【摘要】：目的用新型聚乙烯亞胺納米凝膠(PEI-NP)攜帶小RNA(miRNA)質(zhì)粒沉默大鼠Kupffer細(xì)胞(KC)核因子κB(NF-κB)的表達(dá)。方法凝膠電泳檢測PEI-NP裝載miRNA質(zhì)粒的容量;PEI-NP/miRNA質(zhì)粒轉(zhuǎn)染RAW264.7細(xì)胞后分別采用CCK-8法檢測細(xì)胞毒性,annexinⅤ-FITC/PI雙染色結(jié)合流式細(xì)胞術(shù)檢測RAW264.7細(xì)胞凋亡和轉(zhuǎn)染率,實時定量PCR(qRT-PCR)檢測RAW264.7細(xì)胞NF-κB mRNA水平,Western blot法檢測RAW264.7細(xì)胞NF-κB蛋白水平,透射電鏡觀察陽離子聚合物在RAW264.7細(xì)胞和肝組織KC內(nèi)的分布;免疫組織化學(xué)技術(shù)檢測KC內(nèi)NF-κB的表達(dá)。結(jié)果 PEI-NP/miRNA質(zhì)粒質(zhì)量比為20∶1時載荷率接近100%,體外轉(zhuǎn)染RAW264.7細(xì)胞48 h,轉(zhuǎn)染率為(33.63±1.94)%。與對照組相比,PEI-NP/miRNA轉(zhuǎn)染的RAW264.7細(xì)胞NF-κB蛋白含量顯著降低,透射電鏡觀察到RAW264.7細(xì)胞內(nèi)有大量PEI-NP。肝臟組織透射電鏡只在KC內(nèi)觀察到PEI-NP分布;與對照組相比,免疫組織化學(xué)染色結(jié)果顯示,KC內(nèi)NF-κB蛋白水平明顯降低。結(jié)論 PEI-NP攜帶miRNA質(zhì)粒能成功轉(zhuǎn)染體外培養(yǎng)的巨噬細(xì)胞和體內(nèi)肝臟KC并沉默NF-κB的表達(dá)。
[Abstract]:Aim to silence the expression of (KC) nuclear factor 魏 B (NF- 魏 B) in rat Kupffer cells using a novel polyvinyleneimine nanogel (PEI-NP) carrying small RNA (miRNA) plasmid. Methods the capacity of PEI-NP loaded with miRNA plasmid was detected by gel electrophoresis. After transfection of PEI-NP/miRNA plasmid into RAW264.7 cells, cytotoxicity was detected by CCK-8 assay, apoptosis and transfection rate of RAW264.7 cells were detected by annexin 鈪，

本文編號：2381170