本文選題：TYK2　切入點：P53　出處：《大連醫(yī)科大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：研究背景:三陰性乳腺癌(Triple-negative breast cancer,TNBC)不表達(dá)雌激素受體(ER)、孕激素受體(PR)和缺乏人類表皮生長因子受體2(HER2),這類腫瘤具有侵襲性表型和預(yù)后較差的特點,雖然部分患者對化療敏感,但很快出現(xiàn)化療抵抗。因此,該類型乳腺癌缺乏具體的針對性治療方法。無受體酪氨酸激酶2(Non-receptor tyrosine kinase 2,TYK2)是JAK家族的一員,參與多種細(xì)胞因子信號通路,和人類癌癥的發(fā)生關(guān)系密切。既往研究表明TYK2在三陰性乳腺癌細(xì)胞系中過表達(dá),但TYK2在參與三陰性乳腺癌發(fā)生、發(fā)展中涉及的相關(guān)分子機制尚不明確。人類大多數(shù)腫瘤與P53基因變異有關(guān),野生型P53是調(diào)節(jié)細(xì)胞周期進(jìn)程的腫瘤抑制基因,在DNA損傷修復(fù),細(xì)胞衰老,細(xì)胞凋亡中起重要作用。而突變型P53(mtP53)能夠表現(xiàn)出促進(jìn)腫瘤細(xì)胞的遷移、侵襲以及抗凋亡等惡性生物學(xué)行為。研究表明P53在三陰性乳腺癌MDA-MB-231細(xì)胞株中呈突變表達(dá),且突變型P53和三陰性乳腺癌較差臨床預(yù)后相關(guān)。近些年來,關(guān)于P53與JAK家族的多個成員相互作用關(guān)系的研究較多,但是探討TYK2與突變型P53關(guān)系的相關(guān)報道較少。研究目的:明確TYK2對乳腺癌MDA-MB-231細(xì)胞株增殖、遷移、凋亡等生物學(xué)行為的影響。進(jìn)一步明確在三陰性乳腺癌中TYK2和mtP53的相互作用。為三陰性乳腺癌的治療提供新的思路及實驗依據(jù)。方法:(1)利用siRNA瞬時轉(zhuǎn)染技術(shù),將TYK2-siRNA及P53-siRNA分別轉(zhuǎn)染至三陰性乳腺癌細(xì)胞株MDA-MB-231中,同時將Negative control轉(zhuǎn)染至MDA-MB-231細(xì)胞中作為陰性對照。采用Western blot檢測三組細(xì)胞中TYK2、P53的蛋白表達(dá)情況。(2)采用MTT方法檢測轉(zhuǎn)染TYK2-siRNA和(或)P53-siRNA后MDA-MB-231細(xì)胞的增殖情況。(3)采用劃痕實驗方法檢測TYK2及P53基因沉默后,乳腺癌MDA-MB-231細(xì)胞的遷移情況。(4)通過流式細(xì)胞儀對轉(zhuǎn)染siRNA的細(xì)胞和NC組的細(xì)胞進(jìn)行凋亡檢測。結(jié)果:1.Western blot結(jié)果提示與NC組比較,在乳腺癌MDA-MB-231細(xì)胞中干擾P53基因后,P53基因被沉默,TYK2表達(dá)也下調(diào)。干擾TYK2基因后,TYK2基因被沉默,P53基因表達(dá)也隨之下調(diào)。2.MTT結(jié)果提示:與NC組比較,聯(lián)合或者單轉(zhuǎn)染siTYK2、siP53后,乳腺癌MDA-MB-231細(xì)胞的增殖率均下降。3.劃痕實驗示:聯(lián)合轉(zhuǎn)染組及單轉(zhuǎn)染siTYK2組和siP53組均較NC組細(xì)胞遷移能力下降。但聯(lián)合組較單轉(zhuǎn)染組遷移能力未見明顯升高。4.流式細(xì)胞術(shù)示:聯(lián)合或者分別轉(zhuǎn)染siTYK2、siP53后,對比NC組,乳腺癌MDA-MB-231細(xì)胞凋亡率升高。結(jié)論:1.沉默TYK2基因的表達(dá)可以抑制乳腺癌MBA-MB-231細(xì)胞的P53基因的表達(dá),而沉默P53基因?qū)θ橄侔㎝BA-MB-231細(xì)胞的TYK2基因的表達(dá)也有抑制作用,提示TYK2和mtpP53在三陰性乳腺癌的發(fā)生和發(fā)展中具有一定的關(guān)聯(lián)作用,兩者可以相互通過某種信號通路或某種基因直接或間接地調(diào)控基因的轉(zhuǎn)錄和翻譯。2.沉默TYK2基因能夠?qū)е氯橄侔㎝DA-MB-231細(xì)胞株凋亡的增加,增殖及遷移能力降低,提示TYK2基因能夠通過某種機制影響MDA-MB-231細(xì)胞株的增殖、遷移及凋亡等生物學(xué)行為。3.沉默mtP53能夠?qū)е氯橄侔㎝DA-MB-231細(xì)胞株凋亡的增加,增殖及遷移能力降低,在三陰性乳腺癌的發(fā)展中起著重要作用。
[Abstract]:Background: three negative breast cancer (Triple-negative breast, cancer, TNBC) is the expression of estrogen receptor (ER), progesterone receptor (PR) and the lack of human epidermal growth factor receptor 2 (HER2), which has the characteristics of tumor invasive phenotype and poor prognosis, although part of the patients are sensitive to chemotherapy, but soon resistance to chemotherapy. Therefore, this type of breast cancer is lack of specific targeted therapies. Non receptor tyrosine kinase 2 (Non-receptor tyrosine 2 kinase, TYK2) is a member of the JAK family, is involved in a variety of cytokine signaling pathways, and human cancer are closely related. Previous study showed that overexpression of TYK2 in three negative breast cancer cells lines, but TYK2 in three negative breast cancer, the molecular mechanism involved in the development is not clear. The most human tumors with P53 gene mutation, wild type P53 is the regulation of cell cycle progression is swollen Tumor suppressor gene, DNA repair, cell senescence, play an important role in apoptosis. Mutant P53 (mtP53) can show to promote tumor cell migration, invasion and apoptosis of malignant biological behavior. The research indicated that the P53 was expressed in three mutation negative breast cancer cell line MDA-MB-231, and mutant P53 and poor clinical outcome three negative breast cancer. In recent years, more research about relationship between multiple members of the JAK family and P53 interaction, but related to the reported TYK2 and mutant P53 relationship less. Objective: To explore the TYK2 on the proliferation and migration of breast cancer cell line MDA-MB-231, the influence of biological behavior of apoptosis. To further clarify the interaction of TYK2 in three negative breast cancer and mtP53. Provide new ideas and experimental evidence for the treatment of three negative breast cancer. Methods: (1) using siRNA to instantaneous staining technique, TYK2-s IRNA and P53-siRNA were transfected with three negative breast cancer cell line MDA-MB-231, and will be used as negative control Negative control was transfected into MDA-MB-231 cells. Using Western blot detection of three groups of TYK2 cells, the expression of P53 protein. (2) transfection and TYK2-siRNA were detected by MTT method (or) the proliferation of MDA-MB-231 cells after P53-siRNA. (3) detected by silencing TYK2 and P53 gene scratch test method, the migration of breast cancer MDA-MB-231 cells. (4) were detected by cell apoptosis and NC group of transfected siRNA cells by flow cytometry. Results: 1.Western blot showed that compared with group NC, the interference of P53 gene in breast cancer MDA-MB-231 cells, P53 genes are silenced, TYK2 expression was down regulated. The interference of TYK2 gene, TYK2 gene was silenced, the expression of P53 gene with the down-regulation of.2.MTT results showed that compared with the NC group, the joint or single turn With siTYK2, siP53,.3. were decreased in wound healing rate of MDA-MB-231 breast cancer cell proliferation: the co transfection group and siTYK2 transfection group and siP53 group were lower than NC group cell migration ability decreased. But the combined group than single transfection group there was no significant increase in the migration ability of.4. flow cytometry showed that the transfection of siTYK2 or combined siP53, NC, contrast group, breast cancer MDA-MB-231 cell apoptosis rate increased. Conclusion: the expression of P53 1. gene expression silencing TYK2 gene can inhibit breast cancer MBA-MB-231 cells, and the expression of TYK2 gene silencing P53 gene on MBA-MB-231 breast cancer cells also inhibited, suggesting that TYK2 and mtpP53 have a certain correlation function in the occurrence and development of three negative breast cancer, both can be through the transcription and translation of.2. gene silence of TYK2 gene regulation a signaling pathway or directly or indirectly can lead to milk Increased apoptosis of adenocarcinoma MDA-MB-231 cells, decreased proliferation and migration, suggesting that the TYK2 gene can affect MDA-MB-231 through a mechanism of cell proliferation, migration and apoptosis of.3. and the biological behavior of mtP53 silencing can lead to increased apoptosis of breast cancer MDA-MB-231 cells, reduce the proliferation and migration ability, plays an important role in the development of the three negative breast cancer.

【學(xué)位授予單位】：大連醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R737.9

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相關(guān)期刊論文 前2條

1 Budhi S Yadav;Priyanka Chanana;Swaty Jhamb;;Biomarkers in triple negative breast cancer:A review[J];World Journal of Clinical Oncology;2015年06期

2 Hanan Ahmed Wahba;Hend Ahmed El-Hadaad;;Current approaches in treatment of triple-negative breast cancer[J];Cancer Biology & Medicine;2015年02期

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本文編號：1577195