| TGFβ Stimulation Assay
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Author:
Date:
2014-12-05
[Abstract] TGFβ is part of a growth factor superfamily which modulates cell growth, differentiation, adhesion, migration, ECM synthesis and apoptosis (Massague, 1998; Siegel and Massague, 2003). Free TGFβ binds to its high affinity TGFβ receptor, a receptor serine/threonine kinase, inducing phosphorylation of Smad2/3 which subsequently forms a complex with Smad4 to translocate to the nucleus where it interacts with multiple co-activators and repressors generating distinct transcriptional responses.
Indeed, TGFβ signaling shows a remarkable cellular context dependency and apparent multifunctionality: e.g. TGFβ is able to inhibit cell proliferation in many epithelial cells but can also enhance proliferation in fibroblasts and cell growth in endothelial cells (Guasch et al., ...
[摘要] TGFβ是调节细胞生长，分化，粘附，迁移，ECM合成和凋亡的生长因子超家族的一部分（Massague，1998; Siegel和Massague，2003）。游离TGFβ结合到其高亲和力TGFβ受体，一种受体丝氨酸/苏氨酸激酶，诱导Smad2/3的磷酸化，随后与Smad4形成复合物转移到细胞核，其中它与多种共激活剂和阻遏物相互作用产生不同的转录反应。实际上，TGFβ信号传导显示出显着的细胞环境依赖性和表观多功能性：例如TGFβ能够抑制许多上皮细胞中的细胞增殖，但也可以增强成纤维细胞中的增殖和内皮细胞中的细胞生长（Guasch等人，2007; Xiao等人，2012年）;它增强干细胞多能性，但促进其他细胞的分化（Park，2011）;在癌症发展中，它抑制恶变前细胞增殖，但同时促进转移到转移表型（Chaudhury和Howe，2009）。
TGFβ刺激测定监测细胞对TGFβ的反应性。在TGFβ刺激时，可以分析短期效应例如Smad2磷酸化和长期效应例如细胞增殖。将描述用于小鼠角质形成细胞的测定，其中TGFβ强烈抑制细胞增殖，但是这两种测定也适用于其它细胞类型。
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| Three-dimensional Invasion Assay
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Author:
Date:
2013-09-05
[Abstract] The invasive ability of cancer cells is a crucial function for cancer metastasis and the surrounding microenvironment of cancer cells in living tissues is three-dimension (3D). Therefore, to establish an in vitro invasion assay in a 3D system to predict cancer invasive ability is valuable in the research for cancer metastasis. Here, we describe a 3D invasion assay for observing the morphology and comparing the invasive ability of cancer cells in artificial 3D environments (Yang et al., 2012). Collagen I gels are used to cover on the top of cancer cells attached on coverslip glass dish and medium containing FBS is added as a chemoattractant. After incubation for a suitable time, the cells are fixed and stained. The invasion index can be calculated and the morphology can ...
[摘要] 癌细胞的侵袭能力是癌症转移的关键功能，活组织中癌细胞的周围微环境是三维的（3D）。 因此，在3D系统中建立体外侵袭测定以预测癌症侵袭能力在癌症转移的研究中是有价值的。 在这里，我们描述了用于观察形态学并比较人造3D环境中癌细胞的侵袭能力的3D侵袭测定（Yang等人，2012）。 胶原I凝胶用于覆盖附着在盖玻片玻璃皿上的癌细胞的顶部，并且添加含有FBS的培养基作为化学引诱物。 在孵育合适的时间后，将细胞固定并染色。 可以计算入侵指数，并且可以用激光共聚焦显微镜成像形态
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| MACS Isolation and Culture of Mouse Liver Mesothelial Cells
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Author:
Date:
2013-07-05
[Abstract] Mesothelial cells (MCs) form a single squamous epithelial cell layer and cover the surfaces of the internal organs, as well as the walls of cavities. The isolation of MCs is of great importance to study their function and characteristics for the understanding of physiology and pathophysiology of the liver. Glycoprotein M6a (GPM6A) was originally identified as a cell surface protein expressed in neurons and recently its expression was reported in epicardium and liver MCs (Wu et al., 2001; Bochmann et al., 2010; Li et al., 2012). Here we describe a method to isolate MCs from the adult mouse liver with anti-GPM6A antibodies. Under the low glucose and serum concentration, primary MCs grow and form epithelial colonies (Figure 1).
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[摘要] 间皮细胞（MCs）形成单个鳞状上皮细胞层，并覆盖内部器官的表面，以及腔壁。 MC的分离对于研究它们的功能和特征对于理解肝脏的生理学和病理生理学是非常重要的。 糖蛋白M6a（GPM6A）最初被鉴定为在神经元中表达的细胞表面蛋白，并且最近在心外膜和肝脏MC中报道了其表达（Wu等人，2001; Bochmann等人， ，2010; Li等人，2012）。 在这里我们描述了一种方法从成年小鼠肝脏用抗GPM6A抗体分离MCs。 在低葡萄糖和血清浓度下，原代MC生长并形成上皮集落（图1）。
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