Fish Skin Mucus Extracts: An Underexplored Source of Antimicrobial Agents.

The slow discovery of new antibiotics combined with the alarming emergence of antibiotic-resistant bacteria underscores the need for alternative treatments. In this regard, fish skin mucus has been demonstrated to contain a diverse array of bioactive molecules with antimicrobial properties, including peptides, proteins, and other metabolites. This review aims to provide an overview of the antimicrobial molecules found in fish skin mucus and its reported in vitro antimicrobial capacity against bacteria, fungi, and viruses. Additionally, the different methods of mucus extraction, which can be grouped as aqueous, organic, and acidic extractions, are presented. Finally, omic techniques (genomics, transcriptomics, proteomics, metabolomics, and multiomics) are described as key tools for the identification and isolation of new antimicrobial compounds. Overall, this study provides valuable insight into the potential of fish skin mucus as a promising source for the discovery of new antimicrobial agents.

Glioblastoma-Derived Small Extracellular Vesicles: Nanoparticles for Glioma Treatment.

Glioblastoma (GBM), characterized by fast growth and invasion into adjacent tissue, is the most aggressive cancer of brain origin. Current protocols, which include cytotoxic chemotherapeutic agents, effectively treat localized disease; however, these aggressive therapies present side effects due to the high doses administered. Therefore, more efficient ways of drug delivery have been studied to reduce the therapeutic exposure of the patients. We have isolated and fully characterized small extracellular vesicles (EVs) from seven patient-derived GBM cell lines. After loading them with two different drugs, Temozolomide (TMZ) and EPZ015666, we observed a reduction in the total amount of drugs needed to trigger an effect on tumor cells. Moreover, we observed that GBM-derived small EVs, although with lower target specificity, can induce an effect on pancreatic cancer cell death. These results suggest that GBM-derived small EVs represent a promising drug delivery tool for further preclinical studies and potentially for the clinical development of GBM treatments.

The Immune System of Marine Organisms as Source for Drugs against Infectious Diseases.

The high proliferation of microorganisms in aquatic environments has allowed their coevolution for billions of years with other living beings that also inhabit these niches. Among the different existing types of interaction, the eternal competition for supremacy between the susceptible species and their pathogens has selected, as part of the effector division of the immune system of the former ones, a vast and varied arsenal of efficient antimicrobial molecules, which is highly amplified by the broad biodiversity radiated, above any others, at the marine habitats. At present, the great recent scientific and technological advances already allow the massive discovery and exploitation of these defense compounds for therapeutic purposes against infectious diseases of our interest. Among them, antimicrobial peptides and antimicrobial metabolites stand out because of the wide dimensions of their structural diversities, mechanisms of action, and target pathogen ranges. This revision work contextualizes the research in this field and serves as a presentation and scope identification of the Special Issue from Marine Drugs journal "The Immune System of Marine Organisms as Source for Drugs against Infectious Diseases".

Formation of Multicolor Nanogels Based on Cationic Polyfluorenes and Poly(methyl vinyl ether-alt-maleic monoethyl ester): Potential Use as pH-Responsive Fluorescent Drug Carriers.

In this study, we employed the copolymer poly(methyl vinyl ether-alt-maleic monoethyl ester) (PMVEMA-Es) and three fluorene-based cationic conjugated polyelectrolytes to develop fluorescent nanoparticles with emission in the blue, green and red spectral regions. The size, Zeta Potential, polydispersity, morphology, time-stability and fluorescent properties of these nanoparticles were characterized, as well as the nature of the interaction between both PMVEMA-Es and fluorescent polyelectrolytes. Because PMVEMA-Es contains a carboxylic acid group in its structure, the effects of pH and ionic strength on the nanoparticles were also evaluated, finding that the size is responsive to pH and ionic strength, largely swelling at physiological pH and returning to their initial size at acidic pHs. Thus, the developed fluorescent nanoparticles can be categorized as pH-sensitive fluorescent nanogels, since they possess the properties of both pH-responsive hydrogels and nanoparticulate systems. Doxorubicin (DOX) was used as a model drug to show the capacity of the blue-emitting nanogels to hold drugs in acidic media and release them at physiological pH, from changes in the fluorescence properties of both nanoparticles and DOX. In addition, preliminary studies by super-resolution confocal microscopy were performed, regarding their potential use as image probes.

Self-assembled systems of water soluble metal 8-hydroxyquinolates with surfactants and conjugated polyelectrolytes.

We have studied the interaction of 8-hydroxyquinoline-5-sulfonate (8-HQS) with the metal ions Al(iii) and Zn(ii) in aqueous solution in the presence of tetraalkylammonium surfactants using UV/vis absorption, fluorescence, NMR spectroscopy and electrical conductivity measurements, complemented by DFT calculations and molecular dynamics (MD) simulations. Under appropriate conditions, complexes between 8-HQS and metal ions form rapidly, and have similar electronic, spectroscopic and photophysical properties to the corresponding metal quinolates, such as Alq3. These interact with the cationic surfactants, leading to marked increases in fluorescence intensity. However, significant differences are seen in the behavior of the two metal ions. With aluminium, a stable [Al(8-QS)3](3-) anion is formed, and interacts, predominantly through electrostatic interactions, with the surfactant, without disrupting the metal ion coordination sphere. In contrast, with Zn(ii), there is a competition between the metal ion and surfactants in the interaction with 8-HQS, although the [Zn(8-QS)2(H2O)2](2-) species is stable at appropriate pH and surfactant concentration. The studies are extended to systems with the conjugated polyelectrolyte (CPE) poly-(9,9-bis(6-N,N,N-trimethylammonium)hexyl)-fluorene-phenylene bromide (HTMA-PFP), which has a similar alkylammonium chain to the surfactants. Mixing metal salt, 8-HQS and HTMA-PFP in the presence of a nonionic surfactant leads to the formation of a metal complex/CPE supramolecular assembly between the conjugated polyelectrolyte and the metal/8-HQS complex, as demonstrated by electronic energy transfer. The potential of these systems in sensing, light harvesting, and electron injection/transport layers in organic semiconductor devices is discussed.

Bovine serum albumin and lysozyme nanofibers as versatile platforms for preserving loaded bioactive compounds.

In this study, the electrospinning procedure was optimized to create polyethylene oxide (PEO) NFs highly enriched in proteins with non-structural functions while preserving their activity. For this purpose, several immune-related proteins of low, medium and high molecular weights were used as molecular models. Initially, the electrospinning parameters were adjusted using 3 % w/w PEO and bovine serum albumin (BSA, 5-20 % w/w). As determined by FESEM, their average diameters ranged from 301 to 752 nm, and those with higher protein content (15-20 %) yielded more uniform NFs in both size and morphology terms. Protein integrity remained stable as determined by SDS-PAGE and FTIR. Similar results were observed for the polypeptide lysozyme (LYZ) when incorporated in NFs under these settings. To further explore the potential of these materials, the antimicrobial peptide piscidin (PIS) and an antibody (Ab, HRP-IgG) were used to produce BSA/PIS, LYZ/PIS and BSA/Ab NFs and evaluate the preservation of their activity. The antibacterial assays showed that, in most bacterial species, the activity of PIS remained consistent after being incorporated into the NFs. Furthermore, the activity of HRP-IgG was maintained within the NFs, with enhanced preservation observed in BSA/Ab NFs. These findings expand the possibilities of protein utilization across various applications through nanomaterials.

Use of the conjugated polyelectrolyte poly{[9,9-bis(6'-N,N,N-trimethylammonium)hexyl]fluorene-phenylene} bromide (HTMA-PFP) as a fluorescent membrane marker.

The present work explores the potential use of the conjugated cationic polyfluorene {[9,9-bis(6'-N,N,N-trimethylammonium)hexyl]fluorene-phenylene} bromide (HTMA-PFP) as a fluorescent membrane marker. To this end, the interaction of the polyelectrolyte with anionic model membranes has been investigated using different biophysical approaches. High affinity interaction was confirmed through alterations in the fluorescence spectrum of HTMA-PFP and by Förster resonance energy transfer (FRET) analysis. Quenching data indicate that once HTMA-PFP interacts with the membrane, it penetrates in the hydrophobic core embedded in the lipid bilayer where it presents high fluorescence quantum yield and photostability. Leakage experiments and dynamic light scattering (DLS) measurements show that the integrity of the lipid vesicles is maintained after polymer incorporation since no vesicle fusion or decomposition into small fragments is detectable. This conclusion is supported by fluorescence microscopy images, which confirm that polyelectrolyte interacts with the vesicle, labeling the lipid membrane without altering its morphology. Further experiments performed as a function of temperature indicate that the polymer is accommodated in the membrane without inducing significant loss of lipid cooperativity and without altering the packing of lipids within the bilayer. Finally, results show that polyelectrolyte fluorescence is sensitive to the large structural changes taking place in the lipid bilayer at the lipid phase transition. All these results confirm the ability of HTMA-PFP to visualize membrane structures and to monitor membrane processes.

Formation and characterization of stable fluorescent complexes between neutral conjugated polymers and cyclodextrins.

Solubilisation and stabilization of conjugated polymers, CPs, in aqueous media remains a challenge for many researches trying to extend the biological and environmental applications of this kind of polymers. A number of different alternatives have been considered to address this problem, which are mostly based on the enhancement of the macromolecule polarity, by appending hydrophilic side chains on the polymer backbone. In this work we have investigated a new strategy in which water solubilization is reached by external addition of classical cyclodextrins (α-, ß- and γ-CDs) to a solution of non-polar CPs. This strategy allows working with such polymers eliminating the need to synthesize new water-soluble species. The polymer selected for the study was poly-[9,9-bis(6'-bromohexyl-2,7-fluoren-dyil)-co-alt-(benzene-1,4-diy)], PFPBr(2), a polyfluorene previously synthesized in our laboratory. Results show that PFPBr(2) forms fluorescent complexes in aqueous media with ß-CD and γ-CD, and much less efficiently with α-CD, probably due to the small size of its cavity. The new PFPBr(2)/CD complexes are stable in time and in a large range of pH, however, at high concentration and temperature, they tend to aggregate and precipitate. In order to increase stabilization and minimize polymer aggregation, complexes were encapsulated inside the pores of silica glasses fabricated using the sol-gel process, obtaining transparent and fluorescent hybrid matrices which were stable in time and temperature. In addition, immobilization of the complexes allows an easy manipulation of the material, thus offering promising applications in the development of biological and chemical sensors.

Phytochemical-Based Nanomaterials against Antibiotic-Resistant Bacteria: An Updated Review.

Antibiotic-resistant bacteria (ARB) is a growing global health threat, leading to the search for alternative strategies to combat bacterial infections. Phytochemicals, which are naturally occurring compounds found in plants, have shown potential as antimicrobial agents; however, therapy with these agents has certain limitations. The use of nanotechnology combined with antibacterial phytochemicals could help achieve greater antibacterial capacity against ARB by providing improved mechanical, physicochemical, biopharmaceutical, bioavailability, morphological or release properties. This review aims to provide an updated overview of the current state of research on the use of phytochemical-based nanomaterials for the treatment against ARB, with a special focus on polymeric nanofibers and nanoparticles. The review discusses the various types of phytochemicals that have been incorporated into different nanomaterials, the methods used to synthesize these materials, and the results of studies evaluating their antimicrobial activity. The challenges and limitations of using phytochemical-based nanomaterials, as well as future directions for research in this field, are also considered here. Overall, this review highlights the potential of phytochemical-based nanomaterials as a promising strategy for the treatment against ARB, but also stresses the need for further studies to fully understand their mechanisms of action and optimize their use in clinical settings.

Viricidal Activity of Thermoplastic Polyurethane Materials with Silver Nanoparticles.

The use of diverse Ag-based nanoparticulated forms has shown promising results in controlling viral propagation. In this study, a commercial nanomaterial consisting of ceramic-coated silver nanoparticles (AgNPs) was incorporated into thermoplastic polyurethane (TPU) plates using an industrial protocol, and the surface composition, ion-release dynamics and viricidal properties were studied. The surface characterization by FESEM-EDX revealed that the molar composition of the ceramic material was 5.5 P:3.3 Mg:Al and facilitated the identification of the embedded AgNPs (54.4 ± 24.9 nm). As determined by ICPMS, the release rates from the AgNP-TPU into aqueous solvents were 4 ppm/h for Ag and Al, and 28.4 ppm/h for Mg ions. Regarding the biological assays, the AgNP-TPU material did not induce significant cytotoxicity in the cell lines employed. Its viricidal activity was characterized, based on ISO 21702:2019, using the Spring viraemia of carp virus (SVCV), and then tested against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The results demonstrated that AgNP-TPU materials exhibited significant (75%) and direct antiviral activity against SVCV virions in a time- and temperature-dependent manner. Similar inhibition levels were found against SARS-CoV-2. These findings show the potential of AgNP-TPU-based materials as a supporting strategy to control viral spread.

New therapy for pancreatic cancer based on extracellular vesicles.

Pancreatic Ductal Adenocarcinoma (PDAC), is the most common aggressive cancer of the pancreas. The standard care of PDAC includes tumor resection and chemotherapy, but the lack of early diagnosis and the limited response to the treatment worsens the patient's condition. In order to improve the efficiency of chemotherapy, we look for more efficient systems of drug delivery. We isolated and fully characterized small Extracellular Vesicles (EVs) from the RWP-1 cell line. Our study indicates that the direct incubation method was the most efficient loading protocol and that a minimum total amount of drug triggers an effect on tumor cells. Therefore, we loaded the small EVs with two chemotherapeutic drugs (Temozolomide and EPZ015666) by direct incubation method and the amount of drug loaded was measured by high-performance liquid chromatography (HPLC). Finally, we tested their antiproliferative effect on different cancer cell lines. Moreover, the system is highly dependent on the drug structure and therefore RWP-1 small EVsTMZ were more efficient than RWP-1 small EVsEPZ015666. RWP-1 derived small EVs represent a promising drug delivery tool that can be further investigated in preclinical studies and its combination with PRMT5 inhibitor can be potentially developed in clinical trials for the treatment of PDAC.

Conjugated polymer microspheres for "turn-off"/"turn-on" fluorescence optosensing of inorganic ions in aqueous media.

The synthesis and characterization of a novel water-compatible microsized material, based on fluorescent conjugated polymers (CPs), and its applicability for optical sensing of inorganic ions of environment interest (copper and cyanide) in water media is here described. Polyfluorene-based fluorescent CPs were synthesized and functionalized with imidazole moieties (selective recognition element) and a terminal double bond (covalently linked to an organic matrix) through a postfunctionalization strategy. Further, microspheres of the novel imidazole-functionalized fluorescent CPs, able to work in water media, were synthesized via a microemulsion and polymerization procedure. The synthesized imidazole-functionalized CP microspheres were then evaluated as fluorescence "turn-Off" sensing materials for Cu(2+) detection in aqueous media. Analyte detection was based on the quenching effect of the Cu(2+), selectively recognized by the imidazole group, on the polymer fluorescence emission. The developed optosensor exhibits a detection limit of 1 µg/L for the determination of Cu(2+) in water with a reproducibility of 4%. The synthesized microsized material was also evaluated for the "turn-on" optosensing of cyanide in water, measuring the recovery of the emission signal from the CP that has been previously deactivated by the presence of quencher species. The "turn-On" optosensor allows the selective determination of free cyanide in aqueous solution with high sensitivity (detection limit of 18 µg/L), obtaining a reproducibility of 2.9%. A high sample throughput (between 7 and 12 samples per hour) was achieved in both cases. Analytical applicability of the fluorescent CP microsphere materials has been successfully demonstrated by tap and mineral water analysis.

L-Menthol-Loadable Electrospun Fibers of PMVEMA Anhydride for Topical Administration.

Poly(methyl vinyl ether-alt-maleic anhydride) (PMVEMA) of 119 and 139 molecular weights (P119 and P139, respectively) were electrospun to evaluate the resulting fibers as a topical delivery vehicle for (L-)menthol. Thus, electrospinning parameters were optimized for the production of uniform bead-free fibers from 12% w/w PMVEMA (±2.3% w/w menthol) solutions, and their morphology and size were characterized by field emission scanning electron microscopy (FESEM). The fibers of P119 (F119s) and P139 (F139s) showed average diameter sizes of approximately 534 and 664 nm, respectively, when unloaded, and 837 and 1369 nm when loaded with menthol. The morphology of all types of fibers was cylindrical except for F139s, which mostly displayed a double-ribbon-like shape. Gas chromatography-mass spectrometry (GC-MS) analysis determined that not only was the menthol encapsulation efficiency higher in F139s (92% versus 68% in F119s) but also that its stability over time was higher, given that in contrast with F119s, no significant losses in encapsulated menthol were detected in the F139s after 10 days post-production. Finally, in vitro biological assays showed no significant induction of cytotoxicity for any of the experimental fibers or in the full functionality of the encapsulated menthol, as it achieved equivalent free-menthol levels of activation of its specific receptor, the (human) transient receptor potential cation channel subfamily M (melastatin) member 8 (TRPM8).

How to change the aggregation in the DNA/surfactant/cationic conjugated polyelectrolyte system through the order of component addition: anionic versus neutral surfactants.

The competitive interaction has been studied between double-stranded DNA (dsDNA), the cationic conjugated polyelectrolyte (CPE) poly[9,9-bis(6-N,N,N-trimethylamonium)hexyl)-fluorene-phenylene)] bromide (HTMA-PFP) and anionic or neutral surfactants (sodium dodecyl sulfonate, SDSu, and n-dodecyl pentaoxyethylene glycol ether, C(12)E(5)) in 4% (v/v) dimethyl sulfoxide (DMSO)-water using UV/visible absorption and fluorescence spectroscopy. Dramatic changes are observed in the spectroscopic behavior of the system depending on the order of addition of the reagents, the surfactant charge, and concentration range. If the neutral C(12)E(5) is added to the HTMA-PFP/dsDNA complex, no significant spectroscopic changes are observed. However, if SDSu is added to the same complex, a dramatic increase of the absorbance and emission intensity is observed for surfactant concentrations above the critical micelle concentration (cmc). In contrast, if dsDNA is added to HTMA-PFP/surfactant systems (with surfactant concentrations above their cmc) no significant changes are observed with SDSu, while a dramatic quenching of polymer emission is observed with C(12)E(5), which can be explained quantitatively in terms of HTMA-PFP/surfactant/DNA complexation and the subsequent polymer aggregation upon charge neutralization. The results are compared with those for the binary systems (HTMA-PFP/DNA and HTMA-PFP/surfactants) and indicate the importance of electrostatic interactions between HTMA-PFP and oppositely charged species in the aggregation processes.

Formation of complexes between the conjugated polyelectrolyte poly{[9,9-bis(6'-N,N,N-trimethylammonium)hexyl]fluorene-phenylene} bromide (HTMA-PFP) and human serum albumin.

The interaction between the conjugated polyelectrolyte poly{[9,9-bis(6'-N,N,N-trimethylammonium)hexyl]fluorene-phenylene} bromide (HTMA-PFP) and human serum albumin (HSA) has been investigated from changes observed in both the spectroscopic properties of HTMA-PFP and the intrinsic fluorescence of HSA. Absorption and fluorescence spectra of HTMA-PFP suggest that HTMA-PFP and HSA form polymer-protein complexes due to electrostatic interactions between the cationic side chains of HTMA-PFP and the negatively charged surface of the protein. Interaction between both macromolecules induces an increase in the fluorescence signal of HTMA-PFP, which suggests that hydrophobic forces also contribute to the polymer-protein complex stabilization. In addition, this interaction causes a decrease in the HSA fluorescence, partially due to static quenching and energy transfer between both macromolecules. Effects of HTMA-PFP on the thermal stability and protein conformation were explored from CD experiments. Results indicate that as polymer is added it binds to HSA and initiates unfolding. This unfolding process induces HTMA-PFP chains to become more extended, disrupting backbone interactions and increasing polymer fluorescence intensity.

Electrospun Nanomaterials: Applications in Food, Environmental Remediation, and Bioengineering.

Among the large number of methods to fabricate nanofibers[…] .

Physico-Chemically Distinct Nanomaterials Synthesized from Derivates of a Poly(Anhydride) Diversify the Spectrum of Loadable Antibiotics.

Recent advances in the field of nanotechnology such as nanoencapsulation offer new biomedical applications, potentially increasing the scope and efficacy of therapeutic drug delivery. In addition, the discovery and development of novel biocompatible polymers increases the versatility of these encapsulating nanostructures, enabling chemical properties of the cargo and vehicle to be adapted to specific physiological requirements. Here, we evaluate the capacity of various polymeric nanostructures to encapsulate various antibiotics of different classes, with differing chemical structure. Polymers were sourced from two separate derivatives of poly(methyl vinyl ether-alt-maleic anhydride) (PMVE/MA): an acid (PMVE/MA-Ac) and a monoethyl ester (PMVE/MA-Es). Nanoencapsulation of antibiotics was attempted through electrospinning, and nanoparticle synthesis through solvent displacement, for both polymers. Solvent incompatibilities prevented the nanoencapsulation of amikacin, neomycin and ciprofloxacin in PMVE/MA-Es nanofibers. However, all compounds were successfully loaded into PMVE/MA-Es nanoparticles. Encapsulation efficiencies in nanofibers reached approximately 100% in all compatible systems; however, efficiencies varied substantially in nanoparticles systems, depending on the tested compound (14%-69%). Finally, it was confirmed that both these encapsulation processes did not alter the antimicrobial activity of any tested antibiotic against Staphylococcus aureus and Escherichia coli, supporting the viability of these approaches for nanoscale delivery of antibiotics.

Fluorene-Based Donor-Acceptor Copolymers Containing Functionalized Benzotriazole Units: Tunable Emission and their Electrical Properties.

Monomers 4,7-dibromo-2H-benzo[d]1,2,3-triazole (m1) and 4,7-(bis(4-bromophenyl)ethynyl)-2H-benzo[d]1,2,3-triazole (m2) have been synthesized in good yields using different procedures. Monomers m1 and m2 have been employed for building new copolymers of fluorene derivatives by a Suzuki reaction under microwave irradiation using the same conditions. In each case different chain lengths have been achieved, while m1 gives rise to polymers for m2 oligomers have been obtained (with a number of monomer units lower than 7). Special interest has been paid to their photophysical properties due to excited state properties of these D-A units alternates, which have been investigated by density functional theory (DFT) calculations using two methods: (i) An oligomer approach and (ii) by periodic boundary conditions (PBC). It is highly remarkable the tunability of the photophysical properties as a function of the different monomer functionalization derived from 2H-benzo[d]1,2,3-triazole units. In fact, a strong modulation of the absorption and emission properties have been found by functionalizing the nitrogen N-2 of the benzotriazole units or by elongation of the π-conjugated core with the introduction of alkynylphenyl groups. Furthermore, the charge transport properties of these newly synthesized macromolecules have been approached by their implementation in organic field-effect transistors (OFETs) in order to assess their potential as active materials in organic optoelectronics.

Interaction between poly(9,9-bis(6'-N,N,N-trimethylammonium)hexyl)fluorene phenylene bromide and DNA as seen by spectroscopy, viscosity, and conductivity: effect of molecular weights and DNA secondary structure.

The interaction between three poly(9,9-bis(6-N,N,N-trimethylammonium)hexyl)fluorene phenylene) bromide (HTMA-PFP) samples of different molecular weights (Mn=14.5, 30.1 and 61.3 kg/mol) and both dsDNA and ssDNA secondary structures has been studied using UV-visible absorption and fluorescence spectroscopies (including steady-state, time-resolved, and anisotropy measurements for the latter), viscosity, and electrical conductivity in 4% (v/v) DMSO-water mixtures. At low nucleic acid concentrations, formation of a 1:1 complex in terms of HTMA-PFP repeat units and DNA bases occurs. This interaction results in quenching of polymer emission. For higher molar ratios of DNA to HTMA-PFP, corresponding to charge neutralization, a second process is observed that is attributed to aggregate formation. From the changes in the absorption spectra, the polymer aggregation constant and the aggregate absorption spectra were calculated by applying an iterative method. Polymer aggregation dramatically quenches HTMA-PFP fluorescence in the region of the electroneutrality point. Under these conditions, the ratio of the emission intensity at 412 nm (maximum) to that at 434 nm (I412/I434) reaches a minimum, the electrical conductivity decreases, and the viscosity of the solution remains constant, showing that the DNA concentration can be determined through various HTMA-PFP physicochemical properties. With respect to the photophysical parameters (emission quantum yield, shape and shift of emission spectra), no significant differences were observed between dsDNA and ssDNA or with conjugated polymer or DNA molecular weight. The two short-lived components in the fluorescence decays are attributed to the presence of aggregates. Aggregates are also suggested to be responsible for the decrease in the fluorescence anisotropy through interchain exciton migration.

Polyfluorene-Based Multicolor Fluorescent Nanoparticles Activated by Temperature for Bioimaging and Drug Delivery.

Multifunctional nanoparticles have been attracting growing attention in recent years because of their capability to integrate materials with different features in one entity, which leads them to be considered as the next generation of nanomedicine. In this work, we have taken advantage of the interesting properties of conjugated polyelectrolytes to develop multicolor fluorescent nanoparticles with integrating imaging and therapeutic functionalities. With this end, thermosensitive liposomes were coated with three recently synthesized polyfluorenes: copoly-((9,9-bis(6'-N,N,N-trimethylammonium)hexyl)-2,7-(fluorene)-alt-1,4-(phenylene)) bromide (HTMA-PFP), copoly-((9,9-bis(6'-N,N,N-trimethylammonium)hexyl)-2,7-(fluorene)-alt-4,7-(2- (phenyl)benzo(d) (1,2,3) triazole)) bromide (HTMA-PFBT) and copoly-((9,9-bis(6'-N,N,N- trimethylammonium)hexyl)-2,7-(fluorene)-alt-1,4-(naphtho(2,3c)-1,2,5-thiadiazole)) bromide (HTMA-PFNT), in order to obtain blue, green and red fluorescent drug carriers, respectively. The stability, size and morphology of the nanoparticles, as well as their thermotropic behavior and photophysical properties, have been characterized by Dynamic Light Scattering (DLS), Zeta Potential, transmission electron microscope (TEM) analysis and fluorescence spectroscopy. In addition, the suitability of the nanostructures to carry and release their contents when triggered by hyperthermia has been explored by using carboxyfluorescein as a hydrophilic drug model. Finally, preliminary experiments with mammalian cells demonstrate the capability of the nanoparticles to mark and visualize cells with different colors, evidencing their potential use for imaging and therapeutic applications.