Susceptibility of insulinoma cells to cadmium and modulation by L-type calcium channels

Abstract

Cadmium (Cd), a toxic metal that induces apoptosis and necrosis in a variety of cells, accumulates in pancreas and may be a cause of diabetes in humans. In the insulinoma cells line HIT-T15 (HIT), we measured internal calcium (Ca) and Cd levels by the fluorescent dye Fura-2 and confirm that L-type voltage-dependent calcium channels (VDCC) play a major role in glucose response and represent a pathway of Cd influx in these cells. Therefore we examined the role of VDCC in acute Cd poisoning by comparing its accumulation and cytotoxic effect in HIT cells and in epithelial-like VDCC-free HeLa cells. Cultures were incubated with 10-300 microM Cd for 15 min-6 h. While negligible at the end of the treatment, HIT cell death was evident after 18-24 h, and it was time-, dose- and serum-dependent. Short (< or = 60 min) Cd treatments with lower doses (< or = 100 microM in serum-free medium) induced delayed apoptotic cell death, as demonstrated by DNA fragmentation on agarose gels and segmentation of DAPI-stained nuclei. Longer incubations and/or higher concentrations caused mainly necrosis. The same treatments were largely harmless in HeLa cells, in which neither death nor DNA fragmentation was observed. The Ca antagonist nimodipine was capable to prevent HIT cell death at lower doses of Cd and to restore the apoptotic condition at higher doses, indicating that reduction of Cd flux through VDCC modulates Cd toxicity. These data demonstrate a specific sensitivity to Cd of insulinoma cells that can be significant for pancreatic beta-cell pathology.

Role of Calcium Channels in Heavy Metal Toxicity

“…VDCCs mediate Cd influx in excitable cells [15–18], including mammalian neurons [19], and have been proposed to participate in Cd uptake also in cells from nonexcitable tissues [20]. In a previous work of my laboratory, we showed that in certain cells Cd permeation occurs mainly through VDCC of the L-type [19, 21]. However, in other studies, the presence of VDCC per se did not seem to enhance sensitivity to Cd; for example, VDCCs expressing (PC12) and nonexpressing (PC18) cells showed comparable LD 50 for Cd [18], and the role of this pathway in the induction of cell death appears questionable.…”

Different membrane modifications revealed by atomic force/lateral force microscopy after doping of human pancreatic cells with Cd, Zn, or Pb

“…Such interpretation is strongly supported by the results obtained by many groups which reported the induction of apoptosis after Cd or Zn treatments in pancreatic cells as well as in several other cell lines at similar, or identical, metal concentrations (De la Fuente et al, 2002;Gavazzo et al, 2005;Koh, 2001;Pathak and Khandelwal, 2006;Watjen and Beyersmann, 2004;Wiseman et al, 2006).…”

“…Interestingly, recent findings (Priel and Hershfinkel, 2006) indicate that such safety systems are unable to respond efficiently in the presence of higher (close to 100 lM), non physiological, concentration of Zn which is still capable of triggering the cell damage. On the other hand, both Zn and Cd ions can cross the membrane of insulinoma cells through the voltage-dependent Ca channels and, in particular, the induction of apoptosis by 100 lM of Cd has been recently demonstrated (Gavazzo et al, 2005).…”

A Review of Diabetes Mellitus and Exposure to the Environmental Toxicant Cadmium with an Emphasis on Likely Mechanisms of Action

“…Using the insulinoma cell line, HIT-T15, Cd accumulation was significantly greater when cells were depolarized in a 20 mM KCl solution. In the presence of the L-type Ca channel inhibitor, nimodipine, Cd accumulation was significantly less [44]. In the same study, nimodipine reduced apoptotsis in HIT-T15 cells exposed to Cd.…”