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1.
Adv Healthc Mater ; : e2304118, 2024 Feb 27.
Article in English | MEDLINE | ID: mdl-38412457

ABSTRACT

The burden of bacterial wound infections has considerably increased due to antibiotic resistance to most of the currently available antimicrobial drugs. Herein, for the first time, a chemical coupling of two cationic N-aryl (pyridyl and aminocinnamyl) chitosan derivatives to antimicrobial peptide dendrimers (AMPDs) of different generations (first, second, and third) via thioether-haloacetyl reaction is reported. The new chitosan-AMPD conjugates show high selectivity by killing Pseudomonas aeruginosa and very low toxicity toward mammalian cells, as well as extremely low hemolysis to red blood cells. Electron microscopy reveals that the new chitosan derivatives coupled to AMPD destroy both the inner and outer membranes of Gram-negative P. aeruginosa. Moreover, chitosan-AMPD conjugates show synergetic effects within extremely low concentrations. The new chitosan-AMPD conjugates can be used as potent antimicrobial therapeutic agents, to eradicate pathogens such as those present in acute and chronic infected wounds.

2.
Carbohydr Polym ; 280: 119025, 2022 Mar 15.
Article in English | MEDLINE | ID: mdl-35027127

ABSTRACT

We report herein a new chemical platform for coupling chitosan derivatives to antimicrobial peptide dendrimers (AMPDs) with different degrees of ramification and molecular weights via thiol-maleimide reactions. Previous studies showed that simple incorporation of AMPDs to polymeric hydrogels resulted in a loss of antibacterial activity and augmented cytotoxicity to mammalian cells. We have shown that coupling AMPDs to chitosan derivatives enabled the two compounds to act synergistically. We showed that the antimicrobial activity was preserved when incorporating AMPD conjugates into various biopolymer formulations, including nanoparticles, gels, and foams. Investigating their mechanism of action using electron and time-lapse microscopy, we showed that the AMPD-chitosan conjugates were internalized after damaging outer and inner Gram-negative bacterial membranes. We also showed the absence of AMPD conjugates toxicity to mammalian cells. This chemical technological platform could be used for the development of new membrane disruptive therapeutics to eradicate pathogens present in acute and chronic wounds.


Subject(s)
Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Antimicrobial Peptides/pharmacology , Chitosan , Dendrimers , Pseudomonas aeruginosa/drug effects , Animals , Anti-Bacterial Agents/toxicity , Antimicrobial Peptides/chemistry , Antimicrobial Peptides/toxicity , Cell Membrane/drug effects , Cells, Cultured , Drug Synergism , Hemolysis , Humans , Microbial Sensitivity Tests , Polymers
3.
Pharmaceuticals (Basel) ; 14(4)2021 Mar 28.
Article in English | MEDLINE | ID: mdl-33800588

ABSTRACT

Hyaluronic acid (HA) has been known to play an important role in wound healing process. However, the effect of molecular weight (MW) of exogenously administered HA on the wound healing process has not been fully understood. In this study, we investigated HA with different MWs on wound healing process using human epidermal keratinocytes and dermal fibroblasts. Cell proliferation and migration ability were assessed by water soluble tetrazolium (WST) assay and wound scratch assay. We examined the effect of HA addition in a full-thickness wound model in mice and the gene expression related to wound healing. Proliferation and migration of HaCaT cells increased with the increase of MW and concentration of HA. Interleukin (IL-1ß), IL-8 and vascular endothelial growth factor (VEGF) as well as matrix metalloproteinase (MMP)-9 and MMP-13 were significantly upregulated by high molecular weight (HMW) HA in keratinocytes. Together with VEGF upregulation and the observed promotion of HaCaT migration, HA with the MW of 2290 kDa may hold potential to improve re-epithelialization, a critical obstacle to heal chronic wounds.

4.
Eur J Pharm Sci ; 131: 69-74, 2019 Apr 01.
Article in English | MEDLINE | ID: mdl-30742979

ABSTRACT

The assessment of interchangeability of nanomedicines and nanosimilars is required for ensuring the safety, quality and efficacy of these complex drugs. Since 2011, regulatory agencies and researchers focused their attention on the characterization of iron sucrose, a colloidal solution parenterally used for the treatment of iron deficiency anemia, as well as on their follow-on versions, the so-called iron sucrose similars (ISSs). The purpose of this study was the evaluation of the size and size distribution of both IS and ISSs after dilution in polypropylene bags containing saline, mimicking the preparation of ready-to-use infusions to be administered in a hospital environment. The assays were carried out for 72 h, determining a general stability of the colloidal solutions investigated. Nevertheless, a dissimilar size and size distribution but also different visual appearance of IS and ISSs were recorded when the drugs diluted for therapeutic use. These results agree with previously published physicochemical and clinical data supporting the lack of exchangeability between IS and ISSs.


Subject(s)
Ferric Oxide, Saccharated , Nanoparticles , Colloids , Infusions, Intravenous , Nanomedicine , Particle Size , Saline Solution, Hypertonic , Solutions
5.
Int J Pharm ; 482(1-2): 61-7, 2015 Mar 30.
Article in English | MEDLINE | ID: mdl-25448555

ABSTRACT

We describe here the establishment and first characterization of a co-culture model of human epithelial sublingual cells (HO-1-u-1 cell line) and human dendritic cells derived from human peripheral blood monocytes (PBMC). Cell culture conditions for HO-1-u-1 cells were optimized. First characterization of phenotypic features by electron microscopy and fluorescence imaging revealed resemblance to sublingual tissue specimen from healthy donors. Successful co-culturing of epithelial and dendritic cells (DCs) was confirmed by confocal laser scanning microscopy. Stimulation of HO-1-u1 cells alone and the epithelial/DC co-culture by incubation with liposomes, virosomes and influenza virus lead reproducibly to the release of inflammatory cytokine GM-CSF. This co-culture model may be suitable for elucidation of mechanisms involved in the immune response at the sublingual epithelium as well as for the evaluation of novel topical vaccines, potentially replacing cumbersome ex vivo and in vivo methods currently in place.


Subject(s)
Dendritic Cells/cytology , Dendritic Cells/immunology , Epithelial Cells/cytology , Epithelial Cells/immunology , Mouth Floor/cytology , Mouth Floor/immunology , Cell Line , Coculture Techniques , Cytokines/immunology , Dendritic Cells/ultrastructure , Epithelial Cells/ultrastructure , Humans , Liposomes , Orthomyxoviridae/immunology , Virosomes/immunology
6.
Tissue Barriers ; 2: e29166, 2014.
Article in English | MEDLINE | ID: mdl-25101232

ABSTRACT

We investigated the role of Toll-like receptor (TLR) 2 in maintaining the integrity of the airway epithelial barrier using the human bronchial epithelial cell line Calu-3. Activation of TLR2 by its ligands, Pam3CysSK4 and Peptidoglycan showed a concentration dependent increase in epithelial barrier function, as measured by transepithelial electrical resistance (TEER). This was confirmed by a decrease in paracellular flux of fluorescein sodium. This TLR2 induced increase in TEER was significantly reduced by pretreatment with polyclonal anti-human TLR2-neutralizing antibody. TLR2 stimulation in Calu-3 cell monolayers resulted in an increased expression of the tight junction proteins claudin-1 and ZO-1, and a decreased expression of occludin, at both the mRNA and protein levels. A pseudosubstrate inhibitor to PKCζ significantly prevented the TLR2 mediated increase in barrier function. It also prevented the increase in claudin-1 in a concentration dependent manner up to 1 µM. TLR2 stimulation led to an increase in phosphorylation of atypical PKC ζ, which was prevented by the pseudosubstrate inhibitor in a concentration dependent manner. Taken together, our observations support a model whereby increased tight junction barrier function induced by activation of TLR2 occurs through increased expression of claudin-1, and through modulation of PKC ζ activity.

7.
Mol Pharm ; 10(8): 2996-3004, 2013 Aug 05.
Article in English | MEDLINE | ID: mdl-23869898

ABSTRACT

We describe here the development of nanoparticles made from poly(lactic-co-glycolic acid) (PLGA) able to deliver an encapsulated antigen with a Toll-Like Receptor-7 (TLR-7) agonist as immunostimulatory signal and coated with a muco-adhesive chitosan-derivate layer. The potential to stimulate an immune response of these vaccine formulations in the absence or presence of the TLR-7 agonist at the systemic and mucosal level were evaluated in mice following subcutaneous or nasal administrations. Intranasally immunized mice developed a high systemic immune response equivalent to mice injected subcutaneously. However, mucosal immune responses were only induced at local and distal sites in mucosally immunized animals. The adjuvant effect of imiquimod on the polarization of the immune response was only detected at local sites, which tends to increase safety of this vaccine delivery system.


Subject(s)
Antigens/chemistry , Immunity, Mucosal/immunology , Immunologic Factors/chemistry , Lactic Acid/chemistry , Nanoparticles/chemistry , Polyglycolic Acid/chemistry , Animals , Antigens/immunology , Cell Survival , Female , Hydrophobic and Hydrophilic Interactions , Immunologic Factors/immunology , Mice , Polylactic Acid-Polyglycolic Acid Copolymer
8.
J Microencapsul ; 29(6): 583-95, 2012.
Article in English | MEDLINE | ID: mdl-22439872

ABSTRACT

In the present work, concomitant use of self-microemulsifying drug delivery systems (SMEDDS) and a novel third-generation P-gp inhibitor, GF120918 (elacridar), for the effective transport of taxanes (paclitaxel and docetaxel) across an in vitro model of the intestinal epithelium and uptake into tumor cells were investigated. On the basis of solubility studies and ternary phase diagrams, different SMEDDS formulations of taxanes were prepared and characterized. In caco-2 cell permeation study, paclitaxel-loaded SMEDDS along with GF120918 showed a four-fold increase in apparent permeability, while docetaxel-loaded SMEDDS in combination with GF120918 showed a nine-fold increase in permeability, as compared to plain drug solution. Cell uptake studies on A549 cells were performed with microemulsions formed from both SMEDDS formulations loaded with rhodamine 123 dye and showed good uptake than plain dye solution. Confocal laser scanning microscopic images further confirmed the higher uptake of both SMEDDS formulations in the presence of GF120918.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/antagonists & inhibitors , Acridines , Antineoplastic Agents, Phytogenic , Drug Delivery Systems , Paclitaxel , Taxoids , Tetrahydroisoquinolines , ATP Binding Cassette Transporter, Subfamily B , Acridines/chemistry , Acridines/pharmacokinetics , Acridines/pharmacology , Antineoplastic Agents, Phytogenic/chemistry , Antineoplastic Agents, Phytogenic/pharmacokinetics , Antineoplastic Agents, Phytogenic/pharmacology , Caco-2 Cells , Docetaxel , Drug Screening Assays, Antitumor , Humans , Paclitaxel/chemistry , Paclitaxel/pharmacokinetics , Paclitaxel/pharmacology , Solubility , Taxoids/chemistry , Taxoids/pharmacokinetics , Taxoids/pharmacology , Tetrahydroisoquinolines/chemistry , Tetrahydroisoquinolines/pharmacokinetics , Tetrahydroisoquinolines/pharmacology
9.
J Drug Target ; 17(8): 662-70, 2009 Sep.
Article in English | MEDLINE | ID: mdl-19694614

ABSTRACT

The purpose of this study was to prepare and characterize nanocarrier systems, which allow the application of pDNA vaccines and adjuvants to mucosal vaccination. Chitosan from a vegetal source (Agaricus bisporus) and of GMP quality was used to synthesize the derivative 6-O-carboxymethyl-N,N,N-trimethylchitosan (CM-TMC). Toll-like receptor-2 (TLR-2) agonist, Pam(3)Cys, was synthesized and coupled to CM-TMC through a polyethylene glycol (PEG) spacer. Successively, Pam(3)Cys decorated nanocarriers were prepared by complexation with plasmid DNA (pDNA) expressing green fluorescence protein (GFP), and characterized with respect to their physicochemical properties and protection of the included plasmid against DNase I enzymatic degradation. In vitro studies using phorbol 12-myristyl 13-acetate (PMA) stimulated macrophage-like THP-1 (mTHP-1) cells were focused on cytotoxicity of both polymers and particles, and their potential to stimulate IL-8 release via the TLR-2 pathway. Our results showed that the TLR-2 functionalized pDNA nanocarriers have the ability to complex and to protect pDNA against enzymatic degradation. pDNA nanocarriers were of around 400 nm in size, and displayed a positive zeta potential of 27.9 +/- 1.6 mV. Chitosan, CM-TMC, and Pam(3)Cys-functionalized CM-TMC polymers displayed cytotoxicity on mTHP1 cells in a concentration-dependent manner, which decreased by 50-fold on complexation with pDNA. In addition, decorated pDNA nanocarriers induced IL-8 secretion by mTHP-1 macrophages, which was increased by 10-fold as compared to nondecorated carriers.


Subject(s)
Lipoproteins/chemistry , Macrophages/metabolism , Nanoparticles , Toll-Like Receptor 2/agonists , Agaricus/chemistry , Cell Line , Cell Line, Tumor , Chitosan/chemistry , DNA/administration & dosage , Dose-Response Relationship, Drug , Green Fluorescent Proteins/genetics , Humans , Interleukin-8/metabolism , Particle Size , Plasmids/administration & dosage , Polyethylene Glycols/chemistry
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