基于3D微肿瘤模型探讨乳腺癌微环境及耐药性  

Exploring breast cancer microenvironment and chemoresistance based on 3D in vitro microtumour models

作　　者：王娟如 宋巧稚 刘晓利 吴正升[1,3] Wang Juanru;Song Qiaozhi;Liu Xiaoli;Wu Zhengsheng(Dept of Pathology,The First Affiliated Hospital of Anhui Medical University,Hefei230022;The First Clinical College,Anhui Medical University,Hefei230032;Dept of Pathology,Anhui Medical University,Hefei230032)

机构地区：[1]安徽医科大学第一附属医院病理科,合肥230022 [2]安徽医科大学第一临床医学院,合肥230032 [3]安徽医科大学病理学教研室,合肥230032

出　　处：《安徽医科大学学报》2024年第11期2004-2012,共9页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金项目(编号:81972472);安徽医科大学基础与临床合作研究提升计划资助项目(编号:2020xkjT012)。

摘　　要：目的利用液滴微流控技术构建3D微肿瘤模型来模拟肿瘤微环境。通过3D微肿瘤的制造、表征和对化疗药物的敏感性测试,探究其作为体外乳腺癌研究模型的可行性。方法将人乳腺癌细胞用水凝胶壳封装在富含胶原蛋白的微胶囊核心中,获得体外乳腺癌微肿瘤;将乳腺癌微肿瘤通过Transwell系统与3D微胶囊化的内皮细胞进行共培养。通过显微镜直接观察微肿瘤的结构和生长特征;采用CCK-8实验检测不同培养模型下细胞的增殖情况以及对阿霉素的药物敏感性;采用流式细胞术比较增殖过程细胞凋亡的差异;采用划痕实验和Transwell实验评估细胞迁移和侵袭能力的差异;采用Western blot法检测细胞上皮-间充质转化(EMT)相关蛋白的表达。结果乳腺癌细胞在水凝胶核-壳微胶囊中生长良好;细胞增殖实验结果显示,3D培养和3D共培养细胞增殖速度显著低于2D培养;与2D培养相比,3D培养和3D共培养细胞迁移和侵袭能力增强并显示出更高的EMT相关蛋白表达;与2D培养相比,3D培养和3D共培养细胞对化疗药物的敏感性显著降低。结论3D培养表现出与体内肿瘤相似的形态结构和生物学特征且对化疗药物耐药性更高。Objective To simulate the tumor microenvironment though the 3D microtumor model which was constructed using droplet microfluidics.To explore its feasibility as a model for in vitro breast cancer research through 3D microtumour fabrication,characterisation and sensitivity testing to chemotherapeutic drugs.Methods Breast cancer cells were encapsulated with hydrogel shells in collagen-rich microencapsulated cores to obtain breast cancer microtumours in vitro;breast cancer microtumours were co-cultured with 3D microencapsulated endothelial cells by Transwell system.The structure and growth characteristics of the microtumours were directly observed by microscopy;the CCK-8 assay was used to detect the proliferation of the cells under different culture models and the drug sensitivity to doxorubicin;flow cytometry was used to compare the differences in apoptosis during the proliferation process;and the differences in the migratory and invasive abilities of the cells were assessed by scratch assay and Transwell assay;the expression of epithelial-mesenchymal transition-related proteins was detected by Western blot.Results Breast cancer cells grew well in hydrogel nucleus-shell microcapsules;cell proliferation assays showed that 3D culture and 3D co-culture cells proliferated at a significantly lower rate than 2D culture;3D culture and 3D co-culture cells had enhanced migration and invasion ability and showed higher expression of EMT-related proteins compared to 2D culture;3D culture and 3D co-culture cells were significantly less sensitive to chemotherapeutic drugs compared to 2D culture.The sensitivity of 3D and 3D co-cultured cells to chemotherapeutic drugs was significantly reduced compared to 2D culture.Conclusion 3D cultures show similar morphology and biology to in vivo tumours and are more resistant to chemotherapeutic agents.

关 键 词：3D培养 共培养 核-壳微胶囊 肿瘤微环境 化疗耐药 

分 类 号：R73-354[医药卫生—肿瘤]