Bioactive Decellularized Extracellular Matrix Derived from 3D Stem Cell Spheroids under Macromolecular Crowding Serves as a Scaffold for Tissue Engineering

Abstract

Scaffolds for tissue engineering aim to mimic the native extracellular matrix (ECM) that provides physical support and biochemical signals to modulate multiple cell behaviors. However, the majority of currently used biomaterials are oversimplified and therefore fail to provide a niche required for the stimulation of tissue regeneration. In the present study, 3D decellularized ECM (dECM) scaffolds derived from mesenchymal stem cell (MSC) spheroids and with intricate matrix composition are developed. Specifically, application of macromolecular crowding (MMC) to MSC spheroid cultures facilitate ECM assembly in a 3D configuration, resulting in the accumulation of ECM and associated bioactive components. Decellularized 3D dECM constructs produced under MMC are able to adequately preserve the microarchitecture of structural ECM components and are characterized by higher retention of growth factors. This results in a stronger proangiogenic bioactivity as compared to constructs produced under uncrowded conditions. These dECM scaffolds can be homogenously populated by endothelial cells, which direct the macroassembly of the structures into larger cell‐carrying constructs. Application of empty scaffolds enhances intrinsic revascularization in vivo, indicating that the 3D dECM scaffolds represent optimal proangiogenic bioactive blocks for the construction of larger engineered tissue constructs.

Modulation of Inherent Niches in 3D Multicellular MSC Spheroids Reconfigures Metabolism and Enhances Therapeutic Potential

“…For a determination of the effect of spheroid size on therapeutic potential, a method that can fabricate spheroids with the desired diameters must be used. Herein, by controlling the number of cells seeded into each well of MC hydrogel-coated plates [5][6][7][8][9], we could precisely manipulate the size of the formed spheroids, which is crucial for ensuring the consistency of the following therapeutic effect after cell transplantation. The harvested MSC spheroids contained abundant matrix components, including fibronectin, laminin, collagen, and HSPG.…”

“…Among the developed methods, we [5][6][7][8][9] and others [2,3,[10][11][12] have reported that culturing MSCs in a three-dimensional (3D) multicellular spheroid configuration can significantly enhance both the viability of administered MSCs and their overall therapeutic functions. Specifically, the spheroid configuration can more closely recapitulate the physiological 3D microenvironment than conventional two-dimensional (2D) cultures [3].…”

“…The MSC spheroids were prepared using a methylcellulose (MC) hydrogel that was not bound by cells. Twelve percent (w/v) MC solution was prepared by dissolving MC powder in phosphate buffered saline (PBS; Thermo Fisher Scientific, St. Louis, MO, USA) [5][6][7][8][9]. After autoclaving, a sterilized MC solution was employed to coat each well of a 96-well culture plate.…”

Cell-Penetrating Delivery of Nitric Oxide by Biocompatible Dinitrosyl Iron Complex and Its Dermato-Physiological Implications

“…To determine the activity of tyrosinase, the cells prepared according to the aforementioned methods were lysed with RIPA buffer (20 mM Tris-HCl, 1 mM EGTA, 150 mM NaCl, and 1% Triton X-100) [69] at 4 • C for 20 min followed by centrifugation at 14,000 rpm for 15 min. A bicinchoninic acid assay (G-Biosciences, St. Louis, MO, USA) was used to quantify the total protein of the supernatant.…”

Decellularization Strategies for Regenerating Cardiac and Skeletal Muscle Tissues

“…To tackle the ongoing challenge of obtaining a renewable and robust supply of human cardiac dECM, one group has looked towards the use of cardiac organoids as a cultured source for dECM. Chiang et al used organoids derived from human umbilical cord blood mesenchymal stem cells to lay down a structural protein niche that was then decellularized to provide a dECM scaffold ( Chiang et al, 2021 ). The resulting dECM was shown to possess both essential microarchitecture and functional bioactivity ( Chiang et al, 2021 ).…”