Bacterial antigen-induced release of white cell- and platelet-derived bioactive substances in vitro

Poor prognosis after resection of primary colorectal cancer may be related to the combination of perioperative blood transfusion and subsequent development of infectious complications. Various white cell- and platelet-derived cancer-growth substances may be involved in this process. Therefore, we studied the in vitro release of substances from white cells and platelets stimulated by bacterial antigens and supernatants from stored red-cell components.

Eight units of whole blood (WB) and 8 U of buffy-coat-depleted red-cell (SAGM) blood were donated by healthy blood donors. Subsequently, one-half of each unit was leucocyte-depleted by filtration, and all 32 half-units were stored under standard conditions for 35 d. Just after storage, and on d 7, 21, and 35 during storage, aliquots of the supernatants were removed from the units and frozen at -80 degrees C. WB from other healthy donors was stimulated for 2 h with sodium chloride (controls), with Escherichia coli (E. coli) lipopolysaccharide (LPS) alone, or with LPS plus supernatants from the WB units (diluted 1:10), or from the SAGM units (diluted 1:20) stored for 0, 7, 21, or 35 d, respectively. Similar assays were performed using Staphylococcus aureus-derived protein A as a stimulatory antigen. The concentration of eosinophil cationic protein (ECP), myeloperoxidase (MPO), histamine (HIS), and plasminogen-activator inhibitor-1 (PAI-1) were determined in supernatants from the stored blood and in assay supernatants by using enzyme-linked immunosorbent assay (ELISA) and radioimmunoassay (RIA) methods.

The extracellular concentration of ECP, MPO, and HIS increased significantly in a storage-time-dependent manner in nonfiltered WB and SAGM blood, and the increase was abrogated by prestorage leukofiltration. Similarly, PA-1 increased significantly in nonfiltered WB, and the increase was abrogated by prestorage leukofiltration. The supernatant concentrations of the four substances were significantly increased in LPS-stimulated (0.5-4 fold) and in protein A-stimulated (0.5-13.5-fold) assays compared with controls. The addition of supernatants from stored nonfiltered WB or SAGM blood significantly increased the assay supernatant of ECP, MPO, HIS, and PAU-1 concentrations storage-time-dependently in LPS-stimulated assays. Prestorage leukofiltration abrogated the additional effect of supernatants from stored blood. Similar results were observed for ECP and HIS through the addition of supernatants from stored blood to protein A-stimulated assays. Protein A stimulation did not lead to increased PA-1 release in assays diluted by supernatants from stored blood. However, the MPO concentrations were significantly (p = 0.004), and independent of storage time and leukofiltration, increased in protein A-stimulated assays diluted by supernatants from stored blood compared with sodium chloride dilution.

Extracellular ECP, MPO, HIS, and PA-1 accumulate during storage of nonfiltered red-cell components, but the accumulation can be prevented by prestorage leukofiltration. In addition, bacterial antigens appear to induce significant release of the substances from white cells and platelets. Addition of supernatants from stored, nonfiltered WB and SAGM blood may increase the substance levels in a storage-time-dependent manner, and prestorage leukofiltration may prevent further increase by supernatants, except for MPO.