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GAP JUNCTION COMMUNICATON IN A TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS
Citation:
Wang, X., D E. House, J. Page, AND C F. Blackman. GAP JUNCTION COMMUNICATON IN A TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS. Presented at Bioelectromagnetic Society Annual Meeting, Quebec, Canada, 06/23-27/2002.
Description:
GAP JUNCTION COMMUNICTION IN TRANSFECTED HUMAN CELL LINE: ACTION OF MELATONIN AND MAGNETIC FIELDS.
OBJECTIVE: We previously showed that functional gap junction communication (GJC), as monitored by dye transfer (DT), could be enhanced in mouse C3H 10T112 cells and in mouse primary hepatocytes by treatment with physiological levels of melatonin (Mel), and that magnetic fields could reduce the Mel- induced effect. The objective of this study is to test the ability of physiological levels of Mel to enhance DT in human-derived HeLa cells that had been transfected with the connexin 32 (Cx 32) gene, thereby rendering them DT -competent. A pilot test was performed to determine if the GJC enhancement caused by Mel could be affected by exposure to magnetic fields as we had previously observed.
METHODS: HeLa cells transfected with the Cx 32 gene were obtained from Mesnil et al. (IARC, Lyon, France). The cells were maintained as per methods described by those estigators. Confluent monolayers of cells were treated with 0, 0.1 , 0.4. 1 or 4 nM Mel in complete growth medium for 24 hours. In a pilot experiment. cells were exposed to 4 nM Mel for 24 hours then exposed to sinusoidal magnetic fields, 38.4 JlT DC and parallel 45-Hz AC at 24.4 JlTrms, for 3hr. Following all treatments. the cells were micro injected with Lucifer yellow dye and the percentage of nearest neighbor cells to which the DT was determined.
RESUL TS: The amount of DT was statistically enhanced (p<.05) for cells treated with 0.1 nM Mel (88.0% +1- 3.8 SE) and with 4 nM Mel (90.3% +1- 3.4 SE) compared to the untreated controls (73.2% +1- 4.7 SE). DT in cells treated with 0.4 and 1 nM Mel (76.4% +1- 4.8 SE and 68.9% +1- 5.5 SE. respectively) was not different from the untreated cells. Fifty cells were tested in each treatment category .In the pilot study, cells treated with 4 nM Mel and subsequently with magnetic fields showed no change in DT compared to similar cells not exposed to magnetic fields.
DISCUSSION: Connexin genes have recently been shown to display tumor suppression properties, presumably through the activity of their protein products that form gap junctions, allowing GJC between adjacent cells. One measure of GJC is the transfer of Lucifer yellow fluorescent dye among cells. Putative and known tumor promoting chemicals can reduce GJC. and it has been hypothesized that such reduction is a significant event in the tumor promotion process. We have shown that melatonin. a hormone with oncostatic properties, can enhance GJC in a cell line (10T112 cells) and in mouse primary hepatocytes, and that magnetic fields can remove that GJC enhancement as measured by Dr. In this presentation we show that human cells transfected with connexin 32 gene so as to perform DT, can also respond to Mel with increased Dr at physiological levels of Mel. However, unlike the other cells showing this property, this Cx 32-transfected, human cell line does not respond to magnetic field exposure by reducing the increased DT caused by Mel. This result could be interpreted as demonstrating that cells requiring a transfected gene to express GJC, although responsive to Mel, may not have all the regulatory elements other cells have to control the DT. An alternative possibility is that only specific connexins are responsive to magnetic field treatment. Since 1 or112 cells express Cx 43, whereas mouse hepatocytes express Cx 26 and Cx 32, it is possible that only Cx 26 and Cx 43 are sensitive to magnetic field treatment. Although more study is needed to establish the basis for this finding, it is apparent that magnetic fields can be used to explore the biochemical control features of GJC regulation.
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