Induction of dedifferentiated clones of LLC-PK1 cells upon long-term exposure to dichlorovinylcysteine.
Dichlorovinylcysteine (DCVC), the key metabolite of the nephrotoxic and nephrocarcinogenic chemicals, trichloroethylene and dichloroacetylene, exerts potent acute cellular toxicity in LLC-PK1 cells (Vamvakas S., Bittner, D., Dekant, W. and Anders, M.W. (1992). Events that precede and that follow S-(1,2-dichlorovinyl)-L-cysteine-induced release of mitochondrial Ca2+ and their association with cytotoxicity to renal cells. Biochem. Pharmacol. 44, 1131-1138). In the present study we investigated whether long-term exposure of LLC-PK1 cells to low, non-cytotoxic concentrations of DCVC results in stable morphological and biochemical dedifferentiation. After 7 weeks exposure to 1 and 5 microM DCVC, morphologically changed single cells were picked under the microscope and cultured in absence of DCVC for 4-8 weeks. In contrast to the physiological cuboidal shape of untreated LLC-PK1 cells, the clones derived from long-term exposure to DCVC consisted of elongated, spindle-shaped cells tending to form irregular borders. Moreover, glucose uptake, pH-dependent ammonia production and dome formation, important indicators of the renal tubule origin of the LLC-PK1 cells, were severely impaired in the clones. In addition to the loss of membrane polarity, the clones exhibited altered composition of the nuclear matrix and intermediate filament proteins by two-dimensional gel electrophoresis, increased poly(ADP-ribosyl)ation of nuclear proteins and enhanced expression of c-fos. The induction of dedifferentiated LLC-PK1 clones with stable characteristics upon long-term exposure to the nephrocarcinogen DCVC may represent a useful in vitro model to study biochemical alterations involved in chronic renal toxicity and carcinogenicity.
Vamvakas
S
S
Institut für Toxikologie, Universität Würzburg, Germany.
Richter
H
H
Bittner
D
D
D000641:Ammonia / Q000378:metabolism; D000818:Animals; D034961:Antigens, Nuclear; D002454:Cell Differentiation / Q000187:drug effects; D002999:Clone Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D003545:Cysteine / Q000031:analogs & derivatives* / Q000633:toxicity; D015870:Gene Expression / Q000187:drug effects; D016762:Genes, fos / Q000235:genetics; D005947:Glucose / Q000378:metabolism; D015151:Immunoblotting; D007381:Intermediate Filament Proteins / Q000032:analysis; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D018374:LLC-PK1 Cells; D009687:Nuclear Proteins / Q000032:analysis; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D013347:Subcellular Fractions / Q000737:chemistry; D013552:Swine; D013997:Time Factors
D034961:Antigens, Nuclear; D007381:Intermediate Filament Proteins; D009687:Nuclear Proteins; C039961:S-(1,2-dichlorovinyl)cysteine; D000641:Ammonia; D011065:Poly(ADP-ribose) Polymerases; D005947:Glucose; D003545:Cysteine