Introduction: In bone tissue engineering, several attempts have been made to generate artificial bone constructs to replace the autograft and allograft treatment and to enhance bone repair and regeneration. However, engineering bone tissue presents some challenges, mainly based on bone tissue’s gradual variation of several biological, mechanical, and structural features. Another challenge is to control the migration of stem cells, which have an inherent ability to migrate. Uncontrolled migration of stem cells can cause to pathological situations such as cancer and inflammatory diseases. Therefore, besides promoting stem cell differentiation, it is crucial to regulate stem cell migration in order to achieve their maximum capacity for differentiation and regeneration.
Methodology: We describe mechanically, biochemically, and topographically graded 3D (nano)composite scaffolds (GradS) generated by a 3D printing technique. We prepared our (nano)composite hydrogels by mixing gelatin methacryloyl (GelMA), alginate, and pH-responsive drug delivery nanomaterials [DexPMO-PDL=dexamethasone (Dex)- and poly-d-lysine (PDL)-functionalized periodic mesoporous organosilicas (PMO)] in different ratios. We prepared three different composite hydrogels increasing GelMA and DexPMO-PDL concentration (GA5, GA10, and GA10-P). GA5 was prepared by mixing GelMA (5% w/v) and Alg (7% w/v), while GA10 was composed of GelMA (10% w/v) and Alg (7% w/v). In order to prepare GA10-P, DexPMO-PDL (0.2% w/v) was mixed with GA10. The prepared (nano)composite hydrogels were used for fabricating a new 3D step-gradient (nano)composite scaffold (GradS) made by 3D bioprinting.
Results: Our results showed that the Dex release from the GA10-P scaffold at pH 7.4 and 6.0 was prolonged and pH dependent. After 7 days of incubation, the amount of Dex released from GA10-P scaffold at pH 6.0 was 2 times higher than at pH 7.4, demonstrating GA10-P’s ability to perform pH responsive drug delivery. Cell experiments were performed on GradS. we only seeded hBMMSC into the GA5 section of the GradS. Fluorescence microscopy images and cell viability kits showed that hBM MSC migrated from the GA5 part toward GA10-P part of the GradS. Since GA10 and GA10-P have higher GelMa and DexPMO-PDL concentrations than GA5, this migration depended on an increase in the GelMA concentration and on the incorporation of DexPMO-PDL into the hydrogel network.
We also investigated the impact of GA5, GA10, GA10-P, and GradS on hBM MSC differentiation toward an osteogenic lineage. Osteogenic differentiation capacity was determined by alizarin red and BCIP/NBT (5-Bromo-4-chloro-3-indolyl phosphate/Nitro blue tetrazolium) colorimetric staining. We observed a proportional increase in the alizarin red- and BCIP/NBT-stained areas with an increase in the GelMA and DexPMO-PDL concentration. This indicates that the osteogenic differentiation capacity of hBM MSC was promoted by an increase in the content of GelMA and DexPMOPDL within the hydrogel network.
Conclusion: The cell experiments demonstrated that the viable number of hBM MSC in the GradS increased as the content of GelMA and DexPMO-PDL increased. This effect was used to control the migration and osteogenic differentiation of hBM MSC toward the GradS section possessing a higher concentration of GelMA and DexPMO-PDL.
Acknowledgements: Dr. N. S. Kehr thanks DFG for funding.
References: Motealleh A. et al., Nano Select doi.org/10.1002/nano.202100113 (2021).