Aberrant redox regulation in human metastatic melanoma cells compared to normal melanocytes

Melanocytes and melanoma cells contain melanin, a complex polymer that modu lates redox changes in these cells. Relative intracellular hydrogen peroxid e levels measured by dichlorodihydrofluorescein are similar in the two cell types, but the levels of superoxide anion measured by dihydroethidium were markedly increased in melanoma cells. Chelator-induced oxidative stress is efficiently suppressed by melanocytes without substantial recruitment of t he transcription factors NF-kappaB and AP-1 as measured by electrophoretic mobility shift assay and quantitated by densitometry or by a change in freq uency of apoptosis as determined by annexin V binding. In contrast, NF-kapp aB in melanoma cells is strongly recruited by changes in redox status and e xhibits a correlative relationship to intracellular hydrogen peroxide (but not superoxide anion). However, the response of the NF-kappaB pathway to in tracellular hydrogen peroxide is anomalous, including downregulation of p65 and I kappaB alpha RNA expression (Northern blot). Additionally, recruitme nt of AP-1 binding in melanoma cells was directly correlated with intracell ular levels of superoxide anion (but not hydrogen peroxide). Neither the de gree of NF-kappaB nor AP-1 binding in melanoma cells was related to the fre quency of apoptosis. The responsiveness of NF-kappaB and AP-1 recruitment t o intracellular levels of hydrogen peroxide and superoxide anion without co ncomitant control of apoptosis provides a general mechanism by which these cells can escape noxious injury (e.g., chemotherapy). The marked enhancemen t of apoptosis in melanoma cells by chelators indicates, however, that this alteration can be circumvented and offers a unique therapeutic window to e xplore. (C) 2001 Elsevier Science Inc.