5′‐Nucleotidase Activates and an Inhibitory Antibody Prevents Neuritic Differentiation of PC12 Cells

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Abstract
Ecto-5'-nucleotidase catalyses the hydrolysis of AMP at the surface of a variety of cells whereas it is absent from others. In addition to its catalytic activity, a function in neural development and also its interaction with extracellular matrix proteins has been reported. In order to further elucidate the biological function of ecto-5'-nucleotidase we have investigated the effect of 5'-nucleotidase on nerve growth factor-induced differentiation of PC12 cells. Furthermore, we compared the effect of an inhibitory versus a non-inhibitory monospecific antibody against the enzyme on neuritic differentiation and survival of PC12 cells that constitutively express the enzyme. When coverslips are coated with the soluble form of ecto-5'-nucleotidase in addition to collagen, there is a considerable increase in nerve growth factor-induced neurite length during the first 24 h of culture. Addition of an antibody to a culture medium that inhibits 5'-nucleotidase activity to 33% of control values dramatically reduces the number of neurites per cell within 3 days of culture. The cells round up, cluster and eventually die. On the contrary, another antibody that had no significant effect on enzyme activity affected neither nerve growth factor-induced neurite formation nor survival of PC12 cells. Addition of adenosine (200 nM, 10 or 20 microM) to the culture medium did not influence PC12 cell differentiation. The effects induced by the inhibitory antibody could be only partially prevented by simultaneous application of adenosine. Our results suggest that 5'-nucleotidase is essential for nerve growth factor-induced neurite outgrowth and survival of PC12 cells.(ABSTRACT TRUNCATED AT 250 WORDS)