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A hydroxyapatite-collagen (HC) composite material can mimic composition and ultra-structures of natural bone and provide adequate bioactive material-tissue interactions. Incorporation of dopamine (DA) is one of keys in increasing the mechanical strength of the HC material to approaching that of cortical bone. In this study, the in vitro osteogenic effects of polydopamine-laced hydroxyapatite collagen calcium silicate (HCCS-PDA) were examined by culturing rat mesenchymal stem cells (rMSCs) on HCCS-PDA and HCCS coated plates. HCCS-PDA group demonstrated less cytotoxic from Live/Dead cytotoxic assay and displayed higher cell attachment, proliferation and mineralization than the HCCS group in vitro. For in vivo bone regeneration, HCCS-PDA or HCCS particulates with or without rMSC aggregates were implanted into rat critical-sized calvarial defects (CSD). After 12 weeks, calvarial bone regeneration was evaluated radiographically, histologically, and histomorphometrically. While the majority of new bone formation occurred around the HCCS-PDA particulates with rMSC aggregates, The HCCS-PDA particulates without rMSC aggregates showed limited osteoconductivity. HCCS with or without rMSC aggregates resulted in less bone formation, indicating a prominent role of DA in effective bone regeneration. Therefore, the HCCS-PDA biomaterial with the aid of rMSCs can be used to develop therapeutic strategies in bone tissue engineering with numerable clinical applications.

Objective: The pore size of the scaffold is a critical factor in repairing large bone defect. Here, we investigated the potential of bone regeneration using novel nanocomposite polydopamine-laced hydroxyapatite collagen calcium silicate (HCCS-PDA) scaffolds with two different pore sizes, 250 and 500 μm.Samples/setting: A total of 12 male Sprague-Dawley rats were implanted with HCCS-PDA scaffold with pore size of either 250 or 500 μm into surgically created critical-sized defect (CSD).Methods: HCCS-PDA scaffolds were fabricated using mould printing technique. The effect of pore size on mechanical strength of the scaffolds was assessed by compression testing. After seeding with rat mesenchymal stem cells (rMSCs), the scaffolds were implanted, and new bone formation was evaluated using microCT and histomorphometric analysis after 8 weeks.Results: MicroCT and histology analysis demonstrated restricted peripheral new bone formation in either dural or periosteal side and limited new bone formation in the 250 μm pore scaffold. Conversely, the 500-μm pore scaffold showed more penetration of new bone into the scaffold and greater bone regeneration in the rat CSD.Conclusion: Based on our results, which demonstrated improved new bone formation in 500 μm pores scaffold, we can conclude that effective scaffold pore size that induces osteointegration and bone regeneration is around 500 μm for HCCS-PDA nanocomposite scaffold. [ABSTRACT FROM AUTHOR]

Bioluminescent imaging (BLI) has emerged as a popular in vivo tracking modality in bone regeneration studies stemming from its clear advantages: non-invasive, real-time, and inexpensive. We recently adopted bioluminescence resonance energy transfer (BRET) principle to improve BLI cell tracking and generated the brightest bioluminescent signal known to date, which thus enables more sensitive real-time cell tracking at deep tissue level. In the present study, we brought BRET-based cell tracking strategy into the field of bone tissue engineering for the first time. We labeled rat mesenchymal stem cells (rMSCs) with our in-house BRET-based GpNLuc reporter and evaluated the cell tracking efficacy both in vitro and in vivo. In scaffold-free spheroid 3D culture system, using BRET-based GpNLuc labeling resulted in significantly better correlation to cell numbers than a fluorescence based approach. In scaffold-based 3D culture system, GpNLuc-rMSCs displayed robust bioluminescence signals with minimal background noise. Furthermore, a tight correlation between BLI signal and cell number highlighted the robust reliability of using BRET-based BLI. In calvarial critical sized defect model, robust signal and the consistency in cell survival evaluation collectively supported BRET-based GpNLuc labeling as a reliable approach for non-invasively tracking MSC. In summary, BRET-based GpNLuc labeling is a robust, reliable, and inexpensive real-time cell tracking method, which offers a promising direction for the technological innovation of BLI and even non-invasive tracking systems, in the field of bone tissue engineering. [ABSTRACT FROM AUTHOR]

Scaffolds that are used for bone repair should provide an adequate environment for biomineralization by mesenchymal stem cells (MSCs). Recently, decellularized pulp matrices (DPM) have been utilized in endodontics for their high regenerative potential. Inspired by the dystrophic calcification on the pulp matrix known as pulp stone, we developed acellular pulp bioscaffolds and examined their potential in facilitating MSCs mineralization for bone defect repair. Pulp was decellularized, then retention of its structural integrity was confirmed by histological, mechanical, and biochemical evaluations. MSCs were seeded and proliferation, osteogenic gene expression, and biomineralization were assessed to verify DPM's osteogenic effects in vitro. MicroCT, energy-dispersive X-ray (EDX), and histological analyses were used to confirm that DPM seeded with MSCs result in greater mineralization on rat critical-sized defects than that without MSCs. Overall, our study proves DPM's potential to serve as a scaffolding material for MSC-mediated bone regeneration for future craniofacial bone tissue engineering. [ABSTRACT FROM AUTHOR]

Although bone marrow–derived mesenchymal stem cells (MSCs) have been extensively explored in bone tissue engineering, only few studies using mesenchymal stem cells from mandible (M-MSCs) have been reported. However, mesenchymal stem cells from mandible have the potential to be as effective as femur-derived mesenchymal stem cells (F-MSCs) in regenerating bone, especially in the orofacial regions, which share embryonic origin, proximity, and accessibility. M-MSCs were isolated and characterized using mesenchymal stem cell–specific markers, colony forming assay, and multi-potential differentiation. In vitro osteogenic potential, including proliferation, osteogenic gene expression, alkaline phosphatase activity, and mineralization, was examined and compared. Furthermore, in vivo bone formations of F-MSCs and M-MSCs in rat critical sized defect were evaluated using microCT and histology. M-MSCs from rat could be successfully isolated and expanded while preserving their MSC's characteristics. M-MSCs demonstrated a comparable proliferation and mineralization potentials and in vivo bone formation as F-MSCs. M-MSCs is a promising cell source candidate for craniofacial bone tissue engineering. [ABSTRACT FROM AUTHOR]

Paget's disease of bone (PDB) is a degenerative disorder affecting the skull and bones. Hyperactive osteoclasts (OCs) initiate bone degradation in the early stage, followed by increased bone formation by osteoblasts (OBs) in trabecular bones during the advanced stage. This OB-OC uncoupling results in bone deformations and irregular trabecular bone patterns. Current mouse models poorly replicate the advanced-stage characteristics of PDB. Optineurin (Gene: OPTN in humans, Optn in mice, protein: OPTN) has been implicated in PDB by genetic analyses. We identified PDB-like cortical lesions associated with OC hyperactivation in an Optn knockout (Optn −/−) mouse model. However, the effects of OPTN dysfunction on OBs and trabecular bone in advanced PDB remain unclear. In this study, we used the Optn −/− mouse model to investigate trabecular bone abnormalities and OB activity in PDB. Micro-computed tomography analysis revealed severe pagetic alterations in craniofacial bones and femurs of aged Optn −/− mice, resembling clinical manifestations of PDB. Altered OB activity was observed in aged Optn −/− mice, implicating compensatory OB response in trabecular bone anomalies. To elucidate the role of OC-OB interactions in PDB, we conducted in vitro experiments using OC conditioned media (CM) to examine the effects on OB osteogenic potential. We found OC CM restored compromised osteogenic induction of Optn −/− bone marrow stromal cells (BMSCs) from young mice, suggesting OCs maintain OB activity through secreted factors. Strikingly, OC CM from aged Optn −/− mice significantly enhanced osteogenic capability of Optn −/− BMSCs, providing evidence for increased OB activity in advanced stages of PDB. We further identified TGF-β/BMP signaling pathway in mediating the effects of OC CM on OBs. Our findings provide insights into Optn's role in trabecular bone abnormalities and OB activity in PDB. This enhances understanding of PDB pathogenesis and may contribute to potential therapeutic strategies for PDB and related skeletal disorders. • Pagetic alterations exhibit in the trabecular bones of aged Optn −/− mice. • Altered osteoblast (OB) activity may contribute to such pagetic phenotypes. • Osteoclast (OC) secretions increase OB activity in aged Optn −/− mice in vitro. • Optn −/− OC secretions activate TGF-β/BMP signaling in aged Optn −/− OB cells. [ABSTRACT FROM AUTHOR]

Abstract: Objective: Cells derived from the neural crest (NC) contribute to the development of several adult tissues, including tooth and periodontal tissue. Here, two transgenic lines, Wnt1-Cre/ZEG and P0-Cre/ZEG, were analysed to determine the fate and distribution of neural crest cells (NCCs) in adult mouse periodontal ligament (PDL). Design: Paraffin-embedded and decalcified histology samples were prepared from Wnt1-Cre/ZEG and P0-Cre/ZEG mice that were 4-, 8-, or 12-weeks old. Expression of GFP (NC-derived cells), NC-markers (Slug, AP-2 alpha, HNK-1, p75NTR and Nestin), and mesenchymal stem cell markers (CD29 and STRO-1) were examined using immunohistochemistry. Results: In four-week-old Wnt1-Cre/ZEG mice, GFP(+) NC-derived cells were specifically detected in the mid-zone of PDL. The GFP(+) cells constituted 1.4% of all cells in PDL, and this percentage decreased as the mice aged. The distribution and prevalence of GFP(+) cells were comparable between Wnt1-Cre/ZEG and P0-Cre/ZEG mice. NC-marker(+) cells were expressed only in GFP(+) cells while MSC markers were detected only in GFP(−) cells. Conclusion: The prevalence and specific distribution of NC-derived cells in adult PDL of Wnt1-Cre/ZEG and P0-Cre/ZEG mouse were examined. Interestingly, various NC markers, including markers for undifferentiated NCCs, were still expressed at high levels in GFP(+) cells. These observations may indicate that labelled cells in the Wnt1-Cre/ZEG and P0-Cre/ZEG mice did not constituted all NC-derived cells, but rather an interesting subset of NC-derived cells. These findings may be useful in understanding the homeostatic character of the PDL and contribute to establishing successful periodontal tissue maintenance. [Copyright &y& Elsevier]