No correlation between CD73 expression and ecto-5'-nucleotidase activity on blood mononuclear cells in vitro - Evidence of CD73 (ecto-5'-nucleotidase) on blood mononuclear cells with distinct antigenic properties

Monoclonal antibodies against ecto-5'-nucleotidase (ecto-5'-NT) have been clustered to CD73. The aim of the present study was to elucidate whether specific ecto-5'-NT activity on blood mononuclear cells (BMC) was correlated with CD73 expression measured by flow cytometry. During culture of CD73-negative lymphocytes the percentage of cells with ecto-5'-NT activity increased without there being a comparable increase in CD73 expression. After 2 days' culture. 9% (3-16, median and range given) demonstrated ecto-5'-NT activity, whereas CD73-positive cells comprised only 1.6% (0.0-3.0). These results show the existence of lymphocytes with ecto-5'-NT activity but without expression of CD73. The ecto-5'-NT activity increased significantly (p<0.02) during culture of unseparated BMC, whereas the number of anti-CD73 binding sites did not change. As a consequence, the number of anti-CD73 binding sites per U ecto-5'-NT activity decreased during culture. Exposure to interleukin-4 or prostaglandin E(2) changed the enzymatic activity but not the number of anti-CD73 binding sites. Treatment of BMC with phosphalidylinositol-specific phospholipase-C released 57% (51%-75%) of the ecto-5'-NT activity into the supernatants, without a detectable decrease in CD73 expression. The fact that ecto-5'-NT was removed from the supernatant after precipitation with anti-CD73 showed that the released ecto-5'-NT activity was due to enzymatic activity of CD73 molecules. The results of this study indicate that CD73 exists on BMC in isoforms with distinct capacities to bind mouse monoclonal anti-CD73.