Receptor-mediated endocytosis of lysozyme in renal proximal tubules of the frog Rana temporaria

Abstract

The mechanism of protein reabsorption in the kidney of lower vertebrates remains insufficiently investigated in spite of raising interest to the amphibian and fish kidneys as a useful model for physiological and pathophysiological examinations. In the present study, we examined the renal tubular uptake and the internalization rote of lysozyme after its intravenous injection in the wintering frog Rana temporaria using immunohisto- and immunocytochemistry and specific markers for some endocytic compartments. The distinct expression of megalin and cubilin in the proximal tubule cells of lysozyme-injected frogs was revealed whereas kidney tissue of control animals showed no positive immunoreactivity. Lysozyme was detected in the apical endocytic compartment of the tubular cells and colocalized with clathrin 10 min after injection. After 20 min, lysozyme was located in the subapical compartment negative to clathrin (endo-somes), and intracellular trafficking of lysozyme was coincided with the distribution of megalin and cubilin. However, internalized protein was retained in the endosomes and did not reach lysosomes within 30 min after treatment that may indicate the inhibition of intra-cellular trafficking in hibernating frogs. For the first time, we provided the evidence that lysozyme is filtered through the glomeruli and absorbed by receptor-mediated clathrin-dependent endocytosis in the frog proximal tubule cells. Thus, the protein uptake in the amphibian mesonephros is mediated by megalin and cubilin that confirms a critical role of endocytic receptors in the renal reabsorption of proteins in amphibians as in mammals.

Targeted PEG-poly(glutamic acid) complexes for inhalation protein delivery to the lung

“…Data on similar experiments on cellular uptake of lysozyme are scarce, but recent in vivo studies reported on absorption of lysozyme in the intestinal tract [49], which appears to be occurring by non-saturable and saturable processes, the latter pointing to an involvement of endocytic pathway(s). Furthermore, it has recently been suggested that receptor-mediated endocytosis of lysozyme in renal proximal tubules may be mediated by megalin and cubilin [50]; the mRNA expression, although at a very low level, of the latter receptor was confirmed in the cell model used in this study. Furthermore it should be noted that complexation of lysozyme with non-targeted copolymers resulted in similar level of internalization as for 'free' lysozyme (Fig.…”

Protein Reabsorption in the Amphibian Kidney: Comparative and Evolutionary Aspects

“…In our studies, the expression of endocytic receptors in PT cells of the frog mesonephros was revealed after injections of YFP [18] and lysozyme [17] using polyclonal antibodies against megalin and cubilin. In 15-30 min, absorbed YFP was colocalized with immunolabeled megalin or cubilin in apical endocytic vesicles (Figure 4A-D).…”

“…Earlier, intestinal absorption and the resulting accumulation of these fluorescent proteins in renal PTs after their intragastric administration were established [16]. Later, the uptake of lysozyme in the amphibian kidney was also demonstrated [17]. Some examples of tubular reabsorption of abovementioned proteins in amphibians and rats are shown below (Figure 2).…”

“…So, lysozyme was retained within endosomal compartment during this time, in contrast to the faster protein traffic in the mammalian kidney. It may be connected with lower metabolic rate and following inhibition of the intracellular transport in hibernating frogs [17].…”

Pathological value of lysozyme staining for renal sarcoidosis

“…Although these changes are not specific for lysozyme-induced tubular injury, enlarged proximal tubular lysosomes are common in CMML nephropathy [ 6 , 8 ]. It is known that proximal tubular cells absorb lysozyme via endocytosis to degrade in lysosomes [ 10 , 11 ]. We think that lysosomes in the proximal tubular cells were processing overloaded lysozyme both in CMML and sarcoidosis.…”