本文選題：喉 + 支架��； 參考：《第四軍醫(yī)大學》2010年碩士論文

【摘要】： 喉移植是全喉切除后恢復喉功能的理想方法,而免疫排斥反應和免疫抑制劑的使用是限制喉移植開展的主要原因�？焖俣鴩乐氐囊浦才懦夥磻桥R床器官移植成功的主要障礙。如能構建低免疫原性生物人工喉則可望解決這一問題。近年來去細胞支架的研究為構建低免疫原性喉提供了希望。去細胞生物支架是用物理或化學等方法去除組織或器官內(nèi)的細胞和可溶性蛋白,僅保留組織結(jié)構中的不溶性基質(zhì)成分,其主要成份為膠原,有助于種子細胞的粘附,增值,生長,分化,可為新組織的生成提供網(wǎng)狀結(jié)構支架[1]。但這種經(jīng)去細胞技術處理的喉支架的免疫原性有待進一步證實。本研究用動脈灌注去細胞技術去除供體喉的肌肉和黏膜而保留其三維空間結(jié)構,構建同種異體生物人工喉軟骨支架并評價其免疫原性,為全喉重建提供理論依據(jù)。 目的 1.通過灌注法構建去細胞全喉支架,探討灌注法構建去細胞軟骨支架的可行性,并對其進行形態(tài)學觀察。 2.通過動物體內(nèi)實驗法評價去細胞喉軟骨支架的免疫原性,為組織工程方法構建全喉的應用提供理論依據(jù)。 方法 1.去細胞全喉支架的構建及其形態(tài)學研究通過頸總動脈灌注離子型去污劑,對離體全喉進行去細胞處理。在去除可溶性基質(zhì)的同時保留喉細胞外基質(zhì)的三維空間結(jié)構及完整的全喉軟骨支架結(jié)構,制備后的去細胞喉支架行大體觀察、掃描電鏡觀察及組織學觀察。 2.去細胞全喉支架的免疫學研究 實驗組:新西蘭大白兔作為喉支架供體,通過雙側(cè)頸總動脈順行灌注去離子劑,構建出去肌細胞全喉支架。青紫藍兔為受體,將去細胞全喉支架植入青紫蘭兔喉肌旁;對照組:新西蘭大白兔新鮮離體喉作供體,青紫蘭兔為受體,將離體喉直接植入青紫蘭兔喉肌旁,各組分別于植入后2,4,12,24周取材進行大體觀察、組織學觀察及淋巴細胞浸潤測定。 結(jié)果 1.去細胞全喉支架的構建及其形態(tài)學研究大體觀察:離體喉經(jīng)十二烷基硫酸鈉(SDS)灌注十六小時后即透明,去細胞喉支架結(jié)構完整,顏色蒼白。掃描電鏡觀察:灌注組喉肌群去細胞程度完全,孔隙多,膠原纖維未受損,且軟骨細胞活力保存完好。去細胞喉冠狀切面HE染色示:喉聲帶、室?guī)�、彈性圓錐等結(jié)構均存在,肌細胞完全被去除,纖維樣結(jié)構仍存在,排列規(guī)整,較完整的保留了細胞外基質(zhì)的方向性。 2.去細胞全喉支架的免疫學研究 各組動物術后均存活至取材時間,實驗組傷口愈合較對照組好。組織學觀察:對照組2周出現(xiàn)免疫排斥反應,4周后軟骨結(jié)構被破壞,喉的基本結(jié)構消失;實驗組2周時出現(xiàn)輕微的炎性反應,4周時形成纖維結(jié)締組織包繞,炎性反應減輕,12周軟骨結(jié)構存在,軟骨周圍大量結(jié)締組織形成,24周后,軟骨結(jié)構消失,被結(jié)締組織取代。淋巴細胞浸潤觀察:各時間點實驗組淋巴細胞浸潤程度均低于對照組(P0.01)。 結(jié)論 1.灌注法能夠清除喉組織的上皮細胞及肌細胞,保留細胞外基質(zhì)及軟骨,達到去細胞要求,構建完整的去細胞全喉支架。 2.去細胞軟骨支架結(jié)構相對完整,免疫排斥輕微,具有良好的生物相容性,是一種可供選擇的喉支架材料。
[Abstract]:The use of immune rejection and immune inhibitor is the main cause of successful laryngeal transplantation . The rapid and severe graft rejection reaction is the main obstacle to the success of clinical organ transplantation . In recent years , the study of cell scaffolds has provided hope for the construction of low immunogenicity throat . In recent years , the research of cell scaffolds has provided hope for the construction of low immunogenicity throat . However , the immunogenicity of the laryngeal scaffold treated by the decell technique is to be further confirmed . This study uses arterial perfusion to remove the muscle and mucous membrane of donor throat to retain its three - dimensional space structure , constructs the allograft artificial laryngeal cartilage scaffold and evaluates its immunogenicity , and provides the theoretical basis for the reconstruction of the whole throat .



Purpose



1 . To study the feasibility of constructing acellular cartilage scaffold by perfusion method , and to observe the morphological observation .



2 . To evaluate the immunogenicity of acellular laryngeal cartilage scaffold by animal experiment method , and provide theoretical basis for tissue engineering method to construct the whole - throat application .



method



1.To remove soluble matrix , the three - dimensional space structure of the extracellular matrix and the whole structure of whole laryngeal cartilage were retained .



2 . Immunologic study of acellular whole - throat scaffold



The experimental group : New Zealand white rabbits were used as the donor of the laryngeal scaffold , and the whole - throat stent of the muscle cells was constructed by the anterograde perfusion of the bilateral common carotid artery . The blue - violet blue rabbits were used as recipients , and the whole - throat stent was transplanted to the laryngeal muscles of the blue - violet rabbit . The control group : New Zealand rabbits were used as donors , and the isolated throat was directly implanted next to the laryngeal muscles of the blue - violet rabbit . All the groups were treated with 2 , 4 , 12 , 24 weeks after implantation .



Results



1 . The structure of acellular whole - throat stent and its morphological study were as follows : After 16 - hour perfusion of sodium dodecyl sulfate ( SDS ) , it was clear that the structure of the acellular laryngeal stent was intact and the color was pale . Scanning electron microscope ( SEM ) showed that the structure of the laryngeal muscle group was complete , the pores were much , the collagen fibers were not damaged , and the viability of the chondrocytes was intact .



2 . Immunologic study of acellular whole - throat scaffold



After 4 weeks , the cartilage structure disappeared and the cartilage structure disappeared and the connective tissue was replaced by connective tissue after 24 weeks . The lymphocyte infiltration in the experimental group was lower than that of the control group ( P0.01 ) .



Conclusion



1 . The perfusion method can eliminate the epithelial cells and muscle cells of the laryngeal tissue , retain the extracellular matrix and the cartilage , meet the requirement of the decell , and construct the complete acellular whole - throat bracket .



2 . The structure of the acellular cartilage scaffold is relatively intact , the immune rejection is mild , and the acellular cartilage scaffold has good biocompatibility , and is an alternative throat bracket material .
【學位授予單位】：第四軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2010
【分類號】：R767.9

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