High Glucose Induced Apoptosis In NIT-1 Cells Detected By Laser Tweezers-Raman Spectroscopy

Objective By observing the changes in the Raman spectra information with the increase of apoptosis rate, to reveal the potential cellular biology which are reflected by the change of spectra and the potential mechanisms of apoptosis in NIT-1 cell induced by high glucose , and to explore a novel approach for study the apoptosis cells.Methods Cells were divided into 7 groups: NG0 and HG0, cells were in the logarithmic phase cells; NG1 and NG2, cells were cultured in normal medium contained glucose 11.1 mmol/L respectively for 2 days and 4 days; HG1 and HG2, cells were cultured after 2 days and 4 days respectively in medium containing high concentration glucose of 25 mmol/L; HG3, cells were the floating cells which floated in the medium after cultured in high concentration glucose medium. All groups'apoptosis rate were measured by the flow cytometry, and obtained spectra by the laser tweezers-Raman spectroscopy system. Raman spectra of cells were dealt with back ground subtraction,soothing,baseline correction,normalization and then analyzing the differences among those 7 groups. Principal component analysis (PCA) was used to identify the spectra of the logarithmic phase and the floating NIT^-1 cells.Results 1.The apoptosis rate of NIT^-1 cells increased significantly in groups which treated with high glucose medium, and it also increased with the elongation of cultured time,P<0.01. 2.The results of the flow cytometry indecated that most of the floating cells were in the state of terminal apoptosis. 3.The Raman spectra show that the area of peaks at 1001 cm^-1 and 1337 cm^-1 decreased significantly in high glucose medium groups when prolonged the cultured time, P<0.05. 4. Compared with logarithmic phase cells, the floating cells were significantly decreased in the area of peaks at 714 cm^-1,781 cm^-1,1001 cm^-1,1086 cm^-1,1125 cm^-1,1337 cm^-1,1447 cm^-1,1655 cm^-1,P<0.01. 5. The logarithmic phase and the floating NIT^-1 cells can be distinguished by PCA.Conclusions 1. High concentration glucose could induce NIT^-1 cell apoptosis. 2. Proteins and nucleic acid are less in apoptotic cells than in normal cells which can be reflected by Raman spectroscopy. 3. Raman spectroscopy system which analysis by PCA can efficient to distinguish normal NIT^-1 cells from late apoptotic NIT^-1 cells.