UPTAKE OF CD-METALLOTHIONEIN INTO LLC-PK1 CELLS - A COMPARATIVE-STUDYWITH CDCL2

The uptake of the Cd-metallothionein complex (CdMT) into LLC-PK1 cells was investigated and compared with that of CdCl2. The cells were incu bated at 37 degrees C for up to 45 min with 1 mu M Cd, as either CdMT or CdCl2 at pH 5.5, 6.4, or 7.4. Under all the experimental conditions described below, the accumulation of Cd from CdMT was markedly lower than that from CdCl2. Cd accumulation at pH 7.4 from CdMT increased li nearly with the time of incubation, whereas Cd accumulation from CdCl2 was saturable, Metabolic inhibitors, 2,4-dinitrophenol (DNP) and carb onylcyanide p-(trifluoromethoxy)-phenylhydrazone (FCCP), and incubatio n at low temperature significantly decreased Cd accumulation from both Cd compounds, Coincubation with 30 mu M ZnCl2, an antagonist of CdCl2 uptake, slightly decreased Cd accumulation from CdMT, but it markedly decreased that from CdCl2. Cd accumulation from CdMT at pH 5.5 was si gnificantly higher than at pH 6.4 or 7.4, whereas Cd accumulation from CdCl2 at pH 5.5 and 6.4 was significantly lower than at pH 7.4. Altho ugh Cd accumulation from CdCl2 at pH 7.4 was significantly decreased b y coincubation with 100 mu M cysteine or glutathione (GSH), this decre ase was not observed at pH 5.5 or 6.4, A small amount of Cd was remove d, by the chelating agent EGTA, from the cell membranes after incubati on with CdMT at pH 6.4 and 7.4, whereas a considerable amount of Cd wa s removed by EGTA after incubation with CdMT at pH 5.5 and with CdCl2 at three pHs. It appears that the CdMT complex is taken up into LLC-PK 1 cells partially ria an energy-dependent process, and the increase in Cd accumulation at low pH is due to the liberation of Cd, High stabil ity and molecular size of the CdMT complex explains why it is not take n up readily into LLC-PK1 cells.