OBJECTIVE: To investigate the feasibility of coralline hydroxyapatite (CHA) as scaffolds in bone tissue engineering. METHODS: The bone marrow stromal cells from 4-month New Zealand rabbits were harvested and cultured in vitro. After multiplied, dexamethasone was used to promote the osteoblastic phenotype of the cells. The cells were harvested and then seeded into CHA. By means of tissue engineering technique, osteoblastic cells/CHA complex were formed. The complex were implanted subcutaneously in nude mice. The CHA alone was implanted as control. Bone regeneration was assessed 6, 8 weeks after implantation by histological and roentgenographic analysis. RESULTS: After six weeks of implantation, x-ray film showed high-density signal, osteoid tissue formed under histological examination. Large amount of new bone were formed and connected to trabecularism 8 weeks after implantation in the experimental group. While in the control group, there were no new bone formation, but amount of fiber tissue grew into the pore of CHA 8 weeks after implantation. CONCLUSION: CHA may be used as a good scaffold material for bone tissue engineering.
In order to investigate the possibility of porous hydroxyapatite ceramics (HAC) in the repair of skull bone defect, twenty-four rabbits were used. The bone defect model was created by operation to obtain a defect in parietal bone in a size of 1 cm x 1 cm. Filled the defect with HAC and methyl-methacrylate-syrene copolymer (MMAS) to fill the defect as control. At 1st, 2nd and 3rd months after operation, behavior of the rabbits was observed and then these animals were sacrificed and specimens were examined under microscope. Results showed as follows: after operation, behavior of all animals were normal. By histological examination, it was found that in HAC group, there were granulation tissue, fibrous tissue and newly formed vessels grew into the pores and the osteoblasts formed osseous trabeculae. There was no inflammatory cell infiltration. In the MMAS grafted asea, there was formation of fibrous membrane. It suggested that HAC might be a good material for bone substitute in repair of skull bone defect.
ObjectiveTo evaluate the effect of poly-amino acid/nano-hydroxyapatite/calcium sulfate (PHC) Cage in lumbar interbody fusion of the goat. MethodsEighteen mature female goats (weighing 29-33 kg) were divided into 3 groups randomly: PHC Cage group (group A), titanium Cage group (group B), and ilium group (group C). A left extraperitoneal approach was used to establish the animal model of discectomy and interbody fusion with Cage or ilium. The general situation was observed for 24 weeks after operation. X-ray films were taken to measure disc space height (DSH) before operation and at 4, 12, and 24 weeks after operation. CT three dimensional reconstuction was performed at 24 weeks after operation to evaluate the interbody fusion according to modified Brantigan grading. The specimens of L3, 4 were harvested for mechanical test, histological, and scanning electron microscope (SEM) observation at 24 weeks after operation. ResultsAll goats survived to the end of experiment. DSH at 4 weeks after operation increased when compared with preoperative one in each group, and then decreased;DSH was significantly lower at 12 and 24 weeks after operation than preoperative one in group C (P<0.05). There was no significant difference in DSH among 3 groups at preoperation and 4 weeks after operation (P>0.05);at 12 and 24 weeks after operation, DSH of groups A and B was significantly higher than that of group C (P<0.05), but no significant difference was found between groups A and B (P>0.05). CT three dimensional reconstuction showed that bony fusion was obtained in all goats of groups A and C, and in 3 goats of group B;according to modified Brantigan grading, the scores of groups A and C were significantlly higher than that of group B (P<0.05), but no significant difference between groups A and C (P>0.05). The biomechanical test showed that there was no significant difference in range of motion between group A and group B (P>0.05), which were significantly lower than that of group C (P<0.05). Microscopy and SEM observations showed that the interface between the Cage and vertebral body in group A was compact without obvious gap, and most conjunctive region was filled with osseous tissue;the interface was filled with soft tissue, and the connection was slack with obvious gap in some region in group B;the interface connection was compact, most region was filled with osseous tissue in group C. ConclusionThe interbody fusion with PHC Cage is effective in goat lumbar interbody fusion model. The interface connection is compact between the Cage and the host bone followed by micro-degradation of PHC Cage, but the long-term degradation need further observation.
Objective To evaluate the osteogenic effect of dextran sulfate/recombinant human bone morphogenetic protein 2/chitosan (DS/rhBMP-2/CS) combined with coralline hydroxyapatite (CHA) in repairing large segmental bone defects by radiographic feature. Methods Fifty-seven 24-week-old male New Zealand rabbits were prepared for establishing right radius bone defect model of 20 mm in length. In which 54 rabbits were randomly divided into 3 groups (n=18), and the CHA, DS/rhBMP-2/CS/CHA, and rhBMP-2/CHA artificial bone grafts were implanted into the bone defect in groups A, B, and C respectively; the remaining 3 rabbits were implanted nothing as blank control group. After operation, the gross condition of the animals was observed; at 4, 8, and 12 weeks after operation, X-ray film observation, Micro-CT scanning, and three-dimensional reconstruction were performed to obtain the volume of the new bone. Results The experimental animals recovered well and were in normal condition. X-ray observation showed that the bone healing in group B was better than that in groups A and C at each time point. At each time point after operation, the X-ray scores of group B were significantly higher than that of group A and group C (P<0.05); the scores of group C at 8 and 12 weeks after operation were significantly higher than that of group B (P<0.05). Micro-CT scanning and three-dimensional reconstruction observation showed that at each time point after operation in group A, the bone defect area had less bone formation and poor osteogenesis; in group B, there were many new bone tissues in bone defect area, and the bone remodeling was well, and gradually closed to normal bone morphology at 12 weeks; in group C, there were many new bone tissues in bone defect area, but the bone formation was general. The new bone volume of group B was significantly higher than that of group A and group C (P<0.05) at each time point after operation, and the score of group C was higher than that of group A at 8 weeks after operation (P<0.05). Conclusion The osteogenic effect of DS/rhBMP-2/CS/CHA sustained-release artificial bone is much better than that of single CHA and rhBMP-2/CHA, which can provide a new idea for treating bone defect by using bone tissue engineering in the future.
Objective Choose polylactide-co-glycolide/hydroxyapatite (PLGA/HA) and porous phosphate calcium (PPC) as the object that we will study, compare their degradabality and choose one as a suitable scaffold for rib reconstruction. Methods All the experiments were divided into PLGA/HA group and CPC group. Degradabality experiment in exvivo: put the two scaffold which have the same size into 0.9% NaCl, keep sterile, then put the container into warm cage，get out and weigh them in 2, 4, 8, 12 and 24 weeks, compare the different speed of the two scaffold. Degradability experiment in vivo: put the two scaffold which have the same size under the skin of the rabbit, and weigh them in 2, 4, 8, 12 and 24 weeks, the tissue around the scaffold was examinzed by HE and the scaffold was examined by electron scanning microscope. Results Micro-CT and Scanning electron microscopy shows that CPC group had better structure （1101.2228±0.6184 mg/ccm vs. 1072.5523±0.7442 mg/ccm）and porosity（70.26％±0.45％ vs.72.82％±0.51％）than PLGA/HA group; The result of degradabality experiment in vitro shows that the speed of the two scaffolds was slow. It is at 24 weeks that the degradability is obvious,and the PLGA/HA group degraded a lot which was 60%. The result of degradabality experiment in vivo shows that the speed of degradabality of PLGA/HA group was faster than that is in the 0.9% Nacl, also was faster than that of CPC group which was 96%.The reponse of tissue around the PLGA/HA was more sever than that of CPC group which is in favour of the growth of cells. Conclusion As for the reconstruction of large defect of rib, CPC is more suitable than PLGA/HA.
Objective To assess the mid-term effectiveness of anterior decompression and fusion with nano-hydroxyapatite/polyamide 66 (n-HA/PA66) cage in treatment of cervical spondylotic myelopathy. Methods A retrospective study was made on 48 patients with cervical spondylotic myelopathy who underwent anterior decompression and fusion with n-HA/PA66 cage between August 2008 and January 2010. There were 33 males and 15 females with an average age of 54.5 years (range, 42-72 years). The disease duration was 3-12 months (mean, 6 months). The affected segments included 35 cases of single segment (C3, 4 in 7, C4, 5 in 18, and C5, 6 in 10) and 13 cases of double segments (C3-5 in 7 and C4-6 in 6). Of 48 patients, 28 was diagnosed as having intervertebral disc protrusion, 12 as having ossification of posterior longitudinal ligament, and 8 as having vertebral osteophyte; 35 patients underwent single segmental anterior corpectomy and fusion, and 13 patients underwent single segmental anterior discectomy and fusion. The pre- and post-operative radiographs (cervical anteroposterior and lateral X-ray films and three-dimensional CT scans) were taken to measure the segmental height and lordosis angle. Brantigan et al assessment standard and visual analogue scale (VAS) and Japanese Orthopaedic Association (JOA) scores were used to evaluate the graft fusion and the improvement of clinical symptoms, respectively. Results All patients were followed up for 46 months on average (range, 36-54 months). No cage breaking, displacement, or sliding was found. At last follow-up, 36 cases were rated as Brantigan grade E, 10 cases as grade D, and 2 cases as grade C; the fusion rate was 96%. Both segmental height and lordosis angle were corrected significantly at immediate and 6 months after operation and last follow-up than those before operation (P lt; 0.05), but no significant difference was found among different time points after operation (P gt; 0.05). At last follow-up, the cage subsidence was (1.3 ± 1.0) mm. The VAS and JOA scores at 6 months after operation and last follow-up were significantly improved when compared with preoperative scores (P lt; 0.05), and the scores at last follow-up were superior to ones at 6 months after operation (P lt; 0.05). Conclusion The mid-term effectiveness of anterior decompression and fusion with the n-HA/PA66 cage in patients with cervical spondylotic myelopathy is satisfactory because it can effectively restore and maintain segmental height and lordosis angle and promote osseous fusion.
Objective To prepare silver-containing hydroxyapatite coating (hydroxyapatite/Ag, HA/Ag) and investigate its antibacterial property and biocompatibil ity in vitro. Methods Vacuum plasma spraying technique was adopted to prepare HA/Ag coating on titanium alloy substrate (3% Ag). After incubating the HA/Ag and the HA coating under staphylococcus aureus and pseudomonas aeruginosa suspensions of 2% tryptic soy broth (TBS) medium for 2, 4 and 7 days, respectively, the biofilm on the coatings was examined by confocal laser scanning microscope, and the bacterial density and viable bacterial percentage of bacterial biofilm were calculated. Meanwhile, the micro-morphology of bacterial biofilm was observed by SEM, the cytotoxicity was detected via MTT and the biocompatibil ity of biofilm was evaluated by acute aemolysis test. Results Compared with HA coating, the bacterial biofilm’s thickness on the surface of HA/Ag coating witnessed no significant difference at 2 days after culture (Pgt; 0.05), but decreased obviously at 4 and 7 days after culture (P lt; 0.01). The bacterial density of the biofilm increased with time, but there was no significant difference between two coatings (P gt; 0.05) at 2, 4 and 7 days after culture. The viable bacterial percentage of the biofilms on the surface of HA/Ag coating decreased obviously compared with that of HA coating at 2, 4 and 7 days after cultureP lt; 0.01). The MTT notified the cytotoxic grade of both coatings was zero. The acute haemolysis assay showed that the hemolytic rate of HA/Ag and HA coating was 0.19% and 0.12%, respectively. Conclusion With good biocompatibil ity, significant antibacterial property against staphylococcus aureus and pseudomonas aeruginosa, no obvious cytotoxicity and no erythrocyte destruction, the vacuum plasma sprayed HA/Ag coating is a promising candidate for the surface of orthopedic metal implants to improve their osseointegration and antibacterial property.
【Abstract】 Objective To investigate the anti-infection and bone repair effects of cationic l i posome-encapsulatedvancomycin combined with the nano-hydroxyapatite/chitosan/konjac glucomannan (n-HA/CS/KGM) composite scaffold invivo. Methods Fifty-one 6-month-old New Zealand white rabbits, weighing 1.5-3.0 kg, were selected to prepare chronicinfectious tibia bone defect model by using Staphylococcus aureus. After 4 weeks, 48 survival rabbits were randomly divided into 4 groups (n=12). After debridement, defect was treated with nothing in group A, with n-HA/CS/KGM composite scaffold in group B, with vancomycin and n-HA/CS/KGM composite scaffold in group C, and with cationic l i posome-encapsulated vancomycin and n-HA/CS/KGM composite scaffold in group D. After 8 weeks of treatment, general observation, X-ray, HE staining, the bacterial culture, and the measurement of the longest diameter of bone defect were done. Results At 4 weeks after modeling, 48 rabbits were diagnosed as having osteomyelitis, including periosteal new bone formation, destruction of bone, and soft tissue swell ing. The Norden score was 3.83 ± 0.52. At 8 weeks after treatment, sinus healed in groups C and D, but sinus was observed in groups A and B; the gross bone pathologieal scores of group D were significantly better than those of groups A and B (P lt; 0.05). Bone defects were repaired completely in group D, the results of the longest diameter of bone defects in group D was significantly better than those in the other 3 groups (P lt; 0.05). New bone formation was observed in groups C and D, but periosteal reactionand marrow low-density shadow were observed in groups A and B; Norden score in group D was significantly better than those in groups A, B, and C (P lt; 0.05). HE staining showed that there were a large number of trabecular bone formation and fibrosis, with no obvious signs of infection in groups C and D, but neutrophil accumulation was observed in groups A and B; Smeltzer scores in groups C and D were significantly better than those in groups A and B (P lt; 0.05). Bacteriological results showed higher negative rate in groups C and D than in groups A and B (P lt; 0.05). Conclusion Cationic l iposome-encapsulated vancomycin and n-HA/CS/KGM composite scaffold can be a good treatment for infectious bone defects in rabbits, providing a new strategy for the therapy of bone defects in chronic infection.
Objective To develop the plastic nano-hydroxyapatite (nano-HA)/poly (3-hydroxybutyrate-hydroxyvalerate) polyethylene glycol(PHBV-PEG) gentamicin (GM) drug delivery system(DDS)(nano-HA/PHBV-PEG-GM-DDS) for treating osteomyelitis and find its releasing character in vivo. Methods The plastic nano-HA/PHBV- PEG-GM-DDS was prepared using nanoHAas the core carrier of GM, nano-HA with PHBV and PEG as coating and plastic fibrin glue(FG) as microsphere scaffold. The morphological features of nano-HA,drug loaded nano-HA and drug loaded nano-HA/PHBVPEG microsphere were examined by electron microscope.The GM concentration in blood, cortex bone and cancellousbone was detected at 12 different time points by the method of K-B after the plastic nano-HA/PHBV-PEGGM-DDS was implanted into the femora of 36 rabbits. Its GM releasing character was assayed in vivo. Results Nano-HA was similar to a blackjack, and its length was less than 60 nm. Drug loaded nano-HA appeared natural crystal condensate, of which surface adsorbed massive GM. The average grain diameter was 200.5 nm. Drug loaded nanoHA/PHBV-PEG microsphere had a shrinkable porous structure, of which surface configuration was consistent. The average grain diameter was 34.5 μm. The GM concentration and the antibacterial annulus was in the linear correlation. The correlation coefficient was 0.998. In cortex and cancellous bone tissue, the GM concentration was about 95.50±16.50 μg/ml and 80.20±13.80 μg/ml from the plastic nano-HA/PHBV-PEG-GM-DDS on the 1st day, then decreased gradually. After 56 days of operation, the GM concentration still exceeded the minimum inhibitory concentrationfor the staphylococcus aureus, but the peak level of serum GM concentration wasunder the nephrotoxicity concentration. Conclusion Plastic nano-HA/PHBV-PEG-GM-DDS was a good drug delivery system with sustained antibiotic effect in vivo. It was an effective method for the treatment of osteomyelitis.
Objective To retrospectively analyze the cl inical effect of l ightbulb operation with nano-hydroxyapatite/ collagen in a consecutive series of patients with osteonecrosis of the femoral head (ONFH). Methods From January 2001to July 2005, 26 patients (35 hips) were treated, 16 males and 10 females, aged 19-54 years old (33.5 on average). The course of disease was 12-36 months (18 months on average). Based on the etiology, 15 cases (22 hips) were steroid induced type, 10 (12 hips) were alcohol induced type and the other one (1 hip ) was idiopathic type. According to the system of Association Research Circulation Osseous (ARCO), there were 6 hi ps of stage IIB, 16 hi ps of stage IIC, 9 hi ps of stage IIIA, 3 hi ps of stage IIIB and 1 hip of stage IIIC. The Harris score was 62.2 ± 7.5. All the patients who had undergone l ightbulb operation with nano-hydroxyapatite/collagen were evaluated both cl inically and radiographically. The bone graft mixture rate of nanohydroxyapatite/ collagen and autogenous bone was 1 ∶ 1, and the mixed bone graft was 6 times of the scraped osteonecrosis volume (30-48 mL). Results The incisions of all 26 patients (35 hi ps) obtained heal ing by first intention. The 2 cases, which got lateral femoral cutaneous nerve injury during the operation, recovered 3-6 months after the operation without any treatment. Another 2 cases got heterotopic ossification 3 months after operation, with no special treatment. All the 26 patients (35 hips) were followed up for 2-7 years (3.5 on average). The patients’ bone heal ing began from the 3rd month after operation. The postoperative Harris score was 85.1 ± 16.2, and there was significant difference compared with the preoperative one (P lt; 0.001). There were 15 hips of excellent, 11 of good, 5 of fair, and 4 of poor which received total hip arthroplasty at the end of the follow-up. According to imaging, 5 hips were progressed from preoperative IIC to IIIA, while the other hips were radiologically stable, with no progress of ONFH. Conclusion Lightbulb operation with nano-hydroxyapatite/collagen provides a surgical treatment to treat early ONFH with satisfactory cl inical outcomes. Nano-hydroxyapatite/collagen is beneficial for the repair and reconstruction of ONFH and suitable for femoral-head-preserving operation for the patients with ONFH of stage II.