Objective To study the mechanism of restenosis of the vein graft and the effect of the grafting injury to the vein graft. Methods One side of the 36 healthy rabbits was randomly chosen as the V-A group, and on the side a 1.5cmlong femoral vein was obtained, and an 0.5-cm-long segment of the obtained femoral vein was separated as the control group. The remaining 1-cm-long femoral vein was inverted and was autogenously implanted into the femoral artery on the same side of the rabbit. The other side of the rabbits was chosen as the V-V group, and on this side a 1-cm-long femoral vein was obtained ex vivo and then was sutured in situ. The vein grafts on both sides were harvested 4 weeks after operation. The specimens from the harvested vein grafts were stained with HE and theelastic fiber Victoria blue for an observation on the histological changes in the walls of the vein grafts, and the specimens were also stained by the immunohistochemistry of the proliferating cell nuclear antigen (PCNA) for an observation on the wall cell proliferation of the vein grafts. The changes in the ultrastructure of the proliferated wall cells of the vein grafts were observed under electron microscope. The two sides of the rabbits were compared. Results The smooth muscle cells of the media developed hyperplasia, but theintima and the media remained unchanged in their thickness (3.50±0.41 μm, 12.23±1.59 μm) in the V-V group, with no difference when compared with the control group (3.40±0.37 μm, 12.14±1.62 μm); however, when compared with the V-A group (25.60±3.21 μm, 21.30±2.47 μm),there was a significant difference in the thickness (Plt;0.01). There were no cells positive for PCNA by the immunohistochemistry examination in the control group. The cells positive for PCNA were found in the intima and the media in both the V-V group and the V-A group; however, the percentageof the cells positive for PCNA in the intima and the media was significantly greater in the V-A group than in the V-V group (16.4%±1.9% and 36.5%±3.7% vs 5.9%±1.3% and 23.4%±3.4%, Plt;0.01). In the V-V group, the endothelial cell could be observed under transmis-sion electron microscope, which was flat and had a processlike villus at its free end, and the endothelial cells were closely arranged andhad hyperplasia of the smooth muscle cells in the media. But in the V-A group,the endothelial cells had an obvious hyperplasia with an irregular shape and a widened space between the cells, and in the intima a great amount of the smooth muscle cells could be observed, which had a broken basement membrane. The smooth muscle cells also had an obvious hyperplasia in the media. The shape and alignment of the endothelial cells in the control group were similar to those in the V-V group, but the hyperplasia of the smooth muscle cells was not observed in the media. Conclusion The grafting injury can cause hyperplasia ofthe vascular wall cells, and if the hemodynamics is changed simultaneously, more serious hyperplasia and cell migration can be observed from the media to the intima, resultingin restenosis of the blood vessels. So, if we can reduce the grafting injury and improve the microcirculation of the vein graft, we may find out the methods ofpreventing restenosis of the vein graft. The animal model of the V-V graftcan help to understand the mechanism of restenosis of the vein graft.
Objective To evaluate the characterization, biocompatibil ity in vitro and in vivo, and antimicrobial activity of an injectable vancomycin-loaded borate glass/chitosan composite (VBC) so as to lay the foundation for its further cl inical application. Methods The sol id phase of VBC was constituted by borate glass and vancomycin, liquid phase was a mixture of chitosan, citric acid, and glucose with the proportion of 1 ∶ 10 ∶ 20. Solid phase and liquid phase was mixed withthe ratio of 2 ∶ 1. Vancomycin-loaded calcium sulfate (VCS) was produced by the same method using calcium sulfate instead of borate glass and sal ine instead of chitosan, as control. High performance liquid chromatography was applied to detect the release rate of antibiotics from VBC and VCS, and minimum inhibitory concentration (MIC) was tested by using an antibiotic tube dilution method. The structure of the VBC and VCS specimens before and 2, 4, 8, 16, and 40 days after immersion in D-Hank’s was examined by scanning electron microscopy, and the phase composition of VBC was analysed by X-ray diffraction after soaked for 40 days. Thirty-three healthy adult New Zealand white rabbits (weighing, 2.25-3.10 kg; male or female) were used to establ ish the osteomyel itis models according to Norden method. After 4 weeks, the models of osteomyel itis were successfully established in 28 rabbits, and they were randomly divided into 4 groups (groups A, B, C, and D). In group A (n=8), simple debridement was performed; in groups B and C (n=8), defect was treated by injecting VCS or VBC after debridement; and in group D (n=4), no treatment was given. The effectiveness of treatment was assessed using radiological and histological techniques after 2 months. Results The releases of vancomycin from VBC lasted for 30 days; the release rate of vancomycin reached 75% at the first 8 days, then could reached more than 90%. The releases of vancomycin from VCS lasted only for 16 days. The MIC of VBC and VCS were both 2 μg/mL. The VCS had a smooth glass crystal surface before immersion, however, it was almost degradated after 4 days. The fairly smooth surface of the VBC pellet became more porous and rougher with time, X-ray diffraction analysis of VBC soaked for 40 days indicated that the borate glass had gradually converted to hydroxyapatite. After 2 months, the best result of treatment was observed in group C, osteomyelitis symptoms disappeared. The X-ray scores of groups A, B, C, and D were 3.50 ± 0.63, 2.29 ± 0.39, 2.00 ± 0.41, and 4.25 ± 0.64, respectively; Smeltzer scores were 6.00 ± 0.89, 4.00 ± 0.82, 3.57 ± 0.98, and 7.25 ± 0.50, respectively. The scores were significantly higher in group D than in groups A, B, and C (P lt; 0.05), and in group A than in groups B and C (P lt; 0.05). The scores were higher in group B than in group C, but no significant difference was found (P gt; 0.05). Conclusion The VBC is effective in treating chronic osteomyelitis of rabbit by providing a sustained release of vancomycin, in addition to stimulating bone regeneration, so it may be a promising biomaterial for treating chronic osteomyelitis.
OBJECTIVE: To establish the animal models of mandibular distraction osteogenesis in rabbits and study its osteogenetic mechanism. METHODS: The right mandibles just anterior to the first molars of 12 rabbits were performed osteotomies, and the mandibles were positioned with distractors. The left mandibles were control group without operation. After 1 week, the distractors were stretched 0.9 mm every day for 10 days progressively. One day, 2, 4, 8 weeks after distraction, the mandibles were studied with gross measurement, X-ray, and histological examination. RESULTS: The right mandible were lengthened 8.3 mm on average without bone nonunion and deformity healing. It was observed that the gaps between the distracted bone edges were first occupied by fibrous tissue. Two weeks after distraction, it was found that the gaps were bridged by callus in X-ray, the new bone and the normal bone could not be differentiated clearly after 8 weeks. In histological sections, there were collagen bundles in early distraction, then those collagen bundles were calcificated and become trabeculaes. No Cartilage was found during distraction. CONCLUSION: It suggests that the rabbit mandible can be lengthened by distraction osteogenesis, and the new bone is formed by intramembranous ossification.
Objective To compare the difference of preparing the acellular larynx scaffold between perfusion method and immersion method, and find better way to make acellular larynx scaffold for tissue engineering. Methods Twenty 6-month-old male New Zealand rabbits, weighing 2.0-2.5 kg, were divided into perfusion group (n=10) and immersion group (n=10) at random. All the larynxes were excised in a sterile fashion. The acellular larynx scaffold was obtained by perfusionmethod and immersion method respectively, and then comparative examinations were performed by the macroscopicview, histological view, scanning electron microscope (SEM), cartilage vital ity assay and toluidine blue staining. ResultsMacroscopic view showed that the larynxes perfused by sodium dodecyl sulphate (SDS) became transparent after 2 hoursof perfusion, but the larynxes immersed by SDS over 16 hours still appeared pink-white. Histology and SEM indicated thatcompared with immersion group, perfusion group showed better acellular effect, more ventages and collagen fibers wereretained, no intact cell or nuclei remained in acellular matrix and chondrocytes were still survival. The porosity was 85.39% ± 3.16% in perfusion group and 34.72% ± 4.51% in immersion group, showing significant difference (P lt; 0.01). The chondrocyte vital ity rate of perfusion group (86.93% ± 1.52%) was higher than that of immersion group (77.73% ± 1.66%), showing significant difference (P lt; 0.01). Toluidine blue staining showed that the chondrocyte heterochromaty was ber in perfusion group than that in immersion group. Conclusion Compared with immersion method, perfusion method is a better way to construct acellular larynx scaffold because it can achieve better acellular effect and retain chondrocyte vital ity at the greatest extent in the acellular larynx scaffold.
Objective To explore the histological changes of bio-derived bone prepared by different methods after implantation, and to provide the scaffold material from xenogeneic animal for tissue engineering. Methods Theextremities of porcine femur were cut into 0.5 cm×0.5 cm×0.5 cm. Then they were divided into 5 groups according to different preparation methods: group A was fresh bone just repeatedly rinsed by saline; group B was degreased; group C was degreased and decalcificated; group D was degreased, acellular and decalcificated; group E wasdegreased and acellular. All the materials were implantated into femoral muscle pouch of rabbit after 25 kGy irradiation sterilization. The cell counting ofinflammatory cells and osteoclasts, HE and Masson staining, material degradation, collagen and new bone formation were observed at 2, 6, and 12 weeks postoperatively. Results The residue level of trace element in biomaterials prepared by different methods is in line with the standards. All the animals survived well. There were no tissue necrosis, fluid accumulation or inflammation at all implantation sites at each time point. The inflammatory cells counting was most in group A, and there was significant difference compared with other groups(P＜0.05). There was no significant difference in osteoclasts counting among all groups. For the index of HE and Masson staining, collagen and new bone formation, groups C and D were best, group E was better, and groups A and B were worse. Conclusion The degreased, acellular and decalcificated porcine bone is better in degradation,bone formation, and lower inflammatory reaction, it can be used better scaffold material for tissue engineered bone.
Objective To explore the possibilityof constructing tissue engineering muscles by combining allogeneic myoblasts with small instestinal submucosa(SIS) in rabbits.Methods A large number of purified myoblasts were obtained with multiprocedure digestion and repeated attachment method from skeletal muscles taken from extremities of immature rabbits which were born 7 days ago. The myoblasts were labeled with BrdU, and then combined with SIS to construct tissue engineering muscles. This kind of tissue engineering muscles were grafted into the gastrocnemius muscle defect (1.5 cm in length, 1.0 cmin width) of fifteen rabbits as the experimental group. The SIS was grafted into the same position in the control group. The rabbits were sacrificed 4, 6, 8 weeks after operation. The tissue engineering muscles were evaluated by macroscopic、histological and immunohistochemical observations, and by quantitative analysis of local immunocyte in the grafting site. Results Allogeneic myoblasts with SIS were combined perfectly in vitro. The SIS was connected tightly to surrounding skeletal muscles and inflammation response was obvious 4 weeks after grafting.The SIS began to break down and inflammation response became slight 6 and 8 weeks after operation. Compared with that of 8th week, the quantitative analysis oflocal immunocyte in 4th and 6th week in both experimental and control group hassignificance(Plt;0.05). Newly formed muscle tissues were found around SIS in the experimental group in 4th, 6th, and 8th week. Expression of BrdU and myosin immunohistochemical staining were positive in the experimental group and negative inthe control group.Conclusion Tissue engineering muscles of rabbits which are constructed by combining allogeneic myoblasts with SIS can survive and proliferate.
ObjectiveTo investigate the effects of different concentrations of osteoprotegerin (OPG) combined with deproteinized bone (DPB) on the bone tunnel after the anterior cruciate ligament (ACL) reconstruction. MethodsThe femoral epiphyseal side was harvested from newborn calf, and allogenic DPB were prepared by hydrogen peroxide-chloroform/methanol method. Then, DPB were immersed in 3 concentrations levels of OPG (30, 60, 100 μg/mL) and 3 concentration ratios (30%, 60%, 100%) of the gel complex were prepared. Sixty healthy New Zealand white rabbits, male or female, weighing (2.7±0.4) kg, were divided randomly into 4 groups (n=15):control group (group A), 30% (group B), 60% (group C), and 100% (group D) OPG/DPB gel complex. The ACL reconstruction models were established by autologous Achilles tendon. Different ratios of OPG/DPB gel complex were implanted in the femoral and tibial bone tunnel of groups B, C, and D, but group A was not treated. The pathology observation (including the percentage of the femoral bone tunnel enlargement) and histological observation were performed and the biomechanical properties were measured at 4, 8, and 12 weeks after operation. ResultsOne rabbit died of infection in groups A and D, 2 rabbits in groups B and C respectively, and were added. General pathology observation showed that the internal orifices of the femoral and tibia tunnels were covered by a little of scar tissue at 4 weeks in all groups. At 8 weeks, white chondroid tissues were observed around the internal orifices of the femoral and tibia tunnels, especially in groups C and D. At 12 weeks, the internal orifices of the femoral and tibia tunnels enlarged in groups A, B, and C, but it was completely closed in group D. At each time point, the rates of the femoral bone tunnel enlargement in groups B, C, and D were significantly lower than that in group A, and group D was significantly lower than groups B and C (P<0.05); group C was significantly lower than group B at 8 weeks, but no significant difference was found at 4 and 12 weeks (P<0.05). Hisological observation showed that fresh fibrous connective tissue was observed in 4 groups at 4 weeks; there was various arrangements of Sharpey fiber in all groups at 8 weeks and the atypical 4-layer structure of bone was seen in group D; at 12 weeks, Sharpey fiber arranged regularly in all groups, with typical 4-layer structure of bone in groups B, C, and D, and an irregular "tidal line" formed, especially in group D. Biomechanics measurement showed that the maximum tensile load in group D was significantly higher than that in groups A and B at 4 weeks (P<0.05), but no significant difference was shown among groups A, B, and C, and between groups C and D (P>0.05); at 8 weeks, it was significantly higher in groups C and group D than group A, and in group D than group B (P<0.05), but there was no significant difference between groups A, C and group B (P>0.05); at 12 weeks, it was significantly higher in groups C and D than groups A and B, and in group D than group C (P<0.05), but difference was not significant between groups A and B (P>0.05). ConclusionDifferent concentrations ratios of OPG/DPB gel complexes have different effects on the bone tunnel after ACL reconstruction. 100% OPG/DPB gel complex has significant effects to prevent the enlargement of bone tunnel and to enhance tendon bone healing.
The damage effects of the pure tumor necrosis factor (TNF) on the normal animals were observed. Eighteeen rabbits were divided into two groups, eight in tested group and ten in control group. 0.5mg per kg of the pure rabbit TNF was given to each animal of the tested group. Results:The symptoms similar to that induced by endotoxin appeared after the TNF injection. The functions of the main organs were markedly damaged. The arterial blood pressure of most animal was low. The weight ratio of the orgen to the body was raised. The pathologic changes were similar to those of the multiple organ failure (MOF) model. Most of the animal died before the end of the experiment. The results suggest that pure TNF could indece multiple organ damages similar to those of MOF.
In this paper,the changes of activities of enzymes relating toenergy metabolism in rabbit's retina during acute ocular hypertension were observed.The activities of succinate dehydrogenase and adenosine triphosphatase were foud to be reduced,while the activities of the lactatic dehydrognease and glucose-6-phosphatase increased.The results reveal the disturbance of metabolism of energy in retina undergone acute ocular hypertension,and suggest that this might be the underlying factors relating to the defects of the functions and structures of the retina. （Chin J Ocul Fundus Dis,1993,9:141-144）
Objective To compare the effect of two types of intermittent pressure on formation of pressure ulcer in rabbit hind l imbs and to investigate the mechanism of gradually changed intermittent pressure produced by waves bed in the prevention of pressure ulcer. Methods Gracil is (3 cm2) in both hind l imbs of 12 adult Japanese white rabbits were randomlyloaded with gradually changed intermittent pressure (50-160 mm Hg, 1 mm Hg=0.133 kPa) and sustained pressure (100 mmHg) serving as the experimental group and the control group, respectively. The experiment was terminated after 4 cycles, and a single cycle included 2 hours of compression and 30 minutes of compression-release. Blood velocity of hind l imbs and blood perfusion of wound were detected by bidirectional doppler blood flow detector and laser doppler perfusion imaging detection system before compression and at every 10 minutes in compression-release period of each cycle (0, 10, 20 and 30 minutes). After the termination, gross observation of the wound was conducted, pathomorphological changes of tissues from compressed area were observed by HE staining, and contents of NO, malondialdehyde (MDA), and superoxide dismutase (SOD) in muscle tissue were measured using colorimetry method. Results No significant difference was evident between two groups in terms of blood flow velocity before compression (P gt; 0.05); the blood flow velocity of two groups decreased significantly at 0 minute in every compressionrelease period of each cycle, and no significant differences were noted between two groups (P gt; 0.05); the blood flow velocity of theexperimental group was higher than that of the control group at 10, 20 and 30 minutes (P lt; 0.05). No significant difference was noted between two groups in terms of wound blood perfusion before compression (P gt; 0.05); the wound blood perfusion of two groups decreased significantly at 0 minute in every compression-release period of each cycle, and no significant differences were noted between two groups (P gt; 0.05); the difference between two groups was not significant at 10 minutes in the first cycle (P gt; 0.05), and the experimental group was higher than the control group at 20 and 30 minutes in the first cycle (P lt; 0.05). In the following 3 cycles, the recovery of perfusion in the experimental group was faster than that of the control group (P lt; 0.05). Gross observation showed the experimental group had less effusion than the control group. The experimental group had intact cutaneous appendage, less inflammatory cell infiltration, and no obvious ulcer formation, whereas the control group had obvious skin ulcer, depletion of cutaneous appendage, and more inflammatory cells infiltration. Significant differences were noted between two groups in terms of NO, MDA, and SOD content (P lt; 0.05). Conclusion Gradually changed intermittent pressure can maintain the blood perfusion of tissue, reduce ischemia-reperfusion injury and cell apoptosis, and prevent the formation of pressure ulcer.