外伤脱位牙保存介质及温度对人牙周膜细胞活性和增殖能力的影响

Effects of storage media and temperature on the viability and proliferation capacity of human periodontal ligament cells

目的:通过体外细胞培养模拟牙脱位保存模型,研究不同保存介质和保存温度对人牙周膜细胞活性和增殖能力的影响。方法:选择3种易于获得的保存介质牛奶、唾液和纯净水,分别在室温和4℃条件下作用于人牙周膜成纤维细胞(hPDLFs)1 h,然后利用5-乙炔基-2′脱氧尿嘧啶核苷(EdU)和细胞增殖检测试剂盒(CCK-8)实验检测作用后hPDLFs细胞的活性和后期增殖能力。结果:EdU实验结果显示,在4℃保存温度下,牛奶处理hPDLFs细胞1 h后,EdU阳性细胞(绿色)最多,约占总细胞的50.9%。CCK-8实验结果显示,4℃下牛奶、唾液、纯净水中hPDLFs活力依次约为62.8%、17.8%和3.3%,室温下则依次为56.2%、15.2%和0.7%。培养24、48、72 h后,细胞保存于4℃牛奶中生长曲线最接近于正常对照,与保存于唾液和纯净水中的细胞相比,细胞增殖速率差异具有统计学意义。结论:hPDLFs在4℃牛奶中表现出了最佳的细胞活力和增殖能力,因此低温牛奶是外伤脱位牙首选的保存液。

Objective:To investigate the effects of storage media and temperature on the activity and proliferation of human periodontal ligament fibroblasts(hPDLFs)by simulating the storage model of an avulsed tooth in vitro cell culture.Methods:Three readily available storage media(milk,saliva,and purified water)were selected and applied to hPDLFs for 1 h at room temperature and 4℃.Then,5-ethynyl-2′-deoxyuridine(EdU)and cell counting kit-8(CCK-8)were used to detect the viability and proliferative capacity of hPDLFs after treatment.Results:The EdU assay showed that after treating the cells with milk at 4℃for 1 h,most cells were EdU-positive(green),accounting for about 50.9%of the total cells.The CCK-8 assay showed that hPDLFs viabilities in milk,saliva,and purified water were 62.8%,17.8%,and 3.3%respectively,at 4℃,and 56.2%,15.2%,and 0.7%,respectively,at room temperature.The growth curve of cells stored in milk at 4℃was the closest to the normal control after 24,48,and 72 h of culture.The proliferation rate of cells stored in milk was statistically significantly different compared with that of cells stored in saliva and purified water.Conclusion:hPDLFs showed the best cell viability and proliferative ability in milk at 4℃.Therefore,low-temperature milk is the preferred preservation solution for avulsed teeth.