SHORT-TERM EFFECTS OF THYROID HORMONE IN PRENATAL DEVELOPMENT AND CELL DIFFERENTIATION

Extranuclear effects of thyroid hormones, L-T3 and L-T4, normally show a time-course of seconds to minutes, and are probably mediated by specific receptors on the plasma membrane. Since the liver is an important target for thyroid hormones, we have investigated their extranuclear effects on chick embryo hepatocytes at 14 and 19 days of embryonic life. At these stages of their development the hepatocytes show different protein expression patterns and activities and, of particular interest here, different levels of deiodinases. Three essential membrane transport activities were studied: the Na+/H+ exchanger, the amino acid transport System A (AIB transport), and the Na+/K+-ATPase; the signal transduction mechanisms involved were examined. We also focused on the possibility that 3,5-diiodo-L-thyronine (3,5-T2) might represent a new hormone able to mimic the extranuclear effects of thyroid hormones.The Na+/H+ exchanger type 1 (NHE-1) is a ubiquitous plasma membrane integral protein showing both housekeeping functions (modulation of intracellular pH and cell volume) and a regulative role, being activated by growth factors, hormones, integrins and hyperosmolarity. Both L-T3 and L-T4 stimulated the Na+/H+ exchanger over a wide concentration range, giving a bell-shape dose response as already found for other hormones (i.e insulin). The maximum increase of intracellular pH was observed at 1 nM hormone concentration, for both 14 and 19 days old cells, and 3,5-T2 was able to mimic the non genomic effect of thyroid hormones, but less efficiently. A different pattern was found for the Na+-dependent amino acid transport System A, which is highly regulated by hormones, growth factors and amino acid deprivation. At variance with the Na+/H+ exchanger, the activation of AIB transport by thyroid hormones appeared related to the proliferative activity and to the stage of embryonal development, being highly significant at 19 days, when the embryo is close to term, and insensitive to thyroid hormones at the 14 days stage. The effect of 3,5-T2 on System A activity was less evident. Yet another behavior was seen for the third transporter studied; since both L-T3, and its analog 3,5-T2 inhibited the Na+/K+-ATPase in chick embryo hepatocytes at different stages of development, in a dose-dependent way. Signal transduction appears to be mediated by interplay among kinases: PKA, PKC, the MAPK pathway, together with second messengers such as calcium, IP3 and DAG. These data allow us to draw the following conclusions on the role of extranuclear effects of thyroid hormones in cell development and differentiation: First, chick embryo hepatocytes are able to respond to thyroid hormones activating the same transduction pathway as growth factors (i.e. EGF). Second, 3,5-T2 is able to mimic thyroid hormone effects on the three transport systems. This capability of 3,5-T2 might be of physiological relevance as a safety factor, in the case of a possible hypothyroidism in the prenatal period. The results obtained so far indicate that in our experimental system thyroid hormones and 3,5-T2 via a short-term mechanism exert modulating effects on membrane transport activities that are important for development and differentiation. Interestingly all three systems are modulated also by nuclear genomic action.