| ObjectiveWith the development of science and technology, more and more bond-relatedmaterials were widely used in medical research and which also contribute to clinicalresearch. However, no matter the quality or quantity of current materials couldn’tsatisfy the scientific demand which change with each passing day, not to mention thebiocompatibility Therefore, the more urgent key to current problem is seeking a newadhesive material which can be effective and mass-produced and be fine inbiocompatibility. According to the strong adhesion characteristics that mussels canadhere to surface of various substances, mussel byssus protein(Mefp) and the keyfactor L-3,4-dihydroxyphenyl alanine (L-DOPA) were chose to adhered gingivalepithelial cell. At the same time, some methods of culture primary gingival epithelialcells were compared. Cell morphology, growth curve, adhesion amount and theconcentration of calcium ion in cell were observed to evaluate the role of musselbyssus protein and L-DOPA in the culture of gingival epithelial cells. The aim of thestudy was seeking an effective way to promote primary culture and subculturegingival epithelial cells. And aim at providing theoretical and technical basis for theclinical research of gingival epithelial cells.Method1. Single dispase enzyme digestion, single tissue slides covering and co-methodswere used to culture gingival epithelial cells of Beagle canines. Cell morphology andgrowth time were observed and compared to optimal the best mean of cell culture.2. Petri dishes were inoculated with a certain amount of cell after mussel byssusproteins, L-DOPA, Poly-L-Lysine (PLL) and distilled water treated, respectively.Then calculate the cell adhesion amount by4,6-Diamidino-2-Phenylindole (DAPI)staining counting. Measure the Optical Density (OD) value by enzyme markinstrument and draw the cell growth curve. Observe cell morphology by scanning electron microscopy (SEM). And analyze the cell proliferation ability by measuringthe concentration of the calcium ion in cell. And all of them were compared with nonetreatment.Result1. Few gingival epithelial cells are adhesive in simple enzyme digestion and thesurvival rate is0.0%. Simple tissue slide covering has more cells adhesive, butfibroblast pollution is serious and the survival rate is only30.5%. Enzyme digestioncombined with tissue slides covering method can get a lot of gingival epithelial cellsat a shorter time and there are none fibroblast contamination. The survival rate is up to91.6%. Chi-square test showed that the survival rate of each group is statisticallysinificant（P<0.05）.2. Compared with the blank group, group treated by mussel byssus protein havea high amount of DAPI-stained cells which are oblate in cell morphology. The48hOD values and the concentrated of calcium ion are both higher（P<0.05）. What’smore, there is a typical S-shaped curve in mussel byssus protein groups. However, allof these are same to poly-lysine treated groups（P>0.05）. Compared with the blankgroup, group treated by L-DOPA has a high amount of DAPI-stained cells. The ODvalues of cells by48h and the concentrated rate of calcium ion are higher too（P<0.05）. And there is a typical S-shaped curve in L-DOPA groups. But thequantity of DAPI-stained cells and concentrated rate of calcium ion（P<0.05） arelarger than groups treated by poly-lysine which are oblate in cell morphology but littlepseudo foots.Conclusions1. Enzyme digestion combined with tissue slides covering method could get a lotof gingival epithelial cells, simply and effectively, in a short time.2. Mussel byssus protein and L-DOPA have same effect on improving theadhesion of gingival epithelial cells with PLL. |
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