发育期牙根端复合体细胞条件培养液对脂肪干细胞增殖、分化的影响

摘要

目的：初步探讨发育期根端复合体（developing apical complex，DAC）条件培养基（DAC condi-tioned medium，DACCM）对脂肪干细胞（adipose tissue-derived stem cells，ADSCs）增殖和分化的影响。方法：取出生20 d SD大鼠分离其下颌磨牙DAC，并采用组织块联合酶消化法培养DAC细胞；经HE、免疫组化染色鉴定DAC后，收集原代培养的DAC细胞上清，分别用过滤和不过滤的方式制备不同条件培养基并用其对ADSCs进行培养；分别于培养3、5、7 d 各时间点，MTT法检测ADSCs增殖能力；用碱性磷酸酶（ALP）试剂盒及反转录酶-聚合酶链反应（RT-PCR）检测ADSCs的ALP蛋白活性及其基因的表达水平。结果：免疫组化染色显示，培养的DAC细胞CK-14、vimentin均为阳性表达；DACCM-未滤培养组ADSCs的增殖活性、ALP蛋白活性以及ALP基因表达水平均高于单纯α-MEM培养组和 DACCM-过滤培养组，差异均有统计学意义（P＜0．05）。结论：不经过滤的DAC细胞上清制备的条件培养基对ADSCs增殖、分化的促进作用更明显。%AIM:To study the effects of the developing apical complex (DAC)cells conditioned medium (DACCM)on the proliferation and differentiation of adipose tissue-derived stem cells (ADSCs).METHODS:DAC cells,separated from the 20 d postnatal SD rat,were cultured by tissue block combined with enzyme digestion method. DACs was identified by HE staining and immumohistochemical staining.DACCM was collected and prepared by filtra-tion and none-filtration (DACCM-F and DACCM-UF)respectively.The proliferation of ADSCs was detected by MTT assay.The mRNA and protein expression of alkaline phosphatase (ALP)were detected by RT-PCR and ALP detection Kit respectively.SPASS13.0 was used for data analysis.RESULTS:DAC cells were obtained from the developing roots of the SD rat.The cells were CK-14 and vimentin positive.The proliferation,ALP mRNA and protein expression of ADSCs cultured in DACCM-UF exceeded those of the cells cultured in α-MEM or DACCM-F(P<0.05 )CON-CLUSION:DACCM-UF may stimulate the proliferation and differentiation of ADSCs.