本文選題：膠質(zhì)瘤 + 血管擬態(tài)　； 參考：《山東大學(xué)》2017年博士論文

【摘要】：研究背景腦膠質(zhì)瘤(Glioma)是中樞神經(jīng)系統(tǒng)中最常見的顱內(nèi)原發(fā)性惡性腫瘤,約占腦內(nèi)原發(fā)腫瘤的50%,其中以高度惡性的膠質(zhì)母細(xì)胞瘤(Glioblastomamultiforme,GBM;WHO Ⅳ級(jí))最為常見。近幾年來,雖然臨床上手術(shù)聯(lián)合放化療等綜合治療手段,在一定程度上改善了惡性膠質(zhì)瘤的治療療效,但膠質(zhì)母細(xì)胞瘤患者的中位總生存期仍然僅有15-18個(gè)月,具有預(yù)后差、復(fù)發(fā)率高的特點(diǎn)。由于膠質(zhì)瘤是一種高度血管化的腦腫瘤,因此,靶向血管內(nèi)皮細(xì)胞的抗血管生成藥物治療膠質(zhì)瘤的應(yīng)用前景備受矚目。然而,近期的數(shù)據(jù)發(fā)現(xiàn),這種看似非常具有潛力的抗血管療法,臨床應(yīng)用后卻出現(xiàn)了療效短暫和療效欠佳的問題。直到1999年血管擬態(tài)(Vasculogenic Mimicry，，VM)概念的提出,人們才意識(shí)到在腫瘤組織中除了由內(nèi)皮細(xì)胞構(gòu)成的血管外,還有其他的血液供應(yīng)形式。血管擬態(tài)是由腫瘤細(xì)胞在特定情況下模擬血管內(nèi)皮細(xì)胞形成的一種功能上類似于正常血管的管道,其內(nèi)可含有紅細(xì)胞,并且可以與常規(guī)的內(nèi)皮血管相通連,構(gòu)成網(wǎng)狀的血液輸送循環(huán),從而為腫瘤提供必要的營養(yǎng)支持。目前已經(jīng)證實(shí),包括膠質(zhì)瘤在內(nèi)的多種惡性腫瘤中均存在血管擬態(tài),并且其陽性率與腫瘤級(jí)別及患者的不良預(yù)后密切相關(guān)。因而,靶向血管擬態(tài)的治療策略很可能是一種非常有前景的抗腫瘤血管治療手段,所以,在對(duì)膠質(zhì)瘤患者進(jìn)行傳統(tǒng)抗血管治療的同時(shí),聯(lián)合應(yīng)用抑制腫瘤血管擬態(tài)形成的藥物可能會(huì)有更好的治療療效。已經(jīng)有越來越多的研究表明,腫瘤微環(huán)境在血管擬態(tài)形成過程中具有不可或缺的作用,但具體分子機(jī)制尚未明確,深入研究腫瘤微環(huán)境將會(huì)促進(jìn)人們對(duì)血管擬態(tài)相關(guān)機(jī)制的了解。免疫細(xì)胞浸潤(rùn)是多種腫瘤微環(huán)境的一個(gè)顯著特征,在腫瘤的侵襲、血管新生以及轉(zhuǎn)移等惡性生物學(xué)行為中發(fā)揮著關(guān)鍵作用,進(jìn)而影響了腫瘤的進(jìn)展。在這群免疫細(xì)胞中巨噬細(xì)胞的浸潤(rùn)比率最高,研究發(fā)現(xiàn),膠質(zhì)母細(xì)胞瘤中浸潤(rùn)的腫瘤相關(guān)巨噬細(xì)胞比例高達(dá)30%,且高表達(dá)CD14、CD68、CD163和CD204等M2型巨噬細(xì)胞標(biāo)志物。既往研究發(fā)現(xiàn),腫瘤局部浸潤(rùn)巨噬細(xì)胞的數(shù)量及活性與患者的預(yù)后密切相關(guān)。此外,有報(bào)道證實(shí),在既往接受抗血管治療后病情復(fù)發(fā)的膠質(zhì)母細(xì)胞瘤患者中,瘤體內(nèi)浸潤(rùn)巨噬細(xì)胞的數(shù)量明顯增加,并且與腫瘤的血管密度呈正相關(guān)。這些結(jié)果表明,在腫瘤進(jìn)展過程中,腫瘤浸潤(rùn)巨噬細(xì)胞對(duì)腫瘤血供的形成發(fā)揮著不容忽視的作用。因此,靶向腫瘤浸潤(rùn)巨噬細(xì)胞的促血管生成作用,也許是克服膠質(zhì)瘤抗血管藥物抵抗的有效途徑。腫瘤微環(huán)境的另一個(gè)重要特征是其內(nèi)存在大量獨(dú)特的免疫介質(zhì),包括細(xì)胞因子以及趨化因子。越來越多的數(shù)據(jù)表明,這類免疫介質(zhì)可以通過調(diào)節(jié)免疫細(xì)胞和腫瘤細(xì)胞的功能,營造出一個(gè)有助于腫瘤進(jìn)展的微環(huán)境。白細(xì)胞介素-6(Interleukin-6,IL-6)就是這樣一種功能多效的免疫介質(zhì),其廣泛存在于多種惡性實(shí)體瘤的炎性微環(huán)境中,通過參與免疫調(diào)節(jié)、促進(jìn)腫瘤細(xì)胞增殖、遷移、存活以及藥物耐藥,引發(fā)腫瘤患者的不良預(yù)后。此外,還有結(jié)果顯示,IL-6與腫瘤抗血管治療效果不佳有著密不可分的關(guān)系。在膠質(zhì)瘤中,IL-6可以通過增強(qiáng)血管內(nèi)皮細(xì)胞的遷移來促進(jìn)腫瘤的侵襲和血管生成,阻斷IL-6同時(shí)合用貝伐單抗可以更有效的阻礙膠質(zhì)瘤的侵襲和生長(zhǎng)。這些結(jié)果表明,腫瘤微環(huán)境中IL-6過表達(dá)會(huì)明顯影響腫瘤的血管生成,但I(xiàn)L-6是否在膠質(zhì)瘤血管擬態(tài)形成過程中同樣扮演著重要的角色,目前仍不清楚。本課題擬在已有研究的基礎(chǔ)上,利用免疫組化的方法初步探索血管擬態(tài)與腫瘤相關(guān)巨噬細(xì)胞在人膠質(zhì)瘤組織中的表達(dá)特點(diǎn)、臨床意義及二者的相關(guān)性。通過體外實(shí)驗(yàn),分析THP-1衍生M2巨噬細(xì)胞和外周血單核衍生M2巨噬細(xì)胞是否會(huì)影響膠質(zhì)瘤血管擬態(tài)的形成,并探討其相關(guān)的作用機(jī)制。第一部分血管擬態(tài)和CD163+巨噬細(xì)胞在膠質(zhì)瘤中的表達(dá)目的檢測(cè)血管擬態(tài)和CD163+巨噬細(xì)胞在膠質(zhì)瘤組織中的表達(dá)情況,并初步分析兩者之間是否存在相關(guān)性,以及其與患者預(yù)后和臨床病理因素之間的關(guān)系。方法1.本研究回顧性納入山東大學(xué)齊魯醫(yī)院行膠質(zhì)瘤切除術(shù)的患者87例,獲取腫瘤組織樣本。采用免疫組織化學(xué)方法檢測(cè)血管擬態(tài)和CD163+巨噬細(xì)胞在膠質(zhì)瘤中的表達(dá)情況,并分析兩者之間的相關(guān)性。2.收集患者的基本信息,包括性別、年齡等基本臨床資料,分析血管擬態(tài)和CD163+巨噬細(xì)胞與膠質(zhì)瘤患者臨床病理因素及預(yù)后的關(guān)系。結(jié)果1.通過免疫組化對(duì)膠質(zhì)瘤組織樣本的連續(xù)切片進(jìn)行CD31/PAS和CD163染色發(fā)現(xiàn),高級(jí)別膠質(zhì)瘤(WHO分級(jí)Ⅲ-Ⅳ級(jí))的血管擬態(tài)(CD31-/PAS+)水平和CD163表達(dá)密度明顯高于低級(jí)別的(WHO分級(jí)Ⅰ-Ⅱ級(jí))(P0.01,P0.01)。相關(guān)性分析結(jié)果顯示,血管擬態(tài)水平與CD163表達(dá)密度在膠質(zhì)瘤組織中呈正相關(guān)(r2=0.416,P0.001)。2.分析膠質(zhì)瘤組織中血管擬態(tài)和CD163的表達(dá)與不同病理因素之間的關(guān)系發(fā)現(xiàn),二者均與腫瘤的級(jí)別有關(guān)(P0.001,P0.001),血管擬態(tài)還與腫瘤的大小有關(guān)(P=0.027);二者與性別、年齡、腫瘤位置、KPS評(píng)分以及腫瘤的生長(zhǎng)方式無關(guān)。3.Kaplan-Meier分析發(fā)現(xiàn),與CD163表達(dá)水平較高的膠質(zhì)瘤患者相比,CD163表達(dá)水平較低的膠質(zhì)瘤患者具有顯著的生存優(yōu)勢(shì)(p=0.006);血管擬態(tài)低水平的膠質(zhì)瘤患者其中位生存時(shí)間明顯長(zhǎng)于血管擬態(tài)高水平的膠質(zhì)瘤患者(P0.001)。進(jìn)一步分析發(fā)現(xiàn),CD163高表達(dá)同時(shí)血管擬態(tài)高水平膠質(zhì)瘤患者的中位生存時(shí)間明顯短于其他組的膠質(zhì)瘤患者(P0.001)。結(jié)論1.血管擬態(tài)水平和CD163+巨噬細(xì)胞浸潤(rùn)數(shù)量均與膠質(zhì)瘤級(jí)別有關(guān),二者在膠質(zhì)瘤組織中的表達(dá)呈正相關(guān)。2.CD163+巨噬細(xì)胞浸潤(rùn)和血管擬態(tài)存在與膠質(zhì)瘤患者的不良預(yù)后有關(guān)。3.血管擬態(tài)水平和CD163+巨噬細(xì)胞浸潤(rùn)數(shù)量與膠質(zhì)瘤患者的臨床病理特征,如性別、年齡、腫瘤位置、KPS評(píng)分以及腫瘤的生長(zhǎng)方式等無關(guān)。第二部分THP-1衍生M2巨噬細(xì)胞調(diào)控膠質(zhì)瘤血管擬態(tài)及機(jī)制研究目的探索THP-1衍生M2巨噬細(xì)胞是否影響膠質(zhì)瘤血管擬態(tài)的形成及其可能的機(jī)制方法1.采用qRT-PCR方法檢測(cè)THP-1衍生巨噬細(xì)胞的表型。2.利用成管實(shí)驗(yàn)檢測(cè)THP-1衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞血管擬態(tài)形成的影響。3.采用qRT-PCR、Western blotting等方法檢測(cè)THP-1衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞血管擬態(tài)相關(guān)標(biāo)志物轉(zhuǎn)錄和蛋白水平的影響。4.采用懸浮芯片和ELISA等技術(shù)檢測(cè)THP-1衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞IL-6分泌的影響。5.利用qRT-PCR實(shí)驗(yàn)技術(shù)檢測(cè)THP-1衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞IL-6和血管擬態(tài)相關(guān)指標(biāo)表達(dá)時(shí)序的影響。6.采用成管實(shí)驗(yàn)、Western blotting等方法檢測(cè)IL-6是否影響膠質(zhì)瘤血管擬態(tài)的形成。7.利用成管實(shí)驗(yàn)、qRT-PCR、ELISA及Western blotting等方法探索THP-1衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞相關(guān)信號(hào)機(jī)制的影響。結(jié)果1.THP-1衍生M2巨噬細(xì)胞表達(dá)高水平的M2型標(biāo)記物(CD163,CD206,IL-10,IL-1RA 和 CCL17)和低水平的 M1 型標(biāo)記物(TNF-α,IL-1β 和 IL-12)。2.與THP-1上清處理組或?qū)φ战M相比,THP-1衍生M2巨噬細(xì)胞的上清明顯促進(jìn)膠質(zhì)瘤細(xì)胞血管擬態(tài)的形成(U251,P0.001;A172,P0.01)和血管擬態(tài)相關(guān)指標(biāo)(MMP-9,MMP-14,LAMC2)的表達(dá)。3.THP-1衍生M2巨噬細(xì)胞上清促進(jìn)膠質(zhì)瘤細(xì)胞IL-6的分泌,并呈現(xiàn)時(shí)間和濃度依賴的特點(diǎn),且IL-6表達(dá)上調(diào)時(shí)間(約6h)要早于血管擬態(tài)相關(guān)指標(biāo)表達(dá)升高的時(shí)間(約12h)。4.IL-6中和抗體(1 μg/ml)可以阻斷THP-1衍生M2巨噬細(xì)胞誘導(dǎo)的膠質(zhì)瘤細(xì)胞血管擬態(tài)形成(U251,P0.01;A172,P0.01)和相關(guān)指標(biāo)表達(dá)的上調(diào)。此外,重組人IL-6(rhIL-6,100ng/ml)能夠促進(jìn)膠質(zhì)瘤細(xì)胞血管擬態(tài)的形成(U251,P0.01;A172,P0.05)和相關(guān)指標(biāo)的表達(dá)。5.THP-1衍生M2巨噬細(xì)胞上清升高膠質(zhì)瘤細(xì)胞PKC信號(hào)通路的磷酸化水平。PKC抑制劑(Bisindolylmaleimide I)可以抑制THP-1衍生M2巨噬細(xì)胞誘導(dǎo)的膠質(zhì)瘤細(xì)胞IL-6的轉(zhuǎn)錄(U251,P0.01;A172,P0.01)和分泌(U251,P0.001;A172,P0.001)水平的上調(diào),進(jìn)而抑制膠質(zhì)瘤細(xì)胞血管擬態(tài)的形成(U251,P0.05;A172,P0.05)和相關(guān)指標(biāo)的表達(dá)。結(jié)論1.THP-1衍生M2巨噬細(xì)胞體外促進(jìn)膠質(zhì)瘤細(xì)胞血管擬態(tài)的形成。2.THP-1衍生M2巨噬細(xì)胞通過激活膠質(zhì)瘤細(xì)胞PKC信號(hào)通路促進(jìn)IL-6的產(chǎn)生,進(jìn)而促進(jìn)血管擬態(tài)的形成。第三部分單核細(xì)胞衍生M2巨噬細(xì)胞調(diào)控膠質(zhì)瘤血管擬態(tài)及機(jī)制研究目的探索人外周血單核細(xì)胞衍生M2巨噬細(xì)胞是否影響膠質(zhì)瘤細(xì)胞血管擬態(tài)的形成及其作用機(jī)制方法1.利用磁珠分選法從人外周血分離CD14+單核細(xì)胞,并用相關(guān)誘導(dǎo)因子將其培養(yǎng)成巨噬細(xì)胞。2.采用qRT-PCR和ELISA技術(shù)檢測(cè)人外周血單核細(xì)胞衍生巨噬細(xì)胞的表型。3.利用qRT-PCR和ELISA實(shí)驗(yàn)方法分析人外周血單核細(xì)胞衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞IL-6分泌的影響。4.采用成管實(shí)驗(yàn)檢測(cè)人外周血單核細(xì)胞衍生M2巨噬細(xì)胞對(duì)膠質(zhì)瘤細(xì)胞血管擬態(tài)形成的影響。5.采用成管實(shí)驗(yàn)檢測(cè)膠質(zhì)瘤細(xì)胞分泌的IL-6對(duì)血管擬態(tài)形成的影響。結(jié)果1.人外周血單核細(xì)胞衍生M2巨噬細(xì)胞表達(dá)高轉(zhuǎn)錄水平的M2型標(biāo)記物(CD163,CD206,CCL18和CCL17)和低轉(zhuǎn)錄水平的M1型標(biāo)記物(TNF-α,IL-1β,IL-6和IL-12)。單核細(xì)胞衍生M2巨噬細(xì)胞分泌高水平的M2型標(biāo)記物(CCL22,CCL18,CCL17 和 IL-10)和低水平的 M1 型標(biāo)記物(IL1-β,IL-6,IL-12 和 IL-8)。2.人外周血單核細(xì)胞衍生M2巨噬細(xì)胞的上清上調(diào)膠質(zhì)瘤細(xì)胞IL-6的轉(zhuǎn)錄(U251,P0.01;A172,P0.001)和分泌(U251,P0.001;A172,P0.001)水平。3.將IL-6中和抗體(1μg/ml)加入共培養(yǎng)體系可以顯著降低膠質(zhì)瘤細(xì)胞管狀結(jié)構(gòu)的形成數(shù)量(U251,P0.01;A172,P0.01)。這表明IL-6中和抗體可以阻斷人外周血單核細(xì)胞衍生M2巨噬細(xì)胞介導(dǎo)的膠質(zhì)瘤細(xì)胞血管擬態(tài)形成的增強(qiáng)作用。結(jié)論1.人外周血單核細(xì)胞衍生M2巨噬細(xì)胞升高膠質(zhì)瘤細(xì)胞IL-6的表達(dá)。2.人外周血單核細(xì)胞衍生M2巨噬細(xì)胞通過增加膠質(zhì)瘤細(xì)胞IL-6的分泌,進(jìn)一步促進(jìn)膠質(zhì)瘤血管擬態(tài)的形成。
[Abstract]:Background brain glioma (Glioma) is the most common primary intracranial malignant tumor in the central nervous system, accounting for about 50% of the primary brain tumors, and highly malignant glioblastoma (Glioblastomamultiforme, GBM; WHO IV) is the most common. The therapeutic effect of malignant glioma is improved to a certain extent, but the median survival period of the patients with glioblastoma is still only 15-18 months, with poor prognosis and high recurrence rate. As glioma is a highly vascularized brain tumor, the application prospect of antiangiogenic drugs targeting vascular endothelial cells in the treatment of glioma Recent data, however, have found that this seemingly very promising antiangio therapy has a short and poor curative effect after clinical applications. It was not until 1999 that the concept of Vasculogenic Mimicry (VM) was put forward, that people realized that in tumor tissue, the blood vessels made up of endothelial cells were found in the tumor tissue. There are other forms of blood supply. Vascular mimicry is a function of a tumor cell that mimics the formation of vascular endothelial cells in a specific condition, a function similar to the normal blood vessel, which contains red cells, and can connect with the conventional endothelial blood vessels to form a reticular circulation of blood, thus providing the necessary camp for the tumor. Support. It has been proved that vascular mimicry exists in various malignant tumors, including glioma, and the positive rate is closely related to the tumor grade and the poor prognosis of the patients. Therefore, the targeted vascular mimicry therapy strategy is likely to be a very promising method of anti swelling tumor vascular treatment, therefore, in the patients with glioma. While traditional antivascular therapy is carried out, the combined application of drugs to inhibit the formation of tumor vasculature may have a better therapeutic effect. More and more studies have shown that the tumor microenvironment plays an indispensable role in the formation of vascular mimicry, but the specific molecular mechanism has not yet been clearly defined and the tumor microenvironment will be deeply studied. Promote people to understand the mechanism related to vascular mimicry. Immune cell infiltration is a significant feature of various tumor microenvironments. It plays a key role in tumor invasion, angiogenesis, metastasis and other malignant biological behaviors, and then affects the progress of the tumor. It is found that the proportion of macrophages infiltrated in glioblastoma is up to 30%, and high expression of the M2 macrophage markers such as CD14, CD68, CD163 and CD204. Previous studies have found that the number and activity of local infiltrating macrophages in the tumor are closely related to the prognosis of the patients. In patients with recurrent glioblastoma, the number of infiltrating macrophages in the tumor increases significantly and is positively correlated with the tumor's blood vessel density. These results suggest that tumor infiltrating macrophages play an unneglecting role in the formation of tumor blood supply during the progression of the tumor. Therefore, the tumor infiltrates macrophages. Angiogenesis may be an effective way to overcome the antiangiogenic resistance of gliomas. Another important feature of the tumor microenvironment is the existence of a large number of unique immune mediators, including cytokines and chemokines. More and more data have shown that these immune mediators can regulate the work of immune cells and tumor cells. -6 (Interleukin-6, IL-6) is a kind of multifunctional immune medium, which is widely used in the inflammatory microenvironment of various malignant solid tumors. By participating in immunoregulation, it promotes tumor cell proliferation, migration, survival and drug resistance and causes cancer patients. In addition, the results show that IL-6 is inextricably related to the poor effect of tumor antiangiogenic therapy. In gliomas, IL-6 can promote tumor invasion and angiogenesis by enhancing the migration of vascular endothelial cells, blocking IL-6 and combining bavavic can more effectively impede the invasion and birth of glioma. These results suggest that the overexpression of IL-6 in the tumor microenvironment will significantly affect the angiogenesis of tumor, but it is still not clear whether IL-6 plays an important role in the formation of glioma angiogenesis. On the basis of the existing research, we should explore the relationship between vascular mimicry and tumor preliminarily by using immunohistochemical method. The expression of macrophages in human glioma tissue, clinical significance and the correlation between the two. Through in vitro experiments, it is analyzed whether THP-1 derived M2 macrophages and peripheral blood mononuclear derived M2 macrophages will affect the formation of glioma vascular mimicry, and explore its related mechanisms. Expression in glioma to detect the expression of vascular mimicry and CD163+ macrophage in glioma tissue, and to analyze the relationship between the two, and the relationship with the prognosis and the clinicopathological factors of the patients. Methods 1. the study was reviewed in the Qilu Hospital of Shandong University. The tumor tissue samples were obtained in 87 cases. Immunohistochemical method was used to detect the expression of vascular mimicry and CD163+ macrophage in glioma, and the correlation between the two.2. was analyzed and the basic information of the patients was collected, including basic clinical data such as sex and age, analysis of vascular mimicry and the presence of CD163+ macrophages and glioma patients. The relationship between pathological factors and prognosis. Results 1. CD31/PAS and CD163 staining showed that the level of vascular mimicry (CD31-/PAS+) and CD163 expression in high grade glioma (WHO grade III - IV) were significantly higher than those of lower grade (WHO grade I - II) (P0.01, P0.01) by immunohistochemical staining. The results showed that the level of vascular mimicry and the expression density of CD163 were positively correlated with the glioma tissue (r2=0.416, P0.001).2. analysis of the relationship between the vascular mimicry and the expression of CD163 in glioma tissue and the different pathological factors. The two were related to the level of the tumor (P0.001, P0.001), and the vascular mimicry was also related to the size of the tumor (P=0.027). .3.Kaplan-Meier analysis of sex, age, tumor location, KPS score, and tumor growth patterns found that the two had significant survival advantages compared to those with a higher level of CD163 expression in glioma patients with lower CD163 expression (p=0.006); the survival time of the patients with a low level of vascular mimicry was in the patients' survival time. Further analysis showed that the median survival time of patients with high expression of CD163 and high level glioma of vascular mimicry was significantly shorter than those of other groups of glioma (P0.001). Conclusion 1. the level of vascular mimicry and the number of macrophage infiltration in CD163+ are related to the grade of glioma, two The expression of.2.CD163+ macrophage infiltration and vascular mimicry in glioma tissues is associated with poor prognosis of patients with glioma, the level of.3. vascular mimicry and the number of CD163+ macrophages and the clinicopathological features of glioma patients, such as sex, age, tumor location, KPS score, and tumor growth patterns. Guan. Second part of THP-1 derived M2 macrophages regulating glioma vascular mimicry and mechanism study to explore whether THP-1 derived M2 macrophages affect the formation of glioma vascular mimicry and its possible mechanism method. 1. qRT-PCR method was used to detect the phenotype of THP-1 derived macrophages for the detection of THP-1 derived M2 macrophage fines Effects of cell on angiogenesis in glioma cells.3. using qRT-PCR, Western blotting and other methods to detect the effect of THP-1 derived M2 macrophages on the transcriptional and protein levels of vascular mimicry related markers in glioma cells.4. uses suspension chips and ELISA techniques to detect the IL-6 secretion of THP-1 derived M2 macrophages to glioma cells. The influence of THP-1 derived M2 macrophages on the expression time sequence of IL-6 and vascular mimicry in glioma cells by using qRT-PCR test technique,.6. was used as a tube test, Western blotting and other methods were used to detect the formation of.7. in the angiogenesis of glioma,.7., qRT-PCR, ELISA, and.5. The effect of THP-1 derived M2 macrophages on the related signal mechanism of glioma cells. Results 1.THP-1 derived M2 macrophages expressed high level M2 markers (CD163, CD206, IL-10, IL-1RA and CCL17) and low level M1 markers. The supernatant of the cell obviously promotes the formation of vascular mimicry in glioma cells (U251, P0.001; A172, P0.01) and vascular mimicry (MMP-9, MMP-14, LAMC2) expression of.3.THP-1 derived M2 macrophage supernatant to promote the secretion of IL-6 in glioma cells, which is characterized by time and concentration dependence, and IL-6 expression (approximately 6h) is earlier than that of vascular mimicry. The increased time (about 12h).4.IL-6 neutralizing antibody (1 mu g/ml) can block the expression of vascular mimicry induced by THP-1 derived M2 macrophages (U251, P0.01; A172, P0.01) and the up-regulated expression of related indexes. In addition, recombinant human IL-6 (rhIL-6100ng/ml) can promote the formation of vascular mimicry in glioma cells (U251, The expression of.01, A172, P0.05) and related indexes,.5.THP-1 derived M2 macrophage supernatant, the phosphorylation level.PKC inhibitor (Bisindolylmaleimide I) of glioma cell PKC signaling pathway can inhibit the transcription of glioma cells induced by THP-1 derived M2 macrophages. The expression of vascular mimicry in glioma cells (U251, P0.05; A172, P0.05) and the expression of related indexes. Conclusion 1.THP-1 derived M2 macrophages promote the formation of vascular mimicry of glioma cells in vitro, and the formation of.2.THP-1 derived M2 macrophages promotes the production of IL-6 by activating the glioma microcell PKC signaling pathway and then promotes the angiogenesis. Formation. Third partial monocyte derived M2 macrophages regulate glioma vascular mimicry and mechanism study to explore whether human peripheral blood mononuclear cells derived M2 macrophages affect the formation and mechanism of glioma vascular mimicry and mechanism method 1. separation of CD14+ mononuclear cells from human peripheral blood by magnetic bead separation, and the use of magnetic beads to separate CD14+ mononuclear cells from human peripheral blood .2. and ELISA techniques were used to detect the phenotypic.3. of human peripheral blood monocyte derived macrophages using qRT-PCR and ELISA techniques. The effects of M2 macrophages derived from human peripheral blood mononuclear cells on the secretion of IL-6 in human glioma cells were analyzed by qRT-PCR and ELISA methods. Effect of nuclear cell derived M2 macrophages on angiogenesis in glioma cells.5. use tube test to detect the effect of IL-6 on angiogenesis in glioma cells. Results 1. human peripheral blood monocyte derived M2 macrophages expressed high transcriptional level M2 markers (CD163, CD206, CCL18 and CCL17) and M of low transcriptional level. Type 1 markers (TNF-, IL-1 beta, IL-6, and IL-12). High level M2 markers secreted by monocyte derived M2 macrophages (CCL22, CCL18, CCL17 and IL-10) and low level M1 type markers 2, P0.001) and secreting (U251, P0.001; A172, P0.001) level.3. added IL-6 neutralization antibody (1 mu) to co culture system to significantly reduce the number of tubular structure of glioma cells (U251, P0.01; A172, P0.01). Conclusion: 1. peripheral blood monocyte derived M2 macrophages increase the expression of IL-6 in human glioma cells.2. human peripheral blood.

【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2017
【分類號(hào)】：R739.41

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

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本文編號(hào)：1853606