【摘要】：目的：探討Ras基因家族(H-ras/N-ras/K-ras)蛋白表達(dá)、基因突變?cè)谄つw病理性瘢痕和瘢痕癌中的意義。 方法：以16例皮膚病理性瘢痕、20例皮膚瘢痕癌組織為研究對(duì)象,以10例正常皮膚組織為對(duì)照。采用免疫組織化學(xué)(SP)法分別檢測(cè)H-ras、N-ras、K-ras蛋白的表達(dá),結(jié)合圖像分析,分別觀測(cè)三組被檢組織中所檢各項(xiàng)指標(biāo)的表達(dá)(平均光密度和陽性面積)。另外對(duì)所選病例的石蠟包埋組織(包括14例瘢痕,14例瘢痕癌)提取DNA,進(jìn)一步做PCR擴(kuò)增及測(cè)序,分析H-ras、N-ras、K-ras基因的第12、13位密碼子點(diǎn)突變情況。所有數(shù)據(jù)運(yùn)用SPSS16.0軟件包進(jìn)行統(tǒng)計(jì)學(xué)分析。 結(jié)果：(1) H-ras、N-ras、K-ras蛋白在正常皮膚表皮和皮膚病理性瘢痕上皮中呈陰性或弱陽性表達(dá),正常皮膚組與瘢痕組比較,差異均無統(tǒng)計(jì)學(xué)意義(P0.05),但瘢痕上皮中陽性表達(dá)細(xì)胞數(shù)量較正常皮膚表皮增多。(2) H-ras、N-ras、K-ras蛋白在瘢痕癌組織中呈強(qiáng)陽性表達(dá),瘢痕癌組的表達(dá)(平均光密度和陽性面積)與正常皮膚組及皮膚瘢痕組比較,差異均有統(tǒng)計(jì)學(xué)意義(P0.05)。(3)在瘢痕癌中,不管是組織學(xué)類型還是細(xì)胞的分化,均顯示分化越高Ras蛋白表達(dá)越強(qiáng),分化越低Ras蛋白表達(dá)越弱,高分化組與低分化組比較,差異均有統(tǒng)計(jì)學(xué)意義(P0.05)。(4)K-ras陽性信號(hào)在正常皮膚組主要定位于細(xì)胞核,在瘢痕組主要定位于細(xì)胞核和細(xì)胞質(zhì),在瘢痕癌組為細(xì)胞質(zhì)的表達(dá),出現(xiàn)自胞核向胞質(zhì)逐漸移位的現(xiàn)象。(5)Ras基因家族點(diǎn)突變分析：從石蠟包埋組織(28例樣本)中提取DNA,并經(jīng)PCR反應(yīng)后,均成功擴(kuò)增出目的片段。經(jīng)測(cè)序后,未發(fā)現(xiàn)H-ras、N-ras、K-ras基因第12、13位密碼子的點(diǎn)突變。 結(jié)論：(1)在皮膚病理性瘢痕上皮中H-ras、N-ras、K-ras蛋白限制性的低表達(dá),可能與皮膚瘢痕上皮增生具有一定的相關(guān)性。(2)瘢痕癌與正常皮膚和皮膚病理性瘢痕相比較,H-ras、N-ras、K-ras蛋白的高表達(dá),可能與皮膚瘢痕癌的發(fā)生有關(guān)。(3)H-ras、N-ras、K-ras蛋白與腫瘤細(xì)胞的分化有相關(guān)性,可能在瘢痕癌演進(jìn)過程中逐漸丟失。(4)K-ras蛋白的移位現(xiàn)象,可能與瘢痕癌的發(fā)生有相關(guān)性。(5)瘢痕癌中尚未發(fā)現(xiàn)Ras基因家族12、13位密碼子的突變,其突變位點(diǎn)有待進(jìn)一步探討。
[Abstract]:Objective: to investigate the significance of Ras gene family (H-ras/N-ras/K-ras) protein expression and gene mutation in pathological scar and scar carcinoma of skin. Methods: 16 cases of skin pathological scar, 20 cases of skin scar carcinoma and 10 cases of normal skin tissue were studied. Immunohistochemical (SP) method was used to detect the expression of H-rasn- N-rasna-K-ras protein. Combined with image analysis, the expression of each index (average optical density and positive area) was observed in the three groups. In addition, DNA, was extracted from paraffin-embedded tissues (including 14 scars and 14 scar carcinomas) from selected cases. Further PCR amplification and sequencing were performed to analyze the point mutation at codon 1213 of H-rasn N-rascino K-ras gene. All the data were analyzed by SPSS16.0 software package. Results: (1) the expression of H-rasln N-rasna-K-ras protein was negative or weakly positive in normal skin epidermis and skin pathological scar epithelium, but there was no significant difference between normal skin group and scar group (P0.05). But the number of positive cells in scar epithelium was higher than that in normal skin epidermis. The expression (mean optical density and positive area) of scar carcinoma group was significantly different from that of normal skin group and skin scar group (P0.05). (3), whether histological type or cell differentiation. The higher the differentiation, the stronger the expression of Ras protein, the lower the expression of Ras protein. The difference was statistically significant (P0.05). (4) K-ras positive signal was mainly located in nucleus in normal skin group, mainly in nucleus and cytoplasm in scar group, and in cytoplasm in scar carcinoma group. (5) Ras gene family point mutation analysis: DNA, was extracted from paraffin embedded tissue (28 samples) and amplified successfully after PCR reaction. After sequencing, there was no point mutation at codon 1213 of H-rasln N-rasn K-ras gene. Conclusion: (1) the low expression of H-rasln N-rascino K-ras protein in the skin pathological scar epithelium. (2) compared with normal skin and skin pathological scar, the expression of H-rasN rasn K-ras protein in scar carcinoma is higher than that in normal skin and skin pathological scar. It may be related to the development of skin scar carcinoma. (3) H-rasN-rascinia K-ras protein is associated with the differentiation of tumor cells, and may be lost gradually during the progression of scar carcinoma. (4) the translocation of K-ras protein. (5) the mutation of Ras gene family at codon 1213 has not been found in scar carcinomas, and its mutation site needs to be further studied.
【學(xué)位授予單位】：遵義醫(yī)學(xué)院
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2011
【分類號(hào)】：R739.5

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