Three-Dimensional Cell Printing a Vascularized Airway-on-a-Chip using Tracheal Mucosa derived Decellularized Extracellular Matrix Bioink

Abstract

Organ-on-chip technologies has a great potential of predicting the pharmaceutical effectiveness of newly developed drugs by target tissue based and phenotypic screening. These devices could produce tissue/organ level functionality through the recapitulation of the multicellular organizations, physicochemical microenvironments, tissue-tissue interfaces, and vascular perfusion for blood flow. Despite the significant advances, the complexity of the manufacturing process for microchips, including multiple polymerization, microchannel assembly, and cell culture inside the microchannel, limits their public application. Three-dimensional (3D) cell printing technology allows to easily and readily fabricate the prototype of the designed product using 3D printer and corresponding software for Computer Aided Design (CAD). Furthermore, direct printing of encapsulated cell suspension had demonstrated the possibility of utilizing 3D printing techniques to fabricate 3D tissue/organ models mimicking natural cell arrangement and tissue structure. This thesis presents a novel method to fabricate human vascularized airway-on-a-chip by 3D cell printing technology. To closely simulate a functional airway mucosa, tracheal mucosa-derived decellularized extracellular matrix (tmdECM) bioink was developed to encapsulate endothelial cells. The cell laden bioink was subsequently printed onto the 3D vascular platform that can reproduce the vascular system which is indispensable to maintain organ viable and functional. Finally, this 3D vascular platform was mounted to the upper polydimethylsiloxane (PDMS) chip containing fully differentiated airway epithelium. The assembly is able to recapitulate the interface of airway epithelium and underlying vascular networks in the lamina propria on the airway. Details on the research are given below. Tracheal mucosa-derived dECM (tmdECM) bioink was developed and applied to mimic the microenvironment of mucosal layer to induce functional tracheal epithelium in vitro and in vivo. Compared to collagen type I hydrogel, this bioink not only achieved a typical pseudostratified ciliated columnar tracheal epithelium more rapidly but also realized the mucociliary clearance function of tracheal epithelium in physiologically relevant level. In order to mimic the vascular system beneath the functional tracheal epithelium, a 3D vascular platform was developed by using 3D cell printing and tmdECM bioink. Two types of 3D vascular platforms for either naturally induced angiogenesis or direct vascularture printing were designed and fabricated. Physical/chemical factors within tmdECM enable endothelial cells to form functional vascular networks on the 3D vascular platform, which exhibited relevant responses to physiological stimulation as well as the barrier function. Finally, the human vascularized airway-on-a-chip (hVA-OC) was developed by assembling the vascular platform and the upper PDMS chip containing fully differentiated airway epithelium. The hVA-OC completely recapitulated the interface of airway epithelium and underlying vascular networks in airway, and the asthmatic airway inflammation and allergen exacerbation was successfully reproduced on the hVA-OC. The approach conducted in this research will bring an innovative chip fabrication method, and be easily applied to various tissues/organs. Moreover, these 3D cell printed tissue/organ model can be expected to have a great impact on the field of organ-on-a-chip for research of cell biology, drug screening, and drug discovery.