克拉霉素脂质体的制备及体内外抗菌评价（英文）
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摘要
本研究的目的在于制备由Tat肽和麦胚凝集素修饰(WGA)修饰的脂质体递送系统,有效发挥抗菌效应。分别对其理化性质、耐甲氧西林金黄色葡萄球菌(MRSA)的最低抑菌浓度(MIC)、杀菌动力学、细胞摄取、生物膜形成抑制以及体内抗菌效果进行评价。研究结果表明,由Tat和WGA双修饰的载克拉霉素脂质体具有最低的MIC值和杀菌曲线,流式细胞实验结果表明双修饰的脂质体可将更多的香豆素6导入细菌内部。此外,该脂质体还能有效抑制MRSA生物膜的形成。体内实验结果表明,经过双修饰脂质体给药,小鼠脓肿部位的MRSA菌落数显著低于其他组(P<0.01),即具有最强的体内抗菌效果。总之,由Tat和WGA修饰的脂质体递送系统希望称为有效的抗耐药菌感染的策略。
In this study, we developed a novel liposomal delivery system modified by Tat peptide and wheat germ agglutinin(WGA) with antimicrobial effect. Physicochemical parameters, in vitro antimicrobial, time-kill study, cellular uptake, biofilm formation inhibition and in vivo antibacterial efficacy against Methicillin-resistant Staphylococcus aureus(MRSA) were investigated. Minimum inhibitory concentrations(MICs) and colony-forming units(CFUs) in the time-kill study for Tat-WGA-modified liposomal clarithromycin(CLA-Tat WGALip) were lower than those of free and other modified liposomal CLA. Flow cytometry analysis disclosed that Tat WGALip delivered more coumarin 6 into bacteria. Furthermore, Tat-WGA-modified liposomal CLA efficiently inhibited the formation of MRSA biofiom. CFU of MRSA in the abscess of mice treated with CLA-Tat WGALip was significantly lower than that of any others(P<0.01). Collectively, liposomal delivery system modified by Tat and WGA could be a promising anti-resistant infection strategy.
In this study, we developed a novel liposomal delivery system modified by Tat peptide and wheat germ agglutinin(WGA) with antimicrobial effect. Physicochemical parameters, in vitro antimicrobial, time-kill study, cellular uptake, biofilm formation inhibition and in vivo antibacterial efficacy against Methicillin-resistant Staphylococcus aureus(MRSA) were investigated. Minimum inhibitory concentrations(MICs) and colony-forming units(CFUs) in the time-kill study for Tat-WGA-modified liposomal clarithromycin(CLA-Tat WGALip) were lower than those of free and other modified liposomal CLA. Flow cytometry analysis disclosed that Tat WGALip delivered more coumarin 6 into bacteria. Furthermore, Tat-WGA-modified liposomal CLA efficiently inhibited the formation of MRSA biofiom. CFU of MRSA in the abscess of mice treated with CLA-Tat WGALip was significantly lower than that of any others(P<0.01). Collectively, liposomal delivery system modified by Tat and WGA could be a promising anti-resistant infection strategy.
引文
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[24]De Oliveira,F.F.;Torres,A.F.;Goncalves,T.B.;Santiago,G.M.;De Carvalho,C.B.;Aguiar,M.B.;Camara,L.M.;Rabenhorst,S.H.;Martins,A.M.;Valenca Junior,J.T.;Nagao-Dias,A.T.Efficacy of Plectranthus amboinicus(Lour.)Spreng in a Murine Model of Methicillin-Resistant Staphylococcus aureus Skin Abscesses.Evid.Based Complement Alternat.Med.2013,2013,291592.
[2]Zhang,X.;Hu,X.;Rao,X.Apoptosis induced by Staphylococcus aureus toxins.Microbiol.Res.2017,205,19–24.
[3]Seilie,E.S.;Wardenburg,J.B.Staphylococcus aureus pore-forming toxins:The interface of pathogen and host complexity.Semin.Cell Dev.Biol.2017,72,101–116.
[4]David,M.Z.;Dryden,M.;Gottlieb,T.;Tattevin,P.;Gould,I.M.Recently approved antibacterials for MRSA and other gram-positive pathogens:the shock of the new.Int.J.Antimicrob.Agents.2017,50,303–307.
[5]Parisi,O.I.;Scrivano,L.;Sinicropi,M.S.;Puoci,F.Polymeric nanoparticle constructs as devices for antibacterial therapy.Curr.Opin.Pharmacol.2017,36,72–77.
[6]Erik,T.;Webster,T.J.Reducing infections through nanotechnology and nanoparticles.Int.J.Nanomedicine.2011,6,1463–1473.
[7]Jain,A.;Shah,S.G.;Chugh,A.Cell Penetrating Peptides as Efficient Nanocarriers for Delivery of Antifungal Compound,Natamycin for the Treatment of Fungal Keratitis.Pharm.Res.2015,32,1920–1930.
[8]Holm,T.;Netzereab,S.;Hansen,M.;Langel,U.;H?llbrink,M.Uptake of cell-penetrating peptides in yeasts.FEBS Lett.2005,579,5217–5222.
[9]Zhu,Y.;Zhang,J.;Meng,F.;Deng,C.;Cheng,R.;Feijen,J.;Zhong,Z.c RGD/TAT dual-ligand reversibly crosslinked micelles loaded with docetaxel penetrate deeply into tumor tissue and show high antitumor efficacy in vivo.ACS Appl.Mater.Interfaces.2017,9,35651–35663.
[10]He,B.;Ma,S.;Peng,G.;He,D.TAT-modified Selfassembled Cationic Peptide Nanoparticles as an Efficient Antibacterial Agent.Nanomedicine.2018,14,365–372.
[11]Ghaffar,K.A.;Hussein,W.M.;Khalil,Z.G.;Capon,R.J.;Skwarczynski,M.;Toth,I.Levofloxacin and indolicidin for combination antimicrobial therapy.Curr.Drug Deliv.2015,12,108–114.
[12]Goldstein,I.J.;Hayes,C.E.The Lectins:CarbohydrateBinding Proteins of Plants and Animals.Adv.Carbohydr.Chem.Biochem.1978,35,127–340.
[13]Foster,T.J.Immune evasion by staphylococci.Nat.Rev.Microbiol.2005,3,948–958.
[14]Torchilin,V.P.Fluorescence microscopy to follow the targeting of liposomes and micelles to cells and their intracellular fate.Adv.Drug Deliv.Rev.2005,57,95–109.
[15]Xie,H.Y.;Xie,M.;Zhang,Z.L.;Long,Y.M.;Liu,X.;Tang,M.L.;Pang,D.W.;Tan,Z.;Dickinson,C.;Zhou,W.Wheat germ agglutinin-modified trifunctional nanospheres for cell recognition.Bioconjug.Chem.2007,18,1749–1755.
[16]Lavelle,E.C.;Grant,G.;Pusztai,A.;Pfüller,U.;O’Hagan,D.T.Mucosal immunogenicity of plant lectins in mice.Immunology.2000,99,30–37.
[17]Gabius,H.Endogenous Lectins in Tumors and the Immune System.Cancer Invest.1987,5,39–46.
[18]Mody,R.;Joshi,S.H.A.;Chaney,W.Use of lectins as diagnostic and therapeutic tools for cancer.J.Pharmacol.Toxicol.Methods.1995,33,1–10.
[19]Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically;Approved StandardNinth Edition.Program Index of CLSI Standards and Guidelines.2012,32,15–50.
[20]Principe,L.;D’Arezzo,S.;Capone,A.;Petrosillo,N.;Visca,P.In vitro activity of tigecycline in combination with various antimicrobials against multidrug resistant Acinetobacter baumannii.Ann Clin.Microbiol.Antimicrob.2009,8,18.
[21]Chakraborty,S.P.;Sahu,S.K.;Pramanik,P.;Roy,S.In vitro antimicrobial activity of nanoconjugated vancomycin against drug resistant Staphylococcus aureus.Int.J.Pharm.2012,436,659–676.
[22]Barbieri,D.S.;Tonial,F.;Lopez,P.V.;Sales Maia,B.H.;Santos,G.D.;Ribas,M.O.;Glienke,C.;Vicente,V.A.Antiadherent activity of Schinus terebinthifolius and Croton urucurana extracts on in vitro biofilm formation of Candida albicans and Streptococcus mutans.Arch.Oral.Biol.2014,59,887–896.
[23]Burt,S.A.;Ojofakunle,V.T.;Woertman,J.;Veldhuizen,E.J.The natural antimicrobial carvacrol inhibits quorum sensing in Chromobacterium violaceum and reduces bacterial biofilm formation at sub-lethal concentrations.Plos.One.2014,9,e93414.
[24]De Oliveira,F.F.;Torres,A.F.;Goncalves,T.B.;Santiago,G.M.;De Carvalho,C.B.;Aguiar,M.B.;Camara,L.M.;Rabenhorst,S.H.;Martins,A.M.;Valenca Junior,J.T.;Nagao-Dias,A.T.Efficacy of Plectranthus amboinicus(Lour.)Spreng in a Murine Model of Methicillin-Resistant Staphylococcus aureus Skin Abscesses.Evid.Based Complement Alternat.Med.2013,2013,291592.