油红O染色法在2D和3D细胞培养原代胰腺星形细胞鉴定中的应用

Application of oil red O staining in identification of primary pancreatic stellate cells cultured in 2D and 3D medium

目的通过油红O染色分别在2D和3D细胞培养中观察及鉴定活化和去活化的原代胰腺星形细胞(PSCs)。方法使用爬出法从新鲜胰腺癌组织中培养原代PSCs,2D培养于普通细胞培养板中进行,3D培养在Matrigel基质胶中进行。使用全反式维甲酸(ATRA)连续处理PSCs 7天,获得去活化PSCs。使用油红O法分别对2D培养和3D培养的活化和去活化PSCs进行染色。结果在2D培养中,未处理的PSCs呈活化状态,星芒状,胞质几乎无油红O着色;ATRA处理的PSCs处于去活化状态,呈椭圆形或多角形,胞质中可见较多橘红色颗粒着色。在3D培养中,Matrigel胶滴呈半球形,胶滴中的PSCs为椭圆形,PSCs未经ATRA处理胞质中即有较多橘红色颗粒,维持去活化状态。结论油红O染色可以在2D和3D细胞培养中鉴定原代PSCs的活化和去活化状态,且PSCs在3D环境中生长可维持去活化状态。

Objective To assess and identify activated and deactivated primary pancreatic stellate cells(PSCs)by oil red O staining in 2D and 3D cell cultures.Methods The climb-out method was used to culture primary PSCs from fresh pancreatic cancer tissue,2D culture in ordinary cell culture plates,and 3D culture in Matrigel.PSCs were deactivated by treatment with all-trans retinoic acid(ATRA)for 7 days.PSCs in 2D and 3D cultures were stained using the oil red O method.Results In 2D culture,activated PSCs were star-shaped and had almost no cytoplasmic oil red O,whereas deactivated cells were oval or polygonal and more orange-colored granules in cytoplasm.In 3D culture,Matrigel drops were hemispherical,and PSCs in the drops were oval.PSCs without ATRA treatment in 3D culture had obviously more orange granules in the cytoplasm.Conclusion Oil red O staining can identify the activation status of primary PSCs in 2D and 3D cell cultures.PSCs in a 3D environment maintains the deactivation status without ATRA treatment.