舒尼替尼阻斷STAT3通路抑制頭頸癌細(xì)胞增殖的實(shí)驗(yàn)研究
發(fā)布時(shí)間：2019-05-11 06:07
【摘要】：目的:舒尼替尼是一種多靶向酪氨酸激酶抑制劑,因其可抑制與血管形成有關(guān)的多種受體酪氨酸激酶的活性,具有較好的抗血管生成作用,被應(yīng)用于治療多種腫瘤。近來(lái)研究發(fā)現(xiàn),舒尼替尼能夠誘導(dǎo)腫瘤細(xì)胞凋亡而不依賴于其抑制血管生成作用。信號(hào)轉(zhuǎn)導(dǎo)及轉(zhuǎn)錄激活因子(signal transduction and activator of transcription,STAT)家族中成員STAT3作為多種酪氨酸激酶信號(hào)通路的重要組成部分,其活化后在包括頭頸癌等多種惡性腫瘤細(xì)胞中表達(dá)較高。STAT3異常活化并持續(xù)高表達(dá)與這些惡性腫瘤凋亡的抑制、腫瘤細(xì)胞增殖、腫瘤微環(huán)境血管形成及轉(zhuǎn)移關(guān)系密切。舒尼替尼能否誘導(dǎo)頭頸癌細(xì)胞凋亡,STAT3在此過(guò)程中是否發(fā)揮作用,目前相關(guān)報(bào)道較少。本實(shí)驗(yàn)通過(guò)使用舒尼替尼分別處理UM-22B和SCC90兩種頭頸癌細(xì)胞系,觀察腫瘤細(xì)胞增殖和凋亡情況,以及STAT1和STAT3的變化,從而進(jìn)一步了解舒尼替尼對(duì)頭頸癌的作用及機(jī)制,為舒尼替尼在臨床治療頭頸癌提供理論基礎(chǔ)。方法:體外培養(yǎng)的SCC90和UM-22B細(xì)胞,經(jīng)過(guò)不同濃度舒尼替尼處理不同時(shí)間后,分別采用MTT試驗(yàn)、倒置顯微鏡觀察等方法,檢測(cè)細(xì)胞增殖抑制情況;并且采用流式細(xì)胞術(shù)檢測(cè)STAT1和STAT3磷酸化水平。結(jié)果:1舒尼替尼對(duì)兩種細(xì)胞增殖的影響:MTT顯示,舒尼替尼對(duì)SCC90細(xì)胞增殖的抑制率最高可達(dá)92.89%,24,48,72h的IC50分別是2.37、1.17和0.93μmol/L;對(duì)UM-22B細(xì)胞抑制率最高可達(dá)82.56%,24,48h的IC50分別是6.30和2.31μmol/L。用藥組不同濃度不同處理時(shí)間其生長(zhǎng)抑制率有顯著差異,抑制率的時(shí)間-濃度依賴性明顯;2形態(tài)學(xué)變化:倒置顯微鏡下,SCC90、UM-22B細(xì)胞均顯示:對(duì)照組細(xì)胞生長(zhǎng)較用藥組迅速,細(xì)胞貼壁生長(zhǎng)呈“鋪路石”狀,細(xì)胞呈扁平狀多角形,胞質(zhì)飽滿,胞質(zhì)近中央部位有圓形細(xì)胞核;用藥組細(xì)胞增殖受到抑制,貼壁細(xì)胞體積變小而呈圓形,連接松解,核顏色較對(duì)照組加深;3舒尼替尼對(duì)STAT1和STAT3磷酸化水平的影響:SCC90細(xì)胞經(jīng)過(guò)0.25,0.5,1μmol/L的舒尼替尼處理2,8,16,24小時(shí)后,各實(shí)驗(yàn)組中p-STAT1陽(yáng)性表達(dá)與對(duì)照組相比,無(wú)統(tǒng)計(jì)學(xué)差異;而p-STAT3陽(yáng)性表達(dá)與對(duì)照組相比明顯降低,并且隨著濃度增高,陽(yáng)性率顯著下降,差異有統(tǒng)計(jì)學(xué)意義。UM-22B細(xì)胞經(jīng)過(guò)0.5,1,2,4μmol/L的舒尼替尼處理30分鐘和2小時(shí)后,各實(shí)驗(yàn)組中p-STAT3陽(yáng)性表達(dá)與對(duì)照組相比降低,并且隨著濃度增高,陽(yáng)性率顯著下降。其中,兩個(gè)時(shí)間段中,0.5和1μmol/L組與對(duì)照組相比無(wú)統(tǒng)計(jì)學(xué)差異,而2和4μmol/L組有顯著性差異。結(jié)論:1舒尼替尼可以抑制頭頸鱗癌UM-22B和SCC90細(xì)胞的增殖。2舒尼替尼可以下調(diào)UM-22B和SCC90細(xì)胞STAT3磷酸化水平。3 STAT1在舒尼替尼抑制頭頸癌細(xì)胞增殖過(guò)程中磷酸化水平未見(jiàn)明顯升高。
[Abstract]:Aim: Schnitinib is a multi-target tyrosine kinase inhibitor. It can inhibit the activity of many receptor tyrosine kinases related to angiogenesis and has a good anti-angiogenesis effect. It has been used in the treatment of many kinds of tumors. Recently, it has been found that Schnitinib can induce apoptosis of tumor cells, independent of its inhibitory effect on angiogenesis. STAT3, a member of the signal transduction and activator of transcription (signal transduction and activator of transcription,STAT) family, is an important component of many tyrosine kinase signaling pathways. The abnormal activation and persistent high expression of STAT3 in various malignant tumor cells, including head and neck cancer, were closely related to the inhibition of apoptosis, proliferation of tumor cells and vascular formation and metastasis in tumor microenvironment. There are few reports on whether Schnitinib can induce apoptosis of head and neck cancer cells and whether STAT3 plays a role in this process. In order to further understand the effect and mechanism of Schnitinib on head and neck carcinoma, the proliferation and apoptosis of UM-22B and SCC90 cell lines, and the changes of STAT1 and STAT3 were observed by the treatment of shunitinib, respectively, in order to understand the effect and mechanism of Schnitinib on head and neck cancer. It provides a theoretical basis for the clinical treatment of head and neck cancer with sulnitinib. Methods: SCC90 and UM-22B cells cultured in vitro were treated with different concentrations of sulnitinib for different time. MTT test and inverted microscope were used to detect the inhibition of cell proliferation. The phosphorylation levels of STAT1 and STAT3 were detected by flow cytometry. Results: 1 the effect of sulnitinib on the proliferation of two kinds of SCC90 cells: MTT showed that the inhibition rate of Schnitinib on the proliferation of SCC90 cells reached 92.89%, 24,48 and 2.37 渭 mol / L, 1.17 渭 mol / L and 0.93 渭 mol / L, respectively. The highest inhibition rate of UM-22B cells was 82.56%, 24 h and 48 h IC50 were 6.30 and 2.31 渭 mol / L, respectively. In the treatment group, the growth inhibition rate was significantly different in different concentration and treatment time, and the time-concentration dependence of inhibition rate was obvious. 2 morphological changes: under inverted microscope, SCC90,UM-22B cells in the control group grew faster than those in the treatment group, the adherent growth of the cells was "paving stone", the cells were flat and polyangular, and the cytoplasm was full. Round nucleus was found near the center of cytoplasm. The proliferation of the cells in the treatment group was inhibited, the volume of the adherent cells became small and round, the junction was loosened, and the color of the nucleus was deeper than that of the control group. (3) the effect of sulnitinib on the phosphorylation of STAT1 and STAT3: there was no significant difference in the positive expression of p-STAT1 between the SCC90 cells and the control group 24 hours after treatment with 0.25 渭 mol / L, 0.51 渭 mol / L and 1 渭 mol / L sulnitinib for 2, 8, 16, 24 hours, and there was no significant difference in the positive expression of p-STAT1 between the experimental groups and the control group. The positive expression of p-STAT3 was significantly lower than that of the control group, and the positive rate decreased significantly with the increase of concentration. UM-22B cells were treated with 0.5, 1, 2, 4 渭 mol / L sulnitinib for 30 minutes and 2 hours. The positive expression of p-STAT3 in each experimental group was lower than that in the control group, and the positive rate decreased significantly with the increase of concentration. Among them, there was no significant difference between 0.5 渭 mol / L group and 1 渭 mol / L group and 4 渭 mol / L group, but there was significant difference between 2 渭 mol / L group and 4 渭 mol / L group. Conclusion: 1 Surnitinib can inhibit the proliferation of UM-22B and SCC90 cells in head and neck squamous cell carcinoma. 2 Surnitinib can down-regulate the STAT3 phosphorylation level of UM-22B and SCC90 cells. 3 STAT1 can inhibit the proliferation of head and neck cancer cells during the proliferation of head and neck cancer cells. There was no significant increase in acidizing level.
【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R739.91
本文編號(hào)：2474300
[Abstract]:Aim: Schnitinib is a multi-target tyrosine kinase inhibitor. It can inhibit the activity of many receptor tyrosine kinases related to angiogenesis and has a good anti-angiogenesis effect. It has been used in the treatment of many kinds of tumors. Recently, it has been found that Schnitinib can induce apoptosis of tumor cells, independent of its inhibitory effect on angiogenesis. STAT3, a member of the signal transduction and activator of transcription (signal transduction and activator of transcription,STAT) family, is an important component of many tyrosine kinase signaling pathways. The abnormal activation and persistent high expression of STAT3 in various malignant tumor cells, including head and neck cancer, were closely related to the inhibition of apoptosis, proliferation of tumor cells and vascular formation and metastasis in tumor microenvironment. There are few reports on whether Schnitinib can induce apoptosis of head and neck cancer cells and whether STAT3 plays a role in this process. In order to further understand the effect and mechanism of Schnitinib on head and neck carcinoma, the proliferation and apoptosis of UM-22B and SCC90 cell lines, and the changes of STAT1 and STAT3 were observed by the treatment of shunitinib, respectively, in order to understand the effect and mechanism of Schnitinib on head and neck cancer. It provides a theoretical basis for the clinical treatment of head and neck cancer with sulnitinib. Methods: SCC90 and UM-22B cells cultured in vitro were treated with different concentrations of sulnitinib for different time. MTT test and inverted microscope were used to detect the inhibition of cell proliferation. The phosphorylation levels of STAT1 and STAT3 were detected by flow cytometry. Results: 1 the effect of sulnitinib on the proliferation of two kinds of SCC90 cells: MTT showed that the inhibition rate of Schnitinib on the proliferation of SCC90 cells reached 92.89%, 24,48 and 2.37 渭 mol / L, 1.17 渭 mol / L and 0.93 渭 mol / L, respectively. The highest inhibition rate of UM-22B cells was 82.56%, 24 h and 48 h IC50 were 6.30 and 2.31 渭 mol / L, respectively. In the treatment group, the growth inhibition rate was significantly different in different concentration and treatment time, and the time-concentration dependence of inhibition rate was obvious. 2 morphological changes: under inverted microscope, SCC90,UM-22B cells in the control group grew faster than those in the treatment group, the adherent growth of the cells was "paving stone", the cells were flat and polyangular, and the cytoplasm was full. Round nucleus was found near the center of cytoplasm. The proliferation of the cells in the treatment group was inhibited, the volume of the adherent cells became small and round, the junction was loosened, and the color of the nucleus was deeper than that of the control group. (3) the effect of sulnitinib on the phosphorylation of STAT1 and STAT3: there was no significant difference in the positive expression of p-STAT1 between the SCC90 cells and the control group 24 hours after treatment with 0.25 渭 mol / L, 0.51 渭 mol / L and 1 渭 mol / L sulnitinib for 2, 8, 16, 24 hours, and there was no significant difference in the positive expression of p-STAT1 between the experimental groups and the control group. The positive expression of p-STAT3 was significantly lower than that of the control group, and the positive rate decreased significantly with the increase of concentration. UM-22B cells were treated with 0.5, 1, 2, 4 渭 mol / L sulnitinib for 30 minutes and 2 hours. The positive expression of p-STAT3 in each experimental group was lower than that in the control group, and the positive rate decreased significantly with the increase of concentration. Among them, there was no significant difference between 0.5 渭 mol / L group and 1 渭 mol / L group and 4 渭 mol / L group, but there was significant difference between 2 渭 mol / L group and 4 渭 mol / L group. Conclusion: 1 Surnitinib can inhibit the proliferation of UM-22B and SCC90 cells in head and neck squamous cell carcinoma. 2 Surnitinib can down-regulate the STAT3 phosphorylation level of UM-22B and SCC90 cells. 3 STAT1 can inhibit the proliferation of head and neck cancer cells during the proliferation of head and neck cancer cells. There was no significant increase in acidizing level.
【學(xué)位授予單位】：河北醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R739.91
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