Objective To investigate an improved large vascular reconstruction method in the canine liver transplantation and see whether it can shorten the anheptic time and thus reduce the harmful effects during the anhepatic phase. Methods Thirty-two mongrel dogs were enrolled and divided into two groups randomly:the donor group (n=16) and the acceptorgroup(n=16). The dogs in the acceptor group were divided into two groups, according to the different reconstruction methods: Group A using the magnetic rings for a large vein reconstruction in the canine liver transplantation (n=10), and Group B using a handsewing large vein reconstruction in the canine liver transplantation (n=6). The operation time, hemodymics change, anastomosis site, and survival were observed. Results The operation time was as follows: In Group A, the total operation time, the inferior vena cava anastomosistime, and the anheptic phase time were significantly shorter than those in Group B (3.24±0.49 h vs 4.12±0.51 h，5.89±2.27 min vs 28.33±6.04 min，3.89±0.73 min vs 12.16±3.72 min),with a significant difference between the two groups (Plt;0.01). The haemodymics changes were as follows: In Group A, MAP dropped during the anhepatic phase, but it soon recovered after reperfusion，and there was only 730.56±150.56 ml of fluid including the donor blood that needed to be transfused, with no pressor agent required. In Group B, blood pressure dropped during the anhepatic phase，but it slowly recovered，and there was 2241.67±390.78 ml of fluid. In Group A, all the stomas had no errhysis, twistor thrombus. The twisted stomas could be corrected by the revolving of the magnetic rings. The endangium at the site of anastomosis was smooth. In Group B, most of the stomas had errthysis. In Group A, 3 dogs survived for more than 7 days, 6dogs survived for 3-6 days, and 1 dog survived for only 12 hours. In Group B, 2 dogs survived for 3-6 days, 3 dogs survived for 1-2 days, and 1 dog survivedfor only 12 hours. Conclusion Using the magnetic rings for a large vascular reconstruction in the canine liver transplantation is an improvedmethod, which can simplify the anastomosis procedures and significantly shortenthe anheptic phase time. However, the magnetic rings have to be placed in the abdomen, so this method remains to be further improved.
To evaluate the effect of technique combination of implant-retented titanium lattice with decalcified dental matrix (DDM) implanting. Methods Six healthy male dogs (weighing of 10-20 kg) were randomly divided into 3 groups. All the premolars were extracted on both sides of the jaw in dogs. After 2 weeks, titanium lattice and implant were implanted in the maxillary premolar region with DDM on one side (experimental group), but without on the other side (control group) of each dog. After 4, 9 and 14 weeks, respectively, 2 animals were individually killed each time, and the samples wereevaluated by general observation, X-ray examination, histological observation and histomorphometric analyses. Results General observation: Among the 6 dogs, there was no postoperative infection or death. The X-ray examination showed that the bone density of the experimental group was greater than the control group at 4 and 9 weeks, and had no significant difference as to the vicinity bone at 14 weeks. On the other hand, the density of the control group was very low under the titanium lattice and around the implant. The experimental group revealed a ridge augment of (1.93 ± 0.24) mm, and control group (-1.02 ± 1.20) mm (P lt; 0.05). Developed bone sponge could be found after 14 weeks. Histological observation showed that in the experimental group, the DDM surface was nearly absorbed at 4 weeks. A few new bones were formed at 9 weeks. The whole DDM was absorbed; the trabecular bone was thick and arranged regularly; and the intergradations of implant were observed at 14 weeks. In the control group, there were some inflammatory fibers around the neck of implant at 4 weeks. The inflammatory condition extended to the root of implant and the titanium lattice at 9 weeks. There was no newly-formed bone under the titanium lattice at 14 weeks. Histomorphometric analyses showed that the implant contact bone ratio approached 1 ∶ 1, and showed no significant difference between the new bone fragment and former bone fragment in the experimental group. Conclusion This augmentation of alveolar ridge evaluated by the study is appl icable, but further study is necessary.
The purpose of the study was to observe effect of chinese medicine “Qing Yi Tang” on the repair of injury of intestinal mucosa in acute necrotizing pancreatitis (ANP). Dogs ANP model were induced by injection of 5% sodium taurocholate (0.5 ml/kg) with 3 000 u/kg trypsin into the pancreatic duct. Diamine oxidase and anylase activity in blood, protein and MDA levels of ileal mucosa were to be determined in ANP and after treatment of “Qing Yi Tang”. Intestinal permeability was also to be studied, LPS and bacteria translocation (BT) were obseved. All animals were sacrificed on day 7, the tissue of ileal mocosa was collected for histological and ultrastructural studies. The results showed that after treatment with Chinese medicine “Qing Yi Tang”, the injury of intestinal mucosa in ANP reduced. The length, height, area and protein of ileal mucosa increased significantly, intestinal permenbility decreased, the levels of LPS reduced in 1-2 times, and organ BT rate also reduce in 50%. The results indicated that chinese medicine “Qing Yi Tang” had good effect on improving repair of intestinal mucosa injury, protecting gut barrier function, reducing the incidence of LPS and bacteria translocation.
【Abstract】 Objective To measure the changes of bone mineral density and bone micro-architecture of thefemoral head that harvested from the three-foot bearing ethanol destroyed canine model for osteonecrosis of femoral head, and discuss the influences of local injection of ethanol and biomechanical loading to the structural properties of the femoral head. Methods Twenty-four Beagles were divided randomly into four-foot bearing canines and three-foot bearing canines. One fore-l imb was fixed randomly in three-foot bearing canines. Osteonecrosis was induced in all experimental animals by local injection of 5 mL pure ethanol into one side of the femoral head. The hind l imbs injected with NS were acted as control group, that of three-foot canines injected with ethanol were acted as three-foot canine group, and that of four-foot canines injected with ethanol were acted as four-foot canine group. The contralateral femoral head was injected into equal amount of NS. Animals were sacrificed at the time intervals of 1, 3, 6, and 12 weeks after the injection of ethanol. Quantitative microcomputedtomography was used to characterize changes in bone micro-architecture and bone mineral density of femoralhead. Results The clear three-dimensional model of trabecular bone of necrotic femoral head were obtained. There were no significant differences among 3 groups according to the time l ine by 1 week after ethanol injection(P gt; 0.05). At 3 weeks after injection of ethanol, in three-foot canine group and four-foot canine group, the volume of BMC, BMD, BVF, and BS/BV increased gradually as the distance to the drill ing canal increased. There were significant differences between 3 regions (P lt; 0.05). At 6 weeks, in three-foot canine group and four-foot canine group, the volume of BMC, BMD,BVF, and Tb.N of region I and II decreased significantly compared with region III (P lt; 0.05). At 12 weeks, there are no differences among 3 groups (P gt; 0.05). There were significant decreases in BMD values, BVF, BS/BV, Tb.N, Tb.Sp and Tb.Th after the injection of pure ethanol. And, the changes were more and more obvious by the time l ine. These changes were differentiable at 3 weeks after injection of ethanol, and became obvious at 6 weeks. These changes were more obvious at the part that near the injection canal. The changes in threefoot canine group were more obvious than that in four-foot canine group. Conclusion Resorption of necrotic compact bone trabecular may weaken the structural properties of the femoral head. Moreover, remodel ing and repairing process of necrotic bone trabecular may be hampered by constant biomechanical loading that presented in three-foot bearing canines, and thereby further weaken the structural properties of the femoral head. Biomechanical loading may be one of the critical reasons that lead to the collapse of femoral head.
Six dogs underwent high selective vagotomy and mucosal antrectomy (HSV+MA). The gross and histological change of dog’s stomach were observed at 4-6 months after operation. It was found that the reconstructed antrum healed well and there was no stasis and distension in the stomach .The appearance of the nerves in muscular layer of the antrum was normal. No serious gastritis and mucosal atrophy was observed. These results indicat that HSV+MA is a reasonable procedure for the treatment of duodenal ulcer.
To evaluate the implantation effect of artificial vascular grafts with recombinant fibrinolytic enzyme factor II (rF II)-immobil ized lumina in animal test. Methods Four mm internal diameter (ID) polyurethane (PU) artificial vascular grafts were prepared by di pping and leaching method. The micro-pore size and morphology of the graft walls were observed by SEM. The graft lumina were immobil ized with rF II. Twenty hybrid male dogs [weighing (20 ± 1) kg] were used for animal model of carotid artery defect and were randomly divided into 3 groups: rF II -immobil ized PU group, no rF II -immobil ized PU group and expanded polytetrafluoroethylene (ePTFE) group. The vascular grafts were implanted for repairing injured segments of carotid artery in dogs. The general health state of animals was recorded. At 30 days and 60 days,the patency rate of every group was calculated. At 60 days IDs were measured, cell prol iferation in neointima was inspected by l ight microscope, morphology on neointima was observed by SEM. Results The ID of the PU vascular grafts was (3.74 ± 0.06) mm, wall thickness was 0.4-0.6 mm, the wall density was 0.25 g/cm3, the porosity was 79.8%, racical compl iance was 8.57%/100 mmHg. In the wall, micropores were well distributed and opened-pores structure was observed. Pore size was (140 ± 41) μm in the outside layer, pore size was (100 ± 3) μm in the inside layer, thickness ratio of outside / inside layers was 2 ∶ 1, the pore size was (40 ± 16) μm on the lumina surface. After operation the wounds on neck healed, all the animals survived and had no compl ication. At 30 days and 60 days after implantation, the patency rate for rF II -immobil ized PU group were 100% and 66.7%, for no rF II -immobil ized PU group were 66.7% and 33.3%, and for ePTFE group were 67.7% and 0 respectively, but at 60 days there were thrombosis at anastamotic sites of some grafts occluded. Before operation the IDs for rF II-immobil ized PU group, no rF II -immobil ized PU group and ePTFE group were (3.74 ± 0.06), (3.74 ± 0.06) and (4.00 ± 0.03) mm, at 60 days after operation the IDs were (4.51 ± 0.05), (4.31 ± 0.24) and (4.43 ± 0.12) mm respectively, showing no statistically significant differences between 3 groups (P gt; 0.05). Histological inspection indicated that at 15 days a layer of plasma protein deposited on the lumina, at 30 days some cells adhered to the lumina, at 60 days neointima could be observed on the lumina. Thickness of the neointima became larger with implantation time. At 60 days neointima thickness at proximal end, middle site and distal end ofgraft were (560 ± 22), (78 ± 5) and (323 ± 31) μm respectively for rF II -immobil ized PU group. The results of SEM showed that neointima surface consisted of flat and long cells which long axes ranged with blood flow direction and was similar to lumina morphology of carotid artery of dog. Conclusion Immobil ization of rF II to lumina of grafts could enhance fibrinolytic activity and inhibited formation of thrombo-embol ia which led to an increase in patency rate after implantation.
Objective To explore an effective method of culturing the canine bladder smooth muscle cells, observe the morphological characteristics of the bladder smooth muscle cells growing on acellular small intestinal submucosa（SIS） and offer an experimental basis for reconstruction of the bladder smooth muscle structure by the tissue engineering techniques. Methods The enzymetreatment method and the explant method were respectively used to isolate and harvest the canine bladder smooth muscle cells, and then a primary culture of these cells was performed. The canine bladder smooth musclecells were seeded on the SIS scaffold, and the composite of the bladder smooth muscle cells and the SIS scaffold were co cultured for a further observation. At 5,7 and 9 days of the co culture, the specimens were taken; the bladder smooth muscle cells growing on the SIS scaffold were observed by the hematoxylin staining, the HE staining, and the scanning electron microscopy. The composite of the bladder smooth muscle cells on the SIS scaffold was used as the experimental group, and the bladder smooth muscle cells with no SIS were used as the control group. In each group, 9 holes were chosen for the seeded bladder smooth muscle cells, and then the cells were collected at 3, 5 and 7 days for the cell counting after the enzyme treatment. Morphological characteristics of the cells were observed under the phase contrast microscope and the transmission electron microscope. Expression of the cell specific marker protein was assessed by the immunohistochemical examinaiton. The proliferation of the cells was assessed by the cell counting after the seeding on the SIS scaffold. Results The primary bladder smooth muscle cells that had been harvested by the enzyme treatment method were rapidly proliferated, and the cells had good morphological characteristics. After the primary culture in vitrofor 5 days, the bladder smooth muscle cells grew in confluence. When the bladder smooth muscle cells were seeded by the explant method, a small amount of the spindleshaped bladder smooth muscle cells emigrated from the explant at 3 days. The cells were characterized by the welldeveloped actin filaments inthe cytoplasm and the dense patches in the cell membrane under the transmissionelectron microscope. The immunohistochemical staining showed the canine bladdersmooth muscle cells with positive reacting α actin antibodies. The bladder smooth muscle cells adhered to the surface of the SIS scaffold, growing and proliferating there. After the culture in vitro for 5 days, the smooth muscle cells covered all the surface of the scaffold, showing a singlelayer cellular structure. The cell counts at 3, 5 and 7 days in the experimental group were（16.85±0.79）×105，（39.74±2.16）×105 and （37.15±2.02）×105, respectively. Thecell counts in the control group were（19.43±0.54）×105，（34.50±1.85）×105 and （33.07±1.31）×105, respectively. There was a significant difference between the two groups at 5 days （P＜0.05）. ConclusionWith the enzyme treatment method, the primarily cultured canine bladder smooth muscle cells can produce a great amount of good and active cells in vitro. The acellular SIS can offer an excellent bio scaffold to support the bladder smooth muscle cells to adhere and grow, which has provided the technical foundation for a further experiment on the tissue engineered bladder reconstruction.
ObjectiveTo compare the difference of rotator cuff healing between different types of injury and between different repair methods, and to explore the animal model to accurately simulate the restorative process after repair of rotator cuff injury. MethodTwelve adult male beagle dogs (weighing, 10-15 kg) were divided into 3 groups (n=4) according to different processing methods:acute rotator cuff injury+Mason-Allen suture repair (group A), huge rotator cuff injury+Mason-Allen suture repair (group B), and huge rotator cuff injury+Mason-Allen combined with autogenous semitendinosus expansion suture repair (group C). The external fixation was used for immobilization after repair. After operation, the general situation of the animals was observed, and the infraspinatus tendon was harvested for gross observation at 6 weeks after operation. The biomechanical test of limit load and histological observation of tendon fibers were carried out. ResultsAll the animals survived to the end of the experiment. All incisions healed well and no infection occurred. Gross observation showed more scar tissues at the end of infraspinatus muscle tendon than normal tendon in group A; no obvious tendon tissue was observed at the end of infraspinatus muscle tendon in group B; the infraspinatus muscle tendon was covered with some white scar tissue, but the tendon and the general direction could be observed in group C. The limit load of groups A, B, and C were (223.75±24.28) , (159.25±34.87) , and (233.25±14.24) N respectively, group B was significantly lower than groups A and C (P<0.05) , and no significant differnce was found between group A and group C (P>0.05) . Histological observation showed normal arrangement of tendon fibers in group A; tendon fibers arranged disorderly in group B and tendon cells were significantly less than those of group A; tendon fibers arranged in neat in group C and tendon cells were more than those of group B. ConclusionsCanine autologous semitendinosus expansion repair of massive rotator cuff injury immobilization model can better simulate the clinical rotator cuff injury healing process, so it can be used as an ideal animal model for related research.
Objective To investigate the result of the transplantation of frozen canine phalangeal joint allografts perforated and incorporated with autogenic bone marrow. Methods A proximal interphalangeal joint defect of 1.5 cm was prepared at bilateral sides of twenty-four adult healthy out-bred dogs. Three different types of allografts were applied to repair the defects: fresh autogenic phalangeal joints (group A,n=16), frozen phalangeal joint allografts perforated and incorporated with fresh autogenic bone marrow(group B, n=16), and frozen phalangeal joint allografts(group C, n=16). Radiographic and histological study wereused to evaluate the survival of transplanted joints. The observation was done 1, 3, 6 and 12 months after operation respectively. Results Based on the radiographic and histological changes of the transplanted joints, the osteoarthropathy of transplanted canine phalangeal joints could be divided into 3 degrees: mild degeneration, moderate degeneration and severe degeneration. Mild degeneration was observed in group A from 3 to 12 months. Mild degeneration was also found in group B from 1 to 6 months, and the endochondral ossification was obvious within the drilled bony holes.However, some joints in group B underwent moderate degeneration 12 months after operation. Group C joints in the first month had moderate degeneration, which progressed to severe egeneration 3 months after operation. Conclusion Transplantation of frozen canine phalangeal joint allografts perforated and incorporated with autogenic bone marrow can effectively delay the degeneration of transplanted osteoarticular allografts at the early and middle stage.
Objective To establish a simple and efficient method to isolate and culture the umbilical vein vascular endothelial cells in canine. Methods Twelve umbilical cords [(13.0 ± 1.5) cm in length] were taken from 12 newborn pups of Beagles. And then the vascular endothelial cells were isolated from these umbilical cords digested by 1% collagenase type I for 5, 7, and 10 minutes respectively (4 umbilical cords in each group). After cultured, the vascular endothelial cells were identified by morphology, immunofluorescence, and flow cytometry. And the growth curvature of umbilical vein vascular endothelial cells was detected by MTT assay. Results Few vascular endothelial cells were collected at 5 and 10 minutes after digestion; many vascular endothelial cells were seen at 7 minutes, and became cobblestone with culture time, with a large nucleus; after passage, cell morphology had no obvious change. Fluorescence microscope results showed that positive von Willebrand factor (vWF) and CD31 cells were observed in most of cells. The flow cytometry test displayed that the positive cell rates of vWF and CD31 were 99.0% ± 0.7% and 98.0% ± 1.2%, respectively. The above results indicated that cultured cells were vascular endothelial cells. MTT assay showed that vascular endothelial cells proliferation increased significantly with culture time. Conclusion Enzyme digestion is a convenient method to isolate vascular endothelial cells from canine umbilical vein, and a large number of cells and high purity of cells can be obtained by the method.