大鼠阴茎海绵体干细胞的分离培养及鉴定
发布时间：2018-10-29 22:16
【摘要】： 目的对大鼠阴茎海绵体细胞进行初步分离培养,检测干细胞标志物干细胞抗原-1(Sca-1)、Oct4和肌源性标志物结蛋白(Desmin)的表达,探索阴茎海绵体中是否存在内源性干细胞及肌源性干细胞。 方法取2月龄SD大鼠,重量200～250g,乙醚吸入麻醉后,无菌条件下取阴茎海绵体组织,用Ⅰ型胶原酶、胰蛋白酶消化分离组织,待细胞基本以单细胞形态存在时,过滤、离心收集细胞沉淀,用含20!胎牛血清的DMEM重悬培养。采用改良的Preplate差速贴壁技术对细胞进行初步分离培养,所获PP1-PP6贴壁细胞,应用流式细胞仪、免疫荧光细胞化学法、western blot检测观察分离细胞中Sca-1、Oct4及Desmin的表达,同时行Sca-1/Oct4、Sca-1/Desmin、Oct4/Desmin免疫荧光双标记。 结果分离培养所获PP1和PP2细胞贴壁快,主要是长梭形的成纤维样细胞,PP3-PP5细胞贴壁能力逐渐减慢,出现短梭形细胞,有的呈多角形,以血管内皮细胞、平滑肌细胞为主,PP6细胞中有体积较小,成圆形的漂浮细胞,这种细胞贴壁很慢,2-3天后开始贴壁成圆形或梭形。流式细胞仪检测:PP3细胞中Sca-1、Oct4阳性表达量分别为0.3!±0.2!、0.2!±0.3!,与同型对照组相比均无统计学差异(p㧐0.05),Desmin阳性表达量为15.3!±1.5!,与同型对照组相比有统计学差异(p㩳0.05);PP6细胞中Sca-1、Oct4、Desmin阳性表达量分别为5.7!±0.7!、2.6!±0.3!、41.1!±1.6!,与PP3相比均有统计学差异(p㩳0.05)。免疫荧光细胞化学显示在PP6中有少量细胞分别表达Sca-1和Oct4,均散在分布,Sca-1以胞浆表达为主,Oct4则表达于胞核;表达Desmin细胞聚集,数量较多,主要表达于胞浆;亦发现有极少量双阳性标记细胞(Sca-1/Oct4、Sca-1/Desmin和Oct4/Desmin),Sca-1/Oct4表达于同一细胞的胞浆及胞核,Sca-1/Desmin表达于同一细胞的胞浆,Oct4/Desmin则表达于同一细胞的胞核及胞浆。Western blot显示PP1-PP5细胞中未检测到Sca-1及Oct4蛋白明显表达,但在PP6细胞中则检测到Sca-1及Oct4蛋白的表达,Desmin蛋白在PP1-PP6细胞中表达逐渐增强。 结论在大鼠阴茎海绵体中发现具有干细胞及肌源性干细胞表型的细胞,提示阴茎海绵体中存在内源性干细胞及肌源性干细胞。
[Abstract]:Objective to investigate the expression of stem cell antigen 1 (Sca-1), Oct4 and desmin (Desmin) in rat penis cavernous cells. To explore the existence of endogenous stem cells and myogenic stem cells in the cavernous body of penis. Methods two month old SD rats, weighing 200,250g, were taken from the corpus cavernosum of penis under aseptic condition after inhaled anesthesia with ether. The isolated tissues were digested by type 鈪，
本文编号：2298986
[Abstract]:Objective to investigate the expression of stem cell antigen 1 (Sca-1), Oct4 and desmin (Desmin) in rat penis cavernous cells. To explore the existence of endogenous stem cells and myogenic stem cells in the cavernous body of penis. Methods two month old SD rats, weighing 200,250g, were taken from the corpus cavernosum of penis under aseptic condition after inhaled anesthesia with ether. The isolated tissues were digested by type 鈪，
本文编号：2298986
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