Department of Pharmacy, University of Copenhagen, DK-2100 Copenhagen, Denmark.
Costerton Biofilm Center, Department of Immunology and Microbiology, University of Copenhagen, DK-2200 Copenhagen, Denmark.
ACS Nano. 2021 Apr 27;15(4):6787-6800. doi: 10.1021/acsnano.0c10378. Epub 2021 Mar 16.
In the present study, we investigated lipid membrane interactions of silica nanoparticles as carriers for the antimicrobial peptide LL-37 (LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPRTES). In doing so, smooth mesoporous nanoparticles were compared to virus-like mesoporous nanoparticles, characterized by a "spiky" external surface, as well as to nonporous silica nanoparticles. For this, we employed a combination of neutron reflectometry, ellipsometry, dynamic light scattering, and ζ-potential measurements for studies of bacteria-mimicking bilayers formed by palmitoyloleoylphosphatidylcholine/palmitoyloleoylphosphatidylglycerol. The results show that nanoparticle topography strongly influences membrane binding and destabilization. We found that virus-like particles are able to destabilize such lipid membranes, whereas the corresponding smooth silica nanoparticles are not. This effect of particle spikes becomes further accentuated after loading of such particles with LL-37. Thus, peptide-loaded virus-like nanoparticles displayed more pronounced membrane disruption than either peptide-loaded smooth nanoparticles or free LL-37. The structural basis of this was clarified by neutron reflectometry, demonstrating that the virus-like nanoparticles induce trans-membrane defects and promote incorporation of LL-37 throughout both bilayer leaflets. The relevance of such effects of particle spikes for bacterial membrane rupture was further demonstrated by confocal microscopy and live/dead assays on bacteria. Taken together, these findings demonstrate that topography influences the interaction of nanoparticles with bacteria-mimicking lipid bilayers, both in the absence and presence of antimicrobial peptides, as well as with bacteria. The results also identify virus-like mesoporous nanoparticles as being of interest in the design of nanoparticles as delivery systems for antimicrobial peptides.
在本研究中，我们研究了作为抗菌肽 LL-37（LLGDFFRKSKEKIGKEFKRIVQRIKDFLRNLVPRTES）载体的硅纳米粒子与脂质膜的相互作用。为此，我们将光滑介孔纳米粒子与具有“刺状”外表面的病毒样介孔纳米粒子以及无孔硅纳米粒子进行了比较。我们采用了中子反射法、椭圆光度法、动态光散射和 ζ-电位测量相结合的方法，研究了由棕榈酰油酰磷脂酰胆碱/棕榈酰油酰磷脂酰甘油模拟的双层形成的细菌类似物。结果表明，纳米粒子的形貌强烈影响着膜的结合和破坏。我们发现，病毒样颗粒能够破坏这种脂质膜，而相应的光滑硅纳米粒子则不能。这种颗粒刺的影响在这些颗粒负载 LL-37 后变得更加明显。因此，负载肽的病毒样纳米颗粒比负载肽的光滑纳米颗粒或游离 LL-37 显示出更明显的膜破坏作用。中子反射测量法澄清了这种效应的结构基础，表明病毒样纳米颗粒诱导跨膜缺陷，并促进 LL-37 穿过双层的两个叶层。通过共聚焦显微镜和活/死试验对细菌的研究进一步证明了这种颗粒刺效应对细菌膜破裂的相关性。总之，这些发现表明，颗粒的形貌影响着纳米颗粒与细菌模拟脂质双层的相互作用，无论是在没有还是存在抗菌肽的情况下，以及与细菌的相互作用。研究结果还表明，病毒样介孔纳米粒子在设计作为抗菌肽输送系统的纳米粒子方面具有重要意义。
