title: The Effect of Millisecond Pulsed Electric Fields (msPEF) on Intracellular Drug Transport with Negatively Charged Large Nanocarriers Made of Solid Lipid Nanoparticles (SLN): In Vitro Study.
abstract: Drug delivery technology is still a dynamically developing field of medicine. The main direction in nanotechnology research (nanocarriers, nanovehicles, etc.) is efficient drug delivery to target cells with simultaneous drug reduction concentration. However, nanotechnology trends in reducing the carrier sizes to several nanometers limit the volume of the loaded substance and may pose a danger of uncontrolled access into the cells. On the other hand, nanoparticles larger than 200 nm in diameter have difficulties to undergo rapid diffusional transport through cell membranes. The main advantage of large nanoparticles is higher drug encapsulation efficiency and the ability to deliver a wider array of drugs. Our present study contributes a new approach with large Tween 80 solid lipid nanoparticles SLN (i.e., hydrodynamic GM-SLN-glycerol monostearate, GM, as the lipid and ATO5-SLNs-glyceryl palmitostearate, ATO5, as the lipid) with diameters DH of 379.4 nm and 547 nm, respectively. They are used as drug carriers alone and in combination with electroporation (EP) induced by millisecond pulsed electric fields. We evaluate if EP can support the transport of large nanocarriers into cells. The study was performed with two cell lines: human colon adenocarcinoma LoVo and hamster ovarian fibroblastoid CHO-K1 with coumarin 6 (C6) as a fluorescent marker for encapsulation. The biological safety of the potential treatment procedure was evaluated with cell viability after their exposure to nanoparticles and EP. The EP efficacy was evaluated by FACS method. The impact on intracellular structure organization of cytoskeleton was visualized by CLSM method with alpha-actin and beta-tubulin. The obtained results indicate low cytotoxicity of both carrier types, free and loaded with C6. The evaluation of cytoskeleton proteins indicated no intracellular structure damage. The intracellular uptake and accumulation show that SLNs do not support transport of C6 coumarin. Only application of electroporation improved the transport of encapsulated and free C6 into both treated cell lines.
authors.affiliation: Department of Medical Biochemistry, Medical University, Chałubińskiego 10 St, 50-368, Wroclaw, Poland. julita.kulbacka@umed.wroc.pl.
authors.forename: Julita
authors.initials: J
authors.lastname: Kulbacka
authors.affiliation: Department of Organic and Pharmaceutical Technology, Faculty of Chemistry, Wrocław University of Science and Technology, Wybrzeże Wyspiańskiego 27, 50-370, Wrocław, Poland.
authors.forename: Agata
authors.initials: A
authors.lastname: Pucek
authors.affiliation: Department of Organic and Pharmaceutical Technology, Faculty of Chemistry, Wrocław University of Science and Technology, Wybrzeże Wyspiańskiego 27, 50-370, Wrocław, Poland.
authors.forename: Kazimiera Anna
authors.initials: KA
authors.lastname: Wilk
authors.affiliation: Department of General Zoology, Zoological Institute, University of Wroclaw, Sienkiewicza 21 St, 50-335, Wroclaw, Poland.
authors.forename: Magda
authors.initials: M
authors.lastname: Dubińska-Magiera
authors.affiliation: Institute of Immunology and Experimental Therapy Polish Academy of Sciences, Rudolfa Weigla 12, 53-114, Wroclaw, Poland.
authors.forename: Joanna
authors.initials: J
authors.lastname: Rossowska
authors.affiliation: Polish-Japanese Academy of Information Technology, Koszykowa 86, 02-008, Warsaw, Poland.
authors.forename: Marek
authors.initials: M
authors.lastname: Kulbacki
authors.affiliation: Department of Biomedical Engineering, Faculty of Fundamental Problems of Technology, Wroclaw University of Science and Technology, Wybrzeże Wyspiańskiego 27, 50-370, Wroclaw, Poland.
authors.forename: Małgorzata
authors.initials: M
authors.lastname: Kotulska
mesh_terms: D000818:Animals; D001692:Biological Transport; D002152:Calorimetry, Differential Scanning; D002460:Cell Line; D002470:Cell Survival; D003374:Coumarins / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D004337:Drug Carriers; D016503:Drug Delivery Systems; D018274:Electroporation; D005434:Flow Cytometry; D006801:Humans; D008055:Lipids / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D015394:Molecular Structure; D053758:Nanoparticles / Q000737:chemistry*; D010316:Particle Size; D013844:Thiazoles / Q000008:administration & dosage / Q000737:chemistry / Q000378:metabolism
chemical_list: D003374:Coumarins; D004337:Drug Carriers; D008055:Lipids; D013844:Thiazoles; C517282:coumarin 6
keywords: Coumarin-6; Drug delivery; Electroporation; Millisecond pulsed electric field; Solid lipid nanocarriers