Pastuszka Martha K, Wang Xiangdong, Lock Lye Lin, Janib Siti Mohd, Cui Honggang, DeLeve Laurie D, MacKay J Andrew
Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA 90033-9121, USA.
Research Center for Liver Diseases and the Division of Gastrointestinal and Liver Diseases, Keck School of Medicine at the University of Southern California, Los Angeles, CA 90033, USA.
J Control Release. 2014 Oct 10;191:15-23. doi: 10.1016/j.jconrel.2014.07.003. Epub 2014 Jul 10.
L4F, an alpha helical peptide inspired by the lipid-binding domain of the ApoA1 protein, has potential applications in the reduction of inflammation involved with cardiovascular disease as well as an antioxidant effect that inhibits liver fibrosis. In addition to its biological activity, amphipathic peptides such as L4F are likely candidates to direct the molecular assembly of peptide nanostructures. Here we describe the stabilization of the amphipathic L4F peptide through fusion to a high molecular weight protein polymer. Comprised of multiple pentameric repeats, elastin-like polypeptides (ELPs) are biodegradable protein polymers inspired from the human gene for tropoelastin. Dynamic light scattering confirmed that the fusion peptide forms nanoparticles with a hydrodynamic radius of approximately 50nm, which is unexpectedly above that observed for the free ELP (~5.1nm). To further investigate their morphology, conventional and cryogenic transmission electron microscopy were used to reveal that they are unilamellar vesicles. On average, these vesicles are 49nm in radius with lamellae 8nm in thickness. To evaluate their therapeutic potential, the L4F nanoparticles were incubated with hepatic stellate cells. Stellate cell activation leads to hepatic fibrosis; furthermore, their activation is suppressed by anti-oxidant activity of ApoA1 mimetic peptides. Consistent with this observation, L4F nanoparticles were found to suppress hepatic stellate cell activation in vitro. To evaluate the in vivo potential for these nanostructures, their plasma pharmacokinetics were evaluated in rats. Despite the assembly of nanostructures, both free L4F and L4F nanoparticles exhibited similar half-lives of approximately 1h in plasma. This is the first study reporting the stabilization of peptide-based vesicles using ApoA1 mimetic peptides fused to a protein polymer; furthermore, this platform for peptide-vesicle assembly may have utility in the design of biodegradable nanostructures.
L4F是一种受载脂蛋白A1（ApoA1）蛋白脂质结合结构域启发而设计的α螺旋肽，在减轻心血管疾病相关炎症以及抑制肝纤维化的抗氧化作用方面具有潜在应用价值。除了其生物活性外，像L4F这样的两亲性肽很可能是指导肽纳米结构分子组装的候选物。在此，我们描述了通过与高分子量蛋白质聚合物融合来稳定两亲性L4F肽的方法。弹性蛋白样多肽（ELP）由多个五聚体重复序列组成，是受人类原弹性蛋白基因启发而合成的可生物降解蛋白质聚合物。动态光散射证实，融合肽形成了流体动力学半径约为50nm的纳米颗粒，这一数值意外高于游离ELP所观察到的半径（约5.1nm）。为了进一步研究它们的形态，使用常规和低温透射电子显微镜揭示它们是单层囊泡。这些囊泡的平均半径为49nm，膜层厚度为8nm。为了评估它们的治疗潜力，将L4F纳米颗粒与肝星状细胞一起孵育。肝星状细胞的激活会导致肝纤维化；此外，载脂蛋白A1模拟肽的抗氧化活性可抑制其激活。与这一观察结果一致，发现L4F纳米颗粒在体外可抑制肝星状细胞的激活。为了评估这些纳米结构在体内的潜力，在大鼠中评估了它们的血浆药代动力学。尽管形成了纳米结构，但游离L4F和L4F纳米颗粒在血浆中的半衰期相似，约为1小时。这是第一项报道使用与蛋白质聚合物融合的载脂蛋白A1模拟肽来稳定基于肽的囊泡的研究；此外，这种肽 - 囊泡组装平台可能在可生物降解纳米结构的设计中具有实用性。
