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The rates of secretion of complement C3, haptoglobin and plasminogen have been determined after pulse labelling with [3H]leucine, and compared to the secretion of prothrombin, albumin and transferrin investigated previously (Kvalvaag, A.H., Tollersrud, O.K. and Helgeland, L. (1988) Biochim. Biophys. Acta 937, 319-327). To study membrane association, rough microsomes were treated with increasing concentrations of saponin, sodium deoxycholate or Triton X-100. All six proteins were quantitated in the soluble and membrane fraction by enzyme immunoassays. At concentrations of saponin from 0.08% to 0.32%, each secretory protein showed a characteristic distribution, almost identical to that obtained with 0.05% sodium deoxycholate or 0.08% Triton X-100. Albumin and transferrin with half-times for secretion (t1/2) 30 and 75 min, respectively, are both almost exclusively found in the luminal fraction (greater than 95%). Prothrombin and plasminogen, which both show an intermediate t1/2 (approx. 55 min), are partially associated with the membranes, as only about 60% was released. Haptoglobin and complement C3 also show some association with the membranes (80-85% released). C3 is secreted at the same rate as prothrombin and plasminogen (t1/2 = 55 min), whereas haptoglobin is secreted more rapidly (t1/2 = 40 min). Accordingly, no correlation between kinetics of secretion and membrane association was demonstrated.

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.

A quantitative enzyme-linked immunosorbent assay was developed and utilized to study the stimulation of haptoglobin biosynthesis during an acute inflammatory challenge. A 10-fold increase in intracellular haptoglobin was measured at the peak of the inflammatory response. The increase in serum haptoglobin levels was concomitant with the intracellular levels, demonstrating the secretory output is also elevated during the inflammatory period. A monospecific antihaptoglobin was produced and used to detect the specific polysomes involved in haptoglobin synthesis. The amount of radioactively labeled antibody bound to the nascent haptoglobin chain was increased approx. 3-fold during the inflammatory response, indicating that new haptoglobin was being synthesized and suggesting an increase in functional haptoglobin mRNA resulting from the inflammatory signal.