Injectable Tissue-engineered Bone Research Articles

Ectopic osteogenesis of an injectable nHAC/CSH loaded with blood-acquired mesenchymal progenitor cells in a nude mice model.

An injectable bone cement, nHAC/CSH, which consists of nano-hydroxyapatite/collagen (nHAC) and calcium sulphate hemihydrate (CaSO4.½H2O; CSH) was investigated as a tissue-engineered scaffold material with blood-acquired mesenchymal progenitor cells (BMPCs) as seeding cells. An in vitro study on the cytocompatability of nHAC/CSH and an in vivo study on the ectopic bone formation of nHAC/CSH loaded with dBMPCs were both conducted. The dBMPCs morphology, proliferation, differentiation and apoptosis assays were conducted using the direct contact and extract method. The cells tests exhibited normal growth and bioactive function in vitro. Studies in vivo showed that this injectable tissue engineered bone (ITB) formed bone structure in the heterotopic site of nude mice. These findings indicate that the ITB composed of nHAC/CSH and dBMPCs may represent a useful strategy for clinical reconstruction of irregular bone defects.

Injectable Bone Tissue Engineering Using Expanded Mesenchymal Stem Cells

Patients suffering from bone defects are often treated with autologous bone transplants, but this therapy can cause many complications. New approaches are therefore needed to improve treatment for bone defects, and stem cell therapy presents an exciting alternative approach. Although extensive evidence from basic studies using stem cells has been reported, few clinical applications using stem cells for bone tissue engineering have been developed. We investigated whether injectable tissue-engineered bone (TEB) composed of mesenchymal stem cells (MSCs) and platelet-rich plasma was able to regenerate functional bone in alveolar deficiencies. We performed these studies in animals and subsequently carried out large-scale clinical studies in patients with long-term follow-up; these showed good bone formation using minimally invasive MSC transplantation. All patients exhibited significantly improved bone volume with no side effects. Newly formed bone areas at 3 months were significantly increased over the preoperation baseline (p < .001) and reached levels equivalent to that of native bone. No significant bone resorption occurred during long-term follow-up. Injectable TEB restored masticatory function in patients. This novel clinical approach represents an effective therapeutic utilization of bone tissue engineering.

Alveolar Bone Regeneration around Immediate Implants Using an Injectable nHAC/CSH Loaded with Autogenic Blood‐Acquired Mesenchymal Progenitor Cells: An Experimental Study in the Dog Mandible

Lack of osseointegration between a dental implant and the walls of the alveolar bone is a common problem in immediate implantation. Injectable tissue-engineered bone (ITB) may be an effective and minimally invasive solution to the problem. In this study, an injectable bone cement, nHAC/CSH, which consists of nano-hydroxyapatite/collagen (nHAC) and calcium sulfate hemihydrate (CaSO4 .½H2 O; CSH) was investigated as a tissue-engineered scaffold material with blood-acquired mesenchymal progenitor cells (BMPC) as seeding cells. The aim of the study was to assess the new bone formation around immediate dental implants using nHAC/CSH loaded with dog blood-acquired mesenchymal progenitor cells (dBMPC) in a canine model. dBMPC were first isolated from peripheral blood of healthy adult dogs. Alizarin red and oil red O staining were then used to evaluate the potential of dBMPC to differentiate into bi-lineage mesenchymal tissues in vitro. Four healthy mongrel dogs were used in this study. The alveolar bone defects around immediate implants of dogs were created. Each defect was randomly assigned to one of the following three groups: (1) the ITB group (dBMPC+nHAC/CSH); (2) injectable bone cement nHAC/CSH; or (3) no materials (controls). Methylene blue staining was used to examine the bone formation after 3 months. Studies in vitro revealed that dBMPC could be induced to osteoblasts and adipocytes. The ITB group (dBMPC+nHAC/CSH) showed significantly more bone-implant contact and bone density than either nHAC/CSH or control groups in the areas with peri-implant defects 3 months after implantation. The results indicate that the ITB composed of nHAC/CSH and dBMPC may represent a useful strategy for the clinical reconstruction of bone defects around immediate implantation. However, further investigation is needed involving the use of human BMPC as well as possible use of stem cells.

The role of simvastatin in the osteogenesis of injectable tissue-engineered bone based on human adipose-derived stromal cells and platelet-rich plasma

An injectable tissue-engineered bone (ITB) composed of human adipose-derived stromal cells (hADSCs) and platelet-rich plasma (hPRP) was preliminarily constructed, but its osteogenic capability needs improving. This study aimed to evaluate if simvastatin can be applied as a bone anabolic agent for this ITB. We found 0.01 μ m, 0.1 μ m, and 1 μ m simvastatin could induce hADSCs’ osteoblastic differentiation in vitro that accompanied with non-inhibition on cell proliferation, high alkaline phosphatase activity, more mineralization deposition and more expression of osteoblast-related genes such as osteocalcin, core binding factor α1, bone morphogenetic protein-2, vascular endothelial growth factor, and basic fibroblast growth factor. Simvastatin at 1 μ m seemed the most optimal concentration due to its high osteocalcin secretion in media ( P < 0.01). Quantitative mineralization assay also showed 1 μ m SIM had the most obvious synergistic effect on hPRP’s induction for matrix mineralization of hADSCs ( P < 0.01). When 1 μ m Simvastatin was applied to this ITB to restore the critical-sized calvarial defects in mice, more bone formation was observed in defected regions, and the peripheries just outside the defect margins by X-ray analysis, and H&E staining. These findings indicate that simvastatin at optimal concentrations can be used to promote this ITB’s osteogenesis. However, simvastatin’s effects on this ITB await long-term investigation.

Injectable Tissue-Engineered Bone Using Autogenous Bone Marrow–Derived Stromal Cells for Maxillary Sinus Augmentation: Clinical Application Report from a 2–6-Year Follow-Up

This clinical study used injectable tissue-engineered bone, along with bone marrow-derived stromal cells (BMDSCs) and platelet-rich plasma (PRP), to conduct maxillary sinus floor augmentation by the simultaneous placement of bone graft and dental implants and to examine the state of regenerated bone after functional loading in 16 sinus augmentations in 12 patients whose alveolar crestal bone height was 2-10 mm. We used PRP as an autologous scaffold-which provides signal molecules-with in vitro expanded BMDSCs to enhance osteogenesis. All 41 dental implants prepared with the materials were clinically stable after second-stage surgery. The height of mineralized tissue at 2 years showed the mean increases of 8.8 +/- 1.6 mm compared to preoperative values, and no adverse effects and remarkable bone absorption were seen in the 2-6-year follow-up time. Although these results are preliminary, injectable tissue-engineered bone would stably predict the success of bone formation and dental implants, reduce patient burden, and provide minimally invasive cell therapy for patients.

Injectable tissue-engineered bone composed of human adipose-derived stromal cells and platelet-rich plasma

This study aimed to evaluate the effects of human platelet-rich plasma (hPRP) on the proliferation and osteogenic differentiation of human adipose-derived stromal cells (hADSCs) and to construct a novel injectable tissue-engineered bone (ITB) composed of hPRP and hADSCs. hADSCs were isolated from liposuction tissues of healthy patients. hPRP was obtained by traditional two-step centrifugation. MTT, alkaline phosphatase (ALP) activity and mineralization assays were used to evaluate the effects of different concentrations of hPRP on cell proliferation and osteogenic differentiation in vitro. hADSCs cultured in optimal concentration of activated hPRP were subcutaneously injected into the inguinal groove of nude mice with hPRP and thrombin. X-ray, H&E staining and immunohistochemical analysis were used to examine the bone formation. Studies in vitro revealed that cell proliferation, ALP activity and mineralization were induced by hPRP and 10–12.5% of hPRP seemed to be the optimal concentration. Studies in vivo showed that this ITB formed bone structure in heterotopic site of nude mice. These findings indicate that the ITB composed of hPRP and hADSCs may represent a prologue for the development of a novel biological solution to bone defect. However, further investigations should be done to fully reveal the characteristics of this ITB.

Particle Size of Hydroxyapatite Granules Calcified from Red Algae Affects the Osteogenic Potential of Human Mesenchymal Stem Cells in vitro

Hydroxyapatite (HA) microparticles as a carrier in an injectable tissue-engineered bone filler are considered promising candidates for the treatment of small bone defects in the craniomaxillofacial region. HA granules calcified from red algae, varying in size, were evaluated in vitro for their suitability to be used as a carrier for human mesenchymal stem cells (hMSCs). Three groups of granules were produced in grain sizes of 10–100, 200–500 and 600–1,000 µm. After seeding and culturing hMSCs under osteogenic differentiation conditions onto HA particles for 3, 6 and 9 days, cellular proliferation (tetrazolium salt, XTT), alkaline phosphatase (ALP)-specific activity and total protein synthesis were investigated. The osteoblastic phenotype of the cells was evaluated by assaying the bone-specific genes osteocalcin, osteopontin and collagen type I. XTT assay revealed significantly higher (p < 0.01) proliferation of cells grown on the smallest grain size after 9 days of culture. Regarding ALP-specific activity, significantly higher levels of activity were detected in cells grown on the smallest grain size. Different grain sizes had no significant effects on the secretion of osteocalcin and osteopontin. Collagen type I production was significantly higher (p < 0.05) in cells grown on the biggest grain size in comparison with the two other grain sizes. These results show that the particle size of HA microparticles affects the osteogenic potential of cultured hMSCs and lead to the conclusion that particle size has differential effects on ALP-specific activity and collagen type I production.

Towards Injectable Cell-based Tissue-Engineered Bone: The Effect of Different Calcium Phosphate Microparticles and Pre-Culturing

Towards Injectable Cell-based Tissue-Engineered Bone: The Effect of Different Calcium Phosphate Microparticles and Pre-Culturing

Clinical case reports of injectable tissue-engineered bone for alveolar augmentation with simultaneous implant placement

This clinical study was undertaken to evaluate the use of tissue-engineered bone, mesenchymal stem cells, platelet-rich plasma, and beta-tricalcium phosphate as grafting materials for maxillary sinus floor augmentation or onlay plasty with simultaneous implant placement in six patients with 3- to 5-mm alveolar crestal bone height. All 20 implants were clinically stable at second-stage surgery and 12 months postloading. A mean increase in mineralized tissue height of 7.3 ± 4.6 mm was evident when comparing the pre- and postsurgical radiographs. Injectable tissue-engineered bone provided stable and predictable results in terms of implant success.— Reprinted with permission of Quintessence Publishing.

Construction of an injectable tissue-engineered bone graft using autologous PRP and radiographic evaluation of its effect on repairing segmental radial defects in rabbits

目的 用自体富血小板血浆(PRP)、骨髓基质干细胞(BMSCs)和纤维蛋白胶(FG)构建可注射型组织工程骨并从放射学方面评价其修复自体兔桡骨缺损的效果. 方法 36只新西兰兔随机分为 A、B、C三组,每组 12只.所有动物术前 1周抽取耳背中央动脉血提取 PRP. A组于术前 1～ 2个月抽取双髂骨处骨髓并培养出 BMSCs,在体外与 FG及自体 PRP构建成可注型组织工程骨,植入自体桡骨 1.5cm节段性骨缺损,为实验组. B、C两组分别植入 PRP+ FG、FG于同样骨缺损处为对照组.另取 4只桡骨同样部位和大小骨缺损旷置作为空白组.术后 4、8、12周时摄前臂正位 X线片.用放射影像学评分和图像分析 X线阻射影比较骨缺损的修复情况. 结果 A、B两组骨缺损放射影像学评分以及新生骨痂的密度在 4、8、12周均优于 C组 (P 0.05),但在新生骨痂密度上, A >B >C(P< 0.05).空白组术后 16周骨缺损无愈合. 结论 PRP对骨缺损愈合有明显促进作用,复合 PRP的可注射型材料与 BMSCs构建组织工程骨可修复兔桡骨节段性骨缺损。

Translational research for injectable tissue-engineered bone regeneration using mesenchymal stem cells and platelet-rich plasma: from basic research to clinical case study.

Translational research involves application of basic scientific discoveries into clinically germane findings and, simultaneously, the generation of scientific questions based on clinical observations. At first, as basic research we investigated tissue-engineered bone regeneration using mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) in a dog mandible model. We also confirmed the correlation between osseointegration in dental implants and the injectable bone. Bone defects made with a trephine bar were implanted with graft materials as follows: PRP, dog MSCs (dMSCs) and PRP, autogenous particulate cancellous bone and marrow (PCBM), and control (defect only). Two months later, dental implants were installed. According to the histological and histomorphometric observations at 2 months after implants, the amount of bone-implant contact at the bone-implant interface was significantly different between the PRP, PCBM, dMSCs/ PRP, native bone, and control groups. Significant differences were also found between the dMSCs/PRP, native bone, and control groups in bone density. These findings indicate that the use of a mixture of dMSCs/ PRP will provide good results in implant treatment compared with that achieved by autogenous PCBM. We then applied this injectable tissue-engineered bone to onlay plasty in the posterior maxilla or mandible in three human patients. Injectable tissue-engineered bone was grafted and, simultaneously, 2-3 threaded titanium implants were inserted into the defect area. The results of this investigation indicated that injectable tissue-engineered bone used for the plasty area with simultaneous implant placement provided stable and predictable results in terms of implant success. We regenerated bone with minimal invasiveness and good plasticity, which could provide a clinical alternative to autogenous bone grafts. This might be a good case of translational research from basic research to clinical application.