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The repair of bone defects, especially for the large segment of bone defects, has always been an urgent problem in orthopedic clinic and attracted researchers' attention. Nowadays, the application of tissue engineering bone in the repair of bone defects has become the research hotspot. With the rapid development of tissue engineering, the novel and functional scaffold materials for bone repair have emerged. In this review, we have summarized the multi-functional roles of osteoclasts in bone remodeling. The development of matrix-based tissue engineering bone has laid a theoretical foundation for further investigation about the novel bone regeneration materials which could perform high bioactivity. From the point of view on preserving pre-osteoclasts and targeting mature osteoclasts, this review introduced the novel matrix-based tissue engineering bone based on osteoclasts in the field of bone tissue engineering, which provides a potential direction for the development of novel scaffold materials for the treatment of bone defects.
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Humans , Bone Regeneration , Bone and Bones , Osteoclasts , Tissue EngineeringABSTRACT
@# Three dimensionally printed composite porous bone tissue engineering scaffolds have become a research focus. Composite polyvinyl alcohol (PVA) has good biocompatibilityand degradability, but it cannot be prepared indepen⁃dently because it cannot resist highmechanical resistance. This material shows many advantages, such as good biocom⁃patibility, degradability and mechanical properties, when compounded with other materials with good mechanical proper⁃ties and good biocompatibility. Therefore, 3D printed composite PVA scaffold material can optimize the performance of PVA scaffolds. This article reviews 3D printing bone scaffold technology, polyvinyl alcohol (PVA), and composite PVA scaffolds for in vivo and in vitro bone formation.
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BACKGROUND: Hydroxyapatite-geltin composite has good biocompatibility and osteoinductivity, and can used be as tissue engineering scaffold to repair bone defects. OBJECTIVE: To observe the effect of three-dimensional (3D) printed hydroxyapatite/gelatin scaffold combined with bone marrow mesenchymal stem cells and umbilical vein endothelial cells in repairing rabbit skull defects. METHODS: Nine male New Zealand white rabbits were taken to establish a skull defect model with a diameter of approximately 0.8 cm, and randomly divided into three groups: blank group: no any treatment; control group: only 3D printed hydroxyapatite-geltin scaffold; experimental group: 3D printed hydroxyapatitegeltin scaffold and bone marrow mesenchymal stem cells and bone marrow mesenchymal stem cells complex group, with three rabbits in each group. At 8 weeks after the operation, CT scan of the skull pyramid beam and histological observation of the skull defect were performed on the white rabbits of each group. Animal experiments were approved by the Ethics Committee of Jining Medical College. RESULTS AND CONCLUSION: (1) CT scan of pyramidal tract: Blank group showed obvious bone defects, and the defect area was slightly radiopaque with the edges of the surrounding normal bone tissue. Control group showed some new bone formation, which was discontinuous and inconsistent with the surrounding bone tissue. Experimental group showed that the new bone tissue was linear and continuous; the thickness was thin; and the defect area merged with the adjacent bone tissue edge. (2) Histological observation: Hematoxylin-eosin staining and Masson trichrome staining showed that fibrous connective tissue formation and a small amount of free bone cells were seen in the defect area of the blank group. A small amount of bone formation was seen in the control group. Bone matrix was deposited at the edge of the material, replacing the material to form small bone trabeculae. The material space of the experimental group was gradually replaced by new bone, and the defect area was filled with new bone and trabecular bone structure-like tissue. (3) The results show that the 3D bionic printing hydroxyapatite-geltin scaffold combined with bone marrow mesenchymal stem cells and bone marrow mesenchymal stem cells can effectively promote the growth of bone tissue and accelerate the repair of bone defects.
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Objective To investigate the effect of prevascularized tissue-engineered bone graft on regeneration of femoral bone defects in rats.Methods Models of femoral bone defect were created at the bilateral hind limbs of 20 healthy female 10 week-old rats which were divided into 2 even groups randomly (n =10).In group A,conventional tissue-engineered bone grafts were transplanted into the femoral bone defects;in group B,tissue-engineered bone grafts and vascular bundles were implanted into the femoral defects.At 1,4 and 8 weeks after operation,3 rats were sacrificed each time in each group to harvest samples.The remaining one in each group served as a spare animal.Regeneration of bone defects and degradation of scaffolds were assessed by radiologic modality and hematein eosin staining.Results At week 1,the new bone ratio (BV/TV) was 5.47％ ± 1.90％ in group A and 8.49％ ± 1.26％ in group B,showing no significant difference (P ＞ 0.05);at weeks 4 & 8,the BV/TV were 17.54％ ±2.04％ and 39.73％ ± 4.01％ in group A,significantly lower than those in group B (25.32％ ± 2.15％ and 53.22％ ± 2.94％) (P ＜ 0.05).At weeks 1 & 4,the scaffold degradation ratios (RSV/SV) were 97.33％ ± 2.52％ and 80.60％ ±4.00％,showing no significant differences from those in group B (95.67％ ±3.51％ and 75.22％ ±6.20％) (P ＞ 0.05).At week 8,the scaffold degradation ratio in group A (65.46％ ±4.51％) was significantly higher than that in group B (50.19％ ±4.91％) (P ＜ 0.05).At week 8,hematein eosin staining showed better integration of scaffolds with the femur,faster degradation of the interior scaffolds and greater osteogenetic activity in group B.Conclusion Prevascularization of tissue-engineered bone graft may increase new bone volume and scaffold degradation rate,promoting repair of femoral bone defects in rats.
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Objective To investigate the clinical efficacy of using the tissue engineering bone loaded with adipose derived stem cells (ADSCs)and perforator flap in the treatment of composite tissue defects.Methods From April,2013 to June,2015,there were 9 cases of traumatic bone and skin composite tissue defects,including 7 males and 2 females,with an averaged age of 43 years old.The ADSCs were isolated,induced and co-cultured with demineralized bone scaffold.The tissue engineering bone and deep inferior epigastric artery perforator (DIEP) flap were adopted for reconstruction of composite tissue defects.Results All 9 patients were followed up for 12-36 months,averaged of 18 months.The bone growth was obviously for 5 cases with bone defects at the middle and lower part of the tibia.They tolerated full weight bearing walking.One case of middle humeral bone defect demonstrated normal bone tissue growth,and the 2/3 of cross section had been restored.One case of humeral bone defect and 1 case of radial bone defect reached bone union.The remaining 1 with skull defect showed new bone growth,but it had not yet achieved complete bone healing.Conclusion The combination of tissue engineering bone and perforator flap is a minimally invasive,easy accessible and effective method for reconstruction of composite tissue defects.
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Objective To study the feasibility of microsurgical technique to denervate sympathetic of femoral artery in rabbit, providing a reliable animal experimental model for further study of the mechanism of neuralization in bone tissue engineering.Methods From July, 2014 to July, 2015, 21 New Zealand white rabbits were divided into 4 groups randomly: the control group (n =3), the 4 weeks group (n =6), the 8 weeks group (n =6) and the 12 weeks group (n =6).Bilateral femoral arteries of the 21 rabbits were exposed.Adventitia of femoral arteries in 3 test groups were removed for about 2cm by microsurgical technique, whereas adventitia of the control group remained intact without any treatment.The arteries samples were collected at 4 weeks, 8 weeks and 12 weeks after treatment.The structure of vascular were indicated by hematoxylin-eosin (HE) staining, and the distribution and volume of the sympathetic fibers were evidenced by glyoxylic acid staining and the expression of tyrosine hydroxylase (TH), the marked protein of sympathetic.Results The adventitia of 3 test groups were invisible or lost most of it while the control group remained intact shown by HE staining.For glyoxylic acid staining, the fluorescence intensity value of the control group, 4 weeks group, 8 weeks and 12 weeks were 0.08124 ± 0.00260, 0.02920 ± 0.00206, 0.02661 ± 0.00233, 0.03094 ± 0.00211, respectively (n =6).The distribution and fluorescence intensity of sympathetic nerve were both significantly reduced in test groups compared to the control group (P ＜ 0.05).And there was no statistical difference among the 3 test groups (P ＞ 0.05).Semi-quantitative analysis of the expression of TH was 0.8626 ± 0.03519, 0.3631 ± 0.03019, 0.3964 ± 0.02239, 0.3487 ± 0.02356 respectively, which showed the same tendency as glyoxylic acid staining test.Conclusion Microsurgical technique is promising as an ideal method for the local denervation of sympathetic nerve from artery system as it can significantly reduce sympathetic fibers on adventitia without regeneration during the experimental period.
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Objective To construct bone marrow stem cell sheets and to investigate its effects in the process of osteogenesis.Methods BMSCs were differentiated into osteoblasts and then seeded into a temperature responsive culture dish to construct BMSC sheets.PLGA scaffolds in which both BMSC suspension and BMSC sheets were added,were implanted into the left side of the dogs' mandible.In the other side,PLGA scaffolds that were not wapped with BMSC sheets were implanted as control.At 16 weeks,the samples were processed for radiological analysis and histological examination.Results Cells in the BMSC sheets grew well.In the experimental side,the optical density of the samples was higher than that of the control side (P＜0.05) and plenty of lamellar bones and Haversian system were observed.Conclusions The formation of lamellar bones can be promoted by PLGA scaffolds and BMSC sheets in the process of tissue engineering bone reconstrution.
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Objective The tissue engineering technique and the microsurgery technology is combined to construct the uncellular tissue engineering complex with vascularization and membrane guided dual effect. Through comparing study of using the simple biomembrane guided bone regeneration technique to construct the uncellular tissue engineering complex to repair the large segment bone defect in the animal body,the bone reparative effect of the tissue engineering bone wrapped by pedical fascial flap with vessels and that wrapped by the simple biomembrane was compared, thus to provide experimental evidence for the clinical application. Methods Twenty-four Newzland 5-month-old rabbits were used to build the bilateral periosteumincluded bone defect modelsin the middle piece of the ulna and the length of the defect was 1 cm. Autologous red bone marrow was implanted in the tissue engineering bone which was prepared by osteoinductive absorbing material including BMP. The prepared tissue engineering bone was implanted in the bone defect area. The right side was wrapped by the simple absorbable biomembrane, whereas the left side was wrapped by pedical fascial flap with blood supply. At the fourth, eighth, twelfth and sixteenth week after the operation each group was examined by the radiograph (x-ray), the light density measurement, gross morphology and histological inspection,bone shape measurement analysis in the repairing area and the biomechanics measurement at the twelfth week. The data was analyzed to test the difference of the bond defect repair. Results The radiograph, gross morphology and histological inspection showed the growth of vessels in the implant area, the quantity and the forming speed of the bone trabecula and, the cartilaginous tissue, the formation of the mature bone structure,remodeling of the diaphysis, recanalization of the cavum ossis and the absorption and the degradation of the implant of the group of pedical fascial flap with blood supply was superior to that of the group of the simple absorbable biomembrane. At the fourth, eighth, twelfth and sixteenth week after the operation the bone trabecula area were( 20. 35 ± 2. 41 ) %, ( 40. 21 ± 1.97 ) %, (66. 67 ± 3.44 ) % and ( 86. 47 ± 3.99) % respectively in the group of pedical fascial flap with blood supply, and were ( 7. 46 ± 2.64 ) %, ( 20. 66 ± 2. 28 ) % , ( 40. 22 ±1.84)% and(58. 18 ± 1.79) respectively in the group of the simple absorbable biomembrane. At the same time point after the operation the light density were 0. 636 ± 0. 012,0. 596 ± 0. 062,0. 552 ± 0. 009 and 0. 451 ±0. 008 respectively in the group of pedical fascial flap with blood supply, and 0. 742 ± 0. 032,0. 713 ± 0. 022,0. 655 ±0. 018 and 0. 606 ±0. 015 respectively in the group of the simple absorbable biomembrane. The units of blood vessel reproductive area in the bone repair junctional zone were ( 18.75 ± 2. 09 ) %, ( 37.41 ± 3.22 ) %,(53. 06 ±2. 18)% and (36.72 ±4. 73)% respectively in the group of pedical fascial flap with blood supply,and (5. 34 ± 1.17 ) %, (9. 48 ± 2.96) %, ( 22.43 ± 2. 21 ) % and ( 26. 27 ± 3. 14 ) % respectively in the group of the simple absorbable biomembrane. The biomechanics intension was 26.62 ± 3.96 in the group of pedical fascial flap with blood supply and 18. 38 ±0. 71 in the group of the simple absorbable biomembrane at the twelfth week after the operation. All of the differences were significant( P ＜0. 05 ). Conclusion The pedical fascial flap with blood supply has significant effect in promoting the tissue engineering bone to vascularize and promoting the bone formation by vascularization. The membrane guided bone regeneration technique restricted not only the growth of the fibrous connective tissue in the reparative process of the large segment bone defect effectively, but also the ability of fast and effective vascularization, thus the chronic creep and substitution process would be needed. Simple application of the biomembrane can compensate the shortcoming of chronic creep of the implanted material by the growth of the external callus.
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@# ObjectiveTo evaluate whether β-tricalcium phosphate (β-TCP) combined with bone marrow mesenchymal stem cells (BMSCs) can be used for lumbar posterolateral spine fusion (PLF) instead of autogenous bone graft. Methods6 crab-eating macaques underwent bilateral PLF at L4-5, and divided into 3 groups that implanted β-TCP/BMSCs composite, autogenous bone, and β-TCP. Monkeys were sacrificed 12 weeks after implantation. Manual palpation, micro computed tomography, peripheral quantitative computed tomography (pQCT), and histology were used to assess bone formation. ResultsManual palpation showed that 75% of β-TCP/BMSCs composite group and autogenous group achieved solid spine fusion, whereas none of β-TCP group fused. Histological analysis showed that all of the β-TCP/BMSCs group achieved massive bone formation. Bone mineral density (BMD) evaluated with pQCT in the β-TCP/BMSCs group increased by additional new bone. Conclusionβ-TCP/BMSCs composite can be used for PLF instead of autogenous bone graft.
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Objective To study the effect of a novel injectable scaffold material chitosan- beta-TCP combining bone marrow mesenchymal stem cells (MSCs) and platelet-rich plasma (PRP) on repairing bone defect of goat. Methods The model of the studies was 12ram diameter circular hole tibia bone defect of goat. 30 Chinese goats were raudomly divided into 5 groups: blank group: nothing was embeded in bone defect; simple material group: the material embeded in bone defect was chitosan-beta-TCP; PRP group: the material was chitesan-beta-TCP combining PRP; MSCs group: the material was chitosan-beta-TCP combining MSCs; PRP/MSCs group:the material was chitosan-beta-TCP combining MSCs and PRP. At 4,8 weeks after operation, the samples were observed, histological and image analysis were used to evaluate the effect of bone regeneration. Results At 8 weeks, the surface of bone defect zone of PRP/MSCs group were coverd by continuous new bones, like normal bone. Histological slice showed the esteoid at boundary of normal bone of MSCs/PRP group obviously increased compare to other groups at the 4th or 8th week after operation respectively. The new bone tissues of bone defect were punctiform or lamellar new bone tissues, in which the proportion of big lamellar new bone tissue obviously increased. Image analysis showed that the areas of balnk group, simple material group, PRP group, MSCs group, PRP/MSCs group were 8.79±3.63,14.49± 3.72,24.18 ± 5.38,24.42 ± 5.10,31.10 ± 3.49 at 4 weeks and 15.41 ± 4.21,25.36 ± 5.37,30.71 ± 4.39, 33.97 ± 4.45,48.60 ± 5.97 at 8 weeks respectively. The effect of bone regeneration of PRP/MSCs group was better than other groups (P < 0.05). Conclusion The injectable tissue-engineering bone constructed with chitosan-beta-TCP, MSCs and PRP possesses good ability on repairing bone defect.
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The vaseularization of tissue-engineered bone is the key problem which the development and employment of large sized tissue-engineered bone.The vascular endothelial cell has a great effect on promoting vascularization in tissue-engineered bone.Vascularizations fall into two modes of vaseulogenesis and angiogenesis according to differences in source of endothelial cells.Co-culture of osteoblasts and vascular endothelial cells has better result than single culture of each kind of cells.Different ways of improving the vascularization,such as searching for new source of vascular endothelial cell,co-culture and in vivo experiment are investigated to meet the challenge of bone tissue engineering.
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Objective To observe the changes of osteoblasts and vascularization during osteogenesis by tissue engineering technique under the electron microscope and study the feasibility of improving vascularization of the tissue engineering bone by using the small intestine submucosa(SIS) as the scaffold. Methods The bone mesenchymal stem cells(BMSCs) were isolated by using the gradient centrifuge method.BMSCs were seeded in the SIS.The scaffold-cell constructs were cultured in vitro for two weeks.There were no cells on the SIS as control.They were implanted subcutaneously in the dorsa of the athymic mice.The implants were harvested after(in vivo) incubation for 4,8 and 12 weeks.The changes of osteoblasts and vascularization were observed under the transmission electron microscope and the scanning electron microscope. Results The BMSCs grew quite well.BMSCs differentiated on the surface of the SIS and secreted a great deal of extracellular matrices.The scaffold-cell constructs formed a lot of bone and vessels in vivo.The scaffold degraded after 12 weeks.No osteoblasts but vascularization and fibroblasts were observed as control. Conclusion SIS can be used as the scaffold for constructing tissue engineering bone as it can improve the formation of bone and vessels in vivo.
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Recombinant human BMP-2 was compounded with chitosan/gelatin/hydroxyapatite(HCG) scaffold and the complex was sterilized by 60Co radiating. Osteoblast isolated from cranial bones of newborn rat was primary cultured and seeded onto the complexes. 3 days after culturing, scanning electron microscope(SEM) was applied to detect the compatibility of the cell with the complex. SEM showed osteoblast attached closely with the complex and grew well in its pores. Then the complexes with osteoblast modification were implanted into athymic nude mice subcutaneously. 8 weeks after implantation, X-ray photograph and histological observation were applied to detect the bone formation of the complexes. Under X-ray a high-density areas consistent with the shape of the implanted complex could be seen. Histological observation also proved there was bone formation in the interspace of the complex. A conclusion was drawn that rhBMP-2 compounded HCG scaffold had good osteogenesis ability in vivo.
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Tissue-engineering bone with porous β-tricalcium phosphate (β-TCP) ceramic and autologous bone marrow mesenchymal stem cells (MSC) was constructed and the effect of this composite on healing of segmental bone defects was investigated. 10-15 ml bone marrow aspirates were harvested from the iliac crestof sheep, and enriched for MSC by density gradient centrifugation over a Percoll cushion (1. 073 g/ml). After cultured and proliferated, tissue-engineering bones were constructed with these cells seeded onto porous β-TCP, and then the constructs were implanted in 8 sheep left metatarsus defect (25 mm in length) as experimental group. Porous β-TCP only were implanted to bridge same size and position defects in 8 sheep as control group, and 25 mm segmental bone defects of left metatarsus were left empty in 4 sheep as blank group. Sheep were sacrificed on the 6th, 12th, and 24th week postoperatively and the implants samples were examined by radiograph, histology, and biomechanical test. The 4 sheep in blank group were sacrificed on the 24th week postoperatively. The results showed that new bone tissues were observed either radiographic or histologically at the defects of experimental group as early as 6th week postoperatively, but not in control group, and osteoid tissue, woven bone and lamellar bone occurred earlier than in control group in which the bone defects were repaired in "creep substitution" way, because of the new bone formed in direct manner without progression through a cartilaginous intermediate. At the 24th week, radiographs and biomechanical test revealed an almost complete repair of the defect of experimental group, only partly in control group. The bone defects in blank group were non-healing at the 24th week. It was concluded that engineering bones constructed with porous β-TCP and autologous MSC were capable of repairing segmental bone defects in sheep metatarsus beyond "creep substitution" way and making it healed earlier. Porous β-TCP being constituted with autologous MSC may be a good option in healing critical segmental bonedefects in clinical practice and provide insight for future clinical repair of segmental defect.