Cultured Ito cells of rat liver express the α 2 ‐macroglobulin gene

Ito cells were isolated from rat liver and kept in culture for up to 13 days. The capability of the Ito cells to synthesize α 2 ‐macroglobulin was analyzed at different times after isolation and by pulse‐chase experiments. Newly synthesized α 2 ‐macroglobulin was determined by immunoprecipitation followed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and fluorography. α 2 ‐Macroglobulin synthesis was hardly detectable in Ito cells and their media 3 days after plating. However, 5–11 days after the isolation of the cells, increasing amounts of α 2 ‐macroglobulin were synthesized. The results of pulse‐chase experiments performed on day 7 showed that radioactively labeled α 2 ‐macroglobulin decreased in the intracellular compartment and increased in the culture medium. α 2 ‐Macroglobulin was identified by immunoprecipitation and sodium dodecyl sulfate/polyacrylamide gel electrophoresis under reducing and non‐reducing conditions. Furthermore, when unlabeled α 2 ‐macroglobulin was added during the immunoprecipitation a competition was observed. Incubation of pancreatic elastase with culture medium of rat Ito cells or rat hepatocytes led to the same cleavage products as found with α 2 ‐macroglobulin. α 2 ‐Macroglobulin‐specific mRNA could be demonstrated by Northern blot analysis of total RNA extracted from rat Ito cells. Under the conditions where α 2 ‐macroglobulin was synthesized in Ito cells, no synthesis of α 1 ‐macroglobulin, α 1 ‐inhibitor 3, α 1 ‐proteinase inhibitor, α 1 ‐acid glycoprotein, α 1 ‐acute‐phase globulin (T‐kininogen) and albumin could be demonstrated. It is concluded that α 2 ‐macroglobulin is a true secretory protein of rat Ito cells in culture. This could be of importance for collagen metabolism in liver diseases.