本文選題：唑來(lái)膦酸納米粒　切入點(diǎn)：M2型巨噬細(xì)胞　出處：《西南醫(yī)科大學(xué)》2017年碩士論文

【摘要】：目的:將U937細(xì)胞誘導(dǎo)為M0巨噬細(xì)胞、M1型巨噬細(xì)胞、M2型巨噬細(xì)胞,了解不同類(lèi)型巨噬細(xì)胞對(duì)唑來(lái)膦酸納米粒的攝取情況;了解唑來(lái)膦酸納米粒對(duì)共培養(yǎng)情況下腫瘤細(xì)胞的遷移、侵襲的影響;唑來(lái)膦酸納米粒對(duì)共培養(yǎng)情況下腫瘤細(xì)胞的增殖抑制情況。方法:1、巨噬細(xì)胞誘導(dǎo):將U937細(xì)胞平鋪至六孔板中,加入佛波酯(PMA)誘導(dǎo)6個(gè)小時(shí);2、M1型巨噬細(xì)胞誘導(dǎo):將U937細(xì)胞平鋪至六孔板中,加入佛波酯(MPA)(200ng/ml)誘導(dǎo)6個(gè)小時(shí)后再加入脂多糖(LPS)(100ng/ml)和IFN-γ(20ng/ml)繼續(xù)誘導(dǎo)66小時(shí)(共72小時(shí))。3、M2型巨噬細(xì)胞誘導(dǎo):將U937細(xì)胞平鋪至六孔板中,加入佛波酯(MPA)(200ng/ml)誘導(dǎo)6個(gè)小時(shí)后再加入IL-4(20ng/ml)和IL-13(20ng/ml)繼續(xù)誘導(dǎo)66小時(shí)(共72小時(shí))。并通過(guò)檢測(cè)Arg-1酶及iNOS的表達(dá)情況,檢測(cè)是否誘導(dǎo)成功。4、攝取實(shí)驗(yàn):選取人體成纖維細(xì)胞、M1型巨噬細(xì)胞、M2巨噬細(xì)胞、SW620結(jié)腸癌細(xì)胞,加入熒光標(biāo)記后的唑來(lái)膦酸納米粒,反應(yīng)3小時(shí)后,采用Dil對(duì)細(xì)胞膜進(jìn)行染色,在熒光顯微鏡下觀察四種細(xì)胞對(duì)唑來(lái)膦酸納米粒的攝取情況。5、CCK-8增殖抑制實(shí)驗(yàn):通過(guò)唑來(lái)膦酸納米粒與不同種類(lèi)細(xì)胞的相互作用,了解其對(duì)細(xì)胞的增殖能力的影響;6、細(xì)胞劃痕實(shí)驗(yàn)及transwell小室遷移實(shí)驗(yàn),設(shè)置sw620結(jié)腸癌組、sw620+m1型巨噬細(xì)胞組、sw620+m2型巨噬細(xì)胞組,每組分為加和不加唑來(lái)膦酸納米粒組(20ng/ul),共6組實(shí)驗(yàn)組,觀察每組各因素對(duì)腫瘤細(xì)胞遷移能力的影響。7、侵襲實(shí)驗(yàn):將sw620結(jié)腸癌細(xì)胞放入transwell小室的上室中,下室依次為空白組(單獨(dú)培養(yǎng)基)、唑來(lái)膦酸納米粒組、m1型巨噬細(xì)胞組、m1型巨噬細(xì)胞+唑來(lái)膦酸納米粒組、m2型巨噬細(xì)胞組、m2型巨噬細(xì)胞+唑來(lái)膦酸納米粒組;并采用吉姆薩染色法計(jì)數(shù),了解各因素對(duì)腫瘤細(xì)胞侵襲能力的影響;結(jié)果:1、arg-1及inos檢測(cè)結(jié)果顯示m2型巨噬細(xì)胞免疫組化呈陽(yáng)性反應(yīng),而其余兩組呈陰性反應(yīng),m1型巨噬細(xì)胞inos表達(dá)(74.4%)遠(yuǎn)高于其余兩組(14.0%及14.7%),證實(shí)m1及m2型巨噬細(xì)胞誘導(dǎo)成功。2、m2型巨噬細(xì)胞攝取納米顆粒的熒光強(qiáng)度遠(yuǎn)高于其他細(xì)胞;3、m1型巨噬細(xì)胞抑制腫瘤細(xì)胞遷移;m2型巨噬細(xì)胞促進(jìn)腫瘤細(xì)胞的遷移,與唑來(lái)膦酸納米粒相互作用可抑制m2巨噬細(xì)胞的這種促進(jìn)作用(p0.01);4、m1型巨噬細(xì)胞抑制腫瘤細(xì)胞侵襲力;m2型巨噬細(xì)胞促進(jìn)腫瘤細(xì)胞的侵襲,與唑來(lái)膦酸納米粒相互作用可抑制m2巨噬細(xì)胞的這種促進(jìn)作用(p0.01)5、納米顆粒通過(guò)抑制m2型巨噬細(xì)胞,影響腫瘤細(xì)胞增殖(p0.01)。結(jié)論:唑來(lái)膦酸納米粒能特異性被m2型巨噬細(xì)胞攝取并抑制m2巨噬細(xì)胞的增殖。m2巨噬細(xì)胞促進(jìn)腫瘤的遷移、侵襲和增殖。唑來(lái)膦酸納米粒通過(guò)抑制m2型巨噬細(xì)胞間接抑制腫瘤細(xì)胞的增殖、遷移及侵襲能力。
[Abstract]:Objective: to study the uptake of zoledronate nanoparticles by different types of macrophages and the migration of tumor cells in co-culture of U937 cells induced as M 0 macrophages and M 1 type macrophages.The effect of zoledronic acid nanoparticles on the proliferation of tumor cells in co-culture.Methods: 1, macrophage induction: U937 cells were tiled into a six-well plate, and PMA was added to induce the induction of M 1 macrophages for 6 hours. U937 cells were tiled into a six-well plate.After 6 hours of induction, we added LPSN 100 ng / ml and IFN- 緯 20 ng / ml) and continued to induce 66 hours (72 hours in all, M 2 macrophage induction: tiling U937 cells into a six-well plate,After 6 hours of induction, IL-4 + 20 ng / ml and IL-13 + 20 ng / ml) were added for 66 hours (72 hours in total).By detecting the expression of Arg-1 enzyme and iNOS, we detected whether the induction was successful or not. The uptake experiment: human fibroblast M 1 macrophage M 2 macrophage was selected as the colon cancer cell line SW620, and the fluorescent labeled zoledronate nanoparticles were added.After 3 hours of reaction, the cell membrane was stained with Dil, and the uptake of zoledronate nanoparticles by four kinds of cells was observed under fluorescence microscope. The inhibition of proliferation of zoledronate nanoparticles was measured by the interaction between zoledronate nanoparticles and different kinds of cells.To understand its effect on cell proliferation, cell scratch test and transwell chamber migration test, we set up sw620 colon cancer group with sw620m1 macrophage group and SW620m2 macrophage group.Each group was divided into two groups with or without zoledronic acid nanoparticles (n = 6). The influence of each factor on the migration ability of tumor cells was observed. The invasion experiment: sw620 colon cancer cells were put into the upper chamber of transwell chamber.The lower chamber was divided into blank group (culture medium alone, zoledronic acid nanoparticles group, M 1 macrophage group, zoledronic acid nanoparticles group, zoledronic acid nanoparticles group, zoledronic acid nanoparticles group, M 2 macrophage group, M 2 macrophage group, zoledronic acid nanoparticles group;The effect of various factors on the invasiveness of tumor cells was investigated by means of Gimsa staining. Results the results of 1: 1 arg-1 and inos showed that the macrophages of type 2 were immunocytochemical positive.However, the other two groups showed negative reaction. The inos expression of M 1 type macrophages was much higher than that of the other two groups (14. 0% and 14. 7%, respectively). It was proved that the fluorescence intensity of M 1 and m 2 macrophages induced successfully by M 2 M 2 macrophages was much higher than that of other cells.Macrophages inhibit the migration of tumor cells and type M 2 macrophages promote the migration of tumor cells.The interaction with zoledronate nanoparticles can inhibit the promotion of m2 macrophage.The interaction with zoledronic acid nanoparticles can inhibit the promotion of m2 macrophages. The nanoparticles can inhibit the proliferation of tumor cells by inhibiting m2 macrophages.Conclusion: zoledronate nanoparticles can be specifically ingested by m2 macrophages and inhibit the proliferation of m2 macrophages. M2 macrophages promote tumor migration, invasion and proliferation.Zoledronate nanoparticles indirectly inhibit the proliferation, migration and invasion of tumor cells by inhibiting m 2 macrophages.
【學(xué)位授予單位】：西南醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R73-36

【參考文獻(xiàn)】

相關(guān)期刊論文 前3條

1 石艷;張惠斌;宗莉;;靶向甘露糖受體的載體甘露糖化殼聚糖的全合成及其細(xì)胞毒性評(píng)價(jià)[J];藥物生物技術(shù);2009年03期

2 李康;郭強(qiáng);王翠妮;陳敏;徐薇;熊思東;;M1和M2型巨噬細(xì)胞表型的比較分析[J];現(xiàn)代免疫學(xué);2008年03期

3 顧其勝;陶偉棟;蔣麗霞;;殼聚糖基復(fù)合材料類(lèi)新型生物醫(yī)用材料[J];上海生物醫(yī)學(xué)工程;2007年01期

相關(guān)碩士學(xué)位論文 前1條

1 劉宇;甘露糖化殼聚糖納米粒與腫瘤組織結(jié)合能力的研究[D];瀘州醫(yī)學(xué)院;2015年

，

本文編號(hào)：1706643