Quiescent LLC-PKi Cells as a Model for aVDiamminedichloroplatinum(II) Nephrotoxicity and Modulation by Thiol Rescue Agents1

Confluent LLC-PKi cells express many characteristics of renal prox imal tubule epithelia. We report here the use of this cell line to investigate the possible mechanisms of m-diamminedichloroplatinum(II) (1)1)1')induced nephrotoxicity and diethyldithiocarbamate (DDTC) ameliora tion. Cells were exposed to platinum-based drug for l h and then incubated in drug-free medium until assayed. There was a 10-h delay between DDP exposure and onset of toxicity which then continued to develop. Viability at 72 h was 97 ±2 (SD), 68 ±3, 33 ±3, and 10 ± 2% of control after treatment with 100, 200, 300, and 400 JIM DDP, respectively. rra/u-Diamminedichloroplatinum(II) was 5-fold less toxic than DDP and diammine(l,l-cyclobutanedicarboxylato)platinum(II) (CBDCA) was nontoxic at concentrations up to 2.0 mM. Incubation of cells with DDTC (3 HIM)for l h immediately following DDP exposure resulted in chelation of 43% of total miraceliular platinum by DDTC and significantly increased 72 h viability; 97 ±2, 88 ±3, and 42 ±2% of control for 200, 300, and 400 *tMDDP, respectively. DDP treatment of 1.1210 cells yielded equivalent total intracellular platinum levels, but Pt(DDTC)2 concentrations were one-tenth those in LLC-PK, cells after subsequent treatment with DDTC (3 HIM). Immediate reduction of protein and RNA, but not DNA, synthesis by DDP was concentration dependent over the same range as viability. DDP-induced increases in unscheduled DNA synthesis also did not correlate with cytotoxicity. The inhibition of protein synthesis was un changed by pretreatment with the RNA synthesis inhibitor actinomycin D. DDTC (3 HIM)exposure produced an immediate and persistent DDP dose modification of 1.6 in protein but not RNA synthesis. CBDCA (0.5 to 1.0 m\i) had no effect on protein, RNA, or DNA synthesis and only slight stimulatory effects on unscheduled DNA synthesis. DDTC alone (3-3000 MM) caused significant reduction in DNA synthesis and unsched uled DNA synthesis. Neither sodium thiosulfate nor 2-mercaptoethanesulfonate had any effect on DDP-induced cytotoxicity or inhibition of protein, RNA, or DNA synthesis when incubated immediately after DDP, even though these drugs achieved intracellular concentrations at which DDTC was protective. These data indicate thai quiescent LLC-PKi cells are a good in vitro model for the study of DDP-induced nephrotoxicity and its modulation by thiol rescue agents and thai DDP inhibilion and DDTC rescue of posttranscriptional processes correlate best with viability in LLC-PKi