| Objective:Tendon injuries are common among the field of hand surgery diseases, mostly of which are multiple traumas, and often need tendon transplant to repair defects. Allogeneic tendon because its source adequate ,don't cause vice damage for patients.,is the ideal transplant choice Amniotic membrane is a translucent film; immune rejection does not occur after allograft, immunosuppressive factors can be secreted to inhibit the inflammatory response after transplantation. This experiment uses amniotic membrane surrounding the tendon in the allograft, which is introduced to restore deep flexor tendon defect of Leghorn toe. The effectiveness of healing and prevention of tendon adhesion and suppress immune rejection will be evaluated and the experimental data will be provided for tendon allograft and prevention of tendon adhesion to in surgical field.Methods: choose96 three months old male Leghorn chickens with an average weight of 1.95±0.063kg. Number, tag and randomly divided into A (amniotic membrane allograft tendon group), B group (allogeneic tendon group), C group (control group). In this experiment, the left firstⅢ,Ⅳtoe foot of Leghorn toe will be used as tendon injury model. A "[" style skin incision with a length of about 3cm shall be made between the plantar side which is near to inter-phalangeal joint to the middle of the distal phalanx. Incite the skin and turned up flap to separate the tenacity and expose flexor superficial and deep tendon tissue. The III and IV tendon of deep flexor digitorum taken from the leghorn chicken shall be interchanged and tendon stitched. A piece of amniotic membrane with a gr area of 2.0cm×1.5cm shall be implanted to A, and the membrane shall be surround the upper and lower tendon graft while pure graft tendon is transplanted to B; Flexor digitorum profundus shall be cross-sectioned in the same area of and then sutured. The firstⅢ,Ⅳtoe of the leghorn shall be dressed after surgical operation, the plantar flexion of ankle joint and flexion of metatarsophalangeal joints are 90 degree. The toes joint extension shall be fixed with gypsum board and tape.Gross anatomical observation:choose 6 chickens respectively from the A, B and C group after 7days, 14days, 21days, 28days after surgical operation. Kill the leghorn chicken and incise along original operation incision and observe the pathological changes of tendon tissues, anastomosis fracture, color of transplanted tendon, sliding tendon, tendon healing, adhesion of tendon, etc.Histological observation:choose two leghorn chicken respectively from A, B and C group 7days, 21days after surgical operation, cut the flexor tendon centered by tendon coincidence with a length of 2 cm along the shallow layer of tendon sheath. Fix it with 10% formalin and dehydrate with ethanol and then dip it in wax after transparent treatment in dimethylbenzene. , slice the tendon and were perform HE and Masson staining before observation with light microscope.TEM:choose two leghorn chicken respectively from A, B and C group 14 days, and 28 days after surgical operation, cut the toe along the original incision, and take a 1mm×1mm×1mm tissue, fasten the tissue with 3% glutaraldehyde fixative and 1% osmium tetroxide fixative gradually, and dehydrate in the refrigerator with a temperature of 2℃refrigerator with ethanol, acetone . Pack and bury the tissue with acetone and 812 epoxy resin and slice with ultra-thin slicing machine slices, double stain with 3% uranyl acetate - lead citrate, and observe under electron microscopy.Biomechanical test:choose 4 leghorn chickens from each group 7d, 14d, 21d, 28d, 35d, 42d respectively after operation. Cut the digitorum flexor tendon of the left III and IV toe for biomechanical test.①determination of tensile breaking strength: Remove theⅢdeep flexor tendon, and fixed on the fixture of biomechanical testing machine. The full-load range is set to 200N, the tendon is dragged to the rupture of the tendon with a speed of 15mm/min and connected to the testing machine, run the computer program, simultaneously measure and record the tensile breaking strength data.②Power dissipation of breaking down tendon adhesion: cut the IV toe at the metatarsophalangeal joint, exposing the proximal deep flexor tendon. Fix the proximal phalanx of toe IV. Fix the tendon to the testing machine fixture, and drag with a speed 15mm / min, the full load range is set to 200N. The tendon shall be stopped after a sliding of 14.5mm, and get the force - displacement curve, the area under the curve of power W1. Stretch 14.5mm again after resumption, get-power W2. Power to break the tendon adhesions (set to W0): W0=W1-W2. Data shall be statistically analyzed.Results:①The gross and histological observation: After 7d, 14d, 21d, 28d , it is showed that C group, compared with A group, B group, has less congestion, edema and inflammatory reaction, infiltration of inflammatory cells is more limited. Compared with B group, A group has weaker tissue congestion, edema and inflammatory reaction and invasion range. 21d, 28d after operation, we can see that loose tendon adhesion in A group and dense tendon adhesion in B and C group.②TEM observation: After 14d, 28d, It is showed that C is stronger in generation of synthesis of fibroblasts compared with A, B group, we can see a lot more regular arrangement of collagen fibrils. B has smaller number of fibroblasts compared with A group, the new arrangement of the collagen fibrils are sparse, disorder, not compact.③Biomechanical test: the maximum tensile strength of tendon : The differences of among A and B, C was statistically significant (P <0.05); comparison after 7d, 14d, A, B group was not statistically significant (P> 0.05).For the rest of the time points, difference between A, B groups was statistically significant (P <0.05) . Power dissipation of breaking down tendon adhesion:The differences of among B and C, A was statistically significant (P <0.05). Inter-group comparison between B and C 28d, 35d, 42d respectively after operation is not statistically significant (P> 0.05). For the rest of the time points, difference between B and C groups is statistically significant (P <0.05).Conclusion: Amniotic membrane as a graft material, not only plays the role of secreting immunosuppressive factors, inhibiting the inflammatory reaction transplanted tendon, it can also be effective in preventing tendon adhesion. Amniotic membrane can be absorbed by organs and will not be left in the body and it provides a new direction for research for clinical transplantation of tendon as well as the prevention of postoperative adhesions.
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