determined immunohistochemical distribution in the kidneys and excretion in the urine of these proteins in normal and hyperoxaluric rats. Various members of the family have been shown to inhibit the formation and retention of calcium oxalate crystals in the kidneys.
Materials and methods: Hyperoxaluria was induced in male Sprague-Dawley rats by administering 0.75% ethylene glycol. The inter-alpha inhibitor family consists of inter-alpha inhibitor, pre-alpha inhibitor, the so-called heavy chains H1, H2 and H3, and the light chain bikunin. Antibodies against these molecules were used to localize various proteins in rat kidneys by immunohistochemical techniques. Urine was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis to determine the expression of various members of the inter-alpha inhibitor family.
Results: In normal kidneys staining for inter-alpha inhibitor and other members of the family was mostly limited to the proximal tubules and generally to their luminal contents. Eight weeks after the induction of hyperoxaluria various sections of renal tubules stained positive for inter-alpha inhibitor, bikunin and H3. Positive staining was observed in the tubular lumina as well as in the cytoplasm of epithelial cells. Crystal associated material was heavily stained. Western blot analysis recognized 7 protein bands in the urine. The urinary expression of H1, H3 and pre-alpha-inhibitor was significantly increased.
Conclusions: Apparently hyperoxaluria and renal calcium oxalate crystal deposition result in the increased expression of crystallization inhibitors, such as inter-alpha-inhibitor related proteins, in the kidneys and urine. Results indicate that kidneys respond to nephrolithic challenges by producing proteins that inhibit crystal formation and retention.