PGE1-independent MDCK cells have elevated intracellular cyclic AMP but retain the growth stimulatory effects of glucagon and epidermal growth factor in serum-free medium

Abstract

Prostaglandin E₁ (PGE₁), a component in the hormone‐supplemented, serumfree medium for the Madin Darby canine kidney (MDCK) cell line, has been proposed to increase MDCK cell growth by increasing intracellular cyclic AMP levels. The association between increased intracellular cyclic AMP and the growth stimulatory effect of PGE₁ has been examined in normal MDCK cells and in PGE₁‐independent variants of MDCK. These variant cells have lost the PGE₁ requirement for long term growth in defined medium. Normal MCDK cells had almost twofold higher intracellular cyclic AMP levels during growth in Medium K‐1 (9.0 pmol/mg protein) than in Medium K‐1 minus PGE₁. Furthermore, PGE₁‐independent clone 1 had higher intracellular cyclic AMP levels in Medium K‐1 minus PGE₁ than normal MDCK cells in Medium K‐1. This latter observation suggests that the PGE₁ requirement for MDCK cell growth is associated with the low intracellular cyclic AMP levels of this cell line. An involvement of cyclic AMP in the growth response to PGE₁ is supported by these observations, as well as by the growth stimulatory effects of other agents that affect cyclic AMP metabolism in MDCK cells. These agents include glucagon, isobutyl methylxanthine (IBMX), and dibutyryl cyclic AMP. The growth of PGE₁‐independent clone 1 was inhibited rather than stimulated by PGE₁. Similarly, PGE₁‐independent cell growth was inhibited by IBMX and dibutyryl cyclic AMP. However, the growth response to one agent which increases cyclic AMP (glucagon) was retained in PGE₁‐independent clone 1. This result suggests that the effect of glucagon is not associated with increases in intracellular cyclic AMP. The growth stimulatory effect of epidermal growth factor (EGF) on normal MDCK cells was also studied. Although EGF does not act via a cyclic AMP‐mediated mechanism, EGF increased normal MDCK cell growth and substituted for PGE₁ in Medium K‐1. Thus, EGF and PGE₁ could possibly affect similar growth‐related functions in MDCK cells, although by different pathways. This possibility was examined further, using PGE₁‐independent clone 1. EGF, like glucagon, was still growth stimulatory to the PGE₁‐independent cells. Consequently, the biochemical pathways by which EGF and PGE₁ increase MDCK cell growth probably do not converge.