PMID- 30844570 OWN - NLM STAT- MEDLINE DCOM- 20200416 LR - 20200416 IS - 1878-7568 (Electronic) IS - 1742-7061 (Linking) VI - 88 DP - 2019 Apr 1 TI - Culture of hybrid spheroids composed of calcium phosphate materials and mesenchymal stem cells on an oxygen-permeable culture device to predict in vivo bone forming capability. PG - 477-490 LID - S1742-7061(19)30167-9 [pii] LID - 10.1016/j.actbio.2019.03.001 [doi] AB - Three-dimensional (3-D) cell culture can better mimic physiological conditions in which cells interact with adjacent cells and the extracellular matrix than monolayer culture. We have developed a 3-D cell culture device, the Oxy chip, which can be used to generate and supply oxygen to cell spheroids to prevent hypoxia. Here, we used the Oxy chip to generate hybrid spheroids comprising calcium phosphate (CaP) particles (hydroxyapatite (HA), beta-tricalcium phosphate (beta-TCP) or octacalcium phosphate (OCP)) and mesenchymal stem cells (MSCs, C3H10T1/2 cells or D1 cells) that can be used to analyze cell differentiation mechanisms. We showed that the 3-D cell-cell and cell-material interactions and oxygenation offered by the Oxy chip promoted osteoblastic differentiation of MSCs. We also used histomorphometric analysis of hematoxylin and eosin staining, quality analyses by muCT and collagen orientation observation with picrosirius red staining in bone regeneration following implantation of three CaPs in a critical-sized defect in mouse calvaria. The in vivo bone formation capacity of the three tested CaP materials was OCP >/= beta-TCP > HA: the newly formed bone by OCP had a structure relatively close to that of the calvaria intact bone. When MSCs were 3-D cultured with the CaP materials using the Oxy chip, the in vitro osteogenic capacity of these materials was highly similar to trends observed in vivo. The in vitro alkaline phosphatase activity of D1 cells had the highest correlation with in vivo bone volume (R = 0.900). Chemical and FTIR spectroscopic analyses confirmed that differentiation of D1 cells could be associated with amorphous calcium phosphate (ACP) precipitation concomitant with OCP hydrolysis. Taken together, hybrid spheroid cultures using the Oxy chip can be used to screen and predict bone forming potential of bone substitute materials. STATEMENT OF SIGNIFICANCE: An oxygen permeable-culture chip (Oxy chip) can be used to induce formation of cell spheroids by mesenchymal stem cells (MSCs). Use of the Oxy chip avoids hypoxia in the spheroid core and enhances MSC osteoblastic differentiation relative to conventional spheroid culture methods. The present study showed that the Oxy chip mimics the in vivo environment associated with bone formation and can be used to generate hybrid spheroids consisting of calcium phosphates and MSCs that are useful for analyzing cell differentiation mechanisms. Bone formation analysis following implantation of calcium phosphate materials in mouse calvaria defects showed positive correlation with the in vitro results. We propose that hybrid spheroids cultured on the Oxy chip can be used to screen and predict the bone forming potential of bone substitute materials. CI - Copyright (c) 2019 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved. FAU - Sato, Tomoya AU - Sato T AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan; Division of Advanced Prosthetic Dentistry, Tohoku University Graduate School of Dentistry, Sendai, Japan. FAU - Anada, Takahisa AU - Anada T AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan; Soft Materials Chemistry, Institute for Materials Chemistry and Engineering, Kyushu University, Fukuoka, Japan. Electronic address: takahisa_anada@ms.ifoc.kyushu-u.ac.jp. FAU - Hamai, Ryo AU - Hamai R AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan. FAU - Shiwaku, Yukari AU - Shiwaku Y AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan; Liaison Center for Innovative Dentistry, Tohoku University Graduate School of Dentistry, Sendai, Japan. FAU - Tsuchiya, Kaori AU - Tsuchiya K AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan. FAU - Sakai, Susumu AU - Sakai S AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan. FAU - Baba, Kazuyoshi AU - Baba K AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan; Department of Orthopedic Surgery, Tohoku University Graduate School of Medicine, Sendai 980-8575, Japan. FAU - Sasaki, Keiichi AU - Sasaki K AD - Division of Advanced Prosthetic Dentistry, Tohoku University Graduate School of Dentistry, Sendai, Japan. FAU - Suzuki, Osamu AU - Suzuki O AD - Division of Craniofacial Function Engineering, Tohoku University Graduate School of Dentistry, Sendai, Japan. Electronic address: suzuki-o@m.tohoku.ac.jp. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20190304 PL - England TA - Acta Biomater JT - Acta biomaterialia JID - 101233144 RN - 0 (Calcium Phosphates) RN - 13767-12-9 (octacalcium phosphate) RN - 9007-49-2 (DNA) RN - EC 3.1.3.1 (Alkaline Phosphatase) RN - S88TT14065 (Oxygen) SB - IM MH - Alkaline Phosphatase/metabolism MH - Animals MH - Calcium Phosphates/*pharmacology MH - Cell Culture Techniques/*instrumentation MH - Cell Differentiation/drug effects MH - Cell Line MH - *Cell Membrane Permeability/drug effects MH - DNA/metabolism MH - Disease Models, Animal MH - Male MH - Mesenchymal Stem Cells/*cytology/drug effects/metabolism MH - Mice, Inbred ICR MH - Osteoblasts/cytology/drug effects/enzymology MH - *Osteogenesis/drug effects MH - Oxygen/*pharmacology MH - Skull/diagnostic imaging/pathology MH - Spectroscopy, Fourier Transform Infrared MH - Spheroids, Cellular/*cytology/drug effects MH - X-Ray Microtomography OTO - NOTNLM OT - Bone regeneration OT - Calcium phosphate materials OT - Hybrid spheroids OT - Octacalcium phosphate OT - Spheroid culture device EDAT- 2019/03/08 06:00 MHDA- 2020/04/17 06:00 CRDT- 2019/03/08 06:00 PHST- 2018/12/17 00:00 [received] PHST- 2019/02/28 00:00 [revised] PHST- 2019/03/03 00:00 [accepted] PHST- 2019/03/08 06:00 [pubmed] PHST- 2020/04/17 06:00 [medline] PHST- 2019/03/08 06:00 [entrez] AID - S1742-7061(19)30167-9 [pii] AID - 10.1016/j.actbio.2019.03.001 [doi] PST - ppublish SO - Acta Biomater. 2019 Apr 1;88:477-490. doi: 10.1016/j.actbio.2019.03.001. Epub 2019 Mar 4.