Study On The Cytotoxicity Of The Two-photon Nucleic Acid Molecular Light Switch MVEC In Vitro

The nucleic acid molecular light switch, an important chromosome probe, has been researched extensively. As a light switch molecule, when it is combined into the DNA, its fluorescent signal can be dramatically enhanced. Thus the chromosome combined with light switch molecules can emit bright fluorescence on a background of dark and the uncombined light switch molecules can not make the biological tissues observed. It is very useful to detect and image the chromosome in living cell.. When the molecular light switch possess two-photon absorption properties, It will be able to play a major role to the fluorescence micro image of the biological specimen .To achieve a two-photon nucleic acid molecular light switch, two molecules with cationic and containing N groups derived from 9-ethylcarbazole, MVEC and BMVEC, have been designed, synthesized and characterized by Institute of Crystal Materials of Shandong University .Moreover the N group in the carbazole has been alkylated to increase the two-photon absorption properties of both molecules. They are a new kind of Two-photon nucleic acid molecular light switch for double-strands DNA. By study the site of their max absorption peaks and fluorescent intensity after they combine with dsDNA,we can achieve some useful structural information of DNA Therefore this kind of dye has the very good application prospect.Moreover, we have confirmed MVEC also could enter the living cell , penetrate the cell membrane and combine with the dsDNA emitting blue fluorescence .It can be directly observed under the fluorescence microscope. Theoretically, MVEC can play a important role on the living cells' chromosome research. However, as living specimen dye. Not only can cause living cell or the organization specificity staining, but also have no cytotoxicity or little cytotoxicity and avoid effecting cell's activity. As a result, the research on the biology characteristic (such as toxicity) of MVEC is a principal mission currently. Up to now, no documents or papers, concerning that if MVEC bear cytotoxicity and how much MVEC effect the cell, are reported. This thesis is carrying on thereby, which will provide biology and medicine circle with the experiment basis by researching the biological characteristic of MVEC on its cytotoxicity .The thesis aims at: researching the dyeing characteristic of dsDNA molecular light switch MVEC, and its proliferation inhibition and apoptosis-induce action on human cervical cancer Hela cell and the mouse lung cancer Lewis cell, as well as its acting mechanism. Method: Observing localization of MVEC in the cells with confocal/ fluorescence microscope; Estimating the cytotoxicity of MVEC and the toxic effects-dose/time relationship by MTT; observing the variance in apoptosis-dependant index of MVEC by flow cytometry Annexin V/PI double staining method and agarose gel electrophoresis.This paper obtained following several significant results: 1st, MVEC can cause dose-dependent and time-dependent cells growth inhibition. 2nd, confirmed the reason that MVEC possess cell toxicity is because it has induced apoptosis of hela cell. Moreover DNA ladder were detected by Agarose gel electroghoresis. 3rd,It is defined that optimum concentration of MVEC in which MVEC takes the lowest cytotoxicity and then gets expanded application, such as cell proliferation experiment, estimation of cell division cycle, detecting cell transplantation in vivo etc.