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  本文選題：皮膚損傷 + 細胞間粘附分子-1　； 參考：《中國醫(yī)科大學》2003年碩士論文

【摘要】： 前言 皮膚損傷愈合是一個極其復雜的生物學過程，通常其包括三個階段：炎癥期、增殖期及組織重構期。多種生物學介質如：生長因子、細胞因子及粘附分子等與皮膚損傷愈合過程密切相關，尤為值得提出的是，粘附分子在炎癥的發(fā)生的起始過程中起著至關重要的作用，這些因子在細胞外基質的形成、血管的發(fā)生、肉芽組織的生成及上皮的再形成方面等均具有重要作用。對于法醫(yī)病理工作者來說，對這些生物物質變化過程的研究，可以為判定損傷活性及時間提供很好的方法。 粘附分子之所以稱之為粘附分子，是因為其首要功能就是將淋巴細胞、單核細胞及粒細胞粘附在血管內皮細胞上，使其穿越血管內皮，到達炎癥反應處。 本研究應用免疫組織化學方法，觀察大鼠皮膚切創(chuàng)后不同時間細胞間粘附分子(ICAM-1)及P選擇素的表達變化，以期確定這兩種物質表達與損傷時間變化的規(guī)律，為皮膚損傷時間判定提供有效的免疫組織化學指標。 材料與方法 1．材料 取雄性健康SD大鼠39只，隨機分為13組，每組3只。于傷后5min-7d將大鼠斷頸處死，于傷口邊緣處取全層皮膚組織。取9只雄性健康SD大鼠，隨機分為3組做為死后傷對照組，取3只雄性健康SD大鼠，取與致傷大鼠相同部位皮膚作為對照。石 蠟包埋，進行常規(guī)石蠟切片，厚度為 5 4m。 2．方法 應用免疫組織化學SABC法，步驟如下： ＊石蠟切片常規(guī)脫蠟至水； 2．3％的過氧化氫一甲醇孵育10分鐘，微波抗原修復； 3．3％正常山羊血清孵育，室溫下 20分鐘。 4．力人適當稀釋的一抗（1＊M－1：1：20O＊選擇素1。10o）， 4℃過夜； 5．滴加生物素標記的山羊抗兔IgG，室溫下20分鐘； 6．滴加試劑SABC，室溫孵育20分鐘； 7．DAB顯色 5則 分鐘，水洗，蘇木素復染、透明、中性樹膠 封片。以上抗體及SABC試劑盒均購自于武漢博士德生物工程有 限公司c 3．統(tǒng)計學分析 采用 SPSS for windowslo．0軟件包，應用方差分析進行統(tǒng)計 學處理。 實驗結果 ICAM－1在正常皮膚對照組的表皮層、毛囊、皮脂腺中呈淡 黃色染色；死后傷組中毛囊及皮脂腺著色較淡或不表達。P選擇 素在正常皮膚對照組及死后傷組中均未見表達。在傷后lh，即可 見大鼠損傷邊緣處表皮ICAM一互呈黃褐色染色，傷后3h組， ICAM一豆在表皮層中呈深黃褐色，尤以基底細胞及棘細胞表達為 明顯，在毛囊皮脂腺的染色亦增強，上述變化一直持續(xù)至傷后＊， 傷后3d組ICAM一二染色轉淡，呈黃褐色染色，在傷后sd及7d 組，ICAM一表達基本恢復至正常狀態(tài)。在傷后10分鐘即可見P 選擇素在皮膚組織中呈陽性表達，主要表達在創(chuàng)緣底部毛細血管 ·2· 內皮細胞及白細胞上，一直持續(xù)至傷后sh組，至傷后7h＊選擇素 已恢復至正常狀態(tài)。除此之外，ICAM一二還在炎癥細胞及纖維母 細胞中表達，且隨損傷時間變化而呈規(guī)律性變化（第1組：損傷時 間sndn 一lh，第＊組：3h一7h，第Ill 組：gb一12h，第IV組：id，第 V組：3d，第VI組：sd－7d人J［CAM一且在第二組（平均值。標準 誤：0．41 LO．73％）中陽性細胞．率極低，第＊組”．79。3．74％）及 第＊組K3．33 LI．10％）均呈顯著性增高，至第IV組門0．58。 z 65％）時達高峰，其后逐漸減少。經統(tǒng)計學方差分析，相鄰兩組 陽性細胞率均存在顯著差異，且各組與第IV組比較陽性細胞率亦 存在顯著差異。h＜0．05或0．of人 討 論 ICAM司屬于粘附分子中的免疫球蛋白超家族，為LFA－1 的配體，ICAM一二與LFA一二的相互作用可介導上皮細胞與淋巴 細胞間的粘附；介導單核巨噬細胞、中性粒細胞的粘附。為細胞粘 附所必需。選擇素P主要參與介導白細胞與內皮細胞的起始粘 附。法醫(yī)病理學上關于 ICAM·一的研究已有很多，主要集中在應 用ICAMJ判定心肌缺血及腦損傷時間可行性方面。 ICAM人在正常皮膚的表皮層、毛囊、皮脂腺中呈淡黃色染 色，由于皮膚經常接受各種外來刺激，需要相對較快的細胞更新， 而在正常皮膚中出現ICAM－l的表達，說明其在維持皮膚生物學 穩(wěn)態(tài)方面有重要作用。本實驗研究證明，皮膚損傷后表皮層 ICAM1表達增高，其表達呈現一定時間規(guī)律性，在損傷后 lh，損 傷邊緣表皮細胞即呈黃褐色染色，傷后 3 h組，ICAM一二在表皮層 中呈深黃褐色，并一直持續(xù)至傷后 id，傷后 3 d組 ICAM－1染色轉 淡，呈黃褐色染色，在傷后sd及7d組，1＊＊M一二表達基本恢復至 正常狀態(tài)。這一方面可以說明角質細胞可表達ICAM－＊另一方 ·3· 面也說明了ICAM一二在角質細胞的再生過程中有著重要作用。 有學者認為，特定粘附分子及其相應配體的表達水平和結合的親 合力是不同類型炎癥發(fā)生過程中重要的分子基礎，在炎癥起始階 段，粘附分子在炎癥因子的刺激作用下而活化，使白細胞粘附、穿 越血管內皮細胞向炎癥部位移行。我們應用免疫組化技術證明皮 膚損傷區(qū)中?
[Abstract]:Foreword









The healing of skin lesions is a very complicated biological process , which usually consists of three stages : inflammation stage , proliferation period and tissue reconstruction period . It is especially worth mentioning that adhesion molecules play a crucial role in the process of inflammation . These factors play an important role in the formation of extracellular matrix , angiogenesis , formation of granulation tissue and the formation of epithelium .









The adhesion molecule is called an adhesion molecule , because its primary function is to adhere lymphocytes , monocytes , and neutrophils to vascular endothelial cells , through the vascular endothelium , to the inflammatory reaction .









In this study , the expression of intercellular adhesion molecule ( ICAM - 1 ) and P - selectin ( P - selectin ) at different time after skin - cutting were observed by immunohistochemical method .









Materials and Methods









1 . Materials









Thirty - nine male healthy SD rats were randomly divided into 13 groups . The rats were sacrificed at 5 min to 7 d after injury . All the skin tissues were taken at the edge of the wound . Nine male healthy SD rats were randomly divided into 3 groups .



















Wax was embedded and paraffin sections were paraffin sections with a thickness of 5 m .









2 . Method









The immunohistochemistry SABC method was applied in the following steps :









removing paraffin sections from normal paraffin sections to water ;










2 . 3 % hydrogen peroxide and methanol are incubated for 10 minutes , and the microwave antigen is repaired ;










3.3 % normal goat serum was incubated at room temperature for 20 minutes .









4 . An anti - ( 1 * M - 1 : 1 : 20 O * selectin 1.10o ) diluted appropriately by a force person ,









overnight at 4 & deg ; C ;










5.dripping biotin - labeled goat anti - rabbit IgG at room temperature for 20 minutes ;










6 . dropwise adding reagent SABC and incubating at room temperature for 20 minutes ;










7 . DAB color development for 5 minutes , water washing , Su - lignin compound dyeing , transparent , neutral gum









The above antibody and SABC kit are purchased from Wuhan Docket Biological Engineering Co . , Ltd .









Limit company c









3 . Statistical Analysis









Statistical analysis was carried out by SPSS for windowsill . 0 software package .









Learn to deal with it .









experimental results









The expression of ICAM - 1 in the epidermis , hair follicle and fat gland of the normal skin control group was pale .









Yellow staining ;
The color of the hair follicle and the skin glands in the injured group were less or not expressed . P selection









In the normal skin control group and the injured group , the expression was not found in the control group and the injured group after injury .









The expression of ICAM - 1 in the epidermis of the injured rat was yellow - brown stain , 3h after injury ,









The expression of ICAM - 1 in the epidermal layer is dark brown , especially in basal cells and ratchet cells .









Obviously , the staining of the hair follicle epithelial glands is also enhanced , and the above - mentioned changes have been continued until after injury * ,









The expression of ICAM - 1 in the 3d group was pale yellow and brown , and sd and 7d after injury .









The expression of ICAM - 1 was restored to the normal state , and the expression of ICAM - 1 was observed in 10 minutes after injury .









The positive expression of selectin in skin tissue was mainly expressed at the bottom of the wound .









路 2 路



















The endothelial cells and the white blood cells were continuously injured in the post - wound sh group , and then to 7 h after injury .









It has been restored to normal state . In addition , ICAM - 1 is also in inflammatory cells and fibers .









Expression in cells and changes in regularity with time of injury ( group 1 : injury )









intersndn - 1 h , group * : 3h - 7 h , group III : gb - 12 h , group IV : id , p .









Group V : 3d , Group VI : sd - 7d Human J &bra; CAM - 1 &ket; and in the second group ( mean .









False : 0.41 LO . 73 % ) positive cells . Very low , group * " . 79 . 3.74 % ) and









There was a significant increase in Group * ( K3 . 33 LI . 10 % ) to the group IV gate 0.58 .









Up to the peak at the time of z 65 % , followed by a gradual decrease . After statistical analysis , the two groups were adjacent .









The positive rate of positive cells was significantly different , and the rate of positive cells in each group was also higher than that in group IV .









There was a significant difference . h & lt ; 0.05 or 0 . of people









discussion









The ICAM - 1 belongs to the immunoglobulin superfamily of adhesion molecules .









The interaction between the ligand , the ICAM - 1 and the lA - 2 can mediate the epithelial and lymph nodes .









intercellular adhesion ;
mediate adhesion of mononuclear phagocytes and neutrophils .









The essential part of selectin P is involved in mediating the initial viscosity of leukocytes and endothelial cells .









There are many researches on ICAM - 1 in forensic pathology , which mainly focus on the study of ICAM - 1 .









ICAMJ was used to determine the temporal feasibility of myocardial ischemia and brain injury .









The ICAM was pale yellow in the epidermis , hair follicle , and the skin of the normal skin .









color , a relatively fast cell update is required because the skin is often subjected to a variety of external stimuli ,









The expression of ICAM - 1 in normal skin indicated that it was in the maintenance of skin biology .









There is an important role in the steady - state aspect of the study . This experimental study demonstrates that skin lesions after skin damage









The expression of ICAM1 increased , and the expression of ICAM1 showed a certain time regularity . After injury , the expression of ICAM1 increased .









The epidermal cells of the wound were yellow - brown stain , 3 h after injury , ICAM - 1 in the epidermal layer .









The medium was dark brown , and persisted to the post - wound id , and ICAM - 1 in the 3 d group after injury was stained .









The expression of 1 * * M was basically recovered in the sd and 7d groups after injury .









Normal status . This aspect may indicate that keratinocytes can express ICAM - *









路 3 路



















The expression of ICAM - 2 plays an important role in the regeneration of keratinocytes .









Some scholars believe that the expression level and binding affinity of specific adhesion molecules and their corresponding ligands









The resultant force is an important molecular basis in the process of the occurrence of different types of inflammation , and in the initial stage of inflammation









The adhesion molecule is activated by the stimulation of an inflammatory factor to cause leukocyte adhesion , penetration ,









The more vascular endothelial cells migrate to the site of inflammation .









Skin damage area ?

【學位授予單位】：中國醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2003
【分類號】：D919

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本文編號：1877012