Effects of colchicine and vincristine on polymorphonuclear leucocyte function in vitro

Abstract

The effect of the drugs colchicine and vincristine on normal polymorphonuclear leucocytes (PMNs) was investigated in vitro. These drugs are known to interfere with the organization of microtubules both in the mitotic spindle of dividing cells, causing metaphase arrest, and in the cytoplasm of other non-dividing cells affecting a number of normal cellular functions. PMNs were obtained from healthy volunteers by centrifugation of heparinized blood over Ficoll Isopaque gradients followed by dextran sedimentation and hypotonic lysis of residual erythrocytes. The preparations were greater than 95% pure PMNs, and cell viability was 99% as assessed by Trypan blue exclusion. Four aspects of PMN function were studied in vitro—chemotaxis, phagocytic activity, hexosemonophosphate (HMP) shunt activity and superoxide production. Chemotaxis was measured in a Boyden chamber system using 3.0 Millipore filters and zymosan activated serum as the chemotactic stimulus. Phagocytosis was measured by the ability of PMNs to ingest heat killed candida albicans and the phagocytic index was expressed as the number of yeast cells per PMN. HMP shunt activity in both resting and stimulated cells was determined by measuring the oxidation of glucose – 1 – 14C with release of 14CO2. Superoxide production by PMNs was measured on the basis of its ability to reduce cytochrome C. Ten separate experiments were performed and the results obtained for PMNs exposed to either colchicine (2.5 × 10-4M) or vincristine (1 × 10-6’M) were compared to controls. Chemotaxis, phagocytic activity and superoxide production were all significantly reduced in the presence of either colchicine or vincristine. Stimulated HMP shunt activity was also reduced, but resting PMNs were not affected. The data indicate that these drugs have multiple effects on PMN function which may be important in the in vivo PMN response. The effect of the drugs colchicine and vincristine on normal polymorphonuclear leucocytes (PMNs) was investigated in vitro. These drugs are known to interfere with the organization of microtubules both in the mitotic spindle of dividing cells, causing metaphase arrest, and in the cytoplasm of other non-dividing cells affecting a number of normal cellular functions. PMNs were obtained from healthy volunteers by centrifugation of heparinized blood over Ficoll Isopaque gradients followed by dextran sedimentation and hypotonic lysis of residual erythrocytes. The preparations were greater than 95% pure PMNs, and cell viability was 99% as assessed by Trypan blue exclusion. Four aspects of PMN function were studied in vitro—chemotaxis, phagocytic activity, hexosemonophosphate (HMP) shunt activity and superoxide production. Chemotaxis was measured in a Boyden chamber system using 3.0 Millipore filters and zymosan activated serum as the chemotactic stimulus. Phagocytosis was measured by the ability of PMNs to ingest heat killed candida albicans and the phagocytic index was expressed as the number of yeast cells per PMN. HMP shunt activity in both resting and stimulated cells was determined by measuring the oxidation of glucose – 1 – 14C with release of 14CO2. Superoxide production by PMNs was measured on the basis of its ability to reduce cytochrome C. Ten separate experiments were performed and the results obtained for PMNs exposed to either colchicine (2.5 × 10-4M) or vincristine (1 × 10-6’M) were compared to controls. Chemotaxis, phagocytic activity and superoxide production were all significantly reduced in the presence of either colchicine or vincristine. Stimulated HMP shunt activity was also reduced, but resting PMNs were not affected. The data indicate that these drugs have multiple effects on PMN function which may be important in the in vivo PMN response.