MEG3 shRNA质粒的纳米载药系统对脑梗死后血管新生的影响
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摘要
目的探讨MEG3短发夹RNA(shRNA)质粒的纳米载药系统对脑梗死后血管新生的影响。方法 60只SD大鼠分为对照(A)组、对照质粒(B)组、MEG3(C)组和纳米(D)组,每组15只。制备脑梗死大鼠模型,利用MEG3 shRNA质粒制备纳米载药系统。A组大鼠尾静脉注射PBS 1ml,B组尾静脉注射对照质粒200μg,C组尾静脉注射MEG3shRNA质粒200μg,D组尾静脉注射纳米载药系统(含有MEG3 shRNA 200μg),每天1次,连续注射10d。三苯基氯化四氮唑染色观察脑梗死体积,HE染色法观察脑组织微血管数量,检测MEG3、HES1和DLL4表达情况。结果 A组没有脑梗死出现。D组脑梗死体积小于B组和C组[(98.3±25.7)mm~3 vs.(235.4±38.2)mm~3和(215.5±29.5)mm~3](P<0.05)。A组微血管数量丰富,B组和C组微血管数量减少,D组微血管数量较C组增加(P<0.05)。与A组比较,B组和C组脑组织MEG3表达升高(P<0.05),D组脑组织MEG3表达较B组和C组降低(P<0.05)。与A组比较,B组和C组HES1和DLL4的mRNA和蛋白表达下降(P<0.05),D组HES1和DLL4的mRNA和蛋白表达较B组和C组升高(P<0.05)。结论 MEG3shRNA质粒的纳米载药系统能够穿透血脑屏障,降低脑梗死体积,促进脑组织血管新生,可能在脑梗死的治疗中发挥重要作用。
Objective To investigate the effect of nano drug loading system containing MEG3 shRNA plasmids on the angiogenesis after cerebral infarction.Methods Sixty SD rats were equally divided into four groups.Rat model with cerebral infarction was established and nano drug loading system was established by MEG3 shRNA plasmids.Group A was injected with PBS 1 ml by caudal vein,group B was injected with control plasmid 200μg,group C was injected with MEG3 shRNA plasmids 200μg and group D was injected with nano drug loading system containing MEG3 shRNA plasmids.The injection of 4 groups was performed once a day for 10 days.The volume of cerebral infarction was observed by triphenyl tetrazolium chloride staining.The number of microvessels in brain tissues was detected by HE staining.The expressions of MEG3,HES1 and DLL4 were detected.Results No cerebral infarction was seen in group A.The volume of cerebral infarction was less in group D than that in groups of B and C [(98.3±25.7)mm3 vs.(235.4±38.2)mm~3 and(215.5±29.5)mm~3](P<0.05).The quantity of microvessels was abundant in group A and less in groups of B and C,which was more in group D than that in group C(P<0.05).The expression of MEG3 was higher in groups of B and C than that in group A,which was lower in group D than that in groups of B and C(P<0.05).The mRNA and protein expressions of HES1 and DLL4 were lower in groups of B and C than those in group A,which were higher in group D than those in groups of B and C(P<0.05).Conclusion Nano drug loading system containing MEG3 shRNA plasmids can penetrate the blood-brain barrier,reduce the cerebral infarction volume and promote the angiogenesis,which may play a role in the treatment of cerebral infarction.
Objective To investigate the effect of nano drug loading system containing MEG3 shRNA plasmids on the angiogenesis after cerebral infarction.Methods Sixty SD rats were equally divided into four groups.Rat model with cerebral infarction was established and nano drug loading system was established by MEG3 shRNA plasmids.Group A was injected with PBS 1 ml by caudal vein,group B was injected with control plasmid 200μg,group C was injected with MEG3 shRNA plasmids 200μg and group D was injected with nano drug loading system containing MEG3 shRNA plasmids.The injection of 4 groups was performed once a day for 10 days.The volume of cerebral infarction was observed by triphenyl tetrazolium chloride staining.The number of microvessels in brain tissues was detected by HE staining.The expressions of MEG3,HES1 and DLL4 were detected.Results No cerebral infarction was seen in group A.The volume of cerebral infarction was less in group D than that in groups of B and C [(98.3±25.7)mm3 vs.(235.4±38.2)mm~3 and(215.5±29.5)mm~3](P<0.05).The quantity of microvessels was abundant in group A and less in groups of B and C,which was more in group D than that in group C(P<0.05).The expression of MEG3 was higher in groups of B and C than that in group A,which was lower in group D than that in groups of B and C(P<0.05).The mRNA and protein expressions of HES1 and DLL4 were lower in groups of B and C than those in group A,which were higher in group D than those in groups of B and C(P<0.05).Conclusion Nano drug loading system containing MEG3 shRNA plasmids can penetrate the blood-brain barrier,reduce the cerebral infarction volume and promote the angiogenesis,which may play a role in the treatment of cerebral infarction.
引文
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[2]Zhou Y,Zhang X,Klibanski A.MEG3noncoding RNA:a tumor suppressor[J].J Mol Endocrinol,2012,48(3):R45-R53.
[3]Yan H,Yuan J,Gao L,et al.Long noncoding RNA MEG3activation of p53mediates ischemic neuronal death in stroke[J].Neuroscience,2016,337:191-199.
[4]Li H,Sun H,Qian ZM.The role of the transferrin-transferrinreceptor system in drug delivery and targeting[J].Trends Pharmacol Sci,2002,23(5):206-209.
[5]Han L,Li J,Huang S,et al.Peptide-conjugated polyamidoamine dendrimer as a nanoscale tumor-targeted T1magnetic resonance imaging contrast agent[J].Biomaterials,2011,32(11):2989-2998.
[6]Zhang L,Geng X,Zhou J,et al.Fabrication of poly(γ-glutamic acid)-based biopolymer as the targeted drug delivery system with enhanced cytotoxicity to APN/CD13over-expressed cells[J].J Drug Target,2015,23(5):453-461.
[7]Thom G,Burrell M,Haqqani AS,et al.Enhanced delivery of galanin conjugates to the brain through bioengineering of the anti-transferrin receptor antibody OX26[J].Mol Pharm,2018,15(4):1420-1431.
[8]Liu Z,Zhao H,Shu L,et al.Preparation and evaluation of Baicalin-loaded cationic solid lipid nanoparticles conjugated with OX26for improved delivery across the BBB[J].Drug Dev Ind Pharm,2015,41(3):353-361.
[9]Liu Z,Zhang L,HE Q,et al.Effect of Baicalin-loaded PEGylated cationic solid lipid nanoparticles modified by OX26antibody on regulating the levels of baicalin and amino acids during cerebral ischemia-reperfusion in rats[J].Int J Pharm,2015,489(1/2):131-138.
[10]Yin KJ,Hamblin M,Chen YE.Angiogenesis-regulating microRNAs and ischemic stroke[J].Curr Vasc Pharmacol,2015,13(3):352-365.
[11]Zhang L,Chang J,Zhao Y,et al.Fabrication of a triptolideloaded and poly-γ-glutamic acid-based amphiphilic nanoparticle for the treatment of rheumatoid arthritis[J].Int J Nanomedicine,2018,13:2051-2064.
[12]Liu ZJ,Shirakawa T,Li Y,et al.Regulation of Notch1and Dll4by vascular endothelial growth factor in arterial endothelial cells:implications for modulating arteriogenesis and angiogenesis[J].Mol Cell Biol,2003,23(1):14-25.
[13]Liu LY,Lin MH,Lai ZY,et al.Motor neuron-derived Thsd7ais essential for zebrafish vascular development via the Notch-dll4signaling pathway[J].J Biomed Sci,2016,23(1):59.
[14]Fang Y,Yu S,Ma Y,et al.Association of Dll4/notch and HIF-1a-VEGF signaling in the angiogenesis of missed abortion[J].PLoS One,2013,8(8):e70667.
[15]Abbas OL,Borman H,Terzi YK,et al.The Notch pathway is a critical regulator of angiogenesis in a skin model of ischemia[J].Vasc Med,2015,20(3):205-211.