| Research background:With the deep research on wound repair,more and more attention has been paid to flap repair.This repair method can not only complete the complete coverage of the wound,but also can solve the wound which base is weak or bone and tendon exposure.And this is traditional surgical can difficultly repaired.therefore,the application of surgical flaps have become increasingly frequent.And the random skin flap as one of the most commonly used methods,the improvement of survival rate has been widely concerned by doctors and scholars.The lack of known vascular and ischemic reperfusion injury(I-R injury)in the random flap still greatly limited the survival of random flap.In recent years,with the improvement of the design of flap: the best ratio of length to width and shape,operation mode,delayed vascular anastomosis flap transplantation,strengthen wound nursing,hyperbaric oxygen therapy and postoperative application of various growth factors and other methods of implementation,the survival rate increased significantly,but the clinical application of skin flap necrosis is It is often seen.How to avoid the occurrence of I-R injury and improve the survival rate of skin flap has become a hot research direction in recent years.Recombinant human growth hormone(rhGH)in previous studies hasbeen shown to have promoted the expression of plasma protein mRNA and serum albumin(ALB)synthesis,glycemic and lipid decomposition,immune regulation,vascular proliferation,reduce protein decomposition,reduce nitrogen loss,promote wound healing for.On this basis,the clinical application of rhGH has been increased and expanded to burn wound and flap survival.The addition of rhGH after flap transplantation can lead to the decrease of inflammatory factor IL-1,IL-6 and tumor necrosis factor-a(TNF-a),and decrease the aggregation of neutrophils.And stimulate the migration of monocytes,promote the generation of superoxide anion,so that the body's resistance can be improved,thereby reducing the inflammatory response.Peng Lixia et al.Study showed that subcutaneous injection of rhGH can promote the expression of vascular endothelial growth factor(VEGF)and angiogenesis in the skin flap,and reduce the necrosis rate of skin flap.However,the choice of dose has not been confirmed by the experiment.Based on the above reasons,the rat model was used to verify the effects of different doses of rhGH on the dorsal random skin flap in rats.Purposes: By subcutaneous injection of rhGH affect the survival of random skin flaps in rats,and different dose,observe different doses of rhGH on rat dorsal random flap influence,to find out the best injection dose,make a preliminary assessment for its clinical application.Method: 64 healthy adult SD rats,we design of pedicle random flapin the back end of rats whichi the size is 2cm*4cm.Using the principle of random assignment,the rats were divided into A(physiological saline group),B(0.05IU/Kg rhGH group),C(0.1 IU/Kg rhGH group),D(0.2IU/Kg rh GH)four.Subcutaneous injection was performed 7 days after operation.After seventh days,the survival rate of the flaps was calculated in the four groups,and the results of all the flaps were observed.HE staining and immunohistochemical staining were performed to observe the expression of VEGF and CD34 in tissues,and the microvessel density(MVD)was calculated.The data were analyzed statistically.Result: 1.The general observation of the skin flap: A group of proximal skin flap survival area was pink,sparse hair.In the middle and distal part of the necrosis,black,hard,elastic significantly reduced,no hair growth,acupuncture without bleeding,much lighter red excretions under the skin flap.B group of proximal flap survival is acceptable,the color is pink,gradually color dull to brown,small black crust attachment,elasticity less reduced,acupuncture visible bleeding,the hair is sparse,there is a small amount of excretions under the flap,until the distal brown color.In group C and group D,the survival of the flaps was similar,most of the flaps survived well,and the proximal and middle segments were pale red,and the elasticity was similar to that of the normal tissues.no obvious swelling and so on.The distal part and a few of the edges of the site,some brown black necrosis area,the area is far less than A,B group,acupuncture bleeding,no obvious accumulation of blood under the flap.2.Flap survival rate: A group(control group)was(57.72 +.2.03)%,B group was(72.75 +.0.97)%,C group was(81.37 +.1.49)%,D group was(+ + 1.59)%.All data were analyzed by one-way ANOVA: P < 0.05,the data of each group were not equal.Then the LSD method was used to compare the data of pairwise comparison: the experimental group was compared with the control group,P < O.05,the difference was statistically significant,that is,subcutaneous injection of rhGH can improve the survival rate of the flap.Group B and C,D between the two groups showed that: P < O.05,the difference was statistically significant,the low dose group is lower than the promoting effect of rhGH on middle and high dose groups in the survival rate of flap;C group and D group were compared with that of P > O.05,there was no significant difference between middle and high doses of rhGH to promote flap survival had no significant difference.3.HE staining: A group microscopically structure confused,the structure of each layer that is unknown,tissue edema,a large number of neutrophils and mononuclear cell infiltration,showed acute inflammation changes,a small amount of thin-walled vessels without obvious tubulao vascular growth,part of the growth of fibrous tissue.In the B group,the dermal tissue was more chaotic,the edema of the tissue and the infiltration of neutrophils and monocytes were lower than that of the A group,and there was a large number of capillaries,and the granulation was clear.In group C and group D,there was a little inflammatory cellinfiltration,and the structure of the group was clear,and the new tubular vessels were formed,red cell filling.4.The expression of CD34 and VEGF in tissue: Using immunohistochemical staining to observe the expression intensity of CD34 and VEGF within the organization,the results clearly visible CD34 staining of CD34 showed brown yellow granules and mainly distributed in the cytoplasm around the capillaries,expressed in B,C,D group was significantly higher than A group,D group the highest,followed by group C.VEGF and immunohistochemistry in the drug intervention group showed a large amount of brown granules distributed into clusters,while the control group with punctiform distribution.The distribution density of D group > C group > B group > A group.5.The detection of microvessel density: CD34 immunohistochemical staining were placed at 100 times light microscope view,each slide in the 6 visual fields count microvessel number and the mean.A group:(5 +1.46),B group(10.55 + 1.78),C group(14.40 + 1.63),D group(1519 +1.46),one-way ANOVA analysis,P < 0.05,the overall average is not completely equal,that there is statistical difference between groups the difference.Compared the data of pairwise comparison with LSD method,drug intervention group and control group P was less than 0.05,there was significant difference,C,D group and B group compared to P was less than 0.05,low dose group microvessel density was low.C,D two groups compared P > 0.05,the difference was not statistically significant.Conclusion: 1)rhGH can promote the expression of CD34 and VEGF of rat dorsal random flap,so as to promote angiogenesis in the skin flap;2)the survival rate of recombinant human growth hormone can improve the rat random skin flap;3)the effect of 0.1-0.2IU/kg was the best dose of recombinant human growth hormone on the skin flap,can as the best dose of drug use. |
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