本文選題：神經(jīng)生長因子　切入點：跨區(qū)皮瓣　出處：《第三軍醫(yī)大學》2017年碩士論文

【摘要】：背景及目的:穿支皮瓣作為創(chuàng)面覆蓋、組織修復技術,在臨床上發(fā)揮著舉足輕重的作用。Taylor提出血管體區(qū)(angiosome)及Choke Vessels是穿支皮瓣臨床運用的解剖學基礎,而相鄰血管體間choke vessels的開放及“潛在供區(qū)”有效循環(huán)的建立成為移植后皮瓣遠端組織成活的關鍵。在跨區(qū)修復較大創(chuàng)面時,穿支皮瓣術中、術后壞死時有發(fā)生,處理棘手。外科延遲技術通過多期手術能夠改善遠端血管體區(qū)在皮瓣正式切取后的血供,有效擴大皮瓣的修復面積。目前多數(shù)研究認為其機制是促進相鄰血管體區(qū)間的choke血管的擴張及側支循環(huán)的建立。這一過程可被某些藥物、細胞及其分泌的生化因子加強甚至替代,從而衍生出化學皮瓣延遲(chemical flap delay)。神經(jīng)營養(yǎng)因子(NGF)是一個多功能多肽分子,可以通過內(nèi)皮祖細胞酪氨酸激酶受體TrkA途徑觸發(fā)內(nèi)皮細胞的增殖、遷移,維持血管內(nèi)皮功能。NGF還直接或者間接調節(jié)VEGF表達促進血管生成。此外,內(nèi)皮組細胞上分布血管生長因子II型受體KDR,VEGF與其結合后發(fā)揮血管生成的作用。NGF能夠上調KDR的表達,與VEGF有協(xié)同作用,此為血管生成的另一途徑。然而目前NGF能否通過以上可能的機制影響皮瓣choke區(qū)微血管形態(tài)變化及潛在供區(qū)微循環(huán)建立,從而替代外科延遲手術改善大面積皮瓣成活,還不十分清楚�；谝陨戏治�,本研究篩序并建立動物模型,通過顯微CT評價NGF對SD大鼠背部跨區(qū)供血穿支皮瓣顯微血管形態(tài)變化的影響,并對其機制進行探討,為提高臨床皮瓣成活提供理論支持。方法:第一部分:利用Micro-CT重建技術觀察神經(jīng)生長因子對大鼠跨區(qū)穿支皮瓣血管形態(tài)變化的影響成年SD大鼠40只,隨機分為實驗組和對照組。通過大體解剖篩選合適的大鼠背部跨區(qū)供血穿支皮瓣模型。切取面積約3cm×10cm的背部跨區(qū)皮瓣。實驗組皮下注射NGF溶液(10 nmol·ml-1·kg-1);對照組同法注射0.1M磷酸鹽緩沖溶液(1ml/kg)。術后第3、7天氧化鉛-明膠溶液血管灌注后,利用顯微CT技術,獲取血管體三維形態(tài),Matlable軟件分析血管容積及總長度等計量參數(shù)變化。術后第7天統(tǒng)計皮瓣存活面積。第二部分:神經(jīng)生長因子對大鼠跨區(qū)穿支皮瓣成活初步機制研究成年SD大鼠44只,隨機分為實驗組和對照組。切取面積約3cm×10cm的背部跨區(qū)皮瓣。實驗組皮下注射NGF溶液(10 nmol·ml-1·kg-1);對照組同法注射0.1M磷酸鹽緩沖溶液(1ml/kg)。術后第3、7天局部取材運用Western Blot技術半定量檢測VEGF及CD34蛋白含量;術后第7天免疫組化染色觀測血管生長因子受體(KDR)及NGF受體(Trk A)的表達。結果:1.設計以髂腰動脈為單穿支血管蒂的跨三個血管體區(qū)的大鼠背部皮瓣模型,具有恒定的壞死面積。結果顯示,血管形態(tài)學分析示NGF和時間因相互作用增加血管容積(F=33.304,P0.05)及總長度(F=8.493,P=0.01);實驗組皮瓣成活面積明顯大于對照組(P0.05)。2.NGF明顯促進皮瓣微血管VEGF及CD34蛋白分泌。第3天,VEGF表達實驗組與對照組無顯著統(tǒng)計學差異(P=0.088);實驗組CD34表達明顯高于對照組(P=0.004);第7天,兩種蛋白分子表達,實驗組顯著均優(yōu)于對照組(P0.05)。NGF能夠誘導皮瓣微血管TrkA的陽性表達。結論:NGF能增加choke區(qū)微血管的密度及長度,促進的跨區(qū)供血皮瓣的成活面積;其機制可能是NGF促進皮瓣微血管VEGF及CD34蛋白分泌,誘導Trk A受體陽性表達,通過TrkA-VEGF途徑,促進choke區(qū)微血管生成,維持其內(nèi)皮功能,最終改善潛在供區(qū)微循環(huán)血供,影響皮瓣成活。
[Abstract]:Background and purpose: to cover the wounds as perforator flap, tissue repair technique in clinical plays a pivotal role in.Taylor of vascular body region (angiosome) and Choke Vessels are the anatomical basis for clinical application of perforator flap, and the effective circulation of adjacent blood vessels between choke and vessels open "potential donor" became a key of the flap the distal tissue survival after transplantation. To repair a larger wound in the cross, perforator flap surgery, postoperative necrosis occurred, treatment difficult. Surgical delay through surgery can improve the body region in the distal flap formally after removal of the blood supply, effectively expand the repair area of skin flap. Most studies suggest that the the mechanism is to promote the expansion of choke vessel body interval and adjacent blood vessels in the establishment of collateral circulation. This process can be some of the drugs, and the secretion of cytokines and biochemical and even Instead, which are derived from chemical (chemical flap delay) flap delay. Neurotrophic factor (NGF) is a multifunctional polypeptide that can migrate through the tyrosine kinase receptor TrkA pathway of endothelial progenitor cells to trigger the proliferation of endothelial cells, vascular endothelial function, maintain.NGF also directly or indirectly regulate the expression of VEGF to promote angiogenesis. In addition, the distribution of vascular endothelial growth factor receptor type II KDR cells, VEGF and its combination play a role in angiogenesis of.NGF can upregulate the expression of KDR and VEGF have synergistic effect, this is another way of angiogenesis. However, whether NGF could influence mechanism through the above choke flap area micro microcirculation morphological changes and potential for the district established, so as to replace the surgical delay surgery to improve the large area of flap survival, is not very clear. Based on the above analysis, this study screen order and establish an animal model by Effect of micro CT NGF evaluation of SD rat dorsal perforator flap extra-territorial blood supply forms microscopic changes, and to explore its mechanism, to provide theoretical support to improve clinical flap survival. Methods: the first part: the effect of using Micro-CT reconstruction technique to observe the nerve growth factor on vascular morphological changes of flap inter zone perforator in adult rat SD rats 40 rats were randomly divided into experimental group and control group. The gross anatomy of the rat back cross blood screening perforator flap model. The appropriate cut area is about 3cm * 10cm back cross flap. The experimental group was injected NGF solution (10 nmol - ml-1 - kg-1); the control group received phosphate buffer solution (0.1M 1ml/kg). 3,7 days after operation of lead oxide gelatin solution after vascular perfusion, using micro CT technology to obtain vascular 3D measurement parameters, analysis of the changes of blood volume and the total length of the Matlable software after operation. Seventh days of statistical flap survival area. The second part: the nerve growth factor on the inter zone perforator flap survival in rats preliminary study on the mechanism of 44 adult SD rats, were randomly divided into experimental group and control group. The cut area is about 3cm * 10cm back cross flap. The experimental group was injected NGF solution (10 nmol - ml-1 - kg-1); control group with injection of 0.1M phosphate buffer solution (1ml/kg). 3,7 days after operation using Western Blot technology based local semi quantitative detection of VEGF and CD34 protein in seventh days after operation; immunohistochemical staining was used to observe the vascular endothelial growth factor receptor (KDR) and NGF receptor (Trk A) expression. Results: 1. in order to design the iliolumbar arteries for the back flap in rats model of cross three vascular body region single perforator pedicled, with constant necrosis area. The results showed that the vascular morphological analysis showed that NGF and the interaction time due to increased vascular volume (F=33.304, P0.05) and F (total length =8.493, P=0.01); the experimental group flap survival area was significantly higher than the control group (P0.05.2.NGF) significantly promote flap microvascular VEGF and secretion of CD34 protein. The expression of VEGF in third days, there was no statistically significant difference between the experimental group and the control group (P=0.088); the expression of CD34 in experimental group was significantly higher than the control group (P=0.004); the seventh day, the expression of two protein molecules, the experimental group was significantly better than the control group (P0.05) were positive expression of.NGF can induce microvascular flap TrkA. Conclusion: NGF can increase the density and length of choke region of microvessels, the reverse flow flap to promote the survival area; the mechanism may be that NGF can promote flap microvascular VEGF and secretion of CD34 protein induced. The positive expression of Trk A receptor by TrkA-VEGF pathway, promote the formation of choke microvascular endothelial function, maintain, and ultimately improve the blood microcirculation of potential donor for effect of flap survival.

【學位授予單位】：第三軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R622

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