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Journal of Nanobiotechnology Oct 2018Engineered inorganic nanoparticles (NPs) are essential components in the development of nanotechnologies. For applications in nanomedicine, particles need to be...
BACKGROUND
Engineered inorganic nanoparticles (NPs) are essential components in the development of nanotechnologies. For applications in nanomedicine, particles need to be functionalized to ensure a good dispersibility in biological fluids. In many cases however, functionalization is not sufficient: the particles become either coated by a corona of serum proteins or precipitate out of the solvent. We show that by changing the coating of magnetic iron oxide NPs using poly-L-lysine (PLL) polymer the colloidal stability of the dispersion is improved in aqueous solutions including water, phosphate buffered saline (PBS), PBS with 10% fetal bovine serum (FBS) and cell culture medium, and the internalization of the NPs toward living mammalian cells is profoundly affected.
METHODS
A multifunctional magnetic NP is designed to perform a near-infrared (NIR)-responsive remote control photothermal ablation for the treatment of breast cancer. In contrast to the previously reported studies of gold (Au) magnetic (FeO) core-shell NPs, a Janus-like nanostructure is synthesized with FeO NPs decorated with Au resulting in an approximate size of 60 nm mean diameter. The surface of trisoctahedral Au-FeO NPs was coated with a positively charged polymer, PLL to deliver the NPs inside cells. The PLL-Au-FeO NPs were characterized by transmission electron microscopy (TEM), XRD, FT-IR and dynamic light scattering (DLS). The unique properties of both Au surface plasmon resonance and superparamagnetic moment result in a multimodal platform for use as a nanothermal ablator and also as a magnetic resonance imaging (MRI) contrast agent, respectively. Taking advantage of the photothermal therapy, PLL-Au-FeO NPs were incubated with BT-474 and MDA-MB-231 breast cancer cells, investigated for the cytotoxicity and intracellular uptake, and remotely triggered by a NIR laser of ~ 808 nm (1 W/cm for 10 min).
RESULTS
The PLL coating increased the colloidal stability and robustness of Au-FeO NPs (PLL-Au-FeO) in biological media including cell culture medium, PBS and PBS with 10% fetal bovine serum. It is revealed that no significant (< 10%) cytotoxicity was induced by PLL-Au-FeO NPs itself in BT-474 and MDA-MB-231 cells at concentrations up to 100 μg/ml. Brightfield microscopy, fluorescence microscopy and TEM showed significant uptake of PLL-Au-FeO NPs by BT-474 and MDA-MB-231 cells. The cells exhibited 40 and 60% inhibition in BT-474 and MDA-MB-231 cell growth, respectively following the internalized NPs were triggered by a photothermal laser using 100 μg/ml PLL-Au-FeO NPs. The control cells treated with NPs but without laser showed < 10% cell death compared to no laser treatment control CONCLUSION: Combined together, the results demonstrate a new polymer gold superparamagnetic nanostructure that integrates both diagnostics function and photothermal ablation of tumors into a single multimodal nanoplatform exhibiting a significant cancer cell death.
Topics: Cell Death; Cell Line, Tumor; Ferric Compounds; Fluorescence; Gold; Humans; Hyperthermia, Induced; Magnetite Nanoparticles; Phototherapy; Polylysine; Polymers; Static Electricity; Temperature; Theranostic Nanomedicine; X-Ray Diffraction
PubMed: 30316298
DOI: 10.1186/s12951-018-0405-7 -
ACS Combinatorial Science Apr 2017Cationic macromolecular carriers can be effective carriers for small molecular compounds, drugs, epitopes, or nucleic acids. Polylysine-based polymeric branched...
Cationic macromolecular carriers can be effective carriers for small molecular compounds, drugs, epitopes, or nucleic acids. Polylysine-based polymeric branched polypeptides have been systematically studied on the level of cells and organisms as well. In the present study, we report our findings on the cellular uptake characteristics of nine structurally related polylysine-based polypeptides with cationic side chains composed of (i) single amino acid (poly[Lys(X)], XK) or (ii) oligo[dl-alanine] (poly[Lys(dl-Ala)], AK) or (iii) oligo[dl-alanine] with an additional amino acid (X) at the terminal position (poly[Lys(X-dl-Ala)] (XAK)) or (iv) at the position next to the polylysine backbone (poly[Lys(dl-Ala-X)] (AXK)). In vitro cytotoxicity and cellular uptake were characterized on HT-29 human colon carcinoma and HepG2 human hepatocarcinoma cell lines. Data indicate that the polycationic polypeptides studied are essentially nontoxic in the concentration range studied, and their uptake is very much dependent on the side chain structure (length, identity of amino acid X, and distance between the terminal positive charges) and also on the cell lines. Our findings in uptake inhibition studies suggest that predominantly macropinocytosis and caveole/lipid raft mediated endocytosis are involved. The efficacy of their internalization is markedly influenced by the hydrophobicity and charge properties of the amino acid X. Interestingly, the uptake properties of the these polypeptides show certain similarities to the entry pathways of several cell penetrating peptides.
Topics: Cations; Cell Line, Tumor; Drug Delivery Systems; Endocytosis; Humans; Peptides; Polylysine; Protein Conformation; Structure-Activity Relationship
PubMed: 28276242
DOI: 10.1021/acscombsci.6b00133 -
Applied and Environmental Microbiology Dec 2014Epsilon-poly-l-lysine (ε-PL) is a natural antimicrobial cationic peptide which is generally regarded as safe (GRAS) as a food preservative. Although its antimicrobial...
Epsilon-poly-l-lysine (ε-PL) is a natural antimicrobial cationic peptide which is generally regarded as safe (GRAS) as a food preservative. Although its antimicrobial activity is well documented, its mechanism of action is only vaguely described. The aim of this study was to clarify ε-PL's mechanism of action using Escherichia coli and Listeria innocua as model organisms. We examined ε-PL's effect on cell morphology and membrane integrity and used an array of E. coli deletion mutants to study how specific outer membrane components affected the action of ε-PL. We furthermore studied its interaction with lipid bilayers using membrane models. In vitro cell studies indicated that divalent cations and the heptose I and II phosphate groups in the lipopolysaccharide layer of E. coli are critical for ε-PL's binding efficiency. ε-PL removed the lipopolysaccharide layer and affected cell morphology of E. coli, while L. innocua underwent minor morphological changes. Propidium iodide staining showed that ε-PL permeabilized the cytoplasmic membrane in both species, indicating the membrane as the site of attack. We compared the interaction with neutral or negatively charged membrane systems and showed that the interaction with ε-PL relied on negative charges on the membrane. Suspended membrane vesicles were disrupted by ε-PL, and a detergent-like disruption of E. coli membrane was confirmed by atomic force microscopy imaging of supported lipid bilayers. We hypothesize that ε-PL destabilizes membranes in a carpet-like mechanism by interacting with negatively charged phospholipid head groups, which displace divalent cations and enforce a negative curvature folding on membranes that leads to formation of vesicles/micelles.
Topics: Anti-Bacterial Agents; Biological Transport; Cell Membrane; Escherichia coli; Listeria; Polylysine
PubMed: 25304506
DOI: 10.1128/AEM.02204-14 -
Nature Structural & Molecular Biology Dec 2019Faulty or damaged messenger RNAs are detected by the cell when translating ribosomes stall during elongation and trigger pathways of mRNA decay, nascent protein...
Faulty or damaged messenger RNAs are detected by the cell when translating ribosomes stall during elongation and trigger pathways of mRNA decay, nascent protein degradation and ribosome recycling. The most common mRNA defect in eukaryotes is probably inappropriate polyadenylation at near-cognate sites within the coding region. How ribosomes stall selectively when they encounter poly(A) is unclear. Here, we use biochemical and structural approaches in mammalian systems to show that poly-lysine, encoded by poly(A), favors a peptidyl-transfer RNA conformation suboptimal for peptide bond formation. This conformation partially slows elongation, permitting poly(A) mRNA in the ribosome's decoding center to adopt a ribosomal RNA-stabilized single-stranded helix. The reconfigured decoding center clashes with incoming aminoacyl-tRNA, thereby precluding elongation. Thus, coincidence detection of poly-lysine in the exit tunnel and poly(A) in the decoding center allows ribosomes to detect aberrant mRNAs selectively, stall elongation and trigger downstream quality control pathways essential for cellular homeostasis.
Topics: HEK293 Cells; Humans; Models, Molecular; Nucleic Acid Conformation; Peptides; Poly A; Polyadenylation; Polylysine; Protein Biosynthesis; RNA Stability; RNA, Messenger; RNA, Transfer; RNA, Transfer, Amino Acyl; Ribosomes
PubMed: 31768042
DOI: 10.1038/s41594-019-0331-x -
The Journal of Reproduction and... Oct 2022In this study, we cryopreserved pig spermatozoa using carboxylated poly-L-lysine (CPLL) as the cryoprotectant to determine its efficacy. Pig spermatozoa were placed in a...
In this study, we cryopreserved pig spermatozoa using carboxylated poly-L-lysine (CPLL) as the cryoprotectant to determine its efficacy. Pig spermatozoa were placed in a freezing extender containing 3% (v/v) glycerol and different CPLL concentrations. The motility indices of the spermatozoa cryopreserved with 0.25% (v/v) CPLL at 6 (59.3), 9 (53.7), and 12 (26.2) h after thawing were significantly higher (P < 0.01 or P < 0.05) than those of the spermatozoa cryopreserved without CPLL (53.7, 40.1, and 17.5 at 6, 9, and 12 h after thawing, respectively). The concentration of CPLL in the freezing extender did not affect the ability of frozen-thawed spermatozoa to fertilize oocytes in vitro. However, the blastocyst formation rate of embryos derived from spermatozoa cryopreserved with 0.25% CPLL (24.6%) was significantly higher (P < 0.01) than that of embryos derived from spermatozoa cryopreserved without CPLL (11.2%). The conception rate of the sows inseminated with spermatozoa cryopreserved with 0.25% CPLL (72.2%) was not significantly different from that of the sows inseminated with spermatozoa stored at 17°C (81.3%). However, the mean number of total piglets born to the former (10.0) was significantly lower (P < 0.05) than that of total piglets born to the latter (13.4). The results showed that CPLL in the freezing extender maintained the motility of frozen-thawed pig spermatozoa and improved the in vitro development of embryos produced by in vitro fertilization. In addition, we have demonstrated that piglets could be obtained with artificial insemination using spermatozoa cryopreserved with CPLL.
Topics: Animals; Cryopreservation; Cryoprotective Agents; Female; Glycerol; Male; Polylysine; Semen Preservation; Sperm Motility; Spermatozoa; Swine
PubMed: 35908977
DOI: 10.1262/jrd.2022-058 -
International Journal of Nanomedicine 2021With increases in implant infections, the search for antibacterial and biofilm coatings has become a new interest for orthopaedists and dentists. In recent years,...
INTRODUCTION
With increases in implant infections, the search for antibacterial and biofilm coatings has become a new interest for orthopaedists and dentists. In recent years, graphene oxide (GO) has been extensively studied for its superior antibacterial properties. However, most of these studies have focused on solutions and there are few antibacterial studies on metal surfaces, especially the surfaces of cobalt-chromium-molybdenum (CoCrMo) alloys. ε-Poly-L-lysine (ε-PLL), as a novel food preservative, has a spectrum of antimicrobial activity; however, its antimicrobial activity after coating an implant surface is not clear.
METHODS
In this study, for the first time, a two-step electrodeposition method was used to coat GO and ε-PLL on the surface of a CoCrMo alloy. Its antibacterial and antibiofilm properties against and were then studied.
RESULTS
The results show that the formation of bacteria and biofilms on the coating surface was significantly inhibited, GO and ε-PLL composite coatings had the best antibacterial and antibiofilm effects, followed by ε-PLL and GO coatings. In terms of classification, the coatings are anti-adhesive and contact-killing/inhibitory surfaces. In addition to oxidative stress, physical damage to GO and electrostatic osmosis of ε-PLL are the main antibacterial and antibiofilm mechanisms.
DISCUSSION
This is the first study that GO and ε-PLL coatings were successfully prepared on the surface of CoCrMo alloy by electrodeposition. It provides a promising new approach to the problem of implant infection in orthopedics and stomatology.
Topics: Anti-Bacterial Agents; Biofilms; Coated Materials, Biocompatible; Escherichia coli; Graphite; Polylysine; Staphylococcus aureus; Surface Properties; Vitallium
PubMed: 34737563
DOI: 10.2147/IJN.S321800 -
The Biochemical Journal Jun 1997The mechanism of protein kinase CK2 (CK2) activity stimulation by polylysine has been studied by surface plasmon resonance (SPR). The kinetics of the polylysine...
The mechanism of protein kinase CK2 (CK2) activity stimulation by polylysine has been studied by surface plasmon resonance (SPR). The kinetics of the polylysine interaction with a peptide substrate of the enzyme, and with the enzyme itself, have been investigated. A peptide containing a threonine (T) residue surrounded by a cluster of negatively charged acidic [arginine (R) and glutamic acid (E)] residues, RRREEETEEE, and specifically phosphorylated by CK2, was selected. Polylysine interacts with both the enzyme and the peptide substrate. The rate constant, the stoichiometry of the polylysine-peptide substrate interaction and the kinetic parameters of the stimulated enzyme were used to calculate the polylysine-dependent stimulation of CK2. The results are in agreement with experimentally determined polylysine-dependent stimulation. The polylysine-enzyme interaction is too slow to account for enzyme stimulation. The behaviour of polylysine is not reproduced by the polyamine spermine. The results are consistent with a substrate-mediated mechanism of CK2 stimulation by polylysine, and they suggest that the CK2 stimulation by polyamines occurs by a different mechanism.
Topics: Amino Acid Sequence; Animals; Casein Kinase II; Cattle; Enzyme Activation; Kinetics; Models, Chemical; Peptide Fragments; Polylysine; Protein Serine-Threonine Kinases; Substrate Specificity; Surface Properties
PubMed: 9210426
DOI: 10.1042/bj3240987 -
PloS One 2020The study of cell aggregation in vitro has a tremendous importance these days. In cancer biology, aggregates and spheroids serve as model systems and are considered as...
The study of cell aggregation in vitro has a tremendous importance these days. In cancer biology, aggregates and spheroids serve as model systems and are considered as pseudo-tumors that are more realistic than 2D cell cultures. Recently, in the context of brain tumors (gliomas), we developed a new poly(ethylene glycol) (PEG)-based hydrogel, with adhesive properties that can be controlled by the addition of poly(L-lysine) (PLL), and a stiffness close to the brain's. This substrate allows the motion of individual cells and the formation of cell aggregates (within one day), and we showed that on a non-adhesive substrate (PEG without PLL is inert for cells), the aggregates are bigger and less numerous than on an adhesive substrate (with PLL). In this article, we present new experimental results on the follow-up of the formation of aggregates on our hydrogels, from the early stages (individual cells) to the late stages (aggregate compaction), in order to compare, for two cell lines (F98 and U87), the aggregation process on the adhesive and non-adhesive substrates. We first show that a spaceless model of perikinetic aggregation can reproduce the experimental evolution of the number of aggregates, but not of the mean area of the aggregates. We thus develop a minimal off-lattice agent-based model, with a few simple rules reproducing the main processes that are at stack during aggregation. Our spatial model can reproduce very well the experimental temporal evolution of both the number of aggregates and their mean area, on adhesive and non-adhesive soft gels and for the two different cell lines. From the fit of the experimental data, we were able to infer the quantitative values of the speed of motion of each cell line, its rate of proliferation in aggregates and its ability to organize in 3D. We also found qualitative differences between the two cell lines regarding the ability of aggregates to compact. These parameters could be inferred for any cell line, and correlated with clinical properties such as aggressiveness and invasiveness.
Topics: Cell Adhesion; Cell Aggregation; Cell Culture Techniques; Cell Line; Cell Proliferation; Humans; Hydrogels; Kinetics; Models, Biological; Polyethylene Glycols; Polylysine
PubMed: 32023245
DOI: 10.1371/journal.pone.0222371 -
The Analyst Nov 2019Atomic force microscopy (AFM) provides an effective, label-free technique enabling the imaging of live bacteria under physiological conditions with nanometre precision.... (Comparative Study)
Comparative Study
Atomic force microscopy (AFM) provides an effective, label-free technique enabling the imaging of live bacteria under physiological conditions with nanometre precision. However, AFM is a surface scanning technique, and the accuracy of its performance requires the effective and reliable immobilisation of bacterial cells onto substrates. Here, we compare the effectiveness of various chemical approaches to facilitate the immobilisation of Escherichia coli onto glass cover slips in terms of bacterial adsorption, viability and compatibility with correlative imaging by fluorescence microscopy. We assess surface functionalisation using gelatin, poly-l-lysine, Cell-Tak™, and Vectabond®. We describe how bacterial immobilisation, viability and suitability for AFM experiments depend on bacterial strain, buffer conditions and surface functionalisation. We demonstrate the use of such immobilisation by AFM images that resolve the porin lattice on the bacterial surface; local degradation of the bacterial cell envelope by an antimicrobial peptide (Cecropin B); and the formation of membrane attack complexes on the bacterial membrane.
Topics: Buffers; Cell Adhesion; Cell Membrane; Cells, Immobilized; Escherichia coli; Gelatin; Microscopy, Atomic Force; Polylysine; Porins; Propylamines; Silanes
PubMed: 31620716
DOI: 10.1039/c9an01185d -
Journal of Nuclear Medicine : Official... Jul 1993Efforts to achieve rapid specific targeting of radioisotopes to disease processes using antibodies conjugated with avidin or streptavidin for pretargeting and...
Efforts to achieve rapid specific targeting of radioisotopes to disease processes using antibodies conjugated with avidin or streptavidin for pretargeting and radiobiotin derivatives for isotope delivery are attracting substantial interest. At present, these approaches appear to be limited by low delivery of radiotracer to the target. As an alternate radiobiotin tracer, biotinylated/iodinated polylysine (BIP) was prepared by conjugating poly-L-lysine (MW approximately 10,200) with biotin succinimide esters and the Bolton-Hunter reagent. This reagent was then radioiodinated with 125I via the lodogen method. BIP was characterized by radio-HPLC and its in vitro binding to streptavidin. The in vivo localization of BIP was evaluated in a rat model in which streptavidin agarose beads were physically localized to precapillary arterioles in the lungs. Biodistribution and blocking studies performed at 4 and 24 hr after BIP injection indicated specific binding and localization of the radiolabeled peptide to the lungs (lung-to-blood ratio approximately 8 at 4 hr postinjection). Comparative studies of BIP and 111In chelated to biotin showed BIP to have two-fold higher lung targeting and lower splenic and hepatic uptake than the 111In biotin derivative. Our study demonstrates: (1) the feasibility of using a small peptide as a biotin carrier for pretargeting (and for solubilizing organic tracers which may otherwise be difficult to administer in vivo) and (2) that BIP and BIP-like compounds may be suitable and simple alternatives to radiometal-labeled biotin for pretargeting and may offer improved targeting to prelocalized streptavidin.
Topics: Animals; Biotin; Chromatography, High Pressure Liquid; Female; Iodine Radioisotopes; Lung; Polylysine; Radioimmunodetection; Rats; Rats, Sprague-Dawley; Tissue Distribution
PubMed: 8315493
DOI: No ID Found