Incorporation of [3H]leucine into haptoglobin and albumin by isolated rat hepatocyte suspensions was studied to assess the role of potential mediators of the acute-phase reaction in promoting haptoglobin synthesis. In vitro, in the presence of insulin, the addition of a hormone mixture containing hydrocortisone, glucagon, somatotropin, and triiodothyronine resulted in a 70% increase in leucine incorporation into haptoglobin relative to albumin at 48 h incubation. A variety of agents, selected because they are considered to play some part in the acute-phase reaction, were added to the medium, and similar measurements of leucine incorporation were made. The specific binding to hepatocytes by asialo- and asialo, agalacto-derivatives of haptoglobin or orosomucoid did not affect synthesis of haptoglobin or albumin. Epinephrine, prostaglandins E1 and E2, bacterial lipopolysaccharide, and sera containing active complement components also failed to stimulate relative haptoglobin synthesis. A partially purified preparation of human leukocytic pyrogen, however, caused a 70% increase in leucine incorporation into haptoglobin relative to albumin in the present of the hormone mixture, suggesting that this substance may affect acute-phase protein synthesis.