Primary cultures of rabbit renal proximal tubule cells. III. Comparative cytotoxicity of inorganic and organic mercury.
The present study further developed primary cultures of rabbit renal proximal tubule cells (RPTC) as an in vitro model to study chemical-induced toxicity by investigating the comparative cytotoxicity of mercuric chloride (HgCl2) and methyl mercury chloride (CH3HgCl) to RPTC. Confluent monolayer cultures of RPTC exposed to HgCl2 and CH3HgCl for 24 hr exhibited a concentration-dependent loss in cell viability at culture medium concentrations greater than 25 and 2.5 microM, respectively. Vital dye exclusion was a more sensitive indicator of cytotoxicity than the amount of lactate dehydrogenase activity, alkaline phosphatase activity, N-acetylglucosaminidase activity, and protein content remaining on the culture dish. On the basis of vital dye exclusion, HgCl2 was less toxic to proximal tubule cells in culture than CH3HgCl after 24 hr of exposure, whether cytotoxicity was based on LC50 values (34.2 microM HgCl2 vs 6.1 microM CH3HgCl) or total cellular mercury uptake (4.6 nmol Hg2+/10(5) cells vs 1.25 nmol CH3Hg+/10(5) cells). Differences in the extent and rate of metal uptake were also evident. Maximum cellular uptake of Hg2+ occurred within 6-24 hr after exposure and was not concentration-dependent, whereas maximum uptake of CH3Hg+ occurred within 3 hr of exposure and was concentration-dependent. The intracellular distribution of both mercurials between acid-soluble and acid-insoluble binding sites also differed. At noncytotoxic concentrations of HgCl2 (0.04-5 microM), intracellular Hg2+ bound increasingly to acid-soluble binding sites as a function of time, from 15-30% after 6 hr of exposure to 40-60% after 72 hr of exposure. However, at subcytotoxic (25 microM) and cytotoxic (34.2 microM) concentrations, Hg2+ binding to acid-soluble binding sites remained constant at approximately 30-40% for 6, 12, 24, and 72 hr after exposure. In contrast, only 20% of total cellular CH3Hg+ was bound to acid-soluble binding sites after exposure to 0.039 to 6.1 microM CH3HgCl for 6, 12, and 24 hr. Total cellular glutathione content was unaffected after exposure to 0.04-5 microM HgCl2 and 0.039-6.1 microM CH3HgCl, but was depleted 6 hr after exposure to 25 and 34.2 microM HgCl2. These results indicate that CH3HgCl was a more potent cytotoxicant to RPTC in primary culture than HgCl2. Furthermore, compared to Hg2+, the low binding of CH3Hg+ to acid-soluble binding sites and the absence of a redistribution of CH3Hg+ from acid-insoluble to acid-soluble binding sites appeared to contribute to its more potent toxicity to cultured cells.(ABSTRACT TRUNCATED AT 400 WORDS)
Aleo
M D
MD
Department of Pharmacology and Therapeutics, State University of New York, Buffalo 14214.
Taub
M L
ML
Kostyniak
P J
PJ
D000818:Animals; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D005978:Glutathione / Q000378:metabolism; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008627:Mercuric Chloride / Q000493:pharmacokinetics / Q000633:toxicity*; D008670:Metals / Q000493:pharmacokinetics / Q000494:pharmacology; D008767:Methylmercury Compounds / Q000493:pharmacokinetics / Q000633:toxicity*; D011817:Rabbits; D014018:Tissue Distribution
D016895:Culture Media, Serum-Free; D008670:Metals; D008767:Methylmercury Compounds; D008627:Mercuric Chloride; D005978:Glutathione; C004925:methylmercuric chloride