海藻酸盐基复合水凝胶培养乳腺癌类器官中钙离子的关键作用  

Key role of calcium ion in sodium alginate based composite hydrogel for breast cancer organoid culture

作　　者：林志光 饶琪 梁珊珊[1,2,3] 王若雨 于炜婷[1,2,3] Lin Zhiguang;Rao Qi;Liang Shanshan;Wang Ruoyu;Yu Weiting(Department of Oncology,Affiliated Zhongshan Hospital of Dalian University,Dalian 116001,Liaoning Province,China;Zhongshan Clinical College of Dalian University,Dalian 116001,Liaoning Province,China;Dalian Key Laboratory of Translational Medicine of Marine Polysaccharide Biomaterials,Dalian 116001,Liaoning Province,China)

机构地区：[1]大连大学附属中山医院肿瘤中心,辽宁省大连市116001 [2]大连大学中山临床学院,辽宁省大连市116001 [3]大连市海洋多糖生物材料转化医学重点实验室,辽宁省大连市116001

出　　处：《中国组织工程研究》2025年第22期4702-4709,共8页Chinese Journal of Tissue Engineering Research

基　　金：国家重点研发计划“干细胞及转化研究”专项课题(2018YFA0108203),课题名称:干细胞在生物反应器中的规模化培养和适应性调控,子课题负责人:于炜婷;2021年大连市高层次人才创新、科技人才创业和重点领域创新团队支持计划项目(2021RD02),课题名称:粒子近距离放疗治疗体系构建,项目负责人:王若雨。

摘　　要：背景:基质胶是当前培养肿瘤类器官的主要材料,但单纯基质胶不足以在体外模拟肿瘤生长的力学环境;在基质胶基础上引入海藻酸盐材料可提升水凝胶刚度,但水凝胶力学性能在长期培养中难以维持稳定。目的:在乳腺癌类器官培养体系中引入少量钙离子,观察其对基质胶-海藻酸钠水凝胶长期力学性能的维持作用。方法:(1)在恒定质量浓度(5 mg/m L)的基质胶中引入不同质量浓度(0,2.5,5 mg/m L)的海藻酸钠,分别制备低、中、高刚度水凝胶,使用流变仪定期测量水凝胶的力学性能。(2)将人三阴性乳腺癌细胞MDA-MB-231重悬于不同刚度的水凝胶预凝胶中,成胶后加入含(或不含)钙离子的乳腺癌类器官因子培养基,进行乳腺癌类器官培养,在设定的时间点,使用流变仪定期测量引入钙离子对水凝胶力学性能的影响,光学显微镜下观察乳腺癌类器官形态变化,统计第13天乳腺癌类器官成球率。乳腺癌类器官培养7 d后,加入不同浓度(0.1,1,10,100 nmol/L)的化疗药物多西他赛干预6 d,检测细胞活力,计算多西他赛的半抑制浓度IC_(50)。结果与结论:(1)海藻酸钠的加入有效提高了水凝胶的力学强度。(2)随着乳腺癌类器官培养时间的延长,水凝胶的力学强度均降低,培养第13天,中、高刚度水凝胶在含钙离子培养环境中的力学性能均显著高于在不含钙离子培养环境中的力学性能(P<0.05),低刚度水凝胶在两种培养环境中的力学性能无明显变化(P>0.05)。在长期培养中(13 d),中、高刚度水凝胶组乳腺癌类器官随水凝胶力学性能的下降由圆形转化为梭形,而引入钙离子后两组乳腺癌类器官形态不随培养时间延长而改变。在培养环境中引入钙离子对低刚度水凝胶组乳腺癌类器官成球率无影响,但可提升中、高刚度水凝胶组乳腺癌类器官成球率。(3)在不含钙离子的培养环境中,随着多西他赛浓度的升高,乳腺癌�BACKGROUND:Matrigel is the best material for the culture of tumor organoids,but matrigel alone is not enough to simulate the mechanical environment of tumor growth in vitro.Although the introduction of sodium alginate material can improve the stiffness of the hydrogel based on matrigel,its mechanical properties of hydrogel are difficult to maintain stability in long-term culture.OBJECTIVE:To introduce a small amount of calcium ions into the medium of breast cancer organoids and to observe its maintenance effect on the long-term mechanical properties of the matrigel-sodium alginate hydrogel.METHODS:(1)Sodium alginate composite hydrogels with low,medium,and high stiffness were prepared by introducing different mass concentrations(0,2.5,and 5 mg/mL)of sodium alginate into the constant mass concentration(5 mg/mL)of matrigel.The mechanical properties of hydrogels were measured regularly by rheometer.(2)Human triple negative breast cancer cells MDA-MB-231 were resuspended in hydrogel pre-gels with different stiffness.After gelling,breast cancer organoid factor medium containing(or without)calcium ions was added for breast cancer organoid culture.At a set time point,rheometer was used to regularly measure the effect of calcium ion introduction on the mechanical properties of hydrogel.The morphologic changes of breast cancer organoids were observed under optical microscope.Rate of breast cancer organoids forming into pellets was calculated on day 13.After 7 days of breast cancer organoid culture,different concentrations of the chemotherapy drug docetaxel(0.1,1,10,and 100 nmol/L)were added for intervention for 6 days.Cell viability was detected and the semi-inhibitory concentration of docetaxel,IC50,was calculated.RESULTS AND CONCLUSION:(1)The introduction of sodium alginate effectively improved the mechanical strength of the composite hydrogel.(2)With the extension of breast cancer organoid culture time,the mechanical strength of hydrogels decreased.On day 13 of culture,the mechanical properties of medium and high stiffn

关 键 词：肿瘤类器官 乳腺癌类器官 海藻酸钠 基质胶 复合水凝胶 基质刚度 药物反应 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学] R73-3