IMMUNOCYTOCHEMICAL INVESTIGATION OF THE IN-VIVO ENDOCYTOSIS BY RENAL TUBULAR EPITHELIAL-CELLS

The internalization and degradation of glomerular filtered serum prote ins by the proximal tubular epithelium has been extensively studied by microperfusion methods. By using a cationic probe that easily travers es the glomerular wall into the urinary space, we have performed a mor pho-cytochemical and quantitative study of the in vivo endocytotic act ivity of the proximal tubular epithelial cell. Bovine serum albumin (B SA) was tagged with dinitrophenol (DNP) and cationized to pi over 8. I t was introduced into the circulation of normal mice for 5, 10, and 30 minutes and the distribution of the labeling was determined by protei n A-gold immunocytochemistry, using specific antiDNP antibodies on tis sue sections of routinely aldehyde-fixed, osmium-postfixed, and Epon-e mbedded kidneys. Cationic BSA-DNP was detected at the endothelial and epithelial sides of the glomerular basement membrane, and over capilla ry and tubular basement membranes. In the proximal tubular epithelial cell, labeling was present over microvilli as well as over endosomal a nd lysosomal compartments, with labeling intensities varying from one compartment to the other. Morphometric evaluations of the labeling dem onstrated a progressive incorporation of the probe from microvilli and endocytic compartments at 5 minutes to endocytic and lysosomal compar tments at 10 and then 30 minutes. When considering labeling densities, no significant differences were found on microvilli and basolateral m embranes between times of circulation; however, the labeling density o ver endosomal and lysosomal compartments was very intense at 10 minute s compared with 5 minutes, decreasing at 30 minutes. Results from this study validate the cationic albumin tagged with DNP as a tool in the study of the quantitative aspects of protein endocytosis at the ultras tructural level, in the kidney tubular epithelium. (C) 1995 Wiley-Liss , Inc.