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  • 學位論文

因藥物引發牙齦增生之免疫反應趨向及男性荷爾蒙接受體之免疫染色分析

Immunohistochemical analysis of cytokine profile and androgen receptor of drug induced gingival overgrowth

指導教授 : 呂炫kun 郭彥彬 傅鍔
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摘要


中文摘要 本研究目的是比較健康(H)者(PD<3mm,年齡 30~47歲)、牙周炎(P)患者(PD>5mm,年齡28~55歲)、因嚴重牙周炎需接受拔牙處置患者(S)(PD>9mm,年齡24~79歲)的牙齦組織與因尼菲迪平(Nifedipine)引發牙齦增生患者之增生牙齦(NIGO),(PD>5mm,年齡51~63歲)中的男性荷爾蒙接受體(Androgen receptor,AR)與Th1/Th2型cytokine profile:Th1-I:L-2,IFN-γ;Th2:IL-4,IL-10,IL-13的表現情形。與病人充分溝通後,分別紀錄病人的臨床牙周參數:牙周囊袋深度(probing depth),探測出血點百分比(bleeding on probing)與牙菌斑百分比。經由牙周手術取出牙齦標本病固定於緩衝弗馬林中。利用免疫染色方法LSAB® method(Dako CO.)標定出組織內的AR與Th1/Th2 cytokine。實驗結果經由顯微鏡下觀察並計算epithelial cell、inflammatory cell、gingival fibroblast AR postive cell與Th1:IL-2,IFN-γ;Th2:IL-4,IL-10,IL-13inflammatory cell呈現陽性數目之百分比,統計方法使用Kruskall-Wallis test 、Mann-Whitney U-test分析。實驗結果顯示AR 、Th1/Th2型cytokine profile:Th1:IL-2,IFN-γ;Th2:IL-4,IL-10,IL-13 positive cell在H、P、S和NIGO組表皮層內的基底層、棘皮層、結締組織內的血管內皮細胞、gingival fibroblast與inflammatory cell呈現免疫染色陽性反應。NIGO組牙齦組織內呈現AR positive 的epithelial cell與gingival fibroblast百分比(58.6+2.3;80.2+10.7)與P組(49.3+4.6;52.5+11.8)和H組(38.5+4.6;37.4+11.3)比較達到統計學上的差異(P<0.05)。就免疫反應趨向Th1/Th2 cytokine profile而言,S組的免疫反應趨向有朝向Th2(IL-4,IL-10,IL-13)方向的趨勢。NIGO組雖然Th2型(IL-4)cytokine免疫染色陽性的inflammatory cell百分比與Th1型cytokine(IL-2,IFN-γ)相近,但整體而言是有朝向Th1型cytokine的方向前進,且IL-2postive cell 的百分比與H、S、P組比較達到統計學上的差異(P<0.05)。P組因為接受過牙周基本治療,雖然有朝向Th2(IL-4,IL-10,IL-13)的方向前進,但Th1型cytokine IFN-γ免疫染色陽性之inflammatory cell百分比仍高於IL-4,IL-10,IL-13的數量並達到統計學上的差異(P<0.05)。結果顯示P組牙周組織可能往Th1 cytokine的方向前進,此點有利於組織朝向repair的階段。此外由臨床牙周參數資料顯示,隨著牙周發炎的嚴重度增加而有偏向Th2(IL-4,IL-10,IL-13)cytokine profile的可能性。 因此,我們推測AR與Th1/Th2型cytokine profile可能為牙周炎與因尼菲迪平引發牙齦增生疾病進程中,為重要致病因子,對於牙齦增生的致病機轉中有關鍵性的影響。 關鍵詞 :牙齦增生,男性荷爾蒙接受體 ,牙周病 ,免疫染色 ,免疫反應趨向(Th1/Th2 cytokine profile)。

並列摘要


Abstract The purpose of this study is try to compare the androgen receptor+ cell (AR) (﹪),Th1/Th2 cytokine profile(﹪):IL-2, IFN-γ, IL-4,IL-10,IL-13 in the patients with healthy periodontium(PD<3mm, age range30-47),inflammatory tissues of patients with adult periodontitis(P) with PD>5mm,(n=15,age range28-55),surgically extracted tooth group(S) with PD>9mm,(n=10,age range24-79)and nifedipine induced ginigval overgrowth(NIGO)with probing depth(PD)>5mm,(n=5,age range51-63).After full ethical approval of patients was achieved , the clinical periodontal parameters - pocket depth(PD), bleeding on probing(BOP), and plaque control record were measured around the selected diseasd periodontal area. Gingival samples were harvested during periodontal surgery and fixed in buffered formalin. Gingival biopsies were further processed for immunohistochemical stain with primary antibody of AR,IL-2,IL-4,IFN-γ,IL-10,IL-13 antibody(Santa Cruz biotechnology, Inc. C.A. U.S.A.)by using LSAB® method(Dako) subsequently . The expression of AR,IL-2,IL-4,IFN-γ,IL-10,IL-13 positive cells was counted using grid scan by semiquantitative method. The Kruskall—Wallis test , Mann-Whitney U-test were employed for the statistical analysis. The results revealed that AR, IL-2 ,IL-4,IFN-γ,IL-10,IL-13 was intensively expressed in the nuclei of basal epithelial cells, spinosal cells ,endothelial cell ,inflammatory cells ,and gingival fibroblasts .Strong expression of AR,IFN-γ,IL-2,IL-4 were also found in the NIGO group. Percentage (﹪)of AR labeled cells were significantly higher in the NIGO group of epithelial cells and gingival fibriblasts(58.6+2.3;80.2+10.7) than in the periodontitis group (56.3+3.3;52.5+11.8)and control(38.5+4.6;37.4+11.3)(P<0.05).In the surgically extracted tooth group ,very strong expression of Th2 (IL-4,Il-10,IL-13)type cytokine were found in the inflammatory cell than the Th1(IL-2,IFN-γ) type cytokine .The immuno expression of NIGO group has trend toward to the Th1 type cytokine(IL-2:p<0.01)expression .In the periodontitis group after periodontal phase I therapy was found strongly expression of IFN-γin the inflammatory cell. It seems that as at the quiesence stage of periodontitis , periodontium has the tendency to develope to the direction of tissue repair . Based on these findings , we postulated that AR and cytokine profile might play an important role in the progression of periodontitis and overgrowth of gingiva in NIGO. Key words: gingival overgrowth, androgen receptor, periodontitis, immunohistochemistry, cytokine profile。

參考文獻


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