The Empirical Study Of The Survival Influence Of Recombinant Hirudin In Rats Dorsal Avulsed Flap Models

Objective:To investigate the protective effect and mechanism of recombinant hirudin on avulsed flap in rats dorsal avulsed flap models and explore the application of recombinant hirudin in clinical treating avulsed flap. Methods:Experiments were performed on108male and female Sprague-Dawley rats weighing250±50g.The rats was random distributed as three groups:two treatment groups(A group:local injecting recombinant hirudin; B group:local injecting low molecular heparin) and one control group(C group:local injecting physiological saline).Every group had36rats. Each rat received intraperitoneal injection1%pentobarbital sodium (40mg/kg), Prone Position after anesthesia and disinfection and spread sterile towels. Dorsal random skin flap was prepared on each animal in the middle of the back line shall prevail, pedicle located at head,4cm×6cm in size. The pedicles received protective immobilization. The distal ends were fixed to fine iron stick by suture, connected with sophisticated measure heavily, dispensed tractive force (8kg) ten minutes to form the rats dorsal avulsed flap models. The far ends were sutured after local freeing. Each group’s flaps were respectively injected recombinant hirudin (2mg/1ml/flap), low molecular heparin (2mg/1ml/flap) and equivalence physiological saline on the three classes of long axis subcutaneous postsurgery(the pedicle,2cm from the pedicle and4cm from the pedicle) at once,12 hours,24hours and48hours. To observe the growth of hair, congestion, swelling and survival area of flap, monitor the level of P-Selectin in the distal flap on12th hour,1st day,3rd day and the level of VEFG in the distal flap on3rd day,5th day,7th day, and observe the inflammation cell aggregation and formation of microthrombus in pathological section on1st day,3rd day,5th day,7th day. Results:1.General Observation:The three groups all emerge cyanosis and tumescence on12th hour and24th hour. Group C is worse than A and B. The circumscription of congestion in A and B is unconspicuous while C is obvious. On5th and7th postsurgery day,the distal end of A and B’s cyanosis and tumescence gradually decline and survive well while C is still cyanosis, furvous, texture harden, local trichomadesis, no digital pressure capillary reaction, tending to necrosis. On14th postsurgery day,the percentage of open area of A and B is respective (86.07±.30)%and (81.46±1.06)%, which is higher than group C ((77.03±1.25)%) w ith a statistical difference of P<0.01.Group A is higher than C in the survival rate of flap with a statistical difference of P<0.05.2.Histology Observation:The12th hour after surgery:There is a large hemorrhage in group C, with obvious hyperaemia and edema and multiplicity neutrophil infiltration. Group B has lamellar bleeding and hyperaemia, obvious edema and more neutrophil infiltration. Group A has punctuate hemorrhage, hyperaemia, edema and manipulus neutrophil infiltration.The1st day:There is a sheet hemorrhage in group C, with obvious hyperaemia and edema, neutrophil accumulation masslike and prophase necrosis. Group B has more obvious hemorrhage, edema and neutrophil infiltration. Group A has hyperaemia, edema and scattered neutrophil.The3rd day:There is a more sheet hemorrhage in group C, with obvious congestion, mass inflammatory granulation tissue and scattered neutrophil. Group B has congestion, edema, inflammatory granulation tissue and scattered neutrophil. Group A has scattered congestion, a few hemorrhage, inflammatory granulation tissue and a few scattered neutrophil.The5th day:Group C has a few lamellar hemorrhage, congestion, edema and inflammatory granulation tissue. Group B has a few congestion, edema and comparatively ripe inflammatory granulation tissue. Group A has a few congestion and ripe inflammatory granulation tissue.The7th day:Group C has microthrombosis, emphraxis capillaries and a few neogenesis capillaries. Group B has a few microthrombosis and neogenesis. Group C has abundant neovessels and ripe granulation tissue.3.P selectin content detection (pg/ml):The content of P selection in group A and B is (129.740±12.302)and (151.584±12.409)(111.030±6.824)and (134.434±8.707).(101.674±4.429) and (118.308±7.535) on the12th hour,1st day and3rd day, while the group C is (196.034±8.442)、(162.087±14.485)、(132.473±8.142),with a statistical difference of P<0.05.4.VEGF content detection (pg/ml):The content of VEFG in group A and B is (212.507±15.096)and (158.733±15.311)、(214.010±13.811)and (167.134±11.455)、(218.093±13.464) and (177.484±8.183) on the3rd,5th,7th day, while group C is (120.454±7.575)、(130.133±19.159)、(133.800±4.882) with a statistical difference of P<0.05.Conclusion:The experiment tentatively certifies that locally injected recombinant hirudin can actively suppress the production of P-Selectin, reduce accumulation of inflammatory cells, prevent information of microthrombus, improve ischemia and anoxia of avulsed flaps, decrease injuries of endotheliocyte, encourage expression of VEGF, stimulate hyperplasia of neogenesis capillaries and elevate survival rate of avulsed flaps.