Microfluidic co-culture of pancreatic tumor spheroids with stellate cells as a novel 3D model for investigation of stroma-mediated cell motility and drug resistance.
Microfluidic co-culture of pancreatic tumor spheroids with stellate cells as a novel 3D model for investigation of stroma-mediated cell motility and drug resistance.
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DOI：
10.1186/s13046-017-0654-6
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发表时间：
2018-01-12
期刊：
影响因子：
--
通讯作者：
Kuh HJ
中科院分区：
文献类型：
--
作者：
Lee JH;Kim SK;Khawar IA;Jeong SY;Chung S;Kuh HJ
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中发挥作用。在微流控（微通道）装置中培养各种细胞已被证明在研究细胞相互作用和药物敏感性方面是有用的。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PSC整合在三维（3D）胶原基质中，通过重现上皮-间质转化和化疗耐药性来模拟体内肿瘤微环境。以聚二甲基硅氧烷（PDMS）为原料，采用软光刻技术制备了7通道微通道板。将PANC-1（一种人胰腺癌细胞系）和PSC（各自在微通道板的指定通道内）包埋在I型胶原中培养。采用免疫荧光染色或蛋白质组学分析EMT相关标志物和因子的表达。使用活/死测定法测量暴露于吉西他滨和紫杉醇后的活力变化。PANC-1细胞在5天内形成3D肿瘤球体，并且当与PSC共培养时球体的数量增加。对PANC-1细胞和PSC的培养条件进行了优化，并通过显示为细胞运动性增加的相互激活来证实它们的适当相互作用。共培养的PSC中α-SMA表达增加。EMT相关标志物（如波形蛋白和TGF-β）的表达在共培养的PANC-1球体中高于单培养球体;许多其他EMT相关因子（包括TIMP 1和IL-8）的表达也是如此。吉西他滨暴露后，未观察到生存期的显著变化。当紫杉醇与吉西他滨组合时，在肿瘤球体中的生长抑制优势突出，这伴随着在PSC中的显著细胞毒性。我们证明了在3D胶原基质中作为肿瘤球体生长的癌细胞和在亚毫米接近度下共培养的PSC参与了在微通道板中诱导EMT和耐药性的相互作用。胰腺肿瘤球体与PSC的微流控共培养可以作为以临床相关方式研究EMT和耐药性的有用模型。本文的在线版本（10.1186/s13046-017-0654-6）包含补充材料，可供授权用户使用。
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. The online version of this article (10.1186/s13046-017-0654-6) contains supplementary material, which is available to authorized users.
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影响因子：
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作者：
Pavón MA;Arroyo-Solera I;Céspedes MV;Casanova I;León X;Mangues R
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