300 MHz proton NMR study of the differentiation of diploid human epidermal keratinocytes.

The nuclear magnetic resonance spin-lattice (T1) and spin-spin (T2) relaxation times are closely related to the molecular motions of the molecules in a liquid sample. T1 and T2 of human epidermal cells were measured at 300 MHz as functions of harvesting methods (i.e., scraping vs trypsinization) and age in culture. It was found that T1 and T2 values have smaller variances when the cell is harvested by trypsinization rather than scraping. The correlation coefficients for both T1 and T2, obtained from cells harvested by scraping. More importantly, this is the first report to monitor in vitro aging through relaxation times measurement. There is a significant increase in the values of T1 and T2 from the third to seventh passages. Human keratinocytes slowed down and even ceased to grow the seventh passage. Therefore, the cellular water molecules of human keratinocytes have higher mobility in a more differentiated state. The factors contributing to the change in relaxation times as cells progress toward senescence are discussed.