發(fā)布時間：2018-07-10 03:28

  本文選題：殼聚糖 + TPP　； 參考：《功能材料》2017年12期

【摘要】：系統(tǒng)比較了不同分子量的殼聚糖與TPP離子交聯(lián)制備的納米粒,用于基因轉(zhuǎn)運載體,評價其對質(zhì)粒和siRNA的轉(zhuǎn)運效果。通過粒度儀測定殼聚糖納米粒(CSNPs)的粒徑、多分散系數(shù)以及Zeta電位;透射電鏡觀察CSNPs的形態(tài);MTT法測定其細胞毒性;通過CSNPs在小鼠肌肉內(nèi)的組織切片觀察其生物相容性。CSNPs分別包載質(zhì)粒和siRNA,凝膠電泳和分光光度法測定其包載能力,熒光顯微鏡、激光共聚焦或流式細胞術(shù)分析其對腫瘤細胞的轉(zhuǎn)染效果;溶血實驗和血凝實驗分析其血液相容性。結(jié)果表明,1 mg/mL的CS(160 k Da)和TPP(質(zhì)量比為10∶1)制備的CSNPs粒徑約100 nm,且分布均一、穩(wěn)定,透射電鏡下為形態(tài)規(guī)則的類球型粒子;其細胞毒性均在0~1級,符合生物醫(yī)用材料毒性標(biāo)準(zhǔn);組織切片沒有發(fā)現(xiàn)明顯的炎癥反應(yīng),生物相容性良好;CSNPs對質(zhì)粒和siRNA的包載率較高,并能成功將其轉(zhuǎn)運至細胞內(nèi),但小粒徑的納米粒轉(zhuǎn)染質(zhì)粒后熒光較弱,而攜帶siRNA的CSNPs轉(zhuǎn)染后熒光較強,且流式細胞術(shù)結(jié)果表明其與商品化轉(zhuǎn)染試劑相比轉(zhuǎn)染率較高;溶血及血凝實驗也發(fā)現(xiàn)材料具有良好的血液相容性。所以,制備的小粒徑CSNPs更適合作為一種安全高效的siRNA轉(zhuǎn)運載體,后續(xù)可以加以修飾應(yīng)用于腫瘤的基因靶向治療中。
[Abstract]:The nanoparticles prepared by cross-linking chitosan with TPP ions with different molecular weights were systematically compared to be used as gene transport vectors to evaluate their translocation effects on plasmids and siRNA. The particle size, polydispersity coefficient and Zeta potential of chitosan nanoparticles (CSNPs) were measured by particle size analyzer. The biocompatibility of CSNPs in mouse muscle was observed by tissue section. CSNPs were encapsulated with plasmid and siRNA respectively. The encapsulation ability of CSNPs was determined by gel electrophoresis and spectrophotometry. Laser confocal or flow cytometry was used to analyze the transfection of tumor cells, and hemolysis and hemagglutination were used to analyze the blood compatibility of tumor cells. The results showed that CSNPs prepared by 1 mg / mL CS (160kDa) and TPP (10:1 by mass ratio) were about 100 nm in size, uniform in distribution, stable in distribution, regular in morphology under transmission electron microscope, and their cytotoxicity was in grade 0 ~ 1, which met the toxicity standard of biomaterials. No obvious inflammatory reaction was found in the tissue section. The biocompatibility of CSNPs was higher than that of the plasmid and siRNA, and could be successfully transported into the cells, but the fluorescence of the plasmid transfected with the small size nanoparticles was weak. The results of flow cytometry showed that the transfection efficiency of CSNPs carrying siRNA was higher than that of commercial transfection reagents, and the hemolysis and hemagglutination experiments also showed that the materials had good blood compatibility. Therefore, the prepared CSNPs are more suitable as a safe and efficient siRNA transport vector, and can be modified for gene targeting therapy.
【作者單位】： 青島大學(xué)醫(yī)學(xué)部檢驗學(xué)系;青島大學(xué)附屬醫(yī)院泌尿外科與男科學(xué)重點實驗室;青島大學(xué)附屬醫(yī)院中心實驗室;青島大學(xué)附屬醫(yī)院檢驗科;
【基金】：國家自然科學(xué)基金資助項目(81401899) 山東省自然科學(xué)基金資助項目(ZR2014HP011) 青島青年科學(xué)家應(yīng)用基礎(chǔ)研究基金資助項目(15-9-1-51-jch) 青島大學(xué)附屬醫(yī)院青年基金會資助項目(2417)
【分類號】：O636.1;R450

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