Gene expression profiling of hypoxia signaling in human hepatocellular carcinoma cells.

Cellular, local, and organismal responses to low O2 availability occur during processes such as anaerobic metabolism and wound healing and pathological conditions such as stroke and cancer. These responses include increases in glycolytic activity, vascularization, breathing, and red blood cell production. These responses are mediated in part by the hypoxia-inducible factors (HIFs), which receive information on O2 levels from a group of iron- and O2-dependent hydroxylases. Hypoxia mimics, such as cobalt chloride, nickel chloride, and deferoxamine, act to simulate hypoxia by altering the iron status of these hydroxylases. To determine whether these mimics are appropriate substitutes for the lower O2 tension evoked naturally, we compared transcriptional responses of a Hep3B cell line using high-density oligonucleotide arrays. A battery of core genes was identified that was shared by all four treatments (hypoxia, cobalt, nickel, and deferoxamine) including glycolytic enzymes, cell cycle regulators, and apoptotic genes. Importantly, cobalt, nickel, and deferoxamine influenced transcription of distinct sets of genes that were not affected by cellular hypoxia. These global responses to hypoxia indicate a balancing act between adaptation and programmed cell death and suggest caution in the use of hypoxia mimics as substitutes for the low O2 tension that occurs in vivo.

The role of hypoxia inducible factor 1alpha in cobalt chloride induced cell death in mouse embryonic fibroblasts.

Cobalt has been widely used in the treatment of anemia and as a hypoxia mimic in cell culture and it is known to activate hypoxic signaling by stabilizing the hypoxia inducible transcription factor 1alpha (HIF1alpha). However, cobalt exposure can lead to tissue and cellular toxicity. These studies were conducted to determine the role of HIF1alpha in mediating cobalt-induced toxicity. Mouse embryonic fibroblasts (MEFs) that were null for the HIF1alpha protein were used to show that HIF1alpha protein plays a major role in mediating cobalt-induced cytotoxicity. Previous work from our lab and others has shown that two BH3 domain containing cell death genes, BNip3 and NIX, are targets of hypoxia signaling. These experiments document that BNip3 and NIX expression is HIF1alpha-dependent, and cobalt induces their expression in a time and dose dependent manner. In addition, their expression is correlated with an increase in BNIP3 and NIX protein. Characteristically, the elevated level of BNIP3 was correlated with an increased presence of chromatin condensation, one marker for cell injury. Interestingly, this increased chromosomal condensation was not coupled to caspase-3 activation as usually seen in a typical apoptotic response. These results show that HIF1alpha is playing a major role in mediating cobalt-induced toxicity in mouse embryonic fibroblasts and may offer a possible mechanism for the underlying pathology of injuries seen in workers exposed to environmental contaminants that can influence the hypoxia signaling system, such as cobalt.