Ultrastructural immunodetection of osteopontin and osteocalcin as major matrix components of renal calculi

Abstract

The organic matrix of renal calculi has long been considered to influence the crystal growth that occurs in these pathological mineral deposits. Recent advances in characterizing individual organic moieties from mineralized tissues in general and the combined use of antibodies raised against these molecules with different immunocytochemical approaches have allowed their precise distribution to be visualized in a variety of normal and pathological mineralized tissues. The present ultrastructural study reports on the epithelial expression and extracellular localization of several noncollagenous proteins in rat and human kidney stones using high-resolution colloidal-gold immunocytochemistry. To this end, we have examined in an ethylene glycol-induced calcium oxalate model of urolithiasis in the rat, and in human kidney stones, the distribution of certain noncollagenous and plasma proteins known to accumulate in bone and other mineralized tissues that include osteopontin, osteocalcin, bone sialoprotein, albumin, and alpha 2HS-glycoprotein. Of these proteins, osteopontin (uropontin) and osteocalcin (or osteocalcin-related gene/protein) were prominent constituents of the calcium oxalate-associated crystal "ghosts" found in the nuclei, lamellae, and striations of the organic matrix of lumenal renal calculi in the rat and of small crystal ghosts found within epithelial cells. Immunocytochemical labeling for both proteins of the content of secretory granules in tubular epithelial cells from treated rats, together with labeling of a similarly textured organic material in the tubular lumen, provides evidence for cosecretion of osteopontin and osteocalcin by epithelial cells, their transit through the urinary filtrate, and ultimately their incorporation into growing renal calculi. In normal rat kidney, osteopontin was localized to the Golgi apparatus of thin loop of Henle cells. In human calcium oxalate monohydrate stones, osteopontin was similarly detected in the lamellae and striations of the organic matrix. Based on these data, it is proposed that during urolithiasis, secretion of osteopontin (uropontin) and osteocalcin (or osteocalcin-related gene/protein), and the subsequent incorporation of these proteins into kidney stone matrix, may influence the nucleation, growth processes, aggregation, and/or tubular adhesion of renal calculi in mammalian kidneys.