Hemopexin from four species inhibits the association of heme with cultured hepatoma cells or primary rat hepatocytes exhibiting a small number of species specific hemopexin receptors

Hemopexin (Hx) binds heme with a very high affinity (Kd<0.1 pmol/L). It has been implicated as a major vehicle for the transport of heme into liver cells, involving a receptor-mediated recycling mechanism. However, previous studies indicated that heme is not taken up by cultured embryonic chick or adult rat hepatocytes by such a mechanism, because heme added as heme hemopexin failed to affect heme-responsive activities of 5-aminolevulinic acid synthase and heme oxygenase. Here, we investigated the importance of hemopexin in hepatic heme uptake in cultured rat hepatocytes and human HepG2 hepatoma cells, and determined the number and species specificity of hemopexin receptors on the rat hepatocytes. We also tested whether there is a difference between heterologous and homologous hemopexins. We found the following: 1) heme is inhibited from associating with hepatocytes by apo hemopexins from rat, human, rabbit, and chicken; 2) heme readily associates with hepatocytes when heme hemopexin preparations are added in which the ratio of heme to hemopexin exceeds 1.0; 3) heme induces heme oxygenase mRNA in rat hepatocytes and this induction is prevented by excess hemopexin; and 4) rat hepatocytes exhibit only about 2,000 hemopexin receptors per cell when using rat hemopexin, and none when using hemopexin of rabbit and human. We conclude that hemopexin plays a limited role in heme uptake by cultured hepatocytes and hepatoma cells, and that heme which exceeds the hemopexin binding capacity is taken up directly from heme-albumin.