RESUMEN
Purpose: To determine the amoebicidal activity of functionalized poly-epsilon-lysine hydrogels (pÉK+) against Acanthamoeba castellanii. Methods: A. castellanii trophozoites and cysts were grown in the presence of pÉK solution (0-2.17 mM), pÉK or pÉK+ hydrogels, or commercial hydrogel contact lens (CL) for 24 hours or 7 days in PBS or Peptone-Yeast-Glucose (PYG) media (nutrient-deplete or nutrient-replete cultures, respectively). Toxicity was determined using propidium iodide and imaged using fluorescence microscopy. Ex vivo porcine corneas were inoculated with A. castellanii trophozoites ± pÉK, pÉK+ hydrogels or commercial hydrogel CL for 7 days. Corneal infection was assessed by periodic acid-Schiff staining and histologic analysis. Regrowth of A. castellanii from hydrogel lenses and corneal discs at 7 days was assessed using microscopy and enumeration. Results: The toxicity of pÉK+ hydrogels resulted in the death of 98.52% or 83.31% of the trophozoites at 24 hours or 7 days, respectively. The toxicity of pÉK+ hydrogels resulted in the death of 70.59% or 82.32% of the cysts in PBS at 24 hours or 7 days, respectively. Cysts exposed to pÉK+ hydrogels in PYG medium resulted in 75.37% and 87.14% death at 24 hours and 7 days. Ex vivo corneas infected with trophozoites and incubated with pÉK+ hydrogels showed the absence of A. castellanii in the stroma, with no regrowth from corneas or pÉK+ hydrogel, compared with infected-only corneas and those incubated in presence of commercial hydrogel CL. Conclusions: pÉK+ hydrogels demonstrated pronounced amoebicidal and cysticidal activity against A. castellanii. pÉK+ hydrogels have the potential for use as CLs that could minimize the risk of CL-associated Acanthamoeba keratitis.
Asunto(s)
Queratitis por Acanthamoeba/tratamiento farmacológico , Acanthamoeba castellanii/efectos de los fármacos , Amebicidas/farmacología , Córnea/parasitología , Infecciones Parasitarias del Ojo/tratamiento farmacológico , Hidrogeles/farmacología , Polilisina/farmacología , Queratitis por Acanthamoeba/parasitología , Amebicidas/toxicidad , Animales , Células Cultivadas , Soluciones para Lentes de Contacto/farmacología , Modelos Animales de Enfermedad , Epitelio Corneal/efectos de los fármacos , Infecciones Parasitarias del Ojo/parasitología , Humanos , Hidrogeles/toxicidad , Microscopía Fluorescente , Polilisina/toxicidad , Porcinos , Trofozoítos/efectos de los fármacosRESUMEN
Bacterial cellulose (BC) holds several unique properties such as high water retention capability, flexibility, biocompatibility, and high absorption capacity. All these features make it a potential material for wound healing applications. However, it lacks antibacterial properties, which hampers its applications for infectious wound healings. This study reported BC-based dressings containing ε-polylysine (ε-PL), cross-linked by a biocompatible and mussel-inspired polydopamine (PDA) for promoting infectious wound healing. BC membranes were coated with PDA by a simple self-polymerization process, followed by treating with different contents of ε-PL. The resulted membranes showed strong antibacterial properties against tested bacteria by both in vitro and in vivo evaluations. The membranes also exhibited hemocompatibility and cytocompatibility by in vitro investigations. Moreover, the functionalized membranes promoted infected wound healing using Sprague-Dawley rats as a model animal. A complete wound healing was observed in the group treated with functionalized membranes, while wounds were still open for control and pure BC groups in the same duration. Histological investigations indicated that the thickness of newborn skin was greater and smoother in the groups treated with modified membranes in comparison to neat BC or control groups. These results revealed that the functionalized membranes have great potential as a dressing material for infected wounds in future clinical applications.
Asunto(s)
Antibacterianos/uso terapéutico , Vendajes , Celulosa/química , Polilisina/uso terapéutico , Infecciones Cutáneas Estafilocócicas/tratamiento farmacológico , Cicatrización de Heridas/efectos de los fármacos , Animales , Antibacterianos/química , Antibacterianos/toxicidad , Celulosa/toxicidad , Escherichia coli/efectos de los fármacos , Indoles/química , Indoles/uso terapéutico , Indoles/toxicidad , Masculino , Ratones , Pruebas de Sensibilidad Microbiana , Células 3T3 NIH , Polilisina/análogos & derivados , Polilisina/toxicidad , Polímeros/química , Polímeros/uso terapéutico , Polímeros/toxicidad , Ratas Sprague-Dawley , Piel/efectos de los fármacos , Piel/patología , Infecciones Cutáneas Estafilocócicas/patología , Staphylococcus aureus/efectos de los fármacos , Infección de Heridas/tratamiento farmacológico , Infección de Heridas/patologíaRESUMEN
Severe traumatic bleeding control and wound-related anti-infection play a crucial role in saving lives and promoting wound healing for both the military and the clinic. In this contribution, an inherent antibacterial and instant swelling ε-poly-lysine/poly (ethylene glycol) diglycidyl ether (EPPE) superabsorbent was developed by a simple mild ring-opening reaction. The as-prepared EPPE1 displayed a porous structure and rough surface and exhibited instant water-triggered expansion with approximately 6300% swelling ratio in deionized water. Moreover, EPPE1 presented efficient pro-coagulation capacity by hemadsorption that can facilitate blood cell gathering and activation in vitro and exhibited a shorter in vivo hemostasis time than that of commercial gelatin sponge and CELOX in both rat tail amputation and noncompressible rat liver lethal defect model. Also, EPPE1 showed excellent antibacterial capacity, prominent biocompatibility, and great biodegradability. Additionally, EPPE1 significantly promotes in vivo wound healing in a full-thickness skin defect model due to its great hemostasis behavior and remarkable bactericidal performance. Hence, EPPE has great potential for serving as an extensively applied hemostatic agent under varied clinical conditions.
Asunto(s)
Antibacterianos/farmacología , Resinas Epoxi/farmacología , Hemostáticos/farmacología , Polilisina/farmacología , Cicatrización de Heridas/efectos de los fármacos , Animales , Antibacterianos/química , Antibacterianos/toxicidad , Resinas Epoxi/síntesis química , Resinas Epoxi/toxicidad , Escherichia coli/efectos de los fármacos , Hemostasis/efectos de los fármacos , Hemostáticos/química , Hemostáticos/toxicidad , Hígado/lesiones , Staphylococcus aureus Resistente a Meticilina/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Polilisina/análogos & derivados , Polilisina/toxicidad , Porosidad , Ratas Sprague-Dawley , Cola (estructura animal)/lesionesRESUMEN
DNA nanostructures (DNs) have garnered a large amount of interest as a potential therapeutic modality. However, DNs are prone to nuclease-mediated degradation and are unstable in low Mg2+ conditions; this greatly limits their utility in physiological settings. Previously, PEGylated oligolysines were found to protect DNs against low-salt denaturation and to increase nuclease resistance by up to â¼400-fold. Here we demonstrate that glutaraldehyde cross-linking of PEGylated oligolysine-coated DNs extends survival by up to another â¼250-fold to >48 h during incubation with 2600 times the physiological concentration of DNase I. DNA origami with cross-linked oligolysine coats are non-toxic and are internalized into cells more readily than non-cross-linked origami. Our strategy provides an off-the-shelf and generalizable method for protecting DNs in vivo.
Asunto(s)
Reactivos de Enlaces Cruzados/metabolismo , ADN/metabolismo , Desoxirribonucleasa I/metabolismo , Glutaral/metabolismo , Polilisina/metabolismo , Supervivencia Celular/efectos de los fármacos , Reactivos de Enlaces Cruzados/química , Reactivos de Enlaces Cruzados/toxicidad , ADN/química , ADN/toxicidad , Glutaral/química , Glutaral/toxicidad , Células HEK293 , Humanos , Hidrólisis , Nanoestructuras/química , Nanoestructuras/toxicidad , Conformación de Ácido Nucleico , Polietilenglicoles/química , Polietilenglicoles/metabolismo , Polietilenglicoles/toxicidad , Polilisina/química , Polilisina/toxicidadRESUMEN
Silver nanoparticles (AgNPs) represent one of the most abundant biocidal nanomaterials contained in more than 30% of nano-enabled consumer products and 75% of nanomedical products. The cumulative exposure of the general population may therefore reach critical and potentially hazardous levels. Due to data gaps on AgNP effects in humans, it is urgent to further evaluate their possible toxicity, particularly in vulnerable systems like the nervous one. As AgNPs may cross the blood brain and placental barriers, this study evaluated the in vitro effect of different AgNPs on neuronal precursor cells. For this purpose, 10â¯nm-sized AgNPs were stabilized with five different coating agents rendering a neutral, positive and negative surface charge. Murine neural stem cells (mNSCs) were used as cellular model to test AgNP neurotoxicity by evaluating the range of toxicity endpoints including cellular viability, apoptosis induction, oxidative stress response, cellular and mitochondrial membrane damages, DNA damage, inflammation response, and neural stem cell regulation. Our results clearly showed that the neurotoxic potential of AgNPs was not dependent on their surface charge or coating agents used for their surface stabilization. All AgNP types exhibited significant toxicity in neuronal precursor cells at an in vitro dose of 5â¯mg Ag/L or lower.
Asunto(s)
Nanopartículas del Metal/toxicidad , Células-Madre Neurales/efectos de los fármacos , Plata/toxicidad , Animales , Apoptosis/efectos de los fármacos , Bovinos , Supervivencia Celular/efectos de los fármacos , Cetrimonio/química , Cetrimonio/toxicidad , Daño del ADN/efectos de los fármacos , Ácido Dioctil Sulfosuccínico/química , Ácido Dioctil Sulfosuccínico/toxicidad , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones Endogámicos C57BL , Estrés Oxidativo/efectos de los fármacos , Polilisina/química , Polilisina/toxicidad , Povidona/química , Povidona/toxicidad , Albúmina Sérica Bovina/química , Albúmina Sérica Bovina/toxicidad , Plata/química , Transcriptoma/efectos de los fármacosRESUMEN
Conjugated polymer hybrid nanoparticles (NPs) loaded with both indocyanine green (ICG) and 1,3-diphenylisobenzofuran (DPBF) are described. The NPs are dually functional in that ICG acts as the photosensitizer, and DPBF as a probe for singlet oxygen (1O2 probe). The nanoparticle core consists of the energy donating host poly(9,9-dioctylfluorenyl-2,7-diyl)-co-(2,5-p-xylene) (PFP). The polymer is doped with the energy acceptor DPBF. Ratiometric fluorometric detection of singlet oxygen is accomplished by measurement of fluorescence at wavelengths of 415 and 458 nm. In addition, the shell of the positively charged polymeric nanoparticles was modified, via electrostatic interaction, with negatively charged PDT drugs ICG. The integrated nanoparticles of type ICG-DPBF-PFP display effective photodynamic performance under 808-nm laser irradiation. The 1O2 sensing behaviors of samples are evaluated based on the ratiometric fluorescent responses produced by DPBF and PFP. 1O2 can be fluorimetically sensed with a detection limit of 28 µM. The multifunctional nanoprobes exhibit effortless cellular uptake, superior photodynamic activity and a rapid ratiometric response to 1O2. Graphical abstractSchematic of a dual-functional nanoplatform for photodynamic therapy (PDT) and singlet oxygen (1O2) feedback. It offers a new strategy for self-monitoring photodynamic ablation. FRET: fluorescence resonance energy transfer. Indocyanine green is attached in the shell of nanoparticles, and 1,3-diphenylisobenzofuran is doped into the energy donating host conjugated polymer.
Asunto(s)
Benzofuranos/química , Verde de Indocianina/química , Nanopartículas/química , Fármacos Fotosensibilizantes/química , Polilisina/química , Oxígeno Singlete/análisis , Benzofuranos/toxicidad , Transferencia Resonante de Energía de Fluorescencia , Células Hep G2 , Humanos , Verde de Indocianina/efectos de la radiación , Verde de Indocianina/toxicidad , Rayos Infrarrojos , Límite de Detección , Nanopartículas/toxicidad , Fotoquimioterapia , Fármacos Fotosensibilizantes/efectos de la radiación , Fármacos Fotosensibilizantes/toxicidad , Polilisina/toxicidad , Oxígeno Singlete/químicaRESUMEN
The principal impediment to gene therapy is the development of efficient, nontoxic gene carriers that can handle and deliver foreign genetic materials into various cell types, including healthy and cancerous cells. Poly-l-lysine (PLL) polymers are one of the most favorable gene carriers among nonviral vectors, and PLL had low transfection and safety issues. The purpose of this study was to measure cellular toxicity, DNA damage, and apoptotic effects of PLL nanoparticles. Neuro2A mammalian cells were cultured and exposed to PLL/DNA complexes at different polymer/DNA ratios (C/P ratio 2 and 6) for 24 h. To evaluate metabolic activity, genotoxicity, and apoptotic influences of PLL nanoparticle, the following experimental methods were employed, in order: 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), DNA damage (COMET analysis) assay, and sub-G1 peak apoptosis assay. Our data indicate that toxicity is concentration dependent and a high concentration of polymer declined the metabolic activity. In addition, largest complexes (C/P 6 in HEPES buffered saline buffer) have slighter negative impact on metabolic activity. In agreement with our cytotoxicity data, apoptotic assay result represented that increase in size of PLL/DNA complexes decrease the number of apoptotic cells. Also, there was a remarkable increase in percent tail DNA of Neuro2A cells treated with higher concentration of PLL and its polyplexes. The present study demonstrated that PLL/DNA complexes caused cytotoxic, apoptotic, and genotoxic effects in a dose-dependent and weight ratio-dependent manner, which also affected the size of polyplexes.
Asunto(s)
ADN/toxicidad , Nanopartículas/toxicidad , Polilisina/toxicidad , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Daño del ADN , Ratones , PlásmidosRESUMEN
Much attention has been drawn to targeted nanodrug delivery systems due to their high therapeutic efficacy in cancer treatment. In this work, doxorubicin (DOX) was incorporated into a zwitterionic arginyl-glycyl-aspartic acid (RGD)-conjugated polypeptide by an emulsion solvent evaporation technique with high drug loading content (45%) and high drug loading efficiency (95%). This zwitterionic nanoformulation showed excellent colloidal stability at high dilution and in serum. The pH-induced disintegration and enzyme-induced degradation of the nanoformulation were confirmed by dynamic light scattering and gel permeation chromatography. Efficient internalization of DOX in the cells and high antitumor activity in vitro was observed. Compared with the free drug, this nanoformulation showed higher accumulation in tumor and lower systemic toxicity in vivo. The DOX-loaded zwitterionic RGD-conjugated polypeptide vesicles show potential application for targeted drug delivery in the clinic.
Asunto(s)
Antineoplásicos/farmacología , Doxorrubicina/farmacología , Portadores de Fármacos/química , Péptidos Cíclicos/química , Ácido Poliglutámico/análogos & derivados , Polilisina/análogos & derivados , Línea Celular Tumoral , Portadores de Fármacos/toxicidad , Liberación de Fármacos , Estabilidad de Medicamentos , Humanos , Nanopartículas/química , Nanopartículas/toxicidad , Péptidos Cíclicos/toxicidad , Ácido Poliglutámico/química , Ácido Poliglutámico/toxicidad , Polilisina/química , Polilisina/toxicidadRESUMEN
The antimicrobial polypeptide ε-poly(l-lysine) (ε-PL) was electrostatically incorporated to poly(acrylic acid) (PAA)/poly(vinyl alcohol) (PVA) electrospun nanofibers. ε-PL loading and distribution was assessed by infrared spectra, ζ-potential measurements and the primary amino reactive dye fluorescamine. Functionalized fibers with 485⯱â¯140â¯nm diameter, could be loaded with 0.57-0.74â¯g ε-PL (g dressing)-1 that released at a constant rate of 5.4⯱â¯2.8â¯mg ε-PL (g dressing day)-1. Such a dressings resulted in two orders of magnitude lower bacterial colonization than non-functionalized PAA-PVA after 14â¯days of incubation. Bacterial impairment was attributed to the damage of cell membranes and the formation of intracellular reactive oxygen species. ε-PL functionalized nanofibers did not display cytotoxicity to human corneal epithelial cells, HCEpC, in 24â¯h MTT assays. However, the viability of rapidly growing tumoral HeLa cells decreased >50% under the same conditions. The prepared biocompatible nanofibrous dressings with durable antibacterial activity show potential application as wound dressings and other biomedical uses.
Asunto(s)
Antibacterianos/farmacología , Vendajes , Materiales Biocompatibles , Polilisina/farmacología , Resinas Acrílicas/química , Antibacterianos/química , Antibacterianos/toxicidad , Bacterias/efectos de los fármacos , Epitelio Corneal/citología , Epitelio Corneal/efectos de los fármacos , Células HeLa , Humanos , Nanofibras , Tamaño de la Partícula , Polilisina/química , Polilisina/toxicidad , Alcohol Polivinílico/química , Especies Reactivas de Oxígeno/metabolismo , Factores de TiempoRESUMEN
High transfection efficiency and low cytotoxicity are the two key factors to be considered in the design of gene carriers. Herein, a novel and versatile gene carrier (PLL-RT) was prepared by introducing "molecular string" RT (i.e., p-toluylsulfonyl arginine) onto the polylysine backbone. The introduction of RT string contributed to the formation of multiple interactions between the polycationic gene carriers and cell membrane or DNA, as well as adopting α-helix conformation, all of which would be beneficial to enhance the gene transfection. In addition, RT string grafted onto other polycations such as hyperbranced PEI25k and dendrimer PAMAM could also acquire improved transfection efficiency and low cytotoxicity. Moreover, PLL-RT presented significant tumor inhibition effect in vivo. This work provided an effective strategy for constructing novel gene carriers with high transfection and low cytotoxicity.
Asunto(s)
ADN/genética , Técnicas de Transferencia de Gen , Polilisina/análogos & derivados , Tosilarginina Metil Éster/análogos & derivados , Animales , Cardiolipinas/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , ADN/metabolismo , Endocitosis/fisiología , Endosomas/metabolismo , Femenino , Humanos , Membranas Artificiales , Ratones Endogámicos BALB C , Conformación Molecular , Neoplasias/terapia , Tamaño de la Partícula , Polilisina/síntesis química , Polilisina/metabolismo , Polilisina/toxicidad , ARN Interferente Pequeño/administración & dosificación , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , ARN Interferente Pequeño/uso terapéutico , Tosilarginina Metil Éster/síntesis química , Tosilarginina Metil Éster/metabolismo , Tosilarginina Metil Éster/toxicidadRESUMEN
Osteoporosis is a major debilitating cause of fractures and decreases the quality of life in elderly patients. Bone homeostasis is maintained by bone forming osteoblasts and bone resorpting osteoclasts. Substantial evidences have shown that targeting osteoclasts using natural products is a promising strategy for the treatment of osteoporosis. In the current study, we investigated the osteoprotective effect of Abietic acid (AA) in in vitro and in vivo models of osteolysis. In vitro experiments demonstrated that, AA suppressed receptor activator of nuclear factor-kappa B ligand (RANKL)-induced osteoclastogenesis and F-actin ring formation in a concentration dependent manner. Mechanistically, AA abrogated RANKL-induced phosphorylation of IKKα/ß (ser 176/180), IkBα (ser 32), and inhibited the nuclear translocation of NF-κB. We also found that, AA attenuated the RANKL-induced phosphorylation of MAPKs and decreased the expression of osteoclast specific genes such as TRAP, DC-STAMP, c-Fos, and NFATc1. Consistent with in vitro results, in vivo Lipoploysaccharide (LPS)-induced osteolysis model showed that AA inhibited the LPS-induced serum surge in cytokines TNF-α and IL-6. µ-CT analysis showed that AA prevented the LPS-induced osteolysis. Furthermore, histopathology and TRAP staining results suggested that AA decreased the number of osteoclasts in LPS-injected mice. Taken together, we demonstrated that the osteoprotective action of AA is coupled with the inhibition of NF-κB and MAPK signaling and subsequent inhibition of NFATc1 and c-Fos activities. Hence, AA may be considered as a promising drug candidate for the treatment of osteoporosis.
Asunto(s)
Abietanos/administración & dosificación , Inflamación/tratamiento farmacológico , Osteogénesis/genética , Osteólisis/tratamiento farmacológico , Ligando RANK/genética , Actinas/genética , Animales , Diferenciación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Humanos , Inflamación/inducido químicamente , Inflamación/genética , Inflamación/patología , Ratones , Quinasas de Proteína Quinasa Activadas por Mitógenos/genética , FN-kappa B/genética , Factores de Transcripción NFATC/genética , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Osteogénesis/efectos de los fármacos , Osteólisis/inducido químicamente , Osteólisis/genética , Osteólisis/patología , Osteoporosis/inducido químicamente , Osteoporosis/tratamiento farmacológico , Osteoporosis/genética , Osteoporosis/patología , Fosfatidiletanolaminas/toxicidad , Fosforilación/efectos de los fármacos , Polilisina/análogos & derivados , Polilisina/toxicidad , Transducción de Señal/efectos de los fármacosRESUMEN
BACKGROUND: The delivery of plasmonic particles, such as gold nanorods, to the tumor microenvironment has attracted much interest in biomedical optics for topical applications as the photoacoustic imaging and photothermal ablation of cancer. However, the systemic injection of free particles still crashes into a complexity of biological barriers, such as the reticuloendothelial system, that prevent their efficient biodistribution. In this context, the notion to exploit the inherent features of tumor-tropic cells for the creation of a Trojan horse is emerging as a plausible alternative. RESULTS: We report on a convenient approach to load cationic gold nanorods into murine macrophages that exhibit chemotactic sensitivity to track gradients of inflammatory stimuli. In particular, we compare a new model of poly-L-lysine-coated particles against two alternatives of cationic moieties that we have presented elsewhere, i.e. a small quaternary ammonium compound and an arginine-rich cell-penetrating peptide. Murine macrophages that are exposed to poly-L-lysine-coated gold nanorods at a dosage of 400 µM Au for 24 h undertake efficient uptake, i.e. around 3 pg Au per cell, retain the majority of their cargo until 24 h post-treatment and maintain around 90% of their pristine viability, chemotactic and pro-inflammatory functions. CONCLUSIONS: With respect to previous models of cationic coatings, poly-L-lysine is a competitive solution for the preparation of biological vehicles of gold nanorods, especially for applications that may require longer life span of the Trojan horse, say in the order of 24 h. This biopolymer combines the cost-effectiveness of small molecules and biocompatibility and efficiency of natural peptides and thus holds potential for translational developments.
Asunto(s)
Macrófagos/metabolismo , Nanotubos/química , Animales , Movimiento Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Citocinas/análisis , Citocinas/metabolismo , Oro/química , Oro/farmacocinética , Oro/toxicidad , Macrófagos/química , Macrófagos/fisiología , Ratones , Nanotubos/toxicidad , Polilisina/química , Polilisina/farmacocinética , Polilisina/toxicidadRESUMEN
Cationic nucleopeptides belong to a family of synthetic oligomers composed by amino acids and nucleobases. Their capability to recognize nucleic acid targets and to cross cellular membranes provided the basis for considering them as novel non-covalent delivery agents for nucleic acid pharmaceuticals. Herein, starting from a 12-mer nucleopeptide model, the number of cationic residues was modulated in order to obtain new nucleopeptides endowed with high solubility in acqueous medium, acceptable bio-stability, low cytotoxicity and good capability to bind nucleic acid. Two candidates were selected to further investigate their potential as nucleic acid carriers, showing higher efficiency to deliver PNA in comparison with RNA. Noteworthy, this study encourages the development of nucleopeptides as new carriers to extend the known strategies for those nucleic acid analogues, especially PNA, that still remain difficult to drive into the cells.
Asunto(s)
Portadores de Fármacos/metabolismo , Ácidos Nucleicos de Péptidos/metabolismo , Polilisina/metabolismo , ARN/metabolismo , Timina/análogos & derivados , Timina/metabolismo , Cationes/síntesis química , Cationes/química , Cationes/metabolismo , Cationes/toxicidad , Línea Celular Tumoral , Permeabilidad de la Membrana Celular , Dicroismo Circular , Portadores de Fármacos/síntesis química , Portadores de Fármacos/química , Portadores de Fármacos/toxicidad , Humanos , Conformación de Ácido Nucleico , Hibridación de Ácido Nucleico , Ácidos Nucleicos de Péptidos/química , Ácidos Nucleicos de Péptidos/genética , Polilisina/síntesis química , Polilisina/química , Polilisina/toxicidad , ARN/química , ARN/genética , Solubilidad , Temperatura , Timina/síntesis química , Timina/toxicidad , Transfección/métodosRESUMEN
Sensing of intracellular singlet oxygen (1O2) is required in order to optimize photodynamic therapy (PDT). An optical nanoprobe is reported here for the optical determination of intracellular 1O2. The probe consists of a porous particle core doped with the commercial 1O2 probe 1,3-diphenylisobenzofuran (DPBF) and a layer of poly-L-lysine. The nanoparticle probes have a particle size of ~80 nm in diameter, exhibit good biocompatibility, improved photostability and high sensitivity for 1O2 in both absorbance (peak at 420 nm) and fluorescence (with excitation/emission peaks at 405/458 nm). Nanoprobes doped with 20% of DPBF are best suited even though they suffer from concentration quenching of fluorescence. In comparison with the commercial fluorescent 1O2 probe SOSG, 20%-doped DPBF-NPs (aged) shows higher sensitivity for 1O2 generated at an early stage. The best nanoprobes were used to real-time monitor the PDT-triggered generation of 1O2 inside live cells, and the generation rate is found to depend on the supply of intracellular oxygen. Graphical abstract A fluorescent nanoprobe featured with refined selectivity and improved sensitivity towards 1O2 was prepared from the absorption-based probe DBPF and used to real-time monitoring of the generation of intracellular 1O2 produced during PDT.
Asunto(s)
Benzofuranos/química , Colorantes Fluorescentes/química , Oxígeno Singlete/metabolismo , Benzofuranos/efectos de la radiación , Benzofuranos/toxicidad , Fluorescencia , Colorantes Fluorescentes/efectos de la radiación , Colorantes Fluorescentes/toxicidad , Células Hep G2 , Humanos , Luz , Nanopartículas/química , Nanopartículas/efectos de la radiación , Nanopartículas/toxicidad , Fotoquimioterapia , Polilisina/química , Polilisina/toxicidad , Oxígeno Singlete/análisis , Oxígeno Singlete/química , Espectrometría de Fluorescencia/métodosRESUMEN
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(Xi)], XiK) or (ii) oligo[dl-alanine] (poly[Lys(dl-Alam)], AK) or (iii) oligo[dl-alanine] with an additional amino acid (X) at the terminal position (poly[Lys(Xi-dl-Alam)] (XAK)) or (iv) at the position next to the polylysine backbone (poly[Lys(dl-Alam-Xi)] (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.
Asunto(s)
Péptidos/síntesis química , Péptidos/metabolismo , Polilisina/síntesis química , Polilisina/metabolismo , Cationes , Línea Celular Tumoral , Sistemas de Liberación de Medicamentos , Endocitosis , Humanos , Péptidos/toxicidad , Polilisina/toxicidad , Conformación Proteica , Relación Estructura-ActividadRESUMEN
Single-walled carbon nanotubes (SWCNTs) are thoroughly purified and dispersed in an aqueous solution of high molecular weight poly-L-lysine (pLlys). Human intestinal epithelial Caco-2/TC7 cells are incubated with the SWCNT dispersions in pLlys, and their effects on cell viability are studied by image flow cytometry. No significant changes are observed in the cell culture wells up to pLlys concentrations of 10 µg ml-1. However, high mortality is detected at pLlys concentrations of 100 µg ml-1. The presence of oxygen-free SWCNTs does not modify the effects of pLlys on cell cultures at any of the tested concentrations (≤1 µg ml-1). In addition, SWCNTs having an 8 wt.% of surface oxygen are tested with identical results. Thus, purified SWCNTs, even bearing oxygen functional groups, act as inert particles in the cell culture medium. This result supports the applicability of SWCNTs as carriers in pharmacological formulations against digestive tract diseases.
Asunto(s)
Coloides/toxicidad , Nanotubos de Carbono/toxicidad , Polilisina/toxicidad , Células CACO-2 , HumanosRESUMEN
In a previous study, a novel biodegradable multiblock copolymer, monomethoxy(poly-ethylene glycol)-poly(d,l-lactide-co-glycolide)-poly(l-lysine) (PEAL), was developed as a new drug carrier material. It is imperative to study the biocompatibility and degradation behavior of PEAL to pave the way for clinical applications. Here, we systematically demonstrated that the PEAL copolymer has the appropriate hydrophilicity and biosafety. The degradation rate of the PEAL films was obtained by observing changes in mass, molecular weight (Mw), Mw distribution and degradation products. The degradation rate was observed to have a highly positive correlation with the pH of the medium and negative correlation with the ratio of lactic acid to glycolic acid (LA/GA). Cytotoxicity tests indicated that the degradation products of the copolymer were non-toxic to cells. In zebrafish embryos, the PEAL nanoparticles had no obvious impact on heart rate, production of reactive oxygen species, mortality, or cell apoptosis, and they were observed to have a long circulation time. Therefore, the PEAL copolymer has great potential for use as a drug carrier material.
Asunto(s)
Portadores de Fármacos/metabolismo , Poliésteres/metabolismo , Polietilenglicoles/metabolismo , Polilisina/metabolismo , Animales , Coagulación Sanguínea/efectos de los fármacos , Plaquetas/citología , Plaquetas/efectos de los fármacos , Línea Celular , Activación de Complemento/efectos de los fármacos , Portadores de Fármacos/química , Portadores de Fármacos/farmacocinética , Portadores de Fármacos/toxicidad , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Nanopartículas/análisis , Nanopartículas/química , Nanopartículas/metabolismo , Nanopartículas/toxicidad , Poliésteres/química , Poliésteres/farmacocinética , Poliésteres/toxicidad , Polietilenglicoles/química , Polietilenglicoles/farmacocinética , Polietilenglicoles/toxicidad , Polilisina/química , Polilisina/farmacocinética , Polilisina/toxicidad , Especies Reactivas de Oxígeno/metabolismo , Pez CebraRESUMEN
Engineered PEG-cleavable catiomers based on poly-L-lysine have been developed as nonviral gene vectors, which have been found to be one of important methods to balance "PEG dilemma." In this protocol, we aim at the standardization of the method and procedure of PEG-cleavable catiomers. Major steps including ring-opening polymerization (ROP) of ε-benzyloxycarbonyl-L-lysine N-carboxyanhydride (zLL-NCA) monomers to yield PEG-cleavable polylysine, examination on bio-stability and bio-efficacy of its gene complexes are described.
Asunto(s)
Disulfuros/química , Portadores de Fármacos/química , Polietilenglicoles/química , Polilisina/química , ARN Interferente Pequeño/química , Transporte Biológico , Línea Celular , Portadores de Fármacos/metabolismo , Portadores de Fármacos/toxicidad , Estabilidad de Medicamentos , Humanos , Polilisina/metabolismo , Polilisina/toxicidad , PolimerizacionRESUMEN
MR imaging (MRI) upon cell labeling is an attractive and clinically translatable tool for longitudinally monitoring the survival and migration of stem cells. The common intracellular delivery of superparamagnetic iron oxide nanoparticles (SPIONs) via poly-L-lysine (PLL) requires a high SPION concentration and a long incubation period for appropriate cell labeling, which may negatively affect the viability and function of stem cells. In this study, we determined the performance of a new class of cationic polymersomes in transferring SPIONs into green fluorescence protein-modified mesenchymal stem cells (MSCs) for cellular MRI in acute ischemic stroke, compared with PLL-coated SPIONs. The results demonstrated that the polymersomes had comparable labeling efficiency and biological safety as well as a marginal benefit on post-transplantation cell survival; the polymersomes had the advantages of a relatively low SPION concentration and a substantially shorter labeling period compared with PLL-coated SPIONs. After transplantation, MSCs labeled using both methods offered a similar therapeutic effect on stroke, and cellular MRI could track the in vivo distribution and migration behavior of biologically active MSCs; however, MRI overestimated the true size of the cell grafts. SPION-loaded cationic polymersomes can be used as an alternative for the efficient, rapid, and safe labeling of stem cells for cellular MRI.
Asunto(s)
Rastreo Celular/métodos , Imagen por Resonancia Magnética/métodos , Nanopartículas de Magnetita , Polilisina , Trasplante de Células Madre , Accidente Cerebrovascular/terapia , Animales , Cationes , Supervivencia Celular/efectos de los fármacos , Proteínas Fluorescentes Verdes/química , Nanopartículas de Magnetita/química , Nanopartículas de Magnetita/toxicidad , Masculino , Polilisina/química , Polilisina/toxicidad , Ratas , Ratas Sprague-Dawley , Células Madre/citologíaRESUMEN
The present study was designed to evaluate the biocompatibility of nanoparticles polyethylene glycol (PEG)-poly L-lysine (PLL)-poly lactic-co-glycolic acid copolymer (PLGA) (PEG-PLL-PLGA) before clinical application. We applied some tests to assess the safety of PEG-PLL-PLGA nanoparticles (NPs). There was low cytotoxicity of PEG-PLL-PLGA NPs in vitro as detected by MTT assay. Cell apoptosis and intracellular accumulation of PEG-PLL-PLGA were determined by FCM assay. The apoptotic rate induced by nanoparticles and the fluorescence intensity of intracellular daunorubicin (DNR) demonstrated that DNR-PEG-PLL-PLGA could be taken up by the mouse fibroblast cells (L929 cells). Hemolysis test and micronucleus (MN) assay demonstrated that the nanoparticles have no obviously blood toxicity and genotoxicity. DNR-PEG-PLL-PLGA NPs were injected into mice through tail vein to calculate the median lethal dose (LD50), the results showed that they had a wide safe scale. Blood was taken by removing the eyeball of mice to study the influence of DNR-PEG-PLL-PLGA in hepatic and renal functions. The results revealed that there was no significant difference as compared with the control group. Interestingly, the pathologic changes of heart, liver, spleen, lung and kidney were observed in nanoparticles treated mice. Thus, this study demonstrates that PEG-PLL-PLGA NPs appear to be highly biocompatible and safe nanoparticles that can be suitable for further application in the treatment of tumor.