Biologically Relevant In Vitro 3D-Model to Study Bone Regeneration Potential of Human Adipose Stem Cells.

Victor J B Van Santen,Astrid D Bakker,Jenneke Klein-Nulend,Angela P Bastidas Coral,Jolanda M A Hogervorst

doi:10.3390/biom12020169

Victor J B Van Santen, Astrid D Bakker + Show 3 more

Open Access

https://doi.org/10.3390/biom12020169

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Abstract

Standard cell cultures may not predict the proliferation and differentiation potential of human mesenchymal stromal cells (MSCs) after seeding on a scaffold and implanting this construct in a bone defect. We aimed to develop a more biologically relevant in vitro 3D-model for preclinical studies on the bone regeneration potential of MSCs. Human adipose tissue-derived mesenchymal stromal cells (hASCs; five donors) were seeded on biphasic calcium phosphate (BCP) granules and cultured under hypoxia (1% O2) for 14 days with pro-inflammatory TNFα, IL4, IL6, and IL17F (10 mg/mL each) added during the first three days, simulating the early stages of repair (bone construct model). Alternatively, hASCs were cultured on plastic, under 20% O2 and without cytokines for 14 days (standard cell culture). After two days, the bone construct model decreased total DNA (3.9-fold), COL1 (9.8-fold), and RUNX2 expression (19.6-fold) and metabolic activity (4.6-fold), but increased VEGF165 expression (38.6-fold) in hASCs compared to standard cultures. After seven days, the bone construct model decreased RUNX2 expression (64-fold) and metabolic activity (2.3-fold), but increased VEGF165 (54.5-fold) and KI67 expression (5.7-fold) in hASCs compared to standard cultures. The effect of the bone construct model on hASC proliferation and metabolic activity could be largely mimicked by culturing on BCP alone (20% O2, no cytokines). The effect of the bone construct model on VEGF165 expression could be mimicked by culturing hASCs under hypoxia alone (plastic, no cytokines). In conclusion, we developed a new, biologically relevant in vitro 3D-model to study the bone regeneration potential of MSCs. Our model is likely more suitable for the screening of novel factors to enhance bone regeneration than standard cell cultures.

Highlights

Bone tissue engineering strategies developed to treat large bone defects are generally based on the incorporation of mesenchymal stromal cells (MSCs) and bioactive factors into a scaffold, which together form a bioactive bone construct [1]
Our preclinical 3D-model may be more suitable for predicting the success, or lack thereof, of novel bone tissue engineering constructs in a later stage of development than standard practices, while being relatively easy to implement in the workflow and still allowing medium-to-high throughput analysis of the bone regeneration potential of human adipose stem cells
We investigated whether culturing Human adipose tissue-derived mesenchymal stromal cells (hASCs) on biphasic calcium phosphate (BCP) under normoxia without cytokines would affect proliferation and/or differentiation as the complete bone construct model

Summary

Introduction

Bone tissue engineering strategies developed to treat large bone defects are generally based on the incorporation of mesenchymal stromal cells (MSCs) and bioactive factors into a scaffold, which together form a bioactive bone construct [1]. This bone construct serves to contribute to bone regeneration upon implantation. Upon implantation of a tissue engineered bone construct, an inflammatory response reminiscent of the early stage of fracture repair occurs in the bone microenvironment, where inflammation and hypoxia trigger the healing process [5,6,7,8,9] Cytokines, such as tumor necrosis factor-α (TNFα), interleukin-4 (IL4), interleukin-6 (IL6), and interleukin-17F

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