CROSSLINE-LIKE STRUCTURE ACCUMULATES AS FLUORESCENT ADVANCED GLYCATION END-PRODUCTS IN RENAL TISSUES OF RATS WITH DIABETIC NEPHROPATHY

Although formation of characteristic fluorescence (Ex(max)/Em(max):370 /440 nm) in renal tissues has been known to be enhanced in diabetic mo del rats and the natural fluorophores have been thought as advanced gl ycation end products (AGEs), these structures remain unclear to this e late. Since we had already isolated crossline derivatives as the main fluorescent AGE-products formed in a sugar-amine-model system in vitro , we tried to determine whether crossline-like structure can be found in renal tissues of rats with diabetic nephropathy. Specific polyclona l antiserum against crossline-structure and its affinity chromatograph y-purified antibody were prepared. The purified antibody did not recog nize native rat serum albumin (RSA), but it recognized RSA modified by prolonged incubation with high concentration of glucose (AGE-RSA) as well as the crossline-hapten. The immunoreactivity of AGE-RSA was comp letely blocked by co-incubation with crossline-derivative. These resul ts confirm immunochemically that in vitro-prepared AGE-proteins contai n crossline-like structure. Similarly, crossline-like structure in AGE -proteins in vivo was examined immunohistochemically. In streptozotoci n-induced diabetic rats, both crossline-like immunoreactivity and fluo rescence (Ex/Em:370/440 nm) of renal tissues increased, and the levels rose further with progression of diabetic nephropathy. Crossline accu mulation was observed in specific renal tissues such as glomeruli (bas ement membrane and mesangium), tubules, and Bowman's capsules. Althoug h the contribution of crossline-like substance(s) to total fluoroescen ce in diabetic renal tissues has not as yet been quantitatively determ ined, our results provide evidence that crossline accumulation occurs in renal tissues of rats in association with the initiation and/or pro gression of diabetic nephropathy.