It is proposed to investigate the means whereby growth-inhibited non-transformed fibroblasts are able cause growth inhibition of neoplastically transformed fibroblasts. The biological model used is the 10T1/2 cell culture system which closely reflects events occurring in vivo. In this system cAMP enhances cell interactions leading to growth control. The goal is to better understand the physiological control of cell division at the cell population level so that growth control process can be manipulated in normal and neoplastic cells.
The aims of this proposal are to critically examine the role of gap-junctions in transferring growth regulatory information between cells, and to amass evidence for or against the cAMP as the chemical messenger of growth control signals. The first phase of the study will be to determine in neoplastic cells sensitive or resistant to growth inhibition by contacting normal cells, whether cAMP can be transferred to the neoplastic cell and whether the extent of this transfer correlates with the extent of growth inhibition. To detect transferred cAMP we shall utilize a sensitive micro-fluorometric assay for the detection of free catalytic units liberated by the action of cAMP on cAMP dependent protein kinase. We will next examine alterations in protein phosphorylation in general and of gap junctional components in particular by polyacrylamide gel electrophoresis, in homogeneous cultures of normal 10T1/2 cells in which drug induced cAMP elevation also enhances growth control. Correlations between enhanced growth control, protein phosphorylation and gap junctional permeance, measured by a dye-injection technique, will be examined, and the effects of agents blocking communication such as the tumor promoters evaluated. Using the information gained in studies of normal cells, the question of whether similar events occur in growth inhibited neoplastic cells will be examined. Using fluorescent activated cell sorting, neoplastic cells will be separated from normal cells, their position in the cell cycle determined and the phosphorylation state of marker proteins determined. A secondary question to be addressed is whether carcinogen-initiated cells when growth arrested by normal cells are in a biological and biochemical state corresponding to the initiated or neoplastic state. By these studies of routes and means of informational transfer between cells, it is hoped to help identify defects that result in noeplasia. This research should narrow the search for the function of oncogene products, and suggest ways to correct these defects.
| Matsukawa, T; Bertram, J S (1988) Augmentation of postconfluence growth arrest of 10T1/2 fibroblasts by endogenous cyclic adenosine 3':5'-monophosphate. Cancer Res 48:1874-81 |
| Mehta, P P; Bertram, J S; Loewenstein, W R (1986) Growth inhibition of transformed cells correlates with their junctional communication with normal cells. Cell 44:187-96 |
