OBJECTIVES: Treatment of critical-sized bone defects with cells and biomaterials offers an efficient alternative to traditional bone grafts. Chitosan (CS) is a natural biopolymer that acts as a scaffold in bone tissue engineering (BTE). Polyphosphate (PolyP), recently identified as an inorganic polymer, acts as a potential bone morphogenetic material, whereas pigeonite (Pg) is a novel iron-containing ceramic. In this study, we prepared and characterized scaffolds containing CS, calcium polyphosphate (CaPP) and Pg particles for bone formation in vitro and in vivo. MATERIALS AND METHODS: Chitosan/CaPP scaffolds and CS/CaPP scaffolds containing varied concentrations of Pg particles (0.25%, 0.5%, 0.75% and 1%) were prepared and characterized by SEM, XRD, EDAX, FT-IR, degradation, protein adsorption, mechanical strength and biomineralization studies. The cytocompatibility of these scaffolds with mouse mesenchymal stem cells (mMSCs, C3H10T1/2) was determined by MTT assay and fluorescence staining. Cell proliferation on scaffolds was assessed using MUSE™ (Merck-Millipore, Germany) cell analyser. The effect of scaffolds on osteoblast differentiation at the cellular level was evaluated by Alizarin red (AR) and alkaline phosphatase (ALP) staining. At the molecular level, the expression of osteoblast differentiation marker genes such as Runt-related transcription factor-2 (Runx2), ALP, type I collagen-1 (Col-I) and osteocalcin (OC) was determined by real-time reverse transcriptase (RT-PCR) analysis. Bone regeneration was assessed by X-ray radiographs, SEM and EDAX analyses, and histological staining such as haematoxylin and eosin staining and Masson's trichrome staining (MTS) in a rat critical-sized tibial defect model system. RESULTS: The inclusion of iron-containing Pg particles at 0.25% concentration in CS/CaPP scaffolds showed enhanced bioactivity by protein adsorption and biomineralization, compared with that shown by CS/CaPP scaffolds alone. Increased proliferation of mMSCs was observed with CS/CaPP/Pg scaffolds compared with control and CS/CaPP scaffolds. Increase in cell proliferation was accompanied by G0/G1 to G2/M phase transition with increased levels of cyclin(s) A, B and C. Pg particles in CS/CaPP scaffolds enhanced osteoblast differentiation at the cellular and molecular levels, as evidenced by increased calcium deposits, ALP activity and expression of osteoblast marker genes. In vivo implantation of scaffolds in rat critical-sized tibial defects displayed accelerated bone formation after 8 weeks. CONCLUSION: The current findings indicate that CS/CaPP scaffolds containing iron-containing Pg particles serve as an appropriate template to support proliferation and differentiation of MSCs to osteoblasts in vitro and bone formation in vivo and thus support their candidature for BTE applications.
OBJECTIVES: Treatment of critical-sized bone defects with cells and biomaterials offers an efficient alternative to traditional bone grafts. Chitosan (CS) is a natural biopolymer that acts as a scaffold in bone tissue engineering (BTE). Polyphosphate (PolyP), recently identified as an inorganic polymer, acts as a potential bone morphogenetic material, whereas pigeonite (Pg) is a novel iron-containing ceramic. In this study, we prepared and characterized scaffolds containing CS, calcium polyphosphate (CaPP) and Pg particles for bone formation in vitro and in vivo. MATERIALS AND METHODS: Chitosan/CaPP scaffolds and CS/CaPP scaffolds containing varied concentrations of Pg particles (0.25%, 0.5%, 0.75% and 1%) were prepared and characterized by SEM, XRD, EDAX, FT-IR, degradation, protein adsorption, mechanical strength and biomineralization studies. The cytocompatibility of these scaffolds with mouse mesenchymal stem cells (mMSCs, C3H10T1/2) was determined by MTT assay and fluorescence staining. Cell proliferation on scaffolds was assessed using MUSE™ (Merck-Millipore, Germany) cell analyser. The effect of scaffolds on osteoblast differentiation at the cellular level was evaluated by Alizarin red (AR) and alkaline phosphatase (ALP) staining. At the molecular level, the expression of osteoblast differentiation marker genes such as Runt-related transcription factor-2 (Runx2), ALP, type I collagen-1 (Col-I) and osteocalcin (OC) was determined by real-time reverse transcriptase (RT-PCR) analysis. Bone regeneration was assessed by X-ray radiographs, SEM and EDAX analyses, and histological staining such as haematoxylin and eosin staining and Masson's trichrome staining (MTS) in a rat critical-sized tibial defect model system. RESULTS: The inclusion of iron-containing Pg particles at 0.25% concentration in CS/CaPP scaffolds showed enhanced bioactivity by protein adsorption and biomineralization, compared with that shown by CS/CaPP scaffolds alone. Increased proliferation of mMSCs was observed with CS/CaPP/Pg scaffolds compared with control and CS/CaPP scaffolds. Increase in cell proliferation was accompanied by G0/G1 to G2/M phase transition with increased levels of cyclin(s) A, B and C. Pg particles in CS/CaPP scaffolds enhanced osteoblast differentiation at the cellular and molecular levels, as evidenced by increased calcium deposits, ALP activity and expression of osteoblast marker genes. In vivo implantation of scaffolds in rat critical-sized tibial defects displayed accelerated bone formation after 8 weeks. CONCLUSION: The current findings indicate that CS/CaPP scaffolds containing iron-containing Pg particles serve as an appropriate template to support proliferation and differentiation of MSCs to osteoblasts in vitro and bone formation in vivo and thus support their candidature for BTE applications.
Authors: Jaclyn Maurer; Margaret M Harris; Vanessa A Stanford; Timothy G Lohman; Ellen Cussler; Scott B Going; Linda B Houtkooper Journal: J Nutr Date: 2005-04 Impact factor: 4.798
Authors: Werner E G Müller; Xiaohong Wang; Bärbel Diehl-Seifert; Klaus Kropf; Ute Schlossmacher; Ingo Lieberwirth; Gunnar Glasser; Matthias Wiens; Heinz C Schröder Journal: Acta Biomater Date: 2011-03-30 Impact factor: 8.947
Authors: Y Kawazoe; T Shiba; R Nakamura; A Mizuno; K Tsutsumi; T Uematsu; M Yamaoka; M Shindoh; T Kohgo Journal: J Dent Res Date: 2004-08 Impact factor: 6.116
Authors: S Dhivya; A Keshav Narayan; R Logith Kumar; S Viji Chandran; M Vairamani; N Selvamurugan Journal: Cell Prolif Date: 2017-11-21 Impact factor: 6.831
Authors: Veronica Zubillaga; Ana Alonso-Varona; Susana C M Fernandes; Asier M Salaberria; Teodoro Palomares Journal: Int J Mol Sci Date: 2020-02-03 Impact factor: 5.923
Authors: Carlos David Grande Tovar; Jorge Iván Castro; Carlos Humberto Valencia; Diana Paola Navia Porras; José Herminsul Mina Hernandez; Mayra Eliana Valencia; José Daniel Velásquez; Manuel N Chaur Journal: Biomolecules Date: 2019-11-01
Authors: Julia Scheinpflug; Moritz Pfeiffenberger; Alexandra Damerau; Franziska Schwarz; Martin Textor; Annemarie Lang; Frank Schulze Journal: Genes (Basel) Date: 2018-05-10 Impact factor: 4.096
Authors: Carlos David Grande Tovar; Jorge Iván Castro; Carlos Humberto Valencia; Diana Paola Navia Porras; José Herminsul Mina Hernandez; Mayra Eliana Valencia Zapata; Manuel N Chaur Journal: Molecules Date: 2020-03-07 Impact factor: 4.411