發(fā)布時間：2018-04-19 19:33

  本文選題：癌癥 + 細胞表皮間質(zhì)轉(zhuǎn)化��； 參考：《浙江大學》2017年碩士論文

【摘要】：癌癥、心血管疾病、神經(jīng)性疾病是威脅人類生命健康最主要的三大病癥。癌癥的研究是目前生物醫(yī)學領域的熱點。癌癥是由外在因素和內(nèi)在因素共同作用所引發(fā)的疾病。外在因素包括輻射、紫外線、有毒物質(zhì)、抽煙、酒精等,內(nèi)在因素包括DNA突變,染色體異常,抑癌基因失活,原癌基因異�；罨�。在內(nèi)外因素的作用下,造成細胞異常生長,在組織表面形成腫瘤。早期形成的腫瘤是良性腫瘤,發(fā)展到后期,會變成惡性腫瘤。在癌細胞轉(zhuǎn)移前期,一群腫瘤表皮細胞發(fā)生表皮間質(zhì)轉(zhuǎn)化,轉(zhuǎn)變成間充質(zhì)類細胞,這群細胞逃脫胞外基質(zhì)的限制,穿過毛細血管,并通過淋巴或血液系統(tǒng)的循環(huán)轉(zhuǎn)移到其他器官,最后少數(shù)細胞發(fā)生間質(zhì)表皮轉(zhuǎn)化,重新變成表皮細胞,在新的位置形成新的腫瘤,即轉(zhuǎn)移灶。癌細胞轉(zhuǎn)移,意味著癌癥已經(jīng)進入中后期,病人的生存時間所剩無幾。因此,表皮間質(zhì)轉(zhuǎn)化所引發(fā)的癌細胞轉(zhuǎn)移是癌癥發(fā)展過程中的重要一環(huán)。表皮間質(zhì)轉(zhuǎn)化(Epithelial-Mesenchymal Transition,EMT)是良性腫瘤向惡性癌癥發(fā)展過程中的重要步驟。目前已知導致EMT發(fā)生的重要轉(zhuǎn)錄因子包括Snail、Slug、TWIST、鋅指結構蛋白等。在EMT發(fā)生過程中,細胞失去表皮細胞的特性,獲得間充質(zhì)細胞的特性。具體表現(xiàn)為細胞形態(tài)變成梭形,細胞間結構變得松散,細胞失去極性。表皮細胞的標志蛋白如E-鈣黏素(E-cadherin)、β-連環(huán)素(β-catenin)、閉合蛋白(Occludin)等減少,間充質(zhì)細胞的標志蛋白如N-鈣黏素(N-cadherin)、波形蛋白(Vimentin)、纖連蛋白(Fibronectin)等增加。實驗室早期研究發(fā)現(xiàn)BMP(Bone morphogenetic protein)信號通路中的轉(zhuǎn)運蛋白 RanBP3L(Ran-binding Protein 3 Like)能把蛋白 Smad1/5/8 從細胞核內(nèi)轉(zhuǎn)運到細胞質(zhì)中,在此基礎上,我們準備深入研究RanBP3L的其他功能。查找文獻資料,我們發(fā)現(xiàn) RanBP3L 的類似蛋白 RanBP3(Ran-binding Protein 3)能被 Akt/PKB 磷酸化,通過序列比對,我們發(fā)現(xiàn)RanBP3L同樣具有Akt的識別的序列72RVRSSS77。實驗確認RanBP3L的S77位點能被Akt磷酸化,并且該磷酸化修飾能被磷酸酶PPM1A去磷酸化。在乳腺上皮細胞MCF10A中穩(wěn)定表達RanBP3L-WT,我們發(fā)現(xiàn)細胞形態(tài)變成梭形,細胞間結構變得松散,表現(xiàn)出間充質(zhì)細胞的特性。表達RanBP3L-S77D時,間充質(zhì)細胞的特性變得更加明顯。表達RanBP3L-S77A時,與對照組接近,仍具有表皮細胞的特性。綜上所述,我們的實驗確認了 RanBP3L的S77位點能被激酶Akt磷酸化,該磷酸化修飾能被磷酸酶PPM1A去磷酸化調(diào)控。并且發(fā)現(xiàn)RanBP3L能促進細胞發(fā)生表皮間質(zhì)轉(zhuǎn)化,其作用效果與磷酸化狀態(tài)有關。
[Abstract]:Cancer, cardiovascular disease and neuropathy are the three most important diseases threatening human life and health. The research of cancer is a hot spot in the field of biomedicine. Cancer is a disease caused by both external and internal factors. External factors include radiation, ultraviolet radiation, toxic substances, smoking, alcohol and so on. Intrinsic factors include DNA mutation, chromosomal abnormality, inactivation of tumor suppressor gene, abnormal activation of proto-oncogene and so on. Under the action of internal and external factors, the cells grow abnormally and form tumor on the tissue surface. Early tumors are benign tumors that develop into malignant tumors later in life. In the early stages of cancer cell metastasis, a group of tumor epidermal cells undergo epidermal interstitial transformation into mesenchymal cells, which escape the restriction of extracellular matrix and pass through the capillaries. Through the circulation of lymphatic or blood system to other organs, finally a few cells develop interstitial epidermis transformation, then become epidermal cells again, and form new tumor in the new position, that is, metastasis focus. Cancer cell metastasis means that cancer has entered the middle and late stages of the patient's survival time is running out. Therefore, the metastasis of cancer cells induced by epidermal interstitial transformation is an important link in the development of cancer. Epithelial-mesenchymal transition (EMTT) is an important step in the development of benign tumor to malignant cancer. The important transcription factors that have been known to cause the occurrence of EMT include Snail Slug TWIST, zinc finger structural protein and so on. During the development of EMT, the cells lost the characteristics of epidermal cells and obtained the characteristics of mesenchymal cells. Cell morphology becomes fusiform, intercellular structure becomes loose and cells lose polarity. The marker proteins of epidermal cells such as E-cadherin, 尾 -cateninine (Occludinin) were decreased, and those of mesenchymal cells such as N-cadherinine, vimentin and fibronectin were increased. In early laboratory studies, it was found that the transporter RanBP3L(Ran-binding Protein 3 in the BMP(Bone morphogenetic protein signaling pathway can transport Smad1/5/8 from the nucleus to the cytoplasm. On the basis of this, we are prepared to further study the other functions of RanBP3L. We found that RanBP3(Ran-binding Protein 3, a similar protein of RanBP3L, could be phosphorylated by Akt/PKB. By sequence alignment, we found that RanBP3L also had Akt recognized sequence 72 RVRSSS77. The S77 site of RanBP3L can be phosphorylated by Akt, and the phosphorylation modification can be dephosphorylated by phosphatase PPM1A. The stable expression of RanBP3L-WTin in MCF10A of breast epithelial cells was observed. We found that the morphology of the cells became fusiform and the intercellular structure became loose, showing the characteristics of mesenchymal cells. When RanBP3L-S77D was expressed, the characteristics of mesenchymal cells became more obvious. The expression of RanBP3L-S77A was similar to that of the control group and still had the characteristics of epidermal cells. In conclusion, our experiment confirmed that the S77 site of RanBP3L can be phosphorylated by kinase Akt, and the phosphorylation modification can be regulated by dephosphorylation of phosphatase PPM1A. It was also found that RanBP3L could promote epidermal mesenchymal transformation, and its effect was related to phosphorylation state.
【學位授予單位】：浙江大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R730.2
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本文編號：1774421