Immunocytochemical investigation of the in vivo endocytosis by renal tubular epithelial cells

Abstract

The internalization and degradation of glomerular filtered serum proteins by the proximal tubular epithelium has been extensively studied by microperfusion methods. By using a cationic probe that easily traverses the glomerular wall into the urinary space, we have performed a morpho-cytochemical and quantitative study of the in vivo endocytotic activity of the proximal tubular epithelial cell. Bovine serum albumin (BSA) was tagged with dinitrophenol (DNP) and cationized to pI over 8. It was introduced into the circulation of normal mice for 5, 10, and 30 minutes and the distribution of the labeling was determined by protein A-gold immunocytochemistry, using specific antiDNP antibodies on tissue sections of routinely aldehyde-fixed, osmiumpostfixed, and Epon-embedded kidneys. Cationic BSA-DNP was detected at the endothelial and epithelial sides of the glomerular basement membrane, and over capillary and tubular basement membranes. In the proximal tubular epithelial cell, labeling was present over microvilli as well as over endosomal and lysosomal compartments, with labeling intensities varying from one compartment to the other. Morphometric evaluations of the labeling demonstrated a progressive incorporation of the probe from microvilli and endocytic compartments at 5 minutes to endocytic and lysosomal compartments at 10 and then 30 minutes. When considering labeling densities, no significant differences were found on microvilli and basolateral membranes between times of circulation; however, the labeling density over endosomal and lysosomal compartments was very intense at 10 minutes 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 ultrastructural level, in the kidney tubular epithelium.

Sex and Strain Differences in the Brush Border and PAS-Positive Granules and Giant Bodies of the Mouse Renal S3 Segment Cells.

“…From the present TEM and AcPase investigation, it is likely that PAS-positive granules and PAS-positive giant bodies of renal S3 segment cells observed under LM are multilamellar lysosomes, probably secondary lysosomes. The lysosomes present in the proximal tubular epithelium are active endocytic organelles containing numerous hydrolases [1,3,5,[14][15][16], and the changes in size and number of the lysosomes are closely related to the stimulation of androgen and the growth and development of the animal [6,8,11]. The present study revealed sex and strain differences in lysosomal size.…”

Sexual dimorphism of proximal straight tubular cells in mouse kidney

“…The brush border is an important structure for the reabsorption of macromolecules (Sumpio and Maack, 1982;Wall and Maack, 1985;Gómez-Pascual et al, 1995;Bendayan and Londoñ o, 1996). Therefore, sexual dimorphism of the PAS staining of the brush border suggests that there are differences in reabsorptive function in PST between males and females.…”

“…The detection of acid phosphatase reaction demonstrated that these electron-dense myelinoid bodies were lysosomes, probably secondary lysosomes. The lysosomes present in the proximal tubular epithelium are endocytic organelles (Wall and Maack, 1985;Gómez-Pascual et al, 1995;Bendayan and Londoñ o, 1996) containing numerous hydrolase enzymes (Johnson et al, 1986). Moreover, lysosomal enzyme activity levels were reported to be higher in male than in female mouse kidney (Johnson et al, 1986;Koenig et al, 1980), and testosterone increases their activities (Koenig et al, 1980).…”

Horseradish peroxidase binding to intestinal brush-border membranes ofCyprinus carpio. Identification of a putative receptor

“…The existence of anionic sites in different cell membranes, including the apical membrane of mammalian enterocytes, has been demonstrated using cationized ferritin and other chemically modified proteins as ligand [Danon et al, 1972;Triguereo et al, 1989;Sanderson and Walker, 1993;Lehr, 1994]. The ionic interaction of a ligand with these anionic sites is usually associated with a nonreceptor mediated endocytosis that segregates the ligand to a degradative pathway within the cell [Gómez-Pascual et al, 1995]. Our results show that increasing NaCl concentrations in the binding assay only affected the specific binding, suggesting that HRP is not interacting with the broad family of anionic sites.…”