The long-term objective of our work is to discover growth-regulatory differences between neoplastic and nonneoplastic cells that can be exploited in chemotherapy. One aspect of growth regulation in nonneoplastic cells that has been intensively investigated recently is the role of increases in free intracellular calcium (Cai) and pH (pHi). Our previous grant award has helped develop the technology to measure Cai and pHi in individual cells. The short-term objective of our studies is to utilize this technology to determine how the increases in Cai and/or pHi are correlated with -- and, in certain instances, required for -- growth stimulation of quiescent nonneoplastic fibroblasts (BALB/c 3T3 and human cells).
The specific aims i nclude: 1) Examination of the role of ionic signals in the growth stimulation of nonneoplastic cells in monolayer culture. For competence factors, we will focus on platelet-derived growth factor (PDGF), fibroblast growth factor (FGF) and comitogens, such as tumor promoters (TPA) or calcium ionophore (A23187). Our studies have already indicated that a transient Cai increase is required for PDGF-stimulated DNA synthesis in quiescent BALB/c 3T3 cells. The present application focuses on defining what other intracellular signals (e.g., DAG) potentiate this mitogenic effect of Cai increase and what early genes are activated by these specific signals. For progression factors, we will study and compare EGF, insulin, IGF-1 and plasma. We will concentrate on determining the signalling pathways involved in the dependence of progression events on extracellular Ca2+. 2) Definition of the role of ionic signals in the growth-resistance of nonneoplastic cells in suspension culture. We will determine which of the intracellular signals induced by growth factors does not occur in suspended cells. 3) Examination of ionic signals and gene activation in neo-plastic cells. We will determine which of the early intracellular signals are absent or constitutively activated in neoplastic cells (e.g., 3T12, ras- transformants, sis-transformants). These tests will help us to determine whether gene activation stimulated by ionic signals (Cai and pH1) is an important aspect of the normal growth regulation that is controlled by exogenous growth factors and attachment to substratum, and whether deregulation of these signals results in the unrestrained growth of neoplasms.
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