Repair Of Cranial Bone Defects Research Articles

Dynamic Cross-Linking, Self-Healing, Antibacterial Hydrogel for Regenerating Irregular Cranial Bone Defects.

Given the widespread clinical demand, addressing irregular cranial bone defects poses a significant challenge following surgical procedures and traumatic events. In situ-formed injectable hydrogels are attractive for irregular bone defects due to their ease of administration and the ability to incorporate ceramics, ions, and proteins into the hydrogel. In this study, a multifunctional hydrogel composed of oxidized sodium alginate (OSA)-grafted dopamine (DO), carboxymethyl chitosan (CMCS), calcium ions (Ca2+), nanohydroxyapatite (nHA), and magnesium oxide (MgO) (DOCMCHM) was prepared to address irregular cranial bone defects via dynamic Schiff base and chelation reactions. DOCMCHM hydrogel exhibits strong adhesion to wet tissues, self-healing properties, and antibacterial characteristics. Biological evaluations indicate that DOCMCHM hydrogel has good biocompatibility, in vivo degradability, and the ability to promote cell proliferation. Importantly, DOCMCHM hydrogel, containing MgO, promotes the expression of osteogenic protein markers COL-1, OCN, and RUNX2, and stimulates the formation of new blood vessels by upregulating CD31. This study could provide meaningful insights into ion therapy for the repair of cranial bone defects.

A multifunctional polydopamine/genipin/alendronate nanoparticle licences fibrin hydrogels osteoinductive and immunomodulatory potencies for repairing bone defects

Here, we fabricated a hybrid nanoparticle composed of polydopamine nanoparticles (pNPs), alendronate (Al) and genipin (GP) for cranial bone defect repair. Al was crosslinked into pNPs via GP (Al@pNPs), after which hybrid nanoparticles were obtained. By embedding these Al@pNPs into the fibrin hydrogels, a multifunctional bone repair scaffold was fabricated (Al@pNPs/Fg). The Al@pNPs/Fg exhibited three synergistic effects on the bone microenvironment: i) enhanced ectomesenchymal stem cell (EMSC) osteogenic differentiation by activating the piezo 1 channel; ii) inhibited the formation and function of osteoclasts related to the NF-κB signaling pathways; and iii) promoted M2 polarization and anti-inflammatory factor expression under normal and simulated inflammatory conditions. Al@pNPs/Fg ultimately promoted cranial bone defect regeneration in an SD rat model. This simple and low-cost technology provides a new approach to constructing an efficient delivery system and has desirable biological properties, providing a tissue-committed niche for the repair of bone defects.

Enhanced bone regeneration via local low-dose delivery of PTH1-34 in a composite hydrogel.

Introducing bone regeneration-promoting factors into scaffold materials to improve the bone induction property is crucial in the fields of bone tissue engineering and regenerative medicine. This study aimed to develop a Sr-HA/PTH1-34-loaded composite hydrogel system with high biocompatibility. Teriparatide (PTH1-34) capable of promoting bone regeneration was selected as the bioactive factor. Strontium-substituted hydroxyapatite (Sr-HA) was introduced into the system to absorb PTH1-34 to promote the bioactivity and delay the release cycle. PTH1-34-loaded Sr-HA was then mixed with the precursor solution of the hydrogel to prepare the composite hydrogel as bone-repairing material with good biocompatibility and high mechanical strength. The experiments showed that Sr-HA absorbed PTH1-34 and achieved the slow and effective release of PTH1-34. In vitro biological experiments showed that the Sr-HA/PTH1-34-loaded hydrogel system had high biocompatibility, allowing the good growth of cells on the surface. The measurement of alkaline phosphatase activity and osteogenesis gene expression demonstrated that this composite system could promote the differentiation of MC3T3-E1 cells into osteoblasts. In addition, the in vivo cranial bone defect repair experiment confirmed that this composite hydrogel could promote the regeneration of new bones. In summary, Sr-HA/PTH1-34 composite hydrogel is a highly promising bone repair material.

Activation of NLRP3 signaling contributes to cadmium-induced bone defects, associated with autophagic flux obstruction

Cadmium (Cd) is a widespread environmental and industrial pollutant to cause various bone metabolic diseases. Our former study reported that Cd promoted adipogenesis and inhibited osteogenic differentiation of primary bone marrow-derived mesenchymal stem cells (BMSCs) by NF-κB inflammation signaling and oxidative stress, and Cd-induced osteoporosis of long bone and compromised repair of cranial bone defect in vivo. However, the underlying mechanisms of Cd-induced bone damage remain elusive. In this study, we used Sprague Dawley (SD) rat and NLRP3-knockout mouse models to elucidate the exact effects and molecular mechanisms of Cd-induced bone damage and aging. Herein we found that the exposure of Cd preferentially targeted a few specific tissues such as bone and kidney. Cd triggered NLRP3 inflammasome pathways and the accumulation of autophagosomes of primary BMSCs, and also Cd stimulated the differentiation and bone resorption function of primary osteoclasts. Moreover, Cd not only activated ROS/NLRP3/caspase-1/p20/IL-1β pathways, but also influenced Keap1/Nrf2/ARE signaling. The data revealed that autophagy dysfunction and NLRP3 pathways synergistically mediated the impairments of Cd in bone tissues. Loss of NLRP3 function partially alleviated Cd-induced osteoporosis and craniofacial bone defect in the NLRP3-knockout mouse model. Furthermore, we characterized the protective effects and potential therapeutic targets of the combined treatment of anti-aging agents (rapamycin+melatonin+NLRP3 selective inhibitor MCC950) on Cd-induced bone damage and inflammatory aging. These results illuminate that ROS/NLRP3 pathways and autophagic flux obstruction are involved in the Cd-induced toxic actions of bone tissues. Collectively, our study unveils some therapeutic targets and the regulatory mechanism to prevent Cd-caused bone rarefaction. The findings improve the mechanistic understanding of environmental Cd exposure-caused bone metabolism disorders and tissue damage.

Overexpression of fibroblast growth factor receptor 2 in bone marrow mesenchymal stem cells enhances osteogenesis and promotes critical cranial bone defect regeneration.

Background: Reconstruction of cranial bone defects is one of the most challenging problems in reconstructive surgery, and several biological tissue engineering methods have been used to promote bone repair, such as genetic engineering of bone marrow mesenchymal stem cells (BMSCs). Fibroblast growth factor receptor 2 (Fgfr2) is an important regulator of bone construction and can be used as a potential gene editing site. However, its role in the osteogenesis process of BMSCs remains unclear. This article clarifies the function of Fgfr2 in BMSCs and explores the role of Fgfr2-overexpressed BMSCs carried by light-induced porous hydrogel (GelMA) in the repair of cranial bone defects. Methods: Lenti-virus was used to overexpress Fgfr2 in BMSCs, and cell counting kit-8, transwell, and flow cytometry assays were conducted to investigate the proliferation, migration, and characteristics. After 0, 3, 7, and 10 days of osteogenic or chondrogenic induction, the changes in osteogenic and chondrogenic ability were detected by real-time PCR, western blot, alkaline phosphatase staining, alizarin Red staining, and alcian blue staining. To investigate the viability of BMSCs carried by GelMA, calcein and propyl iodide staining were carried out as well. Finally, a critical cranial bone defect model was established in 6-week-old male mice and micro-computerized tomography, masson staining, and immunohistochemistry of OCN were conducted to test the bone regeneration properties of implanting Fgfr2-overexpressed BMSCs with GelMA in cranial bone defects over 6 weeks. Results: Overexpression of Fgfr2 in BMSCs significantly promoted cell proliferation and migration and increased the percentage of CD200+CD105+ cells. After osteogenic and chondrogenic induction, Fgfr2 overexpression enhanced both osteogenic and chondrogenic ability. Furthermore, in cranial bone defect regeneration, BMSCs carried by light-induced GelMA showed favorable biocompatibility, and Fgfr2-overexpressed BMSCs induced superior cranial bone regeneration compared to a normal BMSCs group and an untreated blank group. Conclusion: In vitro, Fgfr2 enhanced the proliferation, migration, and stemness of BMSCs and promoted osteogenesis and chondrogenesis after parallel induction. In vivo, BMSCs with Fgfr2 overexpression carried by GelMA showed favorable performance in treating critical cranial bone defects. This study clarifies the multiple functions of Fgfr2 in BMSCs and provides a new method for future tissue engineering.

A Novel Porous Butyryl Chitin-Animal Derived Hydroxyapatite Composite Scaffold for Cranial Bone Defect Repair.

Bone defects, a common orthopedic problem in clinical practice, are a serious threat to human health. As alternative materials to autologous bone grafts, synthetic cell-free functionalized scaffolds have been the focus of recent research in designing scaffolds for bone tissue engineering. Butyryl chitin (BC) is a derivative of chitin (CT) with improved solubility. It has good biocompatibility, but few studies have investigated its use in bone repair. In this study, BC was successfully synthesized with a degree of substitution of 2.1. BC films were prepared using the cast film method and showed strong tensile strength (47.8 ± 4.54 N) and hydrophobicity (86.4 ± 2.46°), which was favorable for mineral deposition. An in vitro cytological assay confirmed the excellent cell attachment and cytocompatibility of the BC film; meanwhile, in vivo degradation indicated the good biocompatibility of BC. Hydroxyapatite (HA), extracted from bovine cancellous bone, had good cytocompatibility and osteogenic induction activity for the mouse osteoblast cell line MC3T3-E1. With the aim of combining the advantages of BC and HA, a BC-HA composite scaffold, with a good pore structure and mechanical strength, was prepared by physical mixing. Administered into skull defects of rats, the scaffolds showed perfect bone-binding performance and effective structural support, and significantly promoted the regeneration of new bone. These results prove that the BC-HA porous scaffold is a successful bone tissue engineering scaffold and has strong potential to be further developed as a substitute for bone transplantation.

IPSC-neural crest derived cells embedded in 3D printable bio-ink promote cranial bone defect repair

Cranial bone loss presents a major clinical challenge and new regenerative approaches to address craniofacial reconstruction are in great demand. Induced pluripotent stem cell (iPSC) differentiation is a powerful tool to generate mesenchymal stromal cells (MSCs). Prior research demonstrated the potential of bone marrow-derived MSCs (BM-MSCs) and iPSC-derived mesenchymal progenitor cells via the neural crest (NCC-MPCs) or mesodermal lineages (iMSCs) to be promising cell source for bone regeneration. Overexpression of human recombinant bone morphogenetic protein (BMP)6 efficiently stimulates bone formation. The study aimed to evaluate the potential of iPSC-derived cells via neural crest or mesoderm overexpressing BMP6 and embedded in 3D printable bio-ink to generate viable bone graft alternatives for cranial reconstruction. Cell viability, osteogenic potential of cells, and bio-ink (Ink-Bone or GelXa) combinations were investigated in vitro using bioluminescent imaging. The osteogenic potential of bio-ink-cell constructs were evaluated in osteogenic media or nucleofected with BMP6 using qRT-PCR and in vitro μCT. For in vivo testing, two 2 mm circular defects were created in the frontal and parietal bones of NOD/SCID mice and treated with Ink-Bone, Ink-Bone + BM-MSC-BMP6, Ink-Bone + iMSC-BMP6, Ink-Bone + iNCC-MPC-BMP6, or left untreated. For follow-up, µCT was performed at weeks 0, 4, and 8 weeks. At the time of sacrifice (week 8), histological and immunofluorescent analyses were performed. Both bio-inks supported cell survival and promoted osteogenic differentiation of iNCC-MPCs and BM-MSCs in vitro. At 4 weeks, cell viability of both BM-MSCs and iNCC-MPCs were increased in Ink-Bone compared to GelXA. The combination of Ink-Bone with iNCC-MPC-BMP6 resulted in an increased bone volume in the frontal bone compared to the other groups at 4 weeks post-surgery. At 8 weeks, both iNCC-MPC-BMP6 and iMSC-MSC-BMP6 resulted in an increased bone volume and partial bone bridging between the implant and host bone compared to the other groups. The results of this study show the potential of NCC-MPC-incorporated bio-ink to regenerate frontal cranial defects. Therefore, this bio-ink-cell combination should be further investigated for its therapeutic potential in large animal models with larger cranial defects, allowing for 3D printing of the cell-incorporated material.

High-resolution imaging of the osteogenic and angiogenic interface at the site of murine cranial bone defect repair via multiphoton microscopy.

The spatiotemporal blood vessel formation and specification at the osteogenic and angiogenic interface of murine cranial bone defect repair were examined utilizing a high-resolution multiphoton-based imaging platform in conjunction with advanced optical techniques that allow interrogation of the oxygen microenvironment and cellular energy metabolism in living animals. Our study demonstrates the dynamic changes of vessel types, that is, arterial, venous, and capillary vessel networks at the superior and dura periosteum of cranial bone defect, suggesting a differential coupling of the vessel type with osteoblast expansion and bone tissue deposition/remodeling during repair. Employing transgenic reporter mouse models that label distinct types of vessels at the site of repair, we further show that oxygen distributions in capillary vessels at the healing site are heterogeneous as well as time- and location-dependent. The endothelial cells coupling to osteoblasts prefer glycolysis and are less sensitive to microenvironmental oxygen changes than osteoblasts. In comparison, osteoblasts utilize relatively more OxPhos and potentially consume more oxygen at the site of repair. Taken together, our study highlights the dynamics and functional significance of blood vessel types at the site of defect repair, opening up opportunities for further delineating the oxygen and metabolic microenvironment at the interface of bone tissue regeneration.

Preparation of photothermal-sensitive PDGF@ZIF-8-PDA@COL/PLGA-TCP composite scaffolds for bone defect repair

The repair of bone defects has long been a challenging and significant health question. Here, collagen hydrogel incorporating platelet-derived growth factor (PDGF)-loaded photopolymerizable ZIF-8-PDA nanoparticles (PDGF@ZIF-8-PDA@COL hydrogel) was prepared and perfused into 3D printed poly (lactide-co-glycolide)-tricalcium phosphate (PLGA-TCP) scaffolds. The resulting PDGF@ZIF-8-PDA@COL/PLGA-TCP composite scaffolds were applied as a bone substitute for cranial bone defect repair. The photopolymerizable ZIF-8-PDA nanoparticles had a mean size of 226.2 ± 5.3 nm with photothermal conversion capacity. PDGF@ZIF-8-PDA@COL/PLGA-TCP composite scaffolds showed a slower release of PDGF compared to PDGF release from collagen hydrogels. The composite scaffolds exhibited excellent antibacterial properties and good in vitro osteoconductive capacity. The osteoconductive activities of PDGF@ZIF-8-PDA@COL/PLGA-TCP composite scaffolds were also investigated in a rat cranial bone defect model in vivo by micro-CT imaging, hematoxylin and eosin staining, Masson’s trichrome staining, and immunohistochemical staining of osteogenesis-related markers. The PDGF@ZIF-8-PDA@COL/PLGA-TCP composite scaffolds accelerated cranial bone formation and gradually degraded over time. All these results provided strong evidence that PDGF@ZIF-8-PDA@COL/PLGA-TCP composite scaffolds might be a promising system for bone defect repair.

Spatiotemporal blood vessel specification at the osteogenesis and angiogenesis interface of biomimetic nanofiber-enabled bone tissue engineering

While extensive research has demonstrated an interdependent role of osteogenesis and angiogenesis in bone tissue engineering, little is known about how functional blood vessel networks are organized to initiate and facilitate bone tissue regeneration. Building upon the success of a biomimetic composite nanofibrous construct capable of supporting donor progenitor cell-dependent regeneration, we examined the angiogenic response and spatiotemporal blood vessel specification at the osteogenesis and angiogenesis interface of cranial bone defect repair utilizing high resolution multiphoton laser scanning microscopy (MPLSM) in conjunction with intravital imaging. We demonstrate here that the regenerative vasculature can be specified as arterial and venous capillary vessels based upon endothelial surface markers of CD31 and Endomucin (EMCN), with CD31+EMCN− vessels exhibiting higher flowrate and higher oxygen tension (pO2) than CD31+EMCN+ vessels. The donor osteoblast clusters are uniquely coupled to the sprouting CD31+EMCN+ vessels connecting to CD31+EMCN− vessels. Further analyses reveal differential vascular response and vessel type distribution in healing and non-healing defects, associated with changes of gene sets that control sprouting and morphogenesis of blood vessels. Collectively, our study highlights the key role of spatiotemporal vessel type distribution in bone tissue engineering, offering new insights for devising more effective vascularization strategies for bone tissue engineering.

Comparison of Bone Graft Storage Effectiveness Between Cryopreservation and Subcutaneous in Patients Conducted Craniectomy Decompression

Abstract
 Decompression craniectomy is a surgical method used for immediate reduction of intracranial pressure. Repair of cranial bone defects to protect the brain and reconstruct the original cranial brain compartment is called a cranioplasty. In this study researchers will conduct research and compare storage of bone graft in the freezer with a temperature of -20 ° C and subcutaneous to the risk of infection. The research design used was paired clinical trials. Clinical tests were in open label form. The analysis of significance using the Mann Whitnay test showed that the value of p = 0.381. This means that there is no significant difference between the storage group in the subcutie and the storage group in the Cryopreservation (p> 0.05). The analysis of significance using the McNemar test showed that the p value = 0.003. This means that there is a significant relationship between the storage group in the subcutie and the storage group in the Cryopreservation (p <0.05).

Nanoclay promotes mouse cranial bone regeneration mainly through modulating drug binding and sustained release

Nanoclay (Nanosilicates, NS) is appearing as an intriguing 2D nanomaterial for bone tissue engineering with multiple proposed functions, e.g., intrinsic osteoinductivity, improving mechanical properties, and drug release capacity. However, the mechanism of NS for in vivo bone regeneration has been hardly defined so far. This knowledge gap will significantly affect the design/application of NS-based biomaterials. To determine the role of NS in osteoblastic differentiation and bone formation, we used mouse calvarial-derived pre-osteoblasts (MC3T3-E1) and a clinically-relevant mouse cranial bone defect model. Instead of a hydrogel, we prepared biomimetic 3D gelatin nanofibrous scaffolds (GF) and NS-blended composite scaffolds (GF/NS) to determine the essential role of NS in critical low-dose (0.5 µg per scaffold) of BMP2-induced cranial bone regeneration. In contrast to “osteoinductivity”, our data indicated that NS could enable single-dose of BMP2, promoting significant osteoblastic differentiation while multiple-dose of BMP2 (without NS) was required to achieve similar efficacy. Moreover, our release study revealed that direct binding to NS in GF scaffolds provided stronger protection to BMP2 and sustained release compared to GF/NS composite scaffolds. Consistently, our in vivo data indicated that only BMP2/NS direct binding treatment was able to repair the large mouse cranial bone defects after 6 weeks of transplantation while neither BMP2, NS alone, nor BMP2 released from GF/NS scaffolds was sufficient to induce significant cranial bone defect repair. Therefore, we concluded that direct nanoclay-drug binding enabled sustained release is the most critical contribution to the significantly improved bone regeneration compared to other possible mechanisms based on our study.

Injectable BMP-2 gene-activated scaffold for the repair of cranial bone defect in mice.

Tissue engineering using adult human mesenchymal stem cells (MSCs) seeded within biomaterial scaffolds has shown the potential to enhance bone healing. Recently, we have developed an injectable, biodegradable methacrylated gelatin‐based hydrogel, which was especially effective in producing scaffolds in situ and allowed the delivery of high viable stem cells and gene vehicles. The well‐demonstrated benefits of recombinant adeno‐associated viral (rAAV) vector, including long‐term gene transfer efficiency and relative safety, combination of gene and cell therapies has been developed in both basic and translational research to support future bone tissue regeneration clinical trials. In this study, we have critically assessed the applicability of single‐step visible light (VL) photocrosslinking fabrication of gelatin scaffold to deliver rAAV encoding human bone morphogenetic protein‐2 (BMP‐2) gene to address the need for sustained BMP‐2 presence localized within scaffolds for the repair of cranial bone defect in mouse model. In this method, rAAV‐BMP‐2 and human bone marrow‐derived MSCs (hBMSCs) were simultaneously included into gelatin scaffolds during scaffold formation by VL illumination. We demonstrated that the subsequent release of rAAV‐BMP‐2 constructs from the scaffold matrix, which resulted in efficient in situ expression of BMP‐2 gene by hBMSCs seeded within the scaffolds, and thus induced their osteogenic differentiation without the supplement of exogenous BMP‐2. The reparative capacity of this novel stem cell‐seeded and gene‐activated scaffolds was further confirmed in the cranial defect in the severe combined immunodeficiency mice, revealed by imaging, histology, and immunohistochemistry at 6 weeks after cranial defect treatment.

A Biomimetic Zinc Alloy Scaffold Coated with Brushite for Enhanced Cranial Bone Regeneration.

Bone tissue engineering is considered as a promising pathway for bone regeneration and defect reconstruction, in which scaffolds play an important role. Zn alloy, which is a biodegradable metal material that has advantages of metallic and biodegradable characteristics, has its special features, especially the ideal degradation rate and acceptable biocompatibility, which make it worthy to be further investigated for medical applications. In this study, new biodegradable porous Zn alloy scaffolds with Ca-P coating were attempted to repair cranial bone defect, and in vitro and in vivo assays were conducted to evaluate its biocompatibility, osteo-inductivity, and osteo-conductivity. The results indicated that coated Zn alloy possessed good biocompatibility, with no cytotoxicity. It could also promote osteogenic differentiation and calcium deposition of rabbit BMSCs in vitro, and new bone formation around the scaffold in vivo. The biodegradable porous Zn alloy scaffold with Ca-P coating is considered to be promising in cranial bone defect repair.

Vasoactive Intestinal Peptide Stimulates Bone Marrow-Mesenchymal Stem Cells Osteogenesis Differentiation by Activating Wnt/β-Catenin Signaling Pathway and Promotes Rat Skull Defect Repair.

Bone defect regeneration is a complex process that involves the coordination of a variety of different type of cells. As bone tissues are innervated and rich in nerve fibers, the neuropeptides released from various never fibers could regulate bone development, metabolism, and remodeling. Among all the neuropeptides, vasoactive intestinal peptide (VIP) could modulate the functions of both osteoblasts and osteoclasts, and may play a vital role in bone marrow mesenchymal stem cell (BMSC) osteogenesis during bone repair. In this study, we investigated the role of VIP in bone formation and the mechanisms of VIP in mediating BMSC osteogenic differentiation, and its possibility in clinical application of bone defect reconstruction. Our in vitro study results indicated that VIP promoted BMSC osteogenic differentiation by activating Wnt/β-catenin signaling pathway in BMSCs. VIP could also stimulate tube formation of EA.hy926 endothelial cell and increase vascular endothelial growth factor (VEGF) expression in BMSCs. Furthermore, in the rat skull defect model, VIP-conjugated functionalized hydrogel significantly enhanced cranial bone defect repair compared with the control group, with increased bone formation and angiogenesis. Taken together, as a member of neuropeptides, VIP could promote the BMSCs osteogenesis and angiogenesis differentiation in vitro and stimulate bone repair in vivo by activating Wnt/β-catenin signaling pathway. The knowledge obtained from this study emphasized the close association between innervation and bone repair process, and VIP may be a potential therapeutic agent for augmenting bone repair.

Tuning pore features of mineralized collagen/PCL scaffolds for cranial bone regeneration in a rat model.

Porosity is indispensable for a bone tissue-engineered scaffold for facilitating endogenous cell migration and nascent bone ingrowth. In large-sized cranial bone defect repair, porous scaffolds meet great challenges to match cranial bone regeneration and provide sufficient protection with structural integrity. Therefore, the pore features of the scaffolds for cranial bone regeneration should differ from those typical porous scaffolds used in tubular bone repair and be finely tuned. In this study, a series of porous mineralized collagen/PCL scaffolds with different pore features were fabricated via freeze-drying and applied in a Sprague Dawley rat cranial bone calvarial defect model. The pore size for four groups increased from 10-45 μm to 40-130 μm. As scaffold porosity increased, the compressive strength decreased from 2.09 ± 0.12 MPa to 0.51 ± 0.04 MPa. The micro-computed tomography three-dimensional reconstruction images showed that as pore size and porosity increased, the amount of new bone formation had a maximum in group 3 (pore size: 20-100 μm, compressive strength: 1.06 ± 0.03 MPa). In addition, the histological and histomorphometric analyses showed a consistent tendency which confirmed the Micro-CT results. Meanwhile, histological findings including bony bridging, tissue response at the bone-implant interface and fibrous capsule thickness indicated that the dura mater pathway played the most important role in the regenerative process of this calvarial defect model.

Electrospun Fiber Mesh for High-Resolution Measurements of Oxygen Tension in Cranial Bone Defect Repair.

Tissue oxygenation is one of the key determining factors in bone repair and bone tissue engineering. Adequate tissue oxygenation is essential for survival and differentiation of the bone-forming cells and ultimately the success of bone tissue regeneration. Two-photon phosphorescence lifetime microscopy (2PLM) has been successfully applied in the past to image oxygen distributions in tissue with high spatial resolution. However, delivery of phosphorescent probes into avascular compartments, such as those formed during early bone defect healing, poses significant problems. Here, we report a multifunctional oxygen-reporting fibrous matrix fabricated through encapsulation of a hydrophilic oxygen-sensitive, two-photon excitable phosphorescent probe, PtP-C343, in the core of fibers during coaxial electrospinning. The oxygen-sensitive fibers support bone marrow stromal cell growth and differentiation and at the same time enable real-time high-resolution probing of partial pressures of oxygen via 2PLM. The hydrophilicity of the probe facilitates its gradual release into the nearby microenvironment, allowing fibers to act as a vehicle for probe delivery into the healing tissue. In conjunction with a cranial defect window chamber model, which permits simultaneous imaging of the bone and neovasculature in vivo via two-photon laser scanning microscopy, the oxygen-reporting fibers provide a useful tool for minimally invasive, high-resolution, real-time 3D mapping of tissue oxygenation during bone defect healing, facilitating studies aimed at understanding the healing process and advancing design of tissue-engineered constructs for enhanced bone repair and regeneration.

Repair of Cranial Bone Defects in Children Using Synthetic Hydroxyapatite Cranioplasty (CustomBone)

In pediatric cases, the use of autologous bone tissue to repair cranial bone defects is often impossible. The synthetic hydroxyapatite bone substitute (CustomBone) can be a good alternative, especially in case of a large bone defect that has to be repaired. This study focuses on a pediatric series of 30 children who underwent cranioplasty with a CustomBone implant. Patient age ranged from 8 months to 16 years, with a mean age of 7 years and 8 months. The most common indication for cranioplasty was posttraumatic decompressive craniectomy. No complications were reported. Cosmetic results were satisfactory in every patient. Only 1 implant had to be changed after severe head trauma because of an epileptic seizure in the early postoperative period. In all patients, cerebral blood flow improved during the postoperative phase. Complete implant osteointegration is a long process because mean time to begin was 13 months (range, 3-22 months). Mean patient follow-up was 6.7 years. Successful prosthesis integration depends on the accuracy of the preoperative model. The minimum thickness of the implant (4 mm) represents a challenge in very young children, but we used it with success in this series. Moreover, high costs represent another limitation for its use. The CustomBone implant meets all necessary conditions for good clinical outcome: excellent protective properties, restoration of normal intracranial physiology, satisfactory cosmetic results, good integration in the autologous bone, and good resistance in case of trauma.

Strontium-incorporated mineralized PLLA nanofibrous membranes for promoting bone defect repair

Mineralized scaffolds, which are fabricated by electrodeposition, mimic the chemistry of natural bone and have attracted a great amount of attention due to their rapid and simple production. In this study, mineralized electrospun polylactic acid (PLLA) nanofibrous membranes containing different amounts of strontium (Sr) were fabricated by an electrodeposition method for potential use in bone regeneration applications. In vitro assays, including cell proliferation and osteogenic differentiation of rat bone marrow-derived mesenchymal stem cells (BMSCs) grown on these membranes and in vivo cranial bone defect repair assays, were carried out. It was found that mineral crystals could be uniformly deposited onto the electrospun nanofibrous membranes, while the morphologies of the formed crystals were affected by the amount of Sr. By analysis of the X-ray diffraction (XRD) measurements, the formed crystalline phase was dramatically affected by the incorporation of Sr, which drove a conversion from the hydroxyapatite (HA) phase to the dicalcium phosphate dehydrate (DCPD) phase. The release of Sr2+ from the Sr/PLLA nanofibrous membranes was monitored over 20 d, and the release rates of Ca2+ and PO43− from the Sr-incorporated samples were higher compared with those of the mineralized sample without Sr. In vitro cell proliferation experiments demonstrated that mineralized Sr/PLLA nanofibrous membranes could facilitate BMSC proliferation. Furthermore, the mineralized Sr/PLLA nanofibrous membranes induced a higher degree of osteogenic differentiation in the BMSCs compared with those of pure PLLA and mineralized PLLA, as determined by the results of alkaline phosphatase (ALP) activity, alizarin red (AR) and osteocalcin (OCN) staining and the expression of osteogenesis-related genes. More importantly, in vivo cranial defect experiments revealed that mineralized Sr/PLLA nanofibrous membranes promoted bone regeneration. These findings indicated that mineralized Sr/PLLA nanofibrous membranes are a promising material for bone tissue engineering.

Stem cells combined 3D electrospun nanofibrous and macrochannelled matrices: a preliminary approach in repair of rat cranial bones

Repair of cranial bone defects is an important problem in the clinical area. The use of scaffolds combined with stem cells has become a focus in the reconstruction of critical-sized bone defects. Electrospinning became a very attracting method in the preparation of tissue engineering scaffolds in the last decade, due to the unique nanofibrous structure of the electrospun matrices. However, they have a limitation for three dimensional (3D) applications, due to their two-dimensional structure and pore size which is smaller than a cellular diameter which cannot allow cell migration within the structure. In this study, electrospun poly(ε-caprolactone) (PCL) membranes were spirally wounded to prepare 3D matrices composed of nanofibers and macrochannels. Mesenchymal stromal/stem cells were injected inside the scaffolds after the constructs were implanted in the cranial bone defects in rats. New bone formation, vascularisation and intramembranous ossification of the critical size calvarial defect were accelerated by using mesenchymal stem cells combined 3D spiral-wounded electrospun matrices.