本文選題：HepaCAM + PKCε��； 參考：《重慶醫(yī)科大學(xué)》2014年碩士論文

【摘要】：第一部分HepaCAM和PKCε蛋白在腎透明細(xì)胞癌組織中的表達 目的：檢測HepaCAM和PKCε蛋白在腎透明細(xì)胞癌組織中的表達。 方法：免疫組化檢測36例人腎透明細(xì)胞癌組織HepaCAM和PKCε蛋白在癌組織和癌旁組織的表達水平，于倒置顯微鏡下成像，IPP6.0軟件分析平均光密度。 結(jié)果：腎透明細(xì)胞癌組織HepaCAM表達顯著缺失，而PKCε呈高表達狀態(tài)；癌旁組織HepaCAM表達增高，而PKCε低表達。PKCε和HepaCAM表達成反相關(guān)。其中PKCε表達與腫瘤分級分期相關(guān)。差異有統(tǒng)計學(xué)意義（p0.05）。 結(jié)論：在組織水平驗證了PKCε和HepaCAM蛋白的表達狀況和分析了二者的關(guān)系。 第二部分HepaCAM過表達影響PKCε胞漿和胞膜定位 目的：探究感染Ad-GFP-HepaCAM腺病毒后對腎癌786-O細(xì)胞株P(guān)KCε蛋白表達和定位的影響。 方法：感染Ad-GFP-HepaCAM腺病毒、Ad-GFP空載腺病毒后， western blot驗證HepaCAM的表達，檢測PKCε細(xì)胞膜、細(xì)胞漿蛋白和總蛋白的表達情況。細(xì)胞免疫熒光進一步驗證HepaCAM表達對PKCε定位的影響。 結(jié)果：實驗組HepaCAM成功表達后，PKCε細(xì)胞膜蛋白降低、漿蛋白升高，總蛋白無顯著差異。細(xì)胞免疫熒光實驗定位實驗組胞漿PKCε較空白和空載組增多（p0.05）。 結(jié)論：HepaCAM表達能影響PKCε在細(xì)胞膜和細(xì)胞漿中的定位變化，但不影響總蛋白表達。 第三部分PKCε特異性轉(zhuǎn)位抑制劑εV1-2對786-O細(xì)胞增殖和遷移的影響 目的：驗證εV1-2對786-O細(xì)胞株增殖和遷移力的影響；比較HepaCAM與εV1-2抑制克隆形成和細(xì)胞遷移的能力。 方法：應(yīng)用CCK-8實驗摸索PKCε特異性轉(zhuǎn)位抑制劑εV1-2對786-O細(xì)胞的最佳作用濃度和時間點。Western blot實驗驗證不同處理后MMP-9，cyclinD1和p-AKT蛋白表達。平板克隆和劃痕實驗驗證HepaCAM與εV1-2對增殖和遷移力的影響。 結(jié)果：10μmol/L和24h為εV1-2對786-O細(xì)胞的最佳作用濃度和時間點。Western blot證實HepaCAM表達和εV1-2處理后MMP-9，，cyclinD1和p-AKT蛋白表達降低。平板克隆和劃痕實驗顯示HepaCAM表達比εV1-2組更能有效抑制克隆形成和細(xì)胞遷移，且二者合用抑制效應(yīng)更強（p0.05）。 結(jié)論：外源性HepaCAM表達和εV1-2處理都降低了增殖和遷移相關(guān)分子的表達，HepaCAM比εV1-2處理更有效抑制克隆細(xì)胞數(shù)和細(xì)胞遷移力。HepaCAM可能作為PKCε上游分子調(diào)節(jié)腎癌786-O細(xì)胞的增殖和遷移。
[Abstract]:The expression of HepaCAM and PKC 蔚 protein in renal clear cell carcinoma Objective: to detect the expression of HepaCAM and PKC 蔚 protein in renal clear cell carcinoma. Methods: the expression levels of HepaCAM and PKC 蔚 proteins in 36 cases of clear cell carcinoma of kidney were detected by immunohistochemistry. The mean optical density was analyzed with the software of IPP6.0 under inverted microscope. Results: the expression of HepaCAM was significantly absent in renal clear cell carcinoma, but the expression of PKC 蔚 was high, the expression of HepaCAM in adjacent tissues was higher, but the expression of PKC 蔚 was lower. PKC 蔚 and HepaCAM expression were inversely correlated. The expression of PKC 蔚 was correlated with tumor grade and stage. The difference was statistically significant (P 0.05). Conclusion: the expression of PKC 蔚 and HepaCAM protein was verified and the relationship between them was analyzed at tissue level. Part two: overexpression of HepaCAM affects the localization of PKC 蔚 cytoplasm and membrane Aim: to investigate the effect of Ad-GFP-HepaCAM adenovirus infection on the expression and localization of PKC 蔚 protein in 786-O cell line. Methods: Ad-GFP-HepaCAM adenovirus was infected with Ad-GFP. Western blot was used to verify the expression of HepaCAM and to detect the expression of PKC 蔚 cell membrane, cytoplasmic protein and total protein. The effect of HepaCAM expression on the localization of PKC 蔚 was further verified by cellular immunofluorescence. Results: after the successful expression of HepaCAM in the experimental group, the membrane protein of PKC 蔚 decreased, the plasma protein increased, but there was no significant difference in total protein. The number of PKC 蔚 in cytoplasm of the experimental group was higher than that of the blank group and the no-load group by immunofluorescence assay (P 0.05). Conclusion the expression of PKC 蔚 can affect the localization of PKC 蔚 in cell membrane and cytoplasm, but it does not affect the expression of total protein. The effect of PKC 蔚 specific translocation inhibitor 蔚 V1-2 on proliferation and migration of 786-O cells Aim: to investigate the effect of 蔚 V1-2 on proliferation and migration of 786-O cell line and to compare the ability of HepaCAM and 蔚 V1-2 to inhibit clone formation and cell migration. Methods: the optimal concentration and time point of PKC 蔚 specific translocation inhibitor 蔚 V1-2 on 786-O cells were explored by CCK-8 assay. Western blot assay was used to verify the expression of MMP-9 cyclin D1 and p-AKT protein after different treatments. The effects of HepaCAM and 蔚 V1-2 on proliferation and mobility were verified by plate cloning and scratch test. Results the optimal concentration and time point of 10 渭 mol/L and 24 h for 蔚 V1-2 on 786-O cells. Western blot confirmed that the expression of HepaCAM and the expression of MMP-9 cyclin D1 and p-AKT protein decreased after treatment with 蔚 V1-2. Plate cloning and scratch assay showed that HepaCAM expression was more effective than 蔚 V1-2 in inhibiting clone formation and cell migration, and the inhibition effect was stronger than that in 蔚 V1-2 group. Conclusion: both exogenous HepaCAM expression and 蔚 V1-2 treatment can reduce the expression of proliferative and migration-related molecules. HepaCAM is more effective than 蔚 V1-2 in inhibiting the number of cloned cells and cell migration. HepaCAM may be used as upstream of PKC 蔚 to regulate the proliferation and migration of 786-O cells in renal cell carcinoma.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2014
【分類號】：R737.11

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