不同配比冻存液对兔脂肪干细胞液氮冻存效果的研究
时间：2022-08-01 10:16:00
[摘要]目的：探究不同浓度配比冻存液对脂肪干细胞（Adipose-derived Stem Cells，ADSCs）冻存复苏后存活率及增殖活性的影响。方法：无菌条件下取新西兰大白兔的脂肪组织，体外分离、培养ADSCs并传代至第3代，行免疫细胞化学染色对其表面标志物进行鉴定，同时对其行成软骨及成表皮细胞诱导以证实其多项分化能力。将第3代ADSCs置于液氮（-196℃）中冻存，根据冻存液中培养基（DMEM）、胎牛血清（Fetal bovine serum，FBS）及二甲基亚砜（Dimethyl sulfoxide，DMSO）的配比不同分为8组：①含不同浓度FBS配比分组：A组（70% DMEM +20% FBS+10% DMSO）、B组（60% DMEM +30%FBS+10%DMSO）、C组（50%DMEM +40%FBS+10%DMSO）、D组（含有40%DMEM +50%FBS+10%DMSO）；②含不同浓度DMSO配比分组：E组（55%DMEM +40%FBS+5%DMSO）、F组（50%DMEM+40%FBS+10%DMSO）、G组（45%DMEM +40%FBS+15%DMSO）、H组（40%DMEM +40%FBS+20%DMSO）。细胞冻存三个月后，对其进行复苏并观察细胞形态学变化；通过细胞计数计算细胞存活率；采用MTT法绘制复苏后细胞生长曲线，以测定其生长增殖情况。结果：细胞复苏后，通过细胞计数计算结果显示C、D组存活率明显高于A、B组（P
[关键词]脂肪来源干细胞；冻存液；液氮
[中图分类号]Q813.1 [文献标志码]A [文章编号]1008-6455（2015）09-0029-05
Abstract： Objective To investigate the ability to maintain the proliferation of Adipose-derived Stem Cells（ADSCs）in different concentration ratio of frozen liquid. Methods Fat isolated from healthy rabbits was isolated，cultured and then generated in vitro for 3 passage into the ADSCs， P3 ADSCs were identified by detecting of adipose stem cell surface markers expression.The third passage cells were preserved in liquid nitrogen for 3 months，ADSCs were divided into eight group in order to cryopreservation：groupA：70%DMEM+20%FBS+10%DMSO，groupB：60%DMEM+30%FBS+10%DMSO，groupC：50%DMEM+40%FBS+10%DMSO，groupD：40%DMEM+50%FBS+10%DMSO，groupE：55%DMEM+40%FBS+5%DMSO，groupF：50%DMEM+40%FBS+10%DMSO，groupG：45%DMEM+40%FBS+15%DMSO，groupH：40%DMEM+40%FBS+20%DMSO.After cryopreservation three months， observed the cells after recovering，and compared with the group not frozen.The growth curves of the cryopreserved cells were analyzed by MTT assay. Results The results of immunocytochemistry
showed ADSCs were negative for CD34，CD106 and positive for CD29，CD44.Trypan blue staining and microscopic counting of results showed that Group C and Group D had higher cell survival rate than Group A and Group B with significant difference（P
Key words：adipose-derived stem cells；Cryoprotectants；liquid nitrogen
showed ADSCs were negative for CD34，CD106 and positive for CD29，CD44.Trypan blue staining and microscopic counting of results showed that Group C and Group D had higher cell survival rate than Group A and Group B with significant difference（P
Key words：adipose-derived stem cells；Cryoprotectants；liquid nitrogen
