Characterization of gentamicin-induced dysfunctions in vitro: the use of optimized primary cultures of rabbit proximal tubule cells.
Compared to prior studies which frequently pinpoint the impairment of one parameter or function, this paper reports for the first time an extensive characterization of the toxic effects of gentamicin in a single model of primary cultured rabbit proximal tubule cells developed without insulin and glucose. Biochemical, functional and morphological approaches were used. Cellular response pattern was examined after a 72-h exposure during either the exponential growth phase or the stationary confluency phase of the culture to 0.2, 1, and 2.5 mM gentamicin. The biochemical study after gentamicin exposure showed increased activities for N-acetyl-beta-D-glucosaminidase and alkaline phosphatase, decreased activities for sphingomyelinase, cathepsin B, Na+/K(+)-ATPase, lactate dehydrogenase and NADPH cytochrome C reductase. Functional evaluation revealed decreased protein synthesis and alpha-methylglucose transport after gentamicin exposure. Morphometric study made it possible to show that the density of lysosomes, the cell fractional volume of the lysosomal compartment, and the mean size of the lysosomal profiles are increased in the cells. Intracellular accumulation of gentamicin in proximal tubular cells was dose dependent and reached high levels in cultured cells. In conclusion, this model compared to others in the literature allowed us to demonstrate in vitro a close response pattern to the in vivo situation after gentamicin exposure.
Monteil
C
C
INSERM U-295, UER Médecine-Pharmacie, Saint Etienne du Rouvray, France.
Leclere
C
C
Fillastre
J P
JP
Morin
J P
JP
D000118:Acetylglucosaminidase / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D002401:Cathepsin B / Q000378:metabolism; D002478:Cells, Cultured; D005839:Gentamicins / Q000633:toxicity*; D066298:In Vitro Techniques; D007687:Kidney Tubules, Proximal / Q000187:drug effects* / Q000201:enzymology / Q000502:physiology; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009251:NADPH-Ferrihemoprotein Reductase / Q000378:metabolism; D011817:Rabbits; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism
D005839:Gentamicins; D007770:L-Lactate Dehydrogenase; D009251:NADPH-Ferrihemoprotein Reductase; D000469:Alkaline Phosphatase; D013108:Sphingomyelin Phosphodiesterase; D000118:Acetylglucosaminidase; D002401:Cathepsin B; D000254:Sodium-Potassium-Exchanging ATPase