Résumé
Objective To prepare biphasic calcium phosphate/polyvinyl alcohol scaffolds by 3D printing at room temperature and explore the effect of 3D scaffolds on in vitro osteogenic differentiation of the bone marrow mesenchymal stem cells (BMSCs).Methods After biphasic calcium phosphate and polyvinyl alcohol solutions were mixed,the biphasic calcium phosphate/polyvinyl alcohol composite scaffolds were prepared by room temperature 3D printing combined with freeze drying technique.Non-printing scaffolds were prepared by injection molding.The surface microstructure,porosity,elastic modulus and hydrophilicity of the 2 sorts of scaffolds were measured.The cytological experiments were carried out in 3 groups (n =3):printed scaffold group,non-printed scaffold group and blank control group (no scaffold).After the BMSCs were seeded onto the scaffolds for 7 and 14 days,the 3 groups were compared in terms of cellular proliferation,alkaline phosphatase activity and expression levels of osteogenesis-related genes.Results 3D composite scaffolds with controllable pore size and porosity were prepared successfully,with an average porosity of 59.6％ ± 3.6％ and an average elastic modulus of 429.3 ± 54.3 kPa.After culture for 7 and 14 days,the cellular absorbance values in the printed scaffold group (0.987 ± 0.047 and 1.497 ± 0.076) were significantly higher than those in the non-printed scaffold group (0.767 ±0.063 and 1.181 ±0.098) (P ＜ 0.05) which were in turn significantly higher than those in the blank control group (0.532 ±0.046 and 0.895 ± 0.062) (P ＜ 0.05).After culture for 7 and 14 days,the ALP activity and expression levels of osteogenesis-related genes in the printed and non-printed scaffold groups showed no significant between-group differences (P ＞ 0.05),but were significantly higher than those in the blank control group (P ＜ 0.05).Conclusions Tissue-engineered composite biphasic calcium phosphate/polyvinyl alcohol scaffolds with controllable pore size and good connectivity can be prepared by freeze-drying and room temperature 3D printing techniques.Co-culture of the scaffolds and BMSCs in vitro promotes adhesion,proliferation and osteogenic differentiation of the cells.
Résumé
Objective To prepare a bionic artificial bone scaffold using a room temperature three dimensional (3D) printing technique and evaluate its biocompatibility and bioactivity in vitro.Methods A room temperature 3D printing technique was applied to fabricate 3D bionic artificial bone scaffolds using collagen/hydroxyapatite.The physico-chemical structure,porosity and mechanical strength of the scaffolds were assessed.The extract liquid of scaffolds was cocultured with bone mesenchymal stem cells (BMSCs) to evaluate the toxicity of scaffolds.There were 3 experimental groups:blank control with no scaffolds,printed scaffolds group and non-printed scaffolds group.The condition of BMSCs on the scaffolds was observed via scanning electron microscopy(SEM) and immunostaining.3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay and SEM were applied to monitor the proliferation of BMSCs on the scaffolds.At last,alkaline phosphatase (ALP) activity and mRNA expression levels of osteogenesis-related genes were detected to assess the osteoinductive property of the scaffolds.Results The 3D printed scaffolds fabricated in the present study were characterized by highly interconnected pores which were controllable and even in size.The cross section of the scaffolds presented an irregular honeycomb-like microstructure.The porosity of printed 3D scaffolds (71.14％ ± 2.24％) was significantly higher than that of non-printed scaffolds (59.04％ ±2.98％) (P ＜ 0.05).The physico-chemical structures of the materials were preserved after printing without additional cytotoxicity.The MTT results at 7 and 14 days revealed that the printed scaffolds had a significantly more cell numbers than the non-printed scaffolds(P ＜ 0.05).SEM showed that the BMSCs adhered well onto the printed scaffolds and proliferated and migrated through the pores.Compared with the blank control,the printed scaffolds showed obviously better osteogenic outcomes.Conclusion The 3D bionic artificial bone scaffolds of collagen/hydroxyapatite manufactured by a room temperature 3D printing technique can provide a good extracellular matrix for BMSCs to proliferate and differentiate.