【摘要】：目的本實(shí)驗(yàn)主要探討了基質(zhì)細(xì)胞衍生因子-1α(stromal cell derived factor-1α,SDF-1α)受體CXCR7(CXC-chemokine receptor 7,CXCR7)在急性單核細(xì)胞白血病(AML-M5)中的表達(dá),及其對(duì)急性單核細(xì)胞白血病細(xì)胞系THP-1細(xì)胞增殖、凋亡和侵襲等功能的影響。方法采集10例初次診斷AML-M5患者外周血和10例正常人外周血,應(yīng)用Ficoll密度梯度離心法提取外周血單個(gè)核細(xì)胞(PBMC);體外懸浮培養(yǎng)人外周血急性單核細(xì)胞白血病細(xì)胞系THP-1細(xì)胞。(1)采用流式細(xì)胞術(shù)檢測(cè)初次診斷AML-M5患者、正常人PBMC幼稚細(xì)胞表面及THP-1細(xì)胞表面CXCR7表達(dá);(2)采用Western Blot法檢測(cè)初診AML-M5患者PBMC、正常人PBMC及THP-1細(xì)胞CXCR7蛋白表達(dá)水平;(3)采用RT-PCR檢測(cè)初診AML-M5患者PBMC、正常人PBMC及THP-1細(xì)胞CXCR7 m RNA表達(dá)水平;(4)采用CCK8法檢測(cè)SDF-1α/CXCR7趨化軸對(duì)THP-1細(xì)胞體外增殖活性的作用,CXCR7單克隆抗體11G8阻斷THP-1細(xì)胞表面CXCR7表達(dá)后,觀察細(xì)胞體外增殖能力的變化;(5)采用Annexin V/PI雙染標(biāo)記法檢測(cè)CXCR7單克隆抗體對(duì)無(wú)血清培養(yǎng)條件下THP-1細(xì)胞凋亡的影響;(6)采用Transwell法觀察SDF-1α/CXCR7趨化軸對(duì)THP-1細(xì)胞體外侵襲能力的影響,CXCR7單克隆抗體阻斷細(xì)胞表面CXCR7表達(dá)后,觀察細(xì)胞侵襲能力的變化。結(jié)果(1)初次診斷AML-M5患者PBMC幼稚細(xì)胞表面CXCR7表達(dá)高于正常對(duì)照(P0.05),THP-1細(xì)胞表面CXCR7也有高表達(dá);(2)THP-1細(xì)胞、AML-M5患者PBMC中CXCR7蛋白水平明顯高于正常對(duì)照(P0.05);(3)THP-1細(xì)胞、AML-M5患者PBMC中CXCR7 m RNA水平高于正常對(duì)照(P0.05);(4)SDF-1α刺激可增高THP-1細(xì)胞增殖活性,但這種增殖活性可被CXCR7單克隆抗體抑制(P0.01);(5)CXCR7單克隆抗體不影響無(wú)血清培養(yǎng)條件下THP-1細(xì)胞的凋亡(P0.05);(6)SDF-1α/CXCR7趨化軸促進(jìn)THP-1細(xì)胞體外侵襲能力,CXCR7單克隆抗體可明顯抑制SDF-1α誘導(dǎo)的THP-1細(xì)胞侵襲(P0.01)。結(jié)論(1)CXCR7在正常對(duì)照者中幾乎不表達(dá),但在初診急性單核細(xì)胞白血病患者及急性單核細(xì)胞白血病細(xì)胞系THP-1細(xì)胞中均有高表達(dá)。(2)SDF-1α可增強(qiáng)急性單核細(xì)胞白血病細(xì)胞系THP-1細(xì)胞的增殖活性,用CXCR7單克隆抗體11G8阻斷CXCR7與SDF-1α之間相互作用可有效減弱THP-1細(xì)胞增殖活性。(3)在無(wú)血清培養(yǎng)條件下,THP-1細(xì)胞的凋亡活性不受CXCR7單克隆抗體的影響。(4)SDF-1α/CXCR7生物軸參與急性單核細(xì)胞白血病THP-1細(xì)胞的體外侵襲能力,同時(shí)應(yīng)用CXCR4和CXCR7兩種阻斷劑時(shí)能最大程度地降低SDF-1α誘導(dǎo)的THP-1細(xì)胞體外侵襲能力。
[Abstract]:Objective to investigate the expression of stromal cell derived factor-1 偽 (SDF-1 偽) receptor CXCR7 (CXC-chemokine receptor 7 CXCR7) in acute monocytic leukemia (AML-M5) and its effects on proliferation, apoptosis and invasion of THP-1 cell line. Methods Peripheral blood samples were collected from 10 patients with AML-M5 and 10 normal controls. Ficoll density gradient centrifugation was used to extract THP-1 cells from peripheral blood mononuclear cells (PBMC);) cultured in vitro. (1) flow cytometry was used to detect the initial diagnosis of AML-M5 patients. (2) the expression of CXCR7 protein in PBMC and THP-1 cells of PBMC, patients with newly diagnosed AML-M5 was detected by Western Blot assay, and (3) the PBMC and THP-1 cells CXCR7 m of PBMC and THP-1 cells of AML-M5 patients were detected by RT-PCR. (4) the effect of SDF-1 偽 / CXCR7 chemotactic axis on the proliferation of THP-1 cells in vitro was detected by CCK8 assay, and the CXCR7 expression on THP-1 cells was blocked by CXCR7 monoclonal antibody 11G8. (5) to detect the effect of CXCR7 monoclonal antibody on apoptosis of THP-1 cells in serum-free culture by Annexin V/PI double staining method, and (6) to observe the invasion ability of SDF-1 偽 / CXCR7 chemotaxis axis on THP-1 cells in vitro by Transwell assay. After CXCR7 monoclonal antibody blocked the expression of CXCR7 on cell surface, The changes of cell invasion ability were observed. Results (1) the expression of CXCR7 on the surface of immature PBMC cells in AML-M5 patients was higher than that of normal controls (P0.05), and the expression of CXCR7 on the surface of THP-1 cells was also high. (2) CXCR7 protein level in PBMC of AML-M5 patients was significantly higher than that of normal controls (P0.05); (3) THP-1 cells, CXCR7 m RNA water in AML-M5 patients PBMC. The proliferation activity of THP-1 cells was increased by SDF-1 偽 stimulation compared with normal control (P0.05); (4). But the proliferation activity was inhibited by CXCR7 monoclonal antibody (P0.01); (5) CXCR7 monoclonal antibody did not affect the apoptosis of THP-1 cells (P0.05); (6) SDF-1 偽 / CXCR7 chemotactic axis promoted the invasion ability of THP-1 cells in vitro. CXCR7 monoclonal antibody could significantly inhibit SDF-1 偽 -induced invasion of THP-1 cells (P0.01). Conclusion (1) CXCR7 was almost not expressed in normal controls. However, there was high expression of SDF-1 偽 in both newly diagnosed patients with acute monocytic leukemia and in THP-1 cell line. (2) SDF-1 偽 could enhance the proliferation activity of THP-1 cell line. Blocking the interaction between CXCR7 and SDF-1 偽 with CXCR7 monoclonal antibody 11G8 could effectively reduce the proliferation activity of THP-1 cells. (3) in serum-free culture, the apoptotic activity of THP-1 cells was not affected by CXCR7 monoclonal antibody. (4) SDF-1 偽 / CXCR7 biological axis was involved in acute mononuclear cells. The invasiveness of THP-1 cells in vitro, At the same time, CXCR4 and CXCR7 could reduce the invasiveness of THP-1 cells induced by SDF-1 偽 to the maximum extent.
【學(xué)位授予單位】：安徽醫(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)】：R733.71

【參考文獻(xiàn)】

相關(guān)期刊論文 前3條

1 傅磊;李振江;楊桂玲;鄭積富;石慶之;陳三軍;李劍;;干擾CXCR4抑制急性單核細(xì)胞白血病細(xì)胞株SHI-1黏附、侵襲及成瘤能力[J];中國(guó)實(shí)驗(yàn)血液學(xué)雜志;2015年05期

2 Yuan-Lu Huang;Xiao-Li Liu;Na Xu;Ya-Juan Xiao;Guan-Lun Gao;;Expression and function of CXCR2,CXCR7 of acute leukemic cells in rat[J];Asian Pacific Journal of Tropical Medicine;2014年05期

3 常春康;張曦;趙佑山;李曉;;CXCR4受體阻斷劑AMD3100研究進(jìn)展[J];中國(guó)實(shí)驗(yàn)血液學(xué)雜志;2011年03期

，

本文編號(hào)：2283541