Necroptosis, also known as “programmed necrosis”, has emerged as a critical factor in a variety of pathologicaland physiological processes and is considered a cell type-specific tightly regulated process with mechanismsthat may vary rather greatly due to the change of cell line. Here we used HT-29, a human colon cancer cellline, to establish a necroptosis model and elucidate associated mechanisms. We discovered that cobalt chloride,a reagent that could induce hypoxia-inducible factor-1α(HIF1α) expression and therefore mimic the hypoxicmicroenvironment of tumor tissue in some aspects induces necroptosis in HT-29 cells when caspase activityis compromised. On the other hand, apoptosis appears to be the predominant death form when caspases arefunctioning normally. HT-29 cells demonstrated significantly increased RIPK1, RIPK3 and MLKL expressionin response to cobalt chloride plus z-VAD treatment, which was accompanied by drastically increased IL1αand IL6 expression, substantiating the notion that necrosis can induce profound immune reactions. The RIPK1kinase inhibitor necrostatin-1 and the ROS scavenger NAC each could prevent necrosis in HT-29 cells andthe efficiency was enhanced by combined treatment. Thus by building up a necroptosis model in human coloncancer cells, we uncovered that mechanically RIP kinases collaborate with ROS during necrosis promoted bycobalt chloride plus z-VAD, which leads to inflammation. Necroptosis may present a new target for therapeuticintervention in cancer cells that are resistant to apoptotic cell death.
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