本文選題：SCL-1細胞 + LXRs�。� 參考：《第三軍醫(yī)大學》2017年碩士論文

【摘要】：研究背景與目的皮膚鱗狀細胞癌(squamous cell carcinoma,SCC)是一種常見的,發(fā)病率較高的皮膚角質(zhì)形成細胞來源的惡性腫瘤。SCC的病因較復雜,其發(fā)病涉及紫外線長時間照射、環(huán)境與遺傳因素、病毒感染等因素。目前的常用治療手段主要有手術(shù)切除治療。肝臟X受體(LXRs)作為轉(zhuǎn)錄因子的核激素受體,發(fā)現(xiàn)其參與了調(diào)控多種正常細胞和腫瘤細胞的增殖情況。其可降低包括血管平滑肌細胞,子宮內(nèi)膜細胞,胰島細胞,肝細胞,角質(zhì)形成細胞和淋巴細胞的增殖,還對黑素瘤、乳腺癌、肺癌、前列腺癌等多種腫瘤都有著抑制細胞增殖的作用。前期研究也已發(fā)現(xiàn)LXRs在皮膚中有廣泛表達,但目前尚未明確LXRs在皮膚鱗癌發(fā)生發(fā)展中的具體作用。Kruppel樣因子4(KLF4)作為一種目前已知的重要的蛋白質(zhì),存在于多種組織和細胞中,如廣泛表達于肺、腸、皮膚、睪丸、胸膜、心肌等器官或組織。它在體內(nèi)不但參與調(diào)節(jié)不同細胞的生長、分化以及凋亡等生物學過程,還參與了炎癥及多種腫瘤相關(guān)的疾病的發(fā)生發(fā)展過程,其在調(diào)節(jié)細胞增殖上既有抑制腫瘤的作用也有促進腫瘤發(fā)生的作用。目前研究發(fā)現(xiàn)KLF4在鱗狀細胞的增殖上是作為抑癌因子存在的,但具體作用機制仍有待進一步研究。鑒于LXRs及KLF4在皮膚鱗癌發(fā)生和發(fā)展中的可能作用,本實驗觀察了LXRs激動劑T0901317對體外人表皮鱗癌SCL-1細胞增殖和對裸鼠異體移植腫瘤生長的影響,以探討KLF4因子在LXRs途徑抑制皮膚鱗癌SCL-1細胞增殖中的作用。方法1.裸鼠皮膚鱗癌SCL-1細胞荷瘤模型構(gòu)建及腫瘤生長曲線測定:皮下注射SCL-1細胞構(gòu)建裸鼠荷瘤模型,給予T0901317試劑灌胃處理后測量實驗組及對照組異體移植腫瘤生長數(shù)據(jù),對比兩組腫瘤的生長曲線情況。2.SCL-1細胞增殖率檢測:采用CCK-8法檢測不同濃度T0901317(1、10、20μmol/L)作用SCL-1細胞24、48、72h后細胞的增殖率。3.SCL-1細胞的細胞周期檢測:用流式細胞術(shù)檢測不同濃度T0901317(1、10、20μmol/L)作用SCL-1細胞24h后的細胞周期分布情況。4.SCL-1細胞的細胞周期相關(guān)因子的m RNA表達:RT-PCR法檢測不同濃度T0901317(1、10、20μmol/L)作用SCL-1細胞24h后細胞周期相關(guān)因子PCNA、p21、p27、CCND1的基因表達變化。5.SCL-1細胞的細胞周期相關(guān)因子的蛋白表達:用Western blot檢測不同濃度T0901317(1、10、20μmol/L)作用SCL-1細胞24h后細胞周期相關(guān)因子PCNA、p21、p27、CCND1以及KLF4因子的蛋白表達變化。結(jié)果1.裸鼠荷瘤實驗組與對照組生長曲線可見較明顯差異,實驗組腫瘤體積小于對照組腫瘤體積。在體內(nèi)LXRs激動劑作用可一定程度抑制腫瘤的體積大小。2.T0901317在一定范圍內(nèi)呈時間和濃度依賴性地抑制體外培養(yǎng)的表皮鱗癌SCL-1細胞增殖。3.T0901317作用于表皮鱗癌SCL-1細胞24h后,SCL-1細胞的G1期百分比上升,而S期、G2期百分比下降。4.T0901317作用于表皮鱗癌SCL-1細胞24h后,在一定濃度范圍內(nèi)誘導了P21的表達增加,KLF4因子蛋白的表達隨藥物濃度也呈增加趨勢,PCNA的表達趨勢與P21呈相反的趨勢,而其他細胞周期相關(guān)因子表達未見明顯差異。結(jié)論LXRs激動劑可以抑制體內(nèi)及體外的SCL-1細胞的增殖,其可能的機制為抑制細胞周期的進行,即通過調(diào)節(jié)細胞周期負性調(diào)節(jié)因子p21的表達,導致細胞周期靜息甚至停滯于G1期,從而抑制細胞增殖。并且其在作用過程中,KLF4因子可能起重要的作用,但具體機制仍需要進一步的研究闡明。
[Abstract]:Background and objective squamous cell carcinoma (SCC) is a common cause of high incidence of cutaneous keratinocyte derived from the malignant tumor of.SCC, the cause of which is complicated, which involves long ultraviolet radiation, environmental and genetic factors, and disease and other factors. The main methods of treatment are mainly hands. The liver X receptor (LXRs), as a nuclear hormone receptor of the transcription factor, has been found to be involved in regulating the proliferation of various normal and tumor cells. It can reduce the proliferation of vascular smooth muscle cells, endometrium cells, islet cells, hepatocytes, keratinocytes and lymphocytes, and also to melanoma, breast cancer, and lung. A variety of cancers, such as cancer and prostate cancer, have the effect of inhibiting cell proliferation. Previous studies have also found that LXRs is widely expressed in the skin, but the specific role of LXRs in the development of squamous cell carcinoma of the skin is not yet clear,.Kruppel like factor 4 (KLF4), as an important known protein, exists in a variety of tissues and cells. It is widely expressed in lung, intestines, skin, testis, pleura, myocardium and other organs or tissues. It not only participates in the biological process of regulating the growth, differentiation and apoptosis of different cells, but also participates in the process of inflammation and the development of various tumor related diseases. It has the effect of regulating the proliferation of cells as well as inhibiting the tumor. The present study shows that KLF4 exists in the proliferation of squamous cells as tumor suppressor factors, but the specific mechanism remains to be further studied. In view of the possible role of LXRs and KLF4 in the development and development of squamous cell carcinoma of the skin, this experiment observed the proliferation of SCL-1 cells in human epidermal squamous cell carcinoma and the proliferation of LXRs agonist T0901317 in vitro. The effect of KLF4 factor on the proliferation of SCL-1 cells in skin squamous cell carcinoma by LXRs pathway in nude mice. Method 1. SCL-1 cell tumor model construction and tumor growth curve determination in nude mice skin squamous cell carcinoma: subcutaneous injection of SCL-1 cells to construct nude mice bearing tumor model, and to give T0901317 reagents after gavage treatment Test group and control group allograft tumor growth data, compared the growth curve of two groups of tumor growth curve.2.SCL-1 cell proliferation rate detection: CCK-8 method was used to detect the proliferation rate of SCL-1 cell 24,48,72h after 24,48,72h (1,10,20 mu mol/L) 24,48,72h cell proliferation rate.3.SCL-1 cell cycle detection: using flow cytometry to detect different concentrations of T09013 17 (1,10,20 mu mol/L) cell cycle distribution of SCL-1 cells 24h, m RNA expression of cell cycle related factors in.4.SCL-1 cells: RT-PCR method for detecting different concentrations of T0901317 (1,10,20 mu mol/L) action of cell cycle related factors after SCL-1 cells Protein expression of Western blot was used to detect the changes of cell cycle related factors PCNA, p21, p27, CCND1, and KLF4 factor expression after 24h (1,10,20 mu mol/L) of different concentrations of T0901317 (1,10,20 mu mol/L) in SCL-1 cells. Results the growth curve of the 1. nude mice bearing tumor experimental group was significantly different from that of the control group, and the tumor volume of the experimental group was less than that of the control group. The volume of LXRs agonists in the body inhibited the tumor size to a certain extent.2.T0901317 in a certain range of time and concentration dependent inhibition of the proliferation of epidermal squamous cell carcinoma SCL-1 cell proliferation.3.T0901317 in the epidermal squamous cell carcinoma SCL-1 cell 24h, the percentage of G1 phase in SCL-1 cells increased, while S phase, G2 phase percentage decreased.4. After the effect of T0901317 on the SCL-1 cell 24h of epidermal squamous cell carcinoma, the expression of P21 increased in a certain concentration range. The expression of KLF4 factor protein increased with the concentration of drug, and the tendency of PCNA expression was opposite to P21, but the expression of other cell cycle related factors was not significantly different. Conclusion LXRs agonist could inhibit the body. And the proliferation of SCL-1 cells in vitro, its possible mechanism is to inhibit the cell cycle, that is, by regulating the expression of the negative regulatory factor p21 in the cell cycle, resulting in the cell cycle resting even stagnating in the G1 phase, thus inhibiting the proliferation of cells. And in the process of action, the KLF4 may play an important role, but the specific mechanism still needs to be done. Further research clarifies.
【學位授予單位】：第三軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R739.5

【參考文獻】

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1 郭坤;肖生祥;耿松梅;;Klf4在銀屑病和濕疹患者皮損中的表達[J];中國麻風皮膚病雜志;2016年07期

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