Light-activated conjugated polymer nanoparticles to defeat pathogens associated with bovine mastitis.

Bovine mastitis (BM) represents a significant challenge in the dairy industry. Limitations of conventional treatments have prompted the exploration of alternative approaches, such as photodynamic inactivation (PDI). In this study, we developed a PDI protocol to eliminate BM-associated pathogens using porphyrin-doped conjugated polymer nanoparticles (CPN). The PDI-CPN protocol was evaluated in four mastitis isolates of Staphylococcus and in a hyper-biofilm-forming reference strain. The results in planktonic cultures demonstrated that PDI-CPN exhibited a bactericidal profile upon relatively low light doses (∼9.6 J/cm2). Furthermore, following a seven-hour incubation period, no evidence of cellular reactivation was observed, indicating a highly efficient post-photodynamic inactivation effect. The successful elimination of bacterial suspensions encouraged us to test the PDI-CPN protocol on mature biofilms. Treatment using moderate light dose (∼64.8 J/cm2) reduced biofilm biomass and metabolic activity by up to 74% and 88%, respectively. The impact of PDI-CPN therapy on biofilms was investigated using scanning electron microscopy (SEM), which revealed nearly complete removal of the extracellular matrix and cocci. Moreover, ex vivo studies conducted on bovine udder skin demonstrated the efficacy of the therapy in eliminating bacteria from these scaffolds and its potential as a prophylactic method. Notably, the histological analysis of skin revealed no signs of cellular degeneration, suggesting that the protocol is safe and effective for BM treatment. Overall, this study demonstrates the potential of PDI-CPN in treating and preventing BM pathogens. It also provides insights into the effects of PDI-CPN on bacterial growth, metabolism, and survival over extended periods, aiding the development of effective control strategies and the optimization of future treatments.

Reusable antimicrobial antibiotic-free dressings obtained by photopolymerization.

In recent years significant efforts have been made to develop new materials for wound dressing with improved healing properties. However, the synthesis methods usually employed to this end are often complex or require several steps. We describe here the synthesis and characterization of antimicrobial reusable dermatological wound dressings based on N-isopropylacrylamide co-polymerized with [2-(Methacryloyloxy) ethyl] trimethylammonium chloride hydrogels (NIPAM-co-METAC). The dressings were obtained with a very efficient single-step synthesis procedure based on visible light (455 nm) by photopolymerization. To this end, F8BT nanoparticles of the conjugated polymer (poly(9,9-dioctylfluorene-alt-benzothiadiazole) - F8BT) were used as macro-photoinitiators, and a modified silsesquioxane was employed as crosslinker. Dressings obtained by this simple and gentle method show antimicrobial and wound healing properties, without the incorporation of antibiotics or any other additives. The physical and mechanical properties of these hydrogel-based dressings were evaluated, as well as their microbiological properties, through in vitro experiments. Results show that dressings with a molar ratio of METAC of 0.5 or higher exhibit high swelling capacity, appropriate water vapor transmission rate values, stability and thermal response, high ductility and adhesiveness. In addition, biological tests showed that the dressings have significant antimicrobial capacity. The best inactivation performance was found for hydrogels synthesized with the highest METAC content. The dressings were tested several times with fresh bacterial cultures, showing a bacterial kill efficiency of 99.99 % even after three repetitions in a row, employing the same dressing, demonstrating the intrinsic bactericidal property of the materials and their reusability. In addition, the gels show low hemolytic effect, high dermal biocompatibility and noticeable wound healing effects. Overall results demonstrate that some specific hydrogel formulations have potential application as dermatological dressings for wound healing and disinfection.

Sweet light o' mine: Photothermal and photodynamic inactivation of tenacious pathogens using conjugated polymers.

Each year a rising number of infections can not be successfully treated owing to the increasing pandemic of antibiotic resistant pathogens. The global shortage of innovative antibiotics fuels the emergence and spread of drug resistant microbes. Basic research, development, and applications of alternative therapies are urgently needed. Since the 90´s, light-mediated therapies have promised to be the next frontier combating multidrug-resistance microbes. These platforms have demonstrated to be a reliable, rapid, and efficient alternative to eliminate tenacious pathogens while avoiding the emergence of resistance mechanisms. Among the materials showing antimicrobial activity triggered by light, conjugated polymers (CPs) have risen as the most promising option to tackle this complex situation. These materials present outstanding characteristics such as high absorption coefficients, great photostability, easy processability, low cytotoxicity, among others, turning them into a powerful class of photosensitizer (PS)/photothermal agent (PTA) materials. Herein, we summarize and discuss the advances in the field of CPs with applications in photodynamic inactivation and photothermal therapy towards bacteria elimination. Additionally, a section of current challenges and needs in terms of well-defined benchmark experiments and conditions to evaluate the efficiency of phototherapies is presented.

Trojan horse monocyte-mediated delivery of conjugated polymer nanoparticles for improved photodynamic therapy of glioblastoma.

Aim: To assess monocyte-based delivery of conjugated polymer nanoparticles (CPNs) for improved photodynamic therapy (PDT) in glioblastoma (GBM). Materials & methods: Human monocyte cells (THP-1) and murine monocytes isolated from bone marrow (mBMDMs) were employed as stealth CPN carriers to penetrate into GBM spheroids and an orthotopic model of the tumor. The success of PDT, using this cell-mediated targeting strategy, was determined by its effect on the spheroids. Results: CPNs did not affect monocyte viability in the absence of light and did not show nonspecific release after cell loading. Activated monocytes incorporated CPNs in a higher proportion than monocytes in their naive state, without a loss of cellular functionality. In vitro PDT efficacy using cell-mediated delivery was superior to that using non vehiculized CPNs. Conclusion: CPN-loaded monocytes could efficiently deliver CPNs into GBM spheroids and the orthotopic model. Improved PDT in spheroids was confirmed using this delivery strategy.

Photodynamic Inactivation of ESKAPE Group Bacterial Pathogens in Planktonic and Biofilm Cultures Using Metallated Porphyrin-Doped Conjugated Polymer Nanoparticles.

Photodynamic inactivation (PDI) protocols using photoactive metallated porphyrin-doped conjugated polymer nanoparticles (CPNs) and blue light were developed to eliminate multidrug-resistant pathogens. CPNs-PDI protocols using varying particle concentrations and irradiation doses were tested against nine pathogenic bacterial strains including antibiotic-resistant bacteria of the ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogens group. The bactericidal effect was achieved in methicillin-resistant Staphylococus aureus (S. aureus) strains using low light doses (9.6-14.4 J/cm2), while Gram-negative bacteria required a higher light dose (28.8 J/cm2). The bacteria-CPN interaction was studied through flow cytometry, taking advantage of the intrinsic CPN fluorescence, demonstrating that CPNs efficiently bind to the bacterial envelope. Finally, the performance of CPNs-PDI was explored in biofilms; good antibiofilm ability and almost complete eradication were observed for S. aureus and Escherichia coli biofilms, respectively, using confocal microscopy. Overall, we demonstrated that CPNs-PDI is an efficient tool not only to kill superbugs as sessile cells but also to disrupt and eradicate biofilms of highly relevant pathogenic bacterial species.

Cytotoxicity and bioadhesive properties of poly-N-isopropylacrylamide hydrogel.

Several hydrogel surfaces present properties that simulate the mechanical and physicochemical features of extracellular matrix (ECM), providing a platform that mimic the native cellular milieus. Poly-N-isopropylacrylamide (PNIPAM) hydrogels are receiving attention in biomedical field due to their thermosensibility and soft texture. However, more extensive biocompatibility and cellular interactions studies with cell lines are needed. Therefore, the aim of this study is focus on evaluating the biocompatibility of PNIPAM through cytotoxicity, genotoxicity, and proliferation tests in murine preadipose cells (3T3-L1), human embryonic kidney cells (HEK293) and human carcinoma-derived cells (A549) in presence of hydrogel surfaces. Bioadhesive capacity above PNIPAM surfaces was also analyzed. MTT and neutral red uptake assays shown non-cytotoxic effect of PNIPAM in the studied cell lines. Genotoxicity was evaluated by the single-cell gel electrophoresis assay, where DNA damages were not detected. [3H]-thymidine staining allowed to corroborate that cell proliferation had progressed correctly. Adopted morphologies for each cell line over PNIPAM were similar to cell growing observed on polystyrene, indicating that the surfaces favor the cell attachment during 5 days' culture. The good biocompatibility of PNIPAM surfaces make it an interesting scaffold with clinical potential in tissue regeneration engineering, and a possible adipose and kidney tissue-engineered construct.

Assessment of polyaniline nanoparticles toxicity and teratogenicity in aquatic environment using Rhinella arenarum model.

With the rapid growth of nanotechnology and the applications of nanoparticles, environmental exposure to these particles is increasing. However, their impact in human and environmental health is not well studied. Anurans, with life stage comprising embryos, tadpoles and adults, have an extremely permeable skin which makes them excellent indicators of environmental health. This study evaluated the acute toxicity effects of polyaniline nanoparticles (PANI-Np) in different dispersant on embryos and larvae of Rhinella arenarum. The results showed that LC50 of PANI-Np dispersed in polyvinylpyrrolidone (PVP) were 1,500 mg/L, while LC50 by PANI-Np dispersed in PVP+PNIPAM (polyN-isopropylacrilamide) showed a highest toxicity (1,170 mg/L). The embryo teratogenicity increased with increasing exposure concentration in both kinds of PANI-Np although in PANI-Np1, there is an increased teratogenic effect associated with the polymer stabilizer PVP.

Polyaniline nanofibers: acute toxicity and teratogenic effect on Rhinella arenarum embryos.

The fate and effect of nanomaterials in the environment is of paramount importance towards the technological application of the materials. This work shows the ecotoxicological potential of polyaniline (PANI) nanofibers in the larvae Rhinella arenarum by means of AMPHITOX test. Acute toxicity of PANI nanofibers towards embryos of the common South American toad R. arenarum (Anura: bufonidae) was evaluated in the premetamorphosis (stage 25) larvae. The exposure of R. arenarum larvae to at dose of 150, 250 and 400 mg L(-1) resulted in 100% viability within 96 h exposure. The embryos at 2-4 blastomers stage (early life stage teratogenic test) revealed that embryos were not killed and no teratogenic effects were observed when embryos were incubated with PANI nanofibers (150 and 250 mg L(-1)), while only a growth retardation of embryos was induced at levels of 250 mg PANI nanofibers L(-1). On the other hand, at 400 mg L(-1) concentration, a reduction in the body length of larvae and tail malformation was observed. This results suggest that a concentration-dependent toxicity is operative, typified by phenotypes that had abnormal body axes. The presence of PANI nanofibers in gut contents and its excretion by larval stages of R. arenarum was confirmed by UV-visible spectroscopy.