以通用電操控微流體平台驅動可交聯含細胞之水凝膠微液滴於三維組織工程應用

Abstract

在組織工程研究中，如何在體外提供更仿生的環境，以培養出功能性的器官單位(如仿 肝小葉、腎元、胰島、骨單位等），一直是一個重要的研究主題。生物組織或器官是由 多種細胞外間質、適當排列細胞、與生長因子等結構性與階層性構成。近來許多團隊在 可交聯之水凝膠材料中培養細胞，以交聯後之三維結構的水凝膠扮演細胞外間質角色， 提供適當強度訊息、包埋生長因子以提供生化訊息，使細胞生理表現更優於二維培養之 結果。除此之外，可光交聯之水凝膠可調控三維結構之幾何形狀，而達到細胞因形狀而 自我排列、或幹細胞而分化之結果。然而要達成高結構性之仿生組織，現有技術仍有許 多需要突破之處，例如：（1)細胞在水凝膠中須更有、結構性的排列，而非隨意分布；（2) 複雜幾何形狀組織難以塑形，非光交聯水凝膠材料更難；（3)無法提供多種細胞/水凝膠 之任意組合與共養。

在本計畫中，我們提出以「通用電操控微流體平台」幫助以上目標之達成。過去我們已 在此平台上實現三大通用性：（A)通用液體電性(可驅動導電之電解液與介電之油等液 體）；（B)通用液體型態(可驅動微液滴、連續微流體、與任意形狀之液體）；（C)通用操控 尺度(可驅動液體與其中細胞或顆粒）。此計劃將此三大通用性應用於：（A)以介電濕潤或 介電泳產生各種導電特性之可交聯預聚物溶液之定量微液滴並操控之；（B)以介電泳任 意變形各式可交聯預聚物溶液以形成各種不同幾何形狀；（C)以介電泳排列細胞於可交 聯預聚物溶液並以介電濕潤或介電泳組合交聯前後預聚物溶液與水凝膠。進而達到本計 畫之三大操控目標：（1)細胞在交聯預聚物細胞溶液中排列與集中(尺度）；（2)各式可 交聯預聚物細胞溶液(包含熱交聯與化學交聯)之驅動與塑形(100 pm尺度）；（3)多種細胞 /水凝膠之任意組合與共養(cm尺度）。

In tissue engineering studies, it is essential to provide a biomimicking environment for in vitro cultivations of cells and further functional organ units (liver lobules, nephrons, pancreatic islets, and osteons). Biological tissues/organs consist of hierarchical organization of various extracellular matrix (ECM) molecules along with spatial arrangement of cells and soluble factors. Recently, researchers culture cells in a 3 dimensional (3D) cross-linked hydrogel which behaves as ECM offering mechanical and biochemical cues to the cells. Moreover, the shape of the photo-cross- linkable hydrogel is adjustable. The engineered geometry to the hydrogel would effectively enhance the alignment of cells and the differentiation of stem cells. However, there are some issues to be addressed to achieve highly hierarchical tissues-like structures, including it is (1) difficult shaping arbitrary geometry of hydrogels, especially non-photo-cross-linkable hydrogels, (2) complicated to deliver highly hierarchical cell arrangements in engineered hydrogels, and (3) unable to assemble various cross-linked hydrogels containing different cells for co-culture.

In this project, we adapt our investigations of “general electric-actuated microfluidic platform” to the application of tissue engineering based on the three demonstrated features: the platform is general to (A) fluid conductivity, (B) fluid geometry, and (C) object scale. Applying the three features, we propose to achieve: (A) generation and manipulation of various cross-linkable prepolymer solution droplets by electrowetting-on-dielectric (EWOD) and dielectrophoresis (DEP), (B) shaping of prepolymer solution droplets by DEP, (C) arrangement of cells in prepolymer solution droplets and cross-linked hydrogel blocks by EWOD and DEP. Moreover, the platform is proposed to obtain three major goals: (1) concentrating and arranging cells on the jam scale, (2) shaping and actuating prepolymer solution droplets on the 100，s jam scale, and (3) assembling multiple cell/hydrogel pairs for co-culture studies on the cm scale.