Maryam Sadat Khoramgah
1,2,3,4 
, Hossein Ghanbarian
1,4, Javad Ranjbari
1,4, Nilufar Ebrahimi
3,5, Fatemeh Sadat Tabatabaei Mirakabad
1,4, Navid Ahmady Roozbahany
3,6, Hojjat Allah Abbaszadeh
2,3,7* , Simzar Hosseinzadeh
8,9* 1 Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
2 Laser Application in Medical Sciences Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
3 Hearing Disorders Research Center, Loghman Hakim Hospital, Shahid Beheshti University of Medical Sciences, Tehran, Iran
4 Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
5 Department of Biomedical Engineering, East Tehran Branch, Islamic Azad University, Tehran, Iran
6 Private Practice, Bradford ON, Canada
7 Department of Biology and Anatomical Sciences, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
8 Medical Nanotechnology and Tissue Engineering Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
9 Department of Tissue Engineering and Applied Cell Sciences, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
Abstract
Introduction: Treatment of critical-sized bone defects is challenging. Tissue engineering as a state-of-the-art method has been concerned with treating these non-self-healing bone defects. Here, we studied the potentials of new three-dimensional nanofibrous scaffolds (3DNS) with and without human adipose mesenchymal stem cells (ADSCs) for reconstructing rat critical-sized calvarial defects (CSCD).
Methods: Scaffolds were made from 1- polytetrafluoroethylene (PTFE), and polyvinyl alcohol (PVA) (PTFE/ PVA group), and 2- PTFE, PVA, and graphene oxide (GO) nanoparticle (PTFE/ PVA/GO group) and seeded by ADSCs and incubated in osteogenic media (OM). The expression of key osteogenic proteins including Runt-related transcription factor 2 (Runx2), collagen type Iα (COL Iα), osteocalcin (OCN), and osteonectin (ON) at days 14 and 21 of culture were evaluated by western blot and immunocytochemistry methods. Next, 40 selected rats were assigned to five groups (n=8) to create CSCD which will be filled by scaffolds or cell-containing scaffolds. The groups were denominated as the following order: Control (empty defects), PTFE/PVA (PTFE/PVA scaffolds implant), PTFE/PVA/GO (PTFE/PVA/GO scaffolds implant), PTFE/PVA/Cell group (PTFE/PVA scaffolds containing ADSCs implant), and PTFE/PVA/GO/Cell group (PTFE/PVA/GO scaffolds containing ADSCs implant). Six and 12 weeks after implantation, the animals were sacrificed and bone regeneration was evaluated using computerized tomography (CT), and hematoxylin-eosin (H&E) staining.
Results: Based on the in-vitro study, expression of bone-related proteins in ADSCs seeded on PTFE/PVA/GO scaffolds were significantly higher than PTFE/PVA scaffolds and TCPS (P<0.05). Based on the in-vivo study, bone regeneration in CSCD were filled with PTFE/PVA/GO scaffolds containing ADSCs were significantly higher than PTFE/PVA scaffolds containing ADSCs (P<0.05). CSCD filled with cell-seeded scaffolds showed higher bone regeneration in comparison with CSCD filled with scaffolds only (P<0.05).
Conclusion: The data provided evidence showing new freeze-dried nanofibrous scaffolds formed from hydrophobic (PTFE) and hydrophilic (PVA) polymers with and without GO provide a suitable environment for ADSCs due to the expression of bone-related proteins. ADSCs and GO in the implanted scaffolds had a distinct effect on the bone regeneration process in this in-vivo study.