Hemopexin protects cells lacking hemopexin receptors by tightly binding heme abrogating its deleterious effects and preventing nonspecific heme uptake, whereas cells with hemopexin receptors undergo a series of cellular events upon encountering heme-hemopexin. The biochemical responses to heme-hemopexin depend on its extracellular concentration and range from stimulation of cell growth at low levels to cell survival at otherwise toxic levels of heme. High (2-10 microM) but not low (0.01-1 microM) concentrations of heme-hemopexin increase, albeit transiently, the protein carbonyl content of mouse hepatoma (Hepa) cells. This is due to events associated with heme transport since cobalt-protoporphyrin IX-hemopexin, which binds to the receptor and activates signaling pathways without tetrapyrrole transport, does not increase carbonyl content. The N-terminal c-Jun kinase (JNK) is rapidly activated by 2-10 microM heme-hemopexin, yet the increased intracellular heme levels are neither toxic nor apoptotic. After 24 h exposure to 10 microM heme-hemopexin, Hepa cells become refractory to the growth stimulation seen with 0.1-0.75 microM heme-hemopexin but HO-1 remains responsive to induction by heme-hemopexin. Since free heme does not induce JNK, the signaling events, like phosphorylation of c-Jun via activation of JNK as well as the nuclear translocation of NFkappaB, G2/M arrest, and increased expression of p53 and of the cell cycle inhibitor p21(WAF1/CIP1/SDI1) generated by heme-hemopexin appear to be of paramount importance in cellular protection by heme-hemopexin.