O.P. Waldron1, H. Xu2, S. Agrawal1, M. Landmesser1, J.C. El-Mallah1,3, M. Asgardoon1, J. Yang2, D.J. Ravnic1,3,4 1Penn State College of Medicine, Irvin S. Zubar Plastic Surgery Research Laboratory, Hershey, PA, USA 2Pennsylvania State University, Department Of Biomedical Engineering, State College, PA, USA 3Penn State Milton S. Hershey Medical Center, Department Of Surgery, Hershey, PA, USA 4Pennsylvania State University, Huck Institutes Of The Life Sciences, State College, PA, USA
Introduction: Oncologic resection and traumatic injuries can lead to significant bone loss. Current autologous reconstructive solutions are morbid thereby urging engineered replacements. Citrate-based biomaterials show potential in inducing mineralization but their efficacy in vitro and in vivo remains elusive. We hypothesize that increasing SVF seeding density will lead to linear changes in scaffold mineralization and incorporating pro-osteoblastic factors like phosphoserine (p-Ser) and hydroxyapatite (HA) into a citrate (POC) scaffold will enhance our in vitro work by promoting mineralization in vivo.
Methods: An SVF pellet from a human lipectomy sample was seeded onto POC scaffolds at doubling densities (2.5 x 105, 5 x 105, 10 x 105, 20 x 105) in osteogenic media. No SVF seeding was the control. Scaffolds were histologically analyzed on Day 21. POC-HA-pSer scaffolds were implanted subcutaneously into Sprague Dawley rats (n=3) and explanted on Day 14. All samples underwent alizarin red (AR) staining to assess mineralization and hematoxylin and eosin (H&E) to assess cellularization. Statistical analysis was performed using unpaired t-tests and ANOVA.
Results: Mean cell counts for H&E and mean percentage of AR staining were quantified for each SVF density per image field. Doubling initial seeding density led to linear increases in cellularization and mineralization at 21 days. Controls had minimal cellularization and mineralization. In vivo, AR staining was consistent between animals but some differences in cellularization were noted between animals (Figure 1).
Conclusion: POC scaffolds are an attractive biomaterial for bone regeneration but little is known about their propensity to induce mineralization in vitro or in vivo. It appears that there is an optimal biomaterial-progenitor cell density to precipitate mineralization in vitro. Uniquely, in vivo scaffolds still elicit cellularization but the stem cell fraction is unknown. Future studies will determine whether the mineralization of citrate-based biomaterials depends on stem cells or mature cells.