Ｔｏ ｉｎｖｅｓｔｉｇａｔｅ ｔｈｅ ｃｈａｒａｃｔｅｒｉｓｔｉｃ ｏｆ ａｎｔｉｂｏｄｉｅｓ ｔｈａｔ ｉｎｄｕｃｅ ｔｈｅ ｒｅｊｅｃｔｉｏｎ ｏｆ ｘｅｎｏｇｒａｆｔｉｎｇ，ＳＤ ｒａｔｓ ｗｅｒｅ ｉｍｍｕｎｉｚｅｄ ｗｉｔｈ ＮＩＨ ｍｉｃｅ ｓｐｌｅｅｎ ｔｏ ｐｒｅｐａｒｅ ｒａｔ ａｎｔｉｍｏｕｓｅ ａｎｔｉｓｅｒａｓ． Ｔｈｅ ａｎｔｉｂｏｄｉｅｓ ｗｅｒｅ ｐｕｒｉｆｉｅｄ ｂｙ ｃａｐｒｙｌｉｃ ａｃｉｄ ｐｒｅｃｉｐｉｔａｔｉｏｎ ａｎｄ ｈｙｄｒｏｘｙ ｌａｐａｔｉｔｅ（ＨＰＴ）ｃｈｒｏｍａｔｏｇｒａｐｈｙ．Ｔｈｅ ｒｅｓｕｌｔｓ ｓｈｏｗｅｄ ｔｈａｔ ｔｈｅ ｔｉｔｌｅ ｏｆ ｒａｔ ａｎｔｉｍｏｕｓｅ ａｎｔｉｓｅｒａ ｗａｓ １∶６４０，ａｎｄ ｔｈｅ ａｎｔｉｂｏｄｉｅｓ ｐｕｒｉｆｉｅｄ ｂｙ ｃａｐｒｙｌｉｃ ａｃｉｄＨＰＴ ｈａｄ ｒｅｌｉａｂｌｅ ｐｕｒｉｔｙ ａｎｄ ｇｏｏｄ ｉｍｍｕｎｏｃｏｍｐｅｔｅｎｃｅ．Ｔｈｉｓ ｍｅｔｈｏｄ ｃｏｕｌｅ ｂｅ ｕｓｅｄ ｉｎ ｐｒｅｐａｒａｔｉｏｎ ａｎｄ ｒｅｓｅａｒｃｈ ｏｆ ｍａｊｏｒ ｈｉｓｔｏｃｏｍｐａｔｉｂｉｌｉｔｙ ａｎｔｉｂｏｄｉｅｓ．
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 evaluate the internal fixation effect, degradation, and biocompatibility of polylactic-co-glycolic acid/hydroxyapatite (PLGA/HA) absorbable cannulated screws in treatment of lateral femoral condyle fracture of canine so as to provide the theory basis for their further improvement and clinical application. Methods Sixteen adult male Beagles (weighing, 9-12 kg) were selected to prepare the models of bilateral lateral femoral condyle fracture; left fracture was fixed with PLGA/HA absorbable cannulated screws as experimental group and right fracture with metal screws as control group. At 2, 4, 8, and 12 weeks after operation, general observation was done and X-ray films were taken for observing fracture healing; bone mineral density was measured; the histological examination was performed; and the degradation property of absorbable cannulated screws was detected. Results All animals survived to the end of the experiment. General observations showed that no fracture displacement occurred and fracture healed at 12 weeks in 2 groups; no breakage, displacement, or loosening of screws was observed in experimental group. X-ray films results showed that the absorbable cannulated screws could not be found out by X-ray in experimental group, but metal screws could be found out in control group; fracture healed with time in 2 groups. The bone mineral density reached the peak at 8 weeks in 2 groups, and no significant difference was found between 2 groups and among different time points in the same group (P gt; 0.05). Histological examination showed that 2 groups had similar fracture healing process at different time points; no obvious inflammatory reaction was found around absorbable cannulated screws in experimental group. The degradation results of absorbable cannulated screws showed that the intrinsic viscosity and molecular weight distribution obviously decreased at 2 weeks; the number average molecular weight and the weight average molecular weight markedly decreased at 4 weeks; and the maximum shear force did not decrease obviously at 8 weeks, and then decreased significantly. Significant differences were found in all indexes among different time points in the same group (P lt; 0.05). Conclusion PLGA/HA absorbable cannulated screws and metal screws show similar fracture healing process for fixing lateral femoral condyle fracture of canine, and the absorbable canulated screws have good biocompatibility. The maximum shear force of PLGA/HA absorbable cannulated screw has no obvious decrease during 8 weeks after operation, so it can ensure full healing of fracture.
Objective To observe the effect of cationic liposomal ceftazidime (CLC) combined with nano-hydroxyapatite/β-tricalcium phosphate (n-HA/β-TCP) in the treatment of chronic osteomyelitis of rabbits. Methods Thirty healthy New Zealand white rabbits (4-6 months old; weighing, 2-3 kg) were selected to prepare the chronic osteomyelitis models. After 4 weeks, the gross observation, X-ray examination, and bacteriological and histopathological examinations were done; the models were made successfully in 27 rabbits. Of 27 rabbits, 24 were randomly divided into 4 groups (n=6): only debridement was performed in group A; ceftazidime was given (90 mg/kg), twice a day for 8 weeks after debridement in group B; ceftazidime and n-HA/β-TC were implanted after debridement in group C; and CLC and n-HA/β-TCP were implanted after debridement in group D. Before and after treatments, X-ray examination was done, and Norden score was recorded. At 8 weeks after treatment, the specimens were harvested for gross observation and for gross bone pathological score (GBPS) using Rissing standard; half of the specimens was used for histological observation and Smeltzer scoring, the other half for bacteriological examination and calculation of the positive rate of bacteria culture. Results At 8 weeks after treatment, Norden score of group D was significantly lower than that of groups A, B, and C (P lt; 0.05), but no significant difference was found among groups A, B, and C (P gt; 0.05). At 8 weeks after treatment, sinus healed in groups C and D, but sinus was observed in groups A and B; the GBPS scores of groups C and D were significantly lower than those of groups A and B (P lt; 0.05). The Smeltzer scores of groups C and D were significantly lower than those of groups A and B (P lt; 0.05). The positive rates of bacteria culture of groups C (0) and D (0) were significantly lower than those of group A (25.0%) and group B (16.7%) (P lt; 0.05). Conclusion CLC combined with n-HA/β-TCP has good effect in treating chronic osteomyelitis of rabbits, and it has better effect in treating chronic osteomyelitis of rabbits than ceftazidime with n-HA/β-TCP.
Objective To evaluate the application of artificial lamina of multi-amino-acid copolymer (MAACP)/nano-hydroxyapatite (n-HA) in prevention of epidural adhesion and compression of scar tissue after posterior cervical laminectomy. Methods Fifteen 2-year-old male goats [weighing, (30 ± 2) kg] were randomly divided into experimental group (n=9) and control group (n=6). In the experimental group, C4 laminectomy was performed, followed by MAACP/n-HA artificial lamina implantations; in the control group, only C4 laminectomy was performed. At 4, 12, and 24 weeks after operation, 2, 2, and 5 goats in the experimental group and 2, 2, and 2 goats in the control group were selected for observation of wound infection, artificial laminar fragmentation and displacement, and its shape; Rydell’s degree of adhesion criteria was used to evaluate the adhesion degree between 2 groups. X-ray and CT images were observed; at 24 weeks after operation, CT scan was used to measure the spinal canal area and the sagittal diameter of C3, C4, and C5 vertebrea, 2 normal goats served as normal group; and MRI was used to assess adhesion and compression of scar tissue on the dura and the nerve root. Then goats were sacrificed and histological observation was carried out. Results After operation, the wound healed well; no toxicity or elimination reaction was observed. According to Rydell’s degree of adhesion criteria, adhesion in the experimental group was significantly slighter than that in the control group (Z= — 2.52, P=0.00). X-ray and CT scan showed that no dislocation of artificial lamina occurred, new cervical bone formed in the defect, and bony spinal canal was rebuilt in the experimental group. Defects of C4 vertebral plate and spinous process were observed in the control group. At 24 weeks, the spinal canal area and sagittal diameter of C4 in the experimental group and normal group were significantly larger than those in the control group (P lt; 0.05), but no significant difference was found between experimental group and normal group (P gt; 0.05). MRI showed cerebrospinal fluid signal was unobstructed and no soft tissue projected into the spinal canal in the experimental group; scar tissue projected into the spinal canal and the dura were compressed by scar tissue in the control group. HE staining and Masson trichrome staining showed that artificial lamina had no obvious degradation with high integrity, some new bone formed at interface between the artificial material and bone in the experimental group; fibrous tissue grew into defect in the control group. Conclusion The MAACP/n-HA artificial lamina could maintaine good biomechanical properties for a long time in vivo and could effectively prevent the epidural scar from growing in the lamina defect area.
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.
【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 evaluate the short-term effectiveness of nano-hydroxyapatite/polyamide-66 (n-HA/PA66) intervertebral cage for lumbar interbody fusion in the patients with lower lumbar degenerative diseases. Methods Between January and October 2011, 20 patients with lower lumbar degenerative diseases underwent transforaminal lumbar interbody fusion with n-HA/PA66 intervertebral cage. There were 8 males and 12 females, aged 22-80 years (mean, 51 years). The disease duration was 1 to 24 months (mean, 4 months). L4, 5 fusion was performed in 8 cases, L5, S1 fusion in 9 cases, and L4-S1 fusion in 3 cases. Among 20 cases, 3 were diagnosed as having recurrent lumbar disc protrusion, 5 as having lumbar degenerative spondylolisthesis, 9 as having lumbar isthmic spondylolisthesis, and 3 as having lumbar spinal stenosis. The intervertebral height and lordosis were measured on X-ray film to assess the surgical correction and postoperative sustain while osseous fusion was observed on 3-dimensional CT. The Oswestry disability index (ODI) and short-form 36 health survey scale (SF-36) scores were obtained to assess the status of clinical recovery. Results All patients had incision healing by first intention. The pain and numb were relieved in varying degrees after operation. No cerebrospinal leakage, nerve root injury, or wound infection was occurred. All patients were followed up 6-9 months (mean, 7 months). No cage displacement or collapse was found. The intervertebral height and lordosis of single fusion segment were significantly improved at 3 days and 3, 6 months after operation when compared with those at preoperation (P lt; 0.01); there was no significant difference among each time point after operation (P gt; 0.05). The fusion rate was 74% at 3 months after operation and 96% at 6 months after operation, with an average of 4 months (range, 3-9 months) for interbody fusion. The ODI and SF-36 scores were significantly improved at 3 days and 6 months after operation when compared with the scores at preoperation (P lt; 0.01); there was no significant difference among each time point after operation (P gt; 0.05). Conclusion The interbody fusion with n-HA/PA intervertebral cage is effective and safe to treat the lower lumbar degenerative diseases. The n-HA/PA66 intervertebral cage is an ideal device of interbody fusion with high fusion rate, low subsidence rate, and high transmission X-ray, but the long-term effectiveness need further observation.
Objective To prepare collagen-chitosan /nano-hydroxyapatite-collagen-polylactic acid (Col-CS/ nHAC-PLA) biomimetic scaffold and to examine its biocompatibility so as to lay the foundation for its application on the treatment of osteochondral defect. Methods PLA was dissolved in dioxane for getting final concentration of 8%, and the nHAC power was added at a weight ratio of nHAC to PLA, 1 ∶ 1. The solution was poured into a mold and frozen. CS and Col were dissolved in 2% acetum for getting the final concentrations of 2% and 1% respectively, then compounded at a weight ratio of CS to Col, 20 ∶ 1. The solution was poured into the frozen mold containing nHAC-PLA, and then biomimetic osteochondral scaffold of Col-CS/nHAC-PLA was prepared by freeze-drying. Acute systemic toxicity test, intracutaneous stimulation test, pyrogen test, hemolysis test, cytotoxicity test, and bone implant test were performed to evaluate its biocompatibility. Results Col-CS/nHAC-PLA had no acute systemic toxicity. Primary irritation index was 0, indicating that Col-CS/nHAC-PLA had very slight skin irritation. In pyrogen test, the increasing temperature of each rabbit was less than 0.6℃, and the increasing temperature sum of 3 rabbits was less than 1.3℃, which was consistent with the evaluation criteria. Hemolytic rate of Col-CS/nHAC-PLA was 1.38% (far less than 5%). The toxicity grade of Col-CS/nHAC-PLA was classified as grade I. Bone implant test showed that Col-CS/nHAC-PLA had good biocompatibility with the surrounding tissue. Conclusion Col-CS/ nHAC-PLA scaffold has good biocompatibility, which can be used as an alternative osteochondral scaffold.
Objective To investigate the antibacterial and osteogenic capabil ities in vivo of hydroxyapatite (HA)/silver (Ag) coating. Methods HA/Ag coating (Ag qual ity percentage was 3%) and HA coating were deposited to external fixator Schanz screws. The tibial fracture model was establ ished in right hindl imb of 18 adult male Beagle dogs (weighing 15-20 kg). Thetibia was stabil ized with an external fixator and 2 Schanz screws of HA coating at proximal tibia (control group, n=18) and HA/Ag coating at distal tibia (experimental group, n=18), and every screw incision was infected with Staphylococcus aureus. Infection in screw holes and the changes of bone-screw interface were observed by wound grading and X-ray films. Results In control group, wounds infection became worse with time (χ2=13.492, P=0.001), while in experimental group, no obvious change was observed (χ2=0.208, P=0.901). The wound grading of experimental group was significantly better than that of the control group at 1, 2, and 3 weeks (P lt; 0.05). Laser scanning confocal microscope showed that there was bacterial adhesion on the surface of screws in 2 groups, viable becteria mainly in control group and non-viable becteria mainly in experimental group. The scanning electron microscope (SEM) observation results of the fractured sclerous tissue section showed that an obvious transparent boundary between screw and bone in control group, but no obvious boundary in experimental group. The osseointegration ratios were 76.23% ± 15.54% in control group and 93.42% ± 5.53% in experimental group, showing significant difference (t=8.843, P=0.000). The SEM observation showed that HA/Ag coating integrated with new bone and the surface of implant was filled with new bone in experimental group; obvious interspace was seen between the HA coating and new bone in control group. Conclusion HA/Ag coating has good antibacterial and osteogenic capabil ities, so it can take effects in preventing infection in screw holes and loosening of implants.