Loss of regulation of cell cycle events mediated by changes in cytosolic Ca$\sp{2+}$ ion activity has been implicated in the progression of normal cells to neoplasia. In this study, the Ca$\sp{2+}$ buffer 5,5$\sp\prime$-difluoro 1,2-bis(2-aminophenoxy) ethane-N,N,N$\sp\prime N\sp\prime$-tetra-acetic acid (5,5$\sp\prime$-dfBAPTA, abbreviated "dfB") has been used to modulate cell division in transformed and primary mouse keratinocytes. Exogenous application, via the tetra(acetoxymethyl) ester ("AM"), of 18-20 $\mu$M dfB/AM to the growth media of transformed cells inhibits cell division and DNA synthesis, without compromising the cells' viability, as shown by $\sp3$H-thymidine incorporation and flow cytometry. Bulk fluorimetry shows that cells treated with dfB/AM are able to buffer Ca$\sp{2+}$ in proportion to the concentration of dfB/AM applied. Primary cultured cells treated with 18-20 $\mu$M dfB/AM die within 3 hours of treatment. Viable dfB/AM treated cultures of transformed cells have a higher proportion of cells in the G$\sb2$ phase of the cell cycle than do controls, as shown by flow cytometry. This result, in combination with that showing reduced $\sp3$H-thymidine incorporation, suggests that 18-20 $\mu$M dfB/AM inhibits a pre- or mid-mitotic step. Light, electron, and confocal microscopies show 18-20 $\mu$M dfB/AM-treated cells to have prominent, thickened nuclear envelopes along with actin cytoskeletons that are distinguishable from controls. Upon return to medium that does not contain dfB/AM, treated transformed cells gradually resume their pre-treatment growth and division patterns.
| Identifer | oai:union.ndltd.org:UMASS/oai:scholarworks.umass.edu:dissertations-7655 |
| Date | 01 January 1996 |
| Creators | Cishek, Dawn M |
| Publisher | ScholarWorks@UMass Amherst |
| Source Sets | University of Massachusetts, Amherst |
| Language | English |
| Detected Language | English |
| Type | text |
| Source | Doctoral Dissertations Available from Proquest |
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