3D Bioprinting Mesenchymal Stem Cell-Derived Neural Tissues Using a Fibrin-Based Bioink

Abstract

Current treatments for neurodegenerative diseases aim to alleviate the symptoms experienced by patients; however, these treatments do not cure the disease nor prevent further degeneration. Improvements in current disease-modeling and drug-development practices could accelerate effective treatments for neurological diseases. To that end, 3D bioprinting has gained significant attention for engineering tissues in a rapid and reproducible fashion. Additionally, using patient-derived stem cells, which can be reprogrammed to neural-like cells, could generate personalized neural tissues. Here, adipose tissue-derived mesenchymal stem cells (MSCs) were bioprinted using a fibrin-based bioink and the microfluidic RX1 bioprinter. These tissues were cultured for 12 days in the presence of SB431542 (SB), LDN-193189 (LDN), purmorphamine (puro), fibroblast growth factor 8 (FGF8), fibroblast growth factor-basic (bFGF), and brain-derived neurotrophic factor (BDNF) to induce differentiation to dopaminergic neurons (DN). The constructs were analyzed for expression of neural markers, dopamine release, and electrophysiological activity. The cells expressed DN-specific and early neuronal markers (tyrosine hydroxylase (TH) and class III beta-tubulin (TUJ1), respectively) after 12 days of differentiation. Additionally, the tissues exhibited immature electrical signaling after treatment with potassium chloride (KCl). Overall, this work shows the potential of bioprinting engineered neural tissues from patient-derived MSCs, which could serve as an important tool for personalized disease models and drug-screening.

Application of Small Molecules in the Central Nervous System Direct Neuronal Reprogramming

“…Recently, it has been proposed that some in vitro chemical-induced stable neurons could be constructed for brains by a three-dimensional (3D) bioprinting technology, which prints biodegradable materials with cells as 3D tissue ( Ho and Hsu, 2018 ; Yuan et al, 2020 ). In 2021, a lab bioprinted MSC-derived neural tissues successfully using a fibrin-based bioink and the RX1 bioprinter with the aid of SB431542, LDN-193189, purmorphamine, fibroblast growth factor 8 (FGF8), fibroblast growth factor-basic (bFGF), and brain-derived neurotrophic factor (BDNF) ( Restan Perez et al, 2021 ). Even more interesting is that Ho et al achieved cell reprogramming via 3D bioprinting of human fibroblasts in polyurethane hydrogel for the manufacture of neural-like constructs with Forkhead box D3 (FoxD3), a transcription factor ( Ho and Hsu, 2018 ).…”

Protocol for printing 3D neural tissues using the BIO X equipped with a pneumatic printhead

“…The high viscosity, fibrin-based bioink and the crosslinker described in this protocol are the TissuePrint-HV Kit and TissuePrint Crosslink from Axolotl Biosciences. The bioink is adapted from one previously shown to support a variety of cell lines, including mesenchymal stem cells (MSCs) ( Restan Perez et al., 2021 ) , neural progenitor stem cells (NPCs) ( de la Vega et al., 2018 ; Abelseth et al., 2019 ; Sharma et al., 2020 , 2021 ; de la Vega et al, 2021 ), glioblastoma (GBMs) ( Smits et al., 2020 ), and human dermal fibroblasts (HDFs). The recommended passage number range is per the user’s individual cell line needs.…”

“…The fibrin-based bioink described here was adapted from one that has been tested with several different cell lines, including hiPSCs, NPCs, and MSCs ( de la Vega et al., 2018 ; Abelseth et al., 2019 ; Sharma et al., 2020 , 2021 ; Smits et al., 2020 ; Restan Perez et al., 2021 ; de la Vega et al., 2018 ; Abelseth et al., 2019 ; Sharma et al., 2020 , 2021 ; Smits et al., 2020 ; Restan Perez et al., 2021 ). The ability of the bioink to maintain the viability and proliferation of other cell types may vary.…”

Enhancing Stem Cell-Based Therapeutic Potential by Combining Various Bioengineering Technologies

“…They reported significantly enhanced viability and chondrogenic differentiation of loaded BM-MSCs within the fabricated hydrogel cartilage scaffolds ( Costantini et al, 2016 ). Restan Perez et al (2021) also designed neural tissues loaded with patient-derived AD-MSCs using fibrin-based bioink and microfluidic RX1 3D bioprinter, and analyzed the expression of various neural markers, dopamine release, and electrophysiological activity. Daly et al (2016) fabricated hypertrophic cartilage templates loaded with BM-MSCs with vascularization and mineralization using gamma-irradiated alginate bioink incorporating Arg-Gly-Asp adhesion peptides ().…”