Engineering biofunctional in vitro vessel models using a multilayer bioprinting technique

Jan Schöneberg,Fabian Kießling,Dirk Rommel,Andreas Blaeser,Federica De Lorenzi,Horst Fischer,Benjamin Theek,Alexander J C Kuehne

doi:10.1038/s41598-018-28715-0

Abstract

Recent advances in the field of bioprinting have led to the development of perfusable complex structures. However, most of the existing printed vascular channels lack the composition or key structural and physiological features of natural blood vessels or they make use of more easily printable but less biocompatible hydrogels. Here, we use a drop-on-demand bioprinting technique to generate in vitro blood vessel models, consisting of a continuous endothelium imitating the tunica intima, an elastic smooth muscle cell layer mimicking the tunica media, and a surrounding fibrous and collagenous matrix of fibroblasts mimicking the tunica adventitia. These vessel models with a wall thickness of up to 425 µm and a diameter of about 1 mm were dynamically cultivated in fluidic bioreactors for up to three weeks under physiological flow conditions. High cell viability (>83%) after printing and the expression of VE-Cadherin, smooth muscle actin, and collagen IV were observed throughout the cultivation period. It can be concluded that the proposed novel technique is suitable to achieve perfusable vessel models with a biofunctional multilayer wall composition. Such structures hold potential for the creation of more physiologically relevant in vitro disease models suitable especially as platforms for the pre-screening of drugs.

Highlights

The aim of tissue engineering is the replication of functional tissues and organs
Microvalve-based bioprinting approaches, on the other hand, only allow the processing of materials with low viscosities such as fibrinogen or gelatine (
The vascular channels spanning the tissue engineered constructs are mostly designed as hollow tubes with a diameter ranging from a few 100 μm[19,20,21] up to the millimetre range[9,22], their inner wall covered by a single layer of endothelial cells (ECs)

Summary

Introduction

The aim of tissue engineering is the replication of functional tissues and organs. The idea of replacing damaged organs or using tissue engineered constructs as novel in vitro disease models for basic research and drug testing is driving current three-dimensional (3D) bioprinting research. The vascular channels spanning the tissue engineered constructs are mostly designed as hollow tubes with a diameter ranging from a few 100 μm[19,20,21] up to the millimetre range[9,22], their inner wall covered by a single layer of endothelial cells (ECs) These printed structures do not fully resemble small arteries and for many applications the complex cellular arrangement of a blood vessel of this particular size may not be necessary. We present a novel strategy to manufacture vessel models for tissue engineering platforms that resemble the cellular arrangement and responses, as well as the mechanical stability of larger in vivo vessels To achieve this purpose we make use of native hydrogels, in particular fibrin and collagen, with a newly developed multi-layered 3D printing technique. With our custom-designed bioreactor, multi-layered models of vascular channels can be dynamically cultivated for longer periods and used for more reliable 3D in vitro test systems or disease models

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Results

Conclusion