【摘要】：每年,在中國宮頸癌的新發(fā)病例高達10萬之多,并且呈現(xiàn)年輕化趨勢。目前,手術及放療是治療宮頸鱗狀細胞癌的有效方法,但其造成的生理功能損害及器官缺失無法逆轉,因此尋求一種破壞性小,副作用小的療法極為迫切。靶向治療,是在細胞分子水平上,針對已經明確的致癌位點特異性殺傷目的細胞而不波及到正常組織。因此,追求精準和高效是未來治療宮頸癌的方向。自噬,存在于正常細胞中的一種生理功能,已發(fā)現(xiàn)其過度激活可以造成類似于凋亡的程序性死亡。因此,自噬相關基因可能成為治療惡性腫瘤的潛在靶點,即通過調節(jié)腫瘤細胞中的自噬水平,誘導目的細胞死亡,可謂癌癥治療新的曙光。在自噬相關基因中,BECN1作為自噬程序的啟動子,正向調節(jié)自噬功能;mTOR作為逆向調控的樞紐,抑制細胞自噬行為;LC3B是自噬體膜性結構的標記者,其表達水平反映自噬活性的強弱。根據(jù)2014年第4版WHO女性生殖系統(tǒng)腫瘤分類。本實驗以LSIL、HSIL、宮頸鱗狀細胞癌為研究對象;以正常宮頸組織為對照。運用免疫組織化學Super Vision法及FISH技術,縱向檢測自噬相關基因在宮頸病變中的mRNA及蛋白表達程度;正、反橫向探究BECN1、LC3B和mTOR在逐級宮頸病變中的表達差異。以期全方位評估自噬活性與宮頸病變之間的相關性。研究發(fā)現(xiàn),BECN1在LSIL、HSIL及宮頸鱗狀細胞癌中的蛋白表達分別是21.46±19.48、12.93±11.67、6.28±5.89,而正常宮頸組織中為36.66±23.28。提示:隨著宮頸病變的進展,BECN1的表達呈現(xiàn)下調趨勢,且均低于正常宮頸組織的表達水平。其各組間差異具有統(tǒng)計學意義(P0.05)。mRNA的表達也呈現(xiàn)相同趨勢。LC3B在LSIL、HSIL及宮頸鱗狀細胞癌中的蛋白表達分別是22.63±15.09、12.61±8.96、6.99±5.71,在正常宮頸中的表達為34.84±15.91。提示:LC3B在宮頸病變中的表達均低于正常宮頸,并且隨著宮頸病變的進展,表達水平越低,各組間差異具有統(tǒng)計學意義(P0.05)。LC3B的mRNA表達也呈現(xiàn)類似趨勢。mTOR在HSIL及癌組中的蛋白表達分別是24.02±17.48、35.48±19.65,顯著高于正常宮頸組(6.9±6.69),而LSIL中的蛋白表達(12.02±9.48)與正常宮頸相比差異無統(tǒng)計學意義(P0.05)。提示:隨著宮頸病變的進展,mTOR的蛋白表達水平越高,其mRNA的表達趨勢與之相似。經統(tǒng)計學分析,BECN1的表達與淋巴結轉移及腫瘤臨床分期有關,即在有淋巴結轉移的患者中呈低表達,且隨著臨床分期越高,表達越少;LC3B與腫瘤分化程度相關,在中低分化的患者中呈低表達;而mTOR與有無淋巴結轉移相關,且在有淋巴結轉移的患者中呈現(xiàn)明顯高表達。另外在宮頸鱗狀細胞癌組中,LC3B與BECN1的表達呈正相關(r=0.642,P0.01),與mTOR的表達呈負相關(r=-0.418,P0.01);但BECN1與mTOR的表達無明顯相關性。綜上所述,根據(jù)BECN1、LC3B和mTOR在宮頸病變及正常宮頸中的表達情況,反映自噬活性隨著病變的進展而下調,并且在HSIL及宮頸癌組中呈明顯的自噬缺陷。這可能提示,低水平的自噬活性可能是影響宮頸病變進展的因素之一。同時,BECN1、LC3B和mTOR的mRNA與蛋白表達結果趨勢一致,表明可能在遺傳物質轉錄、翻譯階段,已有自噬關鍵基因的相互作用以及干擾。另外,自噬相關基因的表達與患者有無淋巴轉移,腫瘤細胞分化程度及臨床分期有關,推測自噬活性可能影響宮頸癌患者的預后。
[Abstract]:Nowadays, surgery and radiotherapy are effective methods for the treatment of cervical squamous cell carcinoma, but the damage of physiological function and organ loss can not be reversed. Therefore, it is very urgent to seek a treatment with less destructive and side effects. Autophagy, a physiological function of normal cells, has been found to cause apoptosis-like programmed death by over-activation. Thus, autophagy-related genes may become potential targets for the treatment of malignant tumors, that is, by regulating the level of autophagy in tumor cells and inducing the death of target cells, which is a new dawn for cancer treatment. LC3B is a marker of autophagy membrane structure, and its expression level reflects the intensity of autophagy activity. According to the WHO classification of female reproductive system tumors in the 4th edition of 2014, LSIL, HSIL, cervical squamous cell carcinoma were studied in this experiment, and normal cervical tissues were taken as control. Longitudinal detection of autophagy-related genes in cervical lesions mRNA and protein expression; positive and negative cross-sectional study of BECN1, LC3B and mTOR in progressive cervical lesions in order to fully assess the correlation between autophagy and cervical lesions. The expression of BECN 1 was down-regulated with the progression of cervical lesions, and was lower than that of normal cervical tissues. The difference was statistically significant (P 0.05). The expression of mRNA also showed the same trend. LC3B was found in LSIL, HSIL and uterus. The expression of LC3B protein in cervical squamous cell carcinoma was 22.63 (+ 15.09), 12.61 (+ 8.96), 6.99 (+ 5.71) and 34.84 (+ 15.91) respectively in normal cervix. The expression of mTOR protein in HSIL and cancer group was 24.02 [17.48] and 35.48 [19.65] respectively, which was significantly higher than that in normal cervix group (6.9 [6.69]). However, the expression of mTOR protein in LSIL was not significantly different from that in normal cervix (P 0.05). Statistical analysis showed that the expression of BECN1 was related to lymph node metastasis and clinical stage, that is, the expression of BECN1 was low in patients with lymph node metastasis, and the higher the clinical stage, the less the expression of BECN1; LC3B was related to the degree of tumor differentiation, but the expression of mTOR was low in patients with moderate and low differentiation, and the expression of mTOR was related to lymph node metastasis. In addition, the expression of LC3B was positively correlated with BECN1 (r = 0.642, P 0.01) and negatively correlated with the expression of mTOR (r = - 0.418, P 0.01), but there was no significant correlation between BECN1 and mTOR. The expression of BECN1, LC3B and mTOR in normal cervix reflects that autophagy activity is down-regulated with the progression of cervical lesions and shows obvious autophagy deficiency in HSIL and cervical cancer. This may suggest that low level of autophagy activity may be one of the factors affecting the progression of cervical lesions. In addition, the expression of autophagy-related genes is related to lymphatic metastasis, tumor cell differentiation and clinical stage. It is speculated that autophagy activity may affect the prognosis of cervical cancer patients.
【學位授予單位】：河北北方學院
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R737.33

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