Mechanism of the thesaurismosis and altered lysosomal dynamics induced by poly-D-glutamic acid in kidney proximal tubular cells.
In the companion paper, we report that a single injection of poly-D-glutamic acid causes an acute lysosomal storage condition and apparently impairs the lysosomal fission dynamics. The present paper addresses the mechanisms of these two alterations using a combination of in vivo and in vitro biochemical approaches. After a single intravenous injection, 14C-poly-D-glutamic acid was rapidly cleared from the plasma and appeared in the urine. Yet, a small but sizable fraction of the injected polymer was taken up by the kidney cortex through a saturable process (Kuptake, 150 mg/kg body wt; uptakemax 96 micrograms/g cortex). Analytical subcellular fractionation of cortex homogenates demonstrated that at initial stages, the 14C label was predominantly associated with subcellular particles of intermediate size and low equilibrium density, and was therefore slowly transferred to larger particles equilibrating at high density, then codistributing with the lysosomal hydrolases. At a concentration of 10 mg/ml (equivalent to its estimated concentration in lysosomes), poly-D-glutamic acid formed micronic aggregates ( > or = 10 microns) when brought to solution at pH < or = 6 in relation to its decreased ionization (pKa of lateral chains approximately equal to 4.25). Finally, 1 day after the injection of poly-D-glutamic acid, the activities of several lysosomal enzymes (hexosaminidase, cathepsin B, acid sphingomyelinase, and sulfatase B), but not of all of them (eg, acid phosphatase), were increased in the kidney cortex. We propose that poly-D-glutamic acid reaches lysosomes by adsorptive endocytosis and becomes concentrated within these organelles because its withstands hydrolysis until it forms aggregates or precipitates, causing a decrease in the fluidity or the deformability ("gelling") of the lysosomal matrix. This should alter the dynamics of intercommunication of these organelles by impairing their fission without a proportionate effect on their fusion properties. In addition, the data suggest that the presence of poly-D-glutamic acid directly or indirectly slows down the degradation of several lysosomal enzymes.
Kishore
B K
BK
Unité de Pharmacologie Cellulaire et Moléculaire, Université Catholique de Louvain, Brussels, Belgium.
Fuming
L u
Lu
Maldague
P
P
Tulkens
P M
PM
Courtoy
P J
PJ
D000818:Animals; D002250:Carbon Radioisotopes; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000378:metabolism; D007672:Kidney Cortex / Q000737:chemistry / Q000201:enzymology / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000378:metabolism*; D008247:Lysosomes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D008659:Metabolic Diseases / Q000097:blood / Q000139:chemically induced* / Q000652:urine; D011099:Polyglutamic Acid / Q000097:blood / Q000633:toxicity* / Q000652:urine; D051381:Rats; D017207:Rats, Sprague-Dawley; D012995:Solubility; D013347:Subcellular Fractions / Q000378:metabolism
D002250:Carbon Radioisotopes; D011099:Polyglutamic Acid; D006867:Hydrolases