Department of Biomedicine & Health Sciences, College of Medicine, The Catholic University of Korea, Seoul, Republic of Korea.
School of Mechanical Engineering, College of Engineering, Korea University, Seoul, Republic of Korea.
J Exp Clin Cancer Res. 2018 Jan 12;37(1):4. doi: 10.1186/s13046-017-0654-6.
Pancreatic stellate cells (PSCs), a major component of the tumor microenvironment in pancreatic cancer, play roles in cancer progression as well as drug resistance. Culturing various cells in microfluidic (microchannel) devices has proven to be a useful in studying cellular interactions and drug sensitivity. Here we present a microchannel plate-based co-culture model that integrates tumor spheroids with PSCs in a three-dimensional (3D) collagen matrix to mimic the tumor microenvironment in vivo by recapitulating epithelial-mesenchymal transition and chemoresistance.
A 7-channel microchannel plate was prepared using poly-dimethylsiloxane (PDMS) via soft lithography. PANC-1, a human pancreatic cancer cell line, and PSCs, each within a designated channel of the microchannel plate, were cultured embedded in type I collagen. Expression of EMT-related markers and factors was analyzed using immunofluorescent staining or Proteome analysis. Changes in viability following exposure to gemcitabine and paclitaxel were measured using Live/Dead assay.
PANC-1 cells formed 3D tumor spheroids within 5 days and the number of spheroids increased when co-cultured with PSCs. Culture conditions were optimized for PANC-1 cells and PSCs, and their appropriate interaction was confirmed by reciprocal activation shown as increased cell motility. PSCs under co-culture showed an increased expression of α-SMA. Expression of EMT-related markers, such as vimentin and TGF-β, was higher in co-cultured PANC-1 spheroids compared to that in mono-cultured spheroids; as was the expression of many other EMT-related factors including TIMP1 and IL-8. Following gemcitabine exposure, no significant changes in survival were observed. When paclitaxel was combined with gemcitabine, a growth inhibitory advantage was prominent in tumor spheroids, which was accompanied by significant cytotoxicity in PSCs.
We demonstrated that cancer cells grown as tumor spheroids in a 3D collagen matrix and PSCs co-cultured in sub-millimeter proximity participate in mutual interactions that induce EMT and drug resistance in a microchannel plate. Microfluidic co-culture of pancreatic tumor spheroids with PSCs may serve as a useful model for studying EMT and drug resistance in a clinically relevant manner.
胰腺星状细胞（PSCs）是胰腺癌肿瘤微环境的主要组成部分，在癌症进展和耐药性方面发挥作用。在微流控（微通道）设备中培养各种细胞已被证明是研究细胞相互作用和药物敏感性的有用方法。在这里，我们提出了一种基于微通道板的共培养模型，该模型将肿瘤球体与 PSCs 整合在三维（3D）胶原基质中，通过模拟体内上皮-间充质转化和化学抗性来模拟肿瘤微环境。
使用聚二甲基硅氧烷（PDMS）通过软光刻制备 7 通道微通道板。人胰腺癌细胞系 PANC-1 和 PSCs 分别在微通道板的指定通道中培养，嵌入 I 型胶原中。使用免疫荧光染色或蛋白质组分析分析 EMT 相关标志物和因子的表达。使用 Live/Dead 测定法测量暴露于吉西他滨和紫杉醇后细胞活力的变化。
PANC-1 细胞在 5 天内形成 3D 肿瘤球体，当与 PSCs 共培养时，球体数量增加。优化了 PANC-1 细胞和 PSCs 的培养条件，并通过显示细胞迁移增加的相互激活来确认其适当的相互作用。共培养下的 PSCs 中 α-SMA 的表达增加。与单核培养的球体相比，共培养的 PANC-1 球体中 EMT 相关标志物（如波形蛋白和 TGF-β）的表达更高；许多其他 EMT 相关因子的表达也更高，包括 TIMP1 和 IL-8。吉西他滨暴露后，存活率无明显变化。当紫杉醇与吉西他滨联合使用时，肿瘤球体的生长抑制优势明显，同时 PSCs 的细胞毒性明显。
我们证明了在 3D 胶原基质中生长的肿瘤球体中的癌细胞和在毫米级接近度下共培养的 PSCs 参与相互作用，从而在微通道板中诱导 EMT 和耐药性。胰腺肿瘤球体与 PSCs 的微流控共培养可能成为一种有用的模型，可用于以临床相关的方式研究 EMT 和耐药性。
