Raghavan Shreya, Ward Maria R, Rowley Katelyn R, Wold Rachel M, Takayama Shuichi, Buckanovich Ronald J, Mehta Geeta
Department of Materials Science Engineering, University of Michigan, Ann Arbor, USA.
Department of Biomedical Engineering, University of Michigan, Ann Arbor, USA.
Gynecol Oncol. 2015 Jul;138(1):181-9. doi: 10.1016/j.ygyno.2015.04.014. Epub 2015 Apr 22.
Ovarian cancer grows and metastasizes from multicellular spheroidal aggregates within the ascites fluid. Multicellular tumor spheroids are therefore physiologically significant 3D in vitro models for ovarian cancer research. Conventional hanging drop cultures require high starting cell numbers, and are tedious for long-term maintenance. In this study, we generate stable, uniform multicellular spheroids using very small number of ovarian cancer cells in a novel 384 well hanging drop array platform.
We used novel tumor spheroid platform and two ovarian cancer cell lines (A2780 and OVCAR3) to demonstrate the stable incorporation of as few as 10 cells into a single spheroid.
Spheroids had uniform geometry, with projected areas (42.60×10(3)μm-475.22×10(3)μm(2) for A2780 spheroids and 37.24×10(3)μm(2)-281.01×10(3)μm(2) for OVCAR3 spheroids) that varied as a function of the initial cell seeding density. Phalloidin and nuclear stains indicated cells formed tightly packed spheroids with demarcated boundaries and cell-cell interaction within spheroids. Cells within spheroids demonstrated over 85% viability. 3D tumor spheroids demonstrated greater resistance (70-80% viability) to cisplatin chemotherapy compared to 2D cultures (30-50% viability).
Ovarian cancer spheroids can be generated from limited cell numbers in high throughput 384 well plates with high viability. Spheroids demonstrate therapeutic resistance relative to cells in traditional 2D culture. Stable incorporation of low cell numbers is advantageous when translating this research to rare patient-derived cells. This system can be used to understand ovarian cancer spheroid biology, as well as carry out preclinical drug sensitivity assays.
卵巢癌在腹水内由多细胞球状聚集体生长并转移。因此，多细胞肿瘤球体是用于卵巢癌研究的具有生理意义的三维体外模型。传统的悬滴培养需要高起始细胞数量，并且长期维持很繁琐。在本研究中，我们在新型384孔悬滴阵列平台中使用极少量的卵巢癌细胞生成稳定、均匀的多细胞球体。
我们使用新型肿瘤球体平台和两种卵巢癌细胞系（A2780和OVCAR3）来证明仅10个细胞就能稳定地整合到单个球体中。
球体具有均匀的几何形状，投影面积（A2780球体为42.60×10³μm² - 475.22×10³μm²，OVCAR3球体为37.24×10³μm² - 281.01×10³μm²）随初始细胞接种密度而变化。鬼笔环肽和细胞核染色表明细胞形成紧密堆积的球体，边界清晰，球体内存在细胞间相互作用。球体内的细胞显示出超过85%的活力。与二维培养（活力30 - 50%）相比，三维肿瘤球体对顺铂化疗表现出更高的抗性（活力70 - 80%）。
可以在高通量384孔板中从有限数量的细胞生成具有高活力的卵巢癌球体。与传统二维培养中的细胞相比，球体表现出治疗抗性。将该研究转化应用于罕见的患者来源细胞时，低细胞数量的稳定整合具有优势。该系统可用于了解卵巢癌球体生物学，以及进行临床前药物敏感性测定。
