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Induction of acute Heyman nephritis in rats gave a significant increase in the urinary excretion of protein (p less than 0.001) and N-acetyl-beta-D-glucosaminidase (NAG; p less than 0.01) 14 days after injection of antibody. The isoelectric points (IP) of NAG were studied by chromatofocusing of the urine, serum and various lysosomal populations purified from kidney cortex homogenates of normal and nephritic rats. The chromatofocusing profiles for serum NAG (IP = 5.8) were totally different from the patterns found in normal and nephritic urines. The acidic IPs of NAG in normal urine (IP = 5.0) were changed into slightly more basic values in nephritic urine (IP = 5.15). Similar changes were also demonstrated in normal urine after acidification and prolonged incubation. The chromatofocusing profile obtained for NAG in the large, dense lysosomes was almost identical to the pattern observed in nephritic urine and demonstrated IPs for NAG in a slightly more basic pH area than observed for small and medium-sized lysosomes. The difference in IP in normal and nephritic urines may therefore be due to an increased autolytic degradation of NAG or excretion of the enzyme from different populations of lysosomes.

Acute passive Heymann glomerulonephritis in rats induced heavy proteinuria and highly increased urinary activity of N-acetyl-beta-D-glucosaminidase, acid beta-galactosidase and acid phosphatase. The cortical activity of these acid hydrolases was increased essentially in the large lysosomes as demonstrated by subfractionation of the lysosome-rich mitochondrial-lysosomal fraction, by rate zonal centrifugation. Banding density of small lysosomes shifted or reduced to slightly lower value (1.225 g/ml), which is between the banding densities of small 'light' (1.20 g/ml) and small 'dense' lysosomes (1.235 g/ml) in normal rat kidney cortex. Labelled protein reabsorbed in the proximal tubule is recovered in these populations of small lysosomes as well as in the large lysosomes or 'protein droplets'. Glomerulonephritis also induced a new population of small 'light' lysosomes (density 1.185-1.195 g/ml) enriched in cathepsin D. The previously demonstrated morphological, biochemical, and physiological heterogeneity of renal lysosomes was confirmed and emphasized in the kidney cortex of glomerulonephritic rats. The main changes in the lysosomal populations appear to reflect the increased protein reabsorption as confirmed by the proteinuria.

1. Some lysosomal populations in the rat kidney cortex appear to be mechanically weak and are readily disrupted by gentle homogenization, while other populations remain intact even after repeated homogenization. 2. Lysosomes in the rat kidney cortex appear to be resistant to hypertonic media but are readily disrupted under hypotonic conditions. 3. Lysosomes in rat kidney cortex are readily disrupted when incubated in isotonic sucrose at 37 degrees C. 4. Measurement of total and free activity of three acid hydrolases: N-acetyl-beta-D-glucosaminidase (NAG), acid beta-galactosidase and acid beta-glycerophosphatase, indicates that the latency of these enzymes is relatively low in the homogenate (10-29%) and the ML-fraction (14-42%), but high (60-95%) in the purified large lysosomes (protein droplets). 5. The latency of purified small lysosomes is relatively lower (30-60%) than that of large lysosomes, suggesting that small lysosome populations are relatively permeable to the acid hydrolase substrates. 6. Latency variations of acid hydrolases amongst subcellular fractions appear to reflect the heterogeneity of lysosomal populations present in the kidney cortical homogenate.

The effect of food restriction (FR) on the kidney cortex lysosomes prepared by rate and isopycnic zonal centrifugation was studied in rats with passive Heymann glomerulonephritis (PHN). FR reduced the renal mass by 41%, but the capacity for handling of labelled endocytosed proteins by the lysosomes was not different from fed PHN rats. While PHN with heavy proteinuria increased the recovery of lysosomal enzymes in the large lysosomes located in the proximal tubule, no changes were observed in FR-PHN rats in spite of significant proteinuria. The density of the small lysosomes was significantly shifted/reduced (from 1,200 and 1,235 g/ml to 1,185 and 1,225 g/ml, respectively) in both fed and FR-PHN rats, suggesting that the handling of extra loads of protein may enhance the absorptive function of small lysosomes found in the lower part of the nephron. FR reduced the mechanical fragility of lysosomes in the kidney cortex of PHN-rats. The highly increased urinary excretion of lysosomal enzymes in fed PHN rats was not observed in FR-PHN rats. As a conclusion, FR reduces both the fragility of lysosomes and the proportion of digestive enzymes in fragile lysosomes. These lysosomal enzymes may be of pathogenic importance in PHN causing cell damage when liberated from disrupted lysosomes.

1. Experimental proteinuria (262.9 mg protein/24 hr urine) was induced in rats by repeated intraperitoneal injections of BSA. 2. Hypertrophy of the kidney cortex was significant 8 days after the start of the BSA injections, and the activities of lysosomal enzymes in kidney cortex and urine were significantly higher in proteinuric compared to nonproteinuric rats. 3. Lysosome populations in the kidney cortex were examined by rate sedimentation of the homogenate and by rate zonal and isopycnic centrifugation of the lysosome-rich ML fraction. 4. The activity of lysosomal enzymes in the kidney cortex increased slightly, essentially in the large, fragile lysosomes mainly recovered from the proximal tubule. 5. Proteinuria induced a shift/reduction in the density of small lysosomes from 1.235 and 1.20 g/ml to 1.225 and 1.185 g/ml, respectively. 6. Proteinuria induced a new population of small lysosomes (density 1.185 g/ml) enriched in cathepsin D.

Rate sedimentation of the kidney cortical mitochondrial/lysosomal (ML) fraction yields two distinct classes of lysosomes: the large lysosomes or protein droplets and a heterogeneous broad band of smaller lysosomes. The protein droplets which are recovered as a well defined zone of high purity also sediment as a homogeneous band after equilibrium banding at a density of 1.235 g/ml in sucrose. The small lysosomes co-sediment with other subcellular organelles as a broad band, indicated by the distribution of various acid hydrolases, which exhibit subtle heterogeneity among these small lysosomes. The distribution of renin containing granules indicates that in size they represent a distinct subpopulation of small lysosomes. Further fractionation of small lysosomes by equilibrium banding separates two distinct populations at densities 1.20 (small light) and 1.235 g/ml (small dense). Comparison of lysosomal populations fractionated in these studies with the distribution of lysosomal acid hydrolases along the different segments of the nephron suggests that large and small dense lysosomes probably originate from the proximal tubule while the small light lysosomes may contain lysosomes from the distal tubule. Very small, lysosome-like organelles subfractionated from the 'microsomes' may constitute a mixture of small light lysosomes, lysosomal fragments and endocytic vesicles from a variety of cell types. Time course studies with 3H labelled Cd-thionein, following intravenous administration, suggests that uptake in the kidney cortex is very rapid and that catabolism takes place in two distinct phases: rapid breakdown starting in the endosome compartment and slower breakdown in lysosomes. From the association of labelled lysozyme (125I) and Cd-thionein (109Cd) it appears that all the different lysosomal populations identified are at some stage involved with uptake and catabolism of these two proteins.