大鼠脂肪基质细胞分离培养.docVIP

【摘要】 目的 探讨不同大鼠脂肪基质细胞的培养、增殖与纯化方法，初步研究其生物学特性。方法 取大鼠皮下和肠系膜部位的脂肪组织，根据两种组织对胶原酶的敏感性不同，获取基质细胞，观察生长情况，并进行鉴定。结果 不同品系大鼠脂肪含量不同，皮下和肠系膜脂肪基质干细胞产率不同，生物学特性相似，贴壁细胞分两类： 第一类有明显油滴；第二类细胞内无明显油滴。第一类细胞经过传代，逐渐消失，第二类细胞所要获取的脂肪基质细胞，组化鉴定CD49d(+)和CD106(-)，为脂肪基质细胞表型。结论 建立了一种大鼠脂肪基质细胞的培养、增殖与纯化方法，不同部位来源的脂肪组织酶消化时间和细胞产率不同，同时发现所获得细胞分两类，性质有所不同，为脂肪基质细胞的相关研究打下一定的细胞生物学基础。 【关键词】 大鼠 脂肪 肪基质细胞 培养 Isolation，Culture and Identification of Rat Adipose?Derived Stem Cells In Vitro CHEN Lian?feng，LIN Xue，FANG Quan (Department of Cardiology，PUMCH，Chinese Academy of Medical Sciences and Peking Union Medical College，Beijing ，China) 【Abstract】 objective To explore the methodology regarding culture，proliferation and purification of adipose?derived stem cells (ADSCs)，and to study their biological characteristics. Methods Fat tissue was obtained from mesentery and subdermal tissues of rats of different lineages. The tissue samples were digested with collagenase to obtain adipose stromal cells. Cells were isolated and purified based on their different reaction to digestion. Immunohistochemistry was used to characterize and identify the harvested cells. Growth of cells was observed with an inverted contrast phase microscope. Results Despite different productivity，adipose stromal cells from different source tissues and different lineages showed identical biological characteristics. After 2?3 days of culture，floating cells became adherent and grew into clusters. After several passages，these cells separated and changed into spindle?shaped. The adherent cells could be further classified into two types. The first type of larger size contained profuse lipid droplets and highly positive by oil red O staining. The second type was of slimmer?shape，with little intracellular lipid and negative oil red O staining. After passages，remaining floating cells，albeit in a small number，continuesly differentiating into cells of the second type，while cells of first type gradually diminished. Assessed by immunohistochemistry，cells of the second typ