Aim
To investigate the effects of (-)epigallocatechin-3-gallate (EGCG), the main polyphenol in green tea, on cell growth, cell cycle and phosphorylated nuclear factor-κB (pNF-κB) expression in neonatal human dermal fibroblasts (nHDFs).
Conclusion
EGCG, through uptake into cytoplasm, reversibly regulated the cell growth and expression of cell cycle-related proteins and genes in normal fibroblasts.
Methods
The proliferation and cell-cycle of nHDFs were determined using WST-8 cell growth assay and flow cytometry, respectively. The apoptosis was examined using DNA ladder and Annexin V-FITC assays. The expression levels of pNF-κB and cell cycle-related genes and proteins in nHDFs were measured using cDNA microarray analyses and Western blot. The cellular uptake of EGCG was examined using fluorescence (FITC)-labeled EGCG (FITC-EGCG) in combination with confocal microscopy.
Results
The effect of EGCG on the growth of nHDFs depended on the concentration tested. At a low concentration (200 μmol/L), EGCG resulted in a slight decrease in the proportion of cells in the S and G(2)/M phases of cell cycle with a concomitant increase in the proportion of cells in G(0)/G(1) phase. At the higher doses (400 and 800 μmol/L), apoptosis was induced. The regulation of EGCG on the expression of pNF-κB was also concentration-dependent, whereas it did not affect the unphosphorylated NF-κB expression. cDNA microarray analysis showed that cell cycle-related genes were down-regulated by EGCG (200 μmol/L). The expression of cyclins A/B and cyclin-dependent kinase 1 was reversibly regulated by EGCG (200 μmol/L). FITC-EGCG was found to be internalized into the cytoplasm and translocated into the nucleus of nHDFs.