Photodynamic therapy of cationic and anionic BSA-curcumin nanoparticles on amastigotes of Leishmania braziliensis and Leishmania major and Leishmania amazonensis.

Cutaneous leishmaniasis is a neglected disease prevalent in tropical countries, and conventional treatment can cause several serious side effects. Photodynamic therapy (PDT) can be considered a promising treatment alternative, as it is non-invasive therapy that has no side effects and uses accessible and low-cost substances, such as curcumin. This study evaluated the PDT response with cationic and anionic BSA nanoparticles encapsulated with curcumin in macrophages infected with L. braziliensis, L. major, and L. amazonensis. The nanoparticle system was characterized using a steady-state technique, scanning electron microscopy (SEM) study, and its biological activity was evaluated using macrophage cell lines infected with different Leishmania species. All spectroscopy measurements demonstrated that BSA curcumin (BSACur) has good photophysical properties, and confocal microscopy shows that macrophages and protozoa internalized the nanoparticles. The viability test demonstrated that at low concentrations, such as 0.1, 0.7, and 1.0 µmol. L-1, there was a decrease in cell viability after PDT application. Furthermore, a decrease in the number of parasites recovered was observed in the PDT groups. The results allowed us to conclude that curcumin loaded into BSA nanoparticles may have potential application in drug delivery systems for PDT protocols, demonstrating reduced cell viability at lower concentrations than free curcumin.

Hydroxyapatite microspheres used as a drug delivery system for gliosarcoma strain 9l/Lacz treatment by photodynamic therapy protocols.

BACKGROUND: Hydroxyapatite (HAp) presents similarities with the human bone structure and presents properties such as biodegradability, biocompatibility, and osteoconductivity, which favors its use in prostheses implants and enables its use as a vehicle for the delivery of photosensitizers (PS) from systems of release (DDS) for photodynamic therapy applications Methods: In this work was to synthesized hydroxyapatite microspheres (meHAp), encapsulated with chloroaluminium phthalocyanine (ClAlPc), for DDS. meHAp was synthesized using vaterite as a template. The drug was encapsulated by mixing meHAp and a 50.0 mg.mL-1 ClAlPc solution. Photochemical, photophysical, and photobiological studies characterized the system. RESULTS: The images from the SEM analysis showed the spherical form of the particles. All spectroscopic results showed excellent photophysical parameters of the drug studied when served in the meHAp system. The incorporation efficiency was 57.8 %. The trypan blue exclusion test results showed a significant reduction (p < 0.05) in cell viability for the groups treated with PDT at all concentrations above 250 µg.mL-1. In 9 L/lacZ gliosarcoma cells, PDT mediated at concentrations from 250 to 62.5 µg.mL-1 reduced cell viability by more than 98 %. In the cell internalization study, it was possible to observe the internalization of phthalocyanines at 37 °C, with the accumulation of PS in the cytoplasm and inside the nucleus in the two tested concentrations. CONCLUSIONS: From all the results presented throughout the article, the meHAp system shows promise for use as a modified release system (DSD) in photodynamic therapy.

Vaterite microparticle-loaded methylene blue for photodynamic activity in macrophages infected with Leishmania braziliensis.

Calcium carbonate (CaCO3) exhibits a variety of crystalline phases, including the anhydrous crystalline polymorphs calcite, aragonite, and vaterite. Developing porous calcium carbonate microparticles in the vaterite phase for the encapsulation of methylene blue (MB) as a photosensitizer (PS) for use in photodynamic therapy (PDT) was the goal of this investigation. Using an adsorption approach, the PS was integrated into the CaCO3 microparticles. The vaterite microparticles were characterized by scanning electron microscopy (SEM) and steady-state techniques. The trypan blue exclusion method was used to measure the biological activity of macrophages infected with Leishmania braziliensis in vitro. The vaterite microparticles produced are highly porous, non-aggregated, and uniform in size. After encapsulation, the MB-loaded microparticles kept their photophysical characteristics. The carriers that were captured allowed for dye localization inside the cells. The results obtained in this study indicated that the MB-loaded vaterite microparticles show promising photodynamic activity in macrophages infected with Leishmania braziliensis.

Synthesis, characterization, and evaluation of chloroaluminium phthalocyanine incorporated in poly(ε-caprolactone) nanoparticles for photodynamic therapy.

BACKGROUND: The use of nanotechnology has been widely used in biomedical science, including orthopedic implants, tissue engineering, cancer therapy and drug elution from nanoparticle systems, such as poly-caprolactone (PCL) nanoparticles, which stand out mainly for their biocompatibility, being considered as effective carriers for photosensitizing drugs (PS) in photodynamic therapy (PDT) protocols. METHODS: This manuscript describes the synthesis and characterization of PCL nanoparticles for controlled release of the drug chloro-aluminum phthalocyanine (ClAlPc) as a photosensitizer for application in PDT. The PCL-ClAlPc nanoparticles were developed by the nanoprecipitation process. The structure and morphology of the nanoparticles were studied with scanning electron microscopy (SEM) and with Fourier transform infrared (FTIR). The size of nanomaterials was studied using the Dynamic Light Scattering (DLS) method. Photophysical and photochemical characterizations were performed. Subsequently, photobiological studies were also used to characterize the system. RESULTS: The nanoparticles had an average diameter of 384.7 ± 138.6 nm and a polydispersity index of 0.153. SEM analysis revealed that the system formed a spherical shape typical of these delivery systems. Charging efficiency was 82.1% ± 1.2%. The phthalocyanine-loaded PCL nanoparticles maintained their photophysical behavior after encapsulation. Cell viability was determined after the dark toxicity test, and it was possible to observe that there was no evidence of toxicity in the dark, for all concentrations tested. The assay also revealed that adenocarcinoma cells treated with free ClAlPc and in the nanoformulation showed 100% cell death when subjected to PDT protocols. The intracellular location of the photosensitizer indicated a high potential for accumulation in the cytoplasm and nucleus. CONCLUSIONS: From the photophysical, photochemical and photobiological analyzes obtained, it was possible to observe that the development of PCL nanoparticles encapsulated with ClAlPc, by the nanoprecipitation method was adequate and that the in vivo release study is efficient to reduce the release rate and attenuate the burst of PS loaded on PCL nanoparticles. The results reinforce that the use of this system as drug delivery systems is useful in PDT protocols.

Synthesis and characterization of photosensitive gelatin-based hydrogels for photodynamic therapy in HeLa-CCL2 cell line.

BACKGROUND: Hydrogel systems are increasingly gaining visibility involving biomedicine, tissue engineering, environmental treatments, and drug delivery systems. These systems have a three-dimensional network composition and high-water absorption capacity, are biocompatible, allowing them to become an option as photosensitizer carriers (PS) for applications in Photodynamic Therapy (PDT) protocols. METHODS: A nanohydrogel system (NAHI), encapsulated with chloroaluminium phthalocyanine (ClAlPc) was synthesized for drug delivery.. NAHI was synthesized using gelatin as based polymer by the chemical cross-linking technique. The drug was encapsulated by immersing the hydrogel in a 1.0 mg.mL-1 ClAlPc solution. The external morphology of NAHI was examined by scanning electron microscopy (SEM). The degree of swelling of the synthesized system was evaluated to determine the water absorption potential. The produced nanohydrogel system was characterized by photochemical, photophysical and photobiologial studies. RESULTS: The images from the SEM analysis showed the presence of three-dimensional networks in the formulation. The swelling test demonstrated that the nanohydrogel freeze-drying process increases its water holding capacity. All spectroscopic results showed excellent photophysical parameters of the drug studied when served in the NAHI system. The incorporation efficiency was 70%. The results of trypan blue exclusion test have shown significant reduction (p < 0.05) in the cell viability for all groups treated with PDT, in all concentrations tested. In HeLa cells, PDT mediated by 0,5 mg.mL-1 ClAlPc encapsulated in NAHI showed a decrease in survival close to 95%. In the internalization cell study was possible to observe the internalization of phthalocyanine after one hour of incubation, at 37 °C, with the the accumulation of PS in the cytoplasm and inside the nucleus at both concentrations tested. CONCLUSIONS: Given the peculiar performance of the selected system, the resulting nanohydrogel is a versatile platform and display potential applications as controlled delivery systems of photosensitizer for photodynamic therapy application.

Polyelectrolytic gelatin nanoparticles as a drug delivery system for the promastigote form of Leishmania amazonensis treatment.

In this study, phthalocianato[bis(dimethylaminoethanoxy)] silicon (NzPC) was loaded onto gelatin nanoparticles functionalized with polyelectrolytes (polystyrene sulfonate/polyallylamine hydrochloride) by layer-by-layer (LbL) assembly for photodynamic therapy (PDT) application in promastigote form of Leishmania amazonensis treatment. The process yield, and encapsulation efficiency were 80.0% ± 1.8 and EE = 87.0% ± 1.1, respectively. The polyelectrolytic gelatin nanoparticles (PGN) had a mean diameter of 437.4 ± 72.85 nm, narrow distribution size with a polydispersity index of 0.086. The obvious switching of zeta potential indicates successful alternating deposition of the polyanion PSS and polycation PAH directly on the gelatin nanoparticles. Photosensitizer photophysical properties were shown to be preserved after gelatin nanoparticle encapsulation. The impact of the PDT in the viability and morphology of Leishmania amazonensis promastigote in culture medium was evaluated. The PGN-NzPc presented low toxicity at the dark and the PDT was capable of decreasing the viability in more than 80% in 0.1 µmol.L-1 concentration tested. The PDT also triggered significant morphological alterations in the Leishmania promastigotes. These results reinforce the idea that the use of PGN as photosensitizers carriers is useful for PDT of Leishmania promastigotes.

Vaterite submicron particles designed for photodynamic therapy in cells.

BACKGROUND: Calcium carbonate (CaCO3) is one of the most abundant materials in the world. It has several different crystalline phases as present in the minerals: calcite, aragonite and vaterite, which are anhydrous crystalline polymorphs. Regarding the preparation of these microparticles, the most important aspect is the control of the polymorphism, particle size and material morphology. This study aimed to develop porous microparticles of calcium carbonate in the vaterite phase for the encapsulation of chloro-aluminum phthalocyanine (ClAlPc) as a photosensitizer (PS) for application in Photodynamic Therapy (TFD). METHODS: In this study, spherical vaterite composed of microparticles are synthesized by precipitation route assisted by polycarboxylate superplasticizer (PSS). The calcium carbonate was prepared by reacting a mixed solution of Na2CO3 with a CaCl2 solution at an ambient temperature, 25 °C, in the presence of polycarboxylate superplasticizer as a stabilizer. The photosensitizer was incorporated by adsorption technique in the CaCO3 microparticles. The CaCO3 microparticles were studied by scanning electron microscopy, steady-state, and their biological activity was evaluated using in vitro cancer cell lines by trypan blue exclusion method. The intracellular localization of ClAlPc was examined by confocal microscopy. RESULTS: The CaCO3 microparticles obtained are uniform and homogeneously sized, non-aggregated, and highly porous microparticles. The calcium carbonate microparticles show an average size of 3 µm average pore size of about 30-40 nm. The phthalocyanine derivative loaded-microparticles maintained their photophysical behavior after encapsulation. The captured carriers have provided dye localization inside cells. The in vitro experiments with ClAlPc-loaded CaCO3 microparticles showed that the system is not cytotoxic in darkness, but exhibits a substantial phototoxicity at 3 µmol.L-1 of photosensitizer concentration and 10 J.cm-2 of light. These conditions are sufficient to kill about 80 % of the cells. CONCLUSIONS: All the performed physical-chemical, photophysical, and photobiological measurements indicated that the phthalocyanine-loaded CaCO3 microparticles are a promising drug delivery system for photodynamic therapy and photoprocesses.

Antitumor activity of photodynamic therapy performed with nanospheres containing zinc-phthalocyanine.

BACKGROUND: The increasing incidence of cancer and the search for more effective therapies with minimal collateral effects have prompted studies to find alternative new treatments. Among these, photodynamic therapy (PDT) has been proposed as a very promising new modality in cancer treatment with the lowest rates of side effects, revealing itself to be particularly successful when the photosensitizer is associated with nanoscaled carriers. This study aimed to design and develop a new formulation based on albumin nanospheres containing zinc-phthalocyanine tetrasulfonate (ZnPcS4-AN) for use in the PDT protocol and to investigate its antitumor activity in Swiss albino mice using the Ehrlich solid tumor as an experimental model for breast cancer. METHODS: Ehrlich tumor's volume, histopathology and morphometry were used to assess the efficacy of intratumoral injection of ZnPcS4-AN in containing tumor aggressiveness and promoting its regression, while the toxicity of possible treatments was assessed by animal weight, morphological analysis of the liver and kidneys, hemogram, and serum levels of total bilirubin, direct bilirubin, indirect bilirubin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma glutamyl transferase (GGT), alkaline phosphatase, creatinine and urea. In order to evaluate the efficacy of PDT, groups of animals treated with intratumoral injection of doxorubicin (Dox) were also investigated. RESULTS: Intratumoral injection of ZnPcS4-AN was found to be efficient in mediating PDT to refrain tumor aggressiveness and to induce its regression. Although tumor volume reduction was not significant, PDT induced a remarkable increase in the necrosis area seen in the tumor's central region, as in other experimental groups, including tumor and Dox treated groups, but also in the tumor's peripheral region. Further, PDT showed minimal adverse effects. Indeed, the use of ZnPcS4-AN in mediating PDT revealed anti-neoplastic activity similar to that obtained while using intratumoral Dox therapy. CONCLUSIONS: PDT mediated by the new formulation ZnPcS4-AN enhanced the inhibition of tumor growth while producing practically no adverse effects and thus emerges as a very promising nanotechnology-based strategy for solid cancer treatment.

Aluminum-chloride-phthalocyanine encapsulated in liposomes: activity against naturally occurring dog breast cancer cells.

Breast tumors represent the most common malignant tumors. Current treatments for humans and pets rely on tumor excision and adjuvant chemotherapy, which may affect both cancer cells and normal cells. Photodynamic therapy (PDT) is an approved treatment modality for a variety of cancers and was recently recommended as a first-line treatment for non-melanoma skin cancers for humans. The main purpose of the present study was to determine the efficacy of PDT using aluminum-chloride-phthalocyanine that is encapsulated in liposomes and LED as a light source to kill naturally occurring female dog breast cancer in vitro. The cytotoxicity behavior of the encapsulated photosensitizer in the dark and under irradiation using the 670 nm laser were investigated using classical trypan blue and MTT cell viability tests, acridine orange and ethidium bromide staining to label organelles, and cell morphology. Cell morphology was evaluated using light and electron microscopy. Our results demonstrate a reduced cell viability that is associated with morphologic alterations. The neoplasic cell destruction was predominantly mediated via a necrotic process, which was assayed using acridine orange and ethidium bromide staining. These findings were confirmed using light and electronic microscopy. The photosensitizer or laser irradiation alone did not induce cytotoxicity or morphological alterations, indicating the safety and efficacy of PDT with chloro-aluminum-phthalocyanine that was encapsulated in liposomes for the treatment of breast cancer cells in vitro.

Photodynamic therapy disinfection of carious tissue mediated by aluminum-chloride-phthalocyanine entrapped in cationic liposomes: an in vitro and clinical study.

Photodynamic therapy (PDT) is a technique employed in the treatment of several superficial infections, such as caries. PDT uses a non-toxic drug termed photosensitizer (PS) followed by light irradiation. The cytotoxic effects of the therapy are related to the production of reactive species produced after light activation of a photosensitizer, which reacts with surrounding molecules and disrupts several of the cell's functions. Within this context, this study aimed to develop a clinical protocol involving PDT application mediated by aluminum-chloride-phthalocyanine (AlClPc) entrapped in cationic liposomes against cariogenic bacteria in caries lesions. Cationic liposomes were used to delivery AlClPc preferentially to bacterial cells due to the strong anionic superficial charges of these cell types. The results are represented in two fundamental steps: (1) in vitro evaluation of AlClPc delivery to cariogenic bacteria and pulp cells, as well as its potential phototoxicity; (2) a clinical study involving volunteer patients that were treated with the PDT protocol mediated by AlClPc-cationic liposome. The main results showed that the AlClPc-cationic liposome was preferentially absorbed by bacterial cells compared to eukaryotic dental pulp cells, and it was efficient in the reduction of microbial load from bacterial cultures. In addition, the clinical study showed a mean reduction of 82% of total bacterial in the treated cavities after PDT application. Taken together, the results presented in this study showed that the antimicrobial PDT protocol mediated by cationic liposomes containing AlClPc is safety for clinical application and is efficient in the reduction of bacterial load in caries lesions.

Photophysical and complexation studies of chloro-aluminum phthalocyanine with beta-cyclodextrin and hydroxypropyl-beta-cyclodextrin.

A variety of nanostructures are being investigated as functional drug carriers for treatment of a wide range of diseases, most notably cardiovascular defects, autoimmune diseases, and cancer. The aim of this present contribution is to evaluate potentially applicable nanomaterials in the diagnosis and treatment of cancer due to their photophysical and photobiological properties and complexation behavior. The delivery systems consisted of chloro-aluminum phthalocyanine associated with beta-cyclodextrin and hydroxypropyl-beta-cyclodextrin. The preparation of the complex and its stoichiometry in an ethanol/buffer (3:1) solution were studied by spectroscopic techniques, which were defined as 1:2. The inclusion complex in the nanometer scale was observed on the basis of changes to the spectroscopic properties. The singlet oxygen production and complex photophysical parameters were determined by measuring luminescence at 1270 nm and by steady state and time resolved spectroscopic, respectively. The preparation of the complex was tested and analyzed with regard to cellular damage by visible light activation. The inclusion complex showed a higher singlet oxygen quantum yield compared with other systems and other photoactive dyes. There was also a reduction in the fluorescence quantum yield compared with the results obtained for zinc phthalocyanine in organic medium. The results reported clearly that the inclusion complex chloro-aluminum phthalocyanine/cyclodextrin showed some changes in its spectroscopy properties leading to better biodistribution and biocompatibility with a potential application in photodynamic therapy, especially in the case of neoplasy. Additionally, it also has non-oncological applications as a drug delivery system.

Free Rhodium (II) citrate and rhodium (II) citrate magnetic carriers as potential strategies for breast cancer therapy.

BACKGROUND: Rhodium (II) citrate (Rh(2)(H(2)cit)(4)) has significant antitumor, cytotoxic, and cytostatic activity on Ehrlich ascite tumor. Although toxic to normal cells, its lower toxicity when compared to carboxylate analogues of rhodium (II) indicates (Rh(2)(H(2)cit)(4)) as a promising agent for chemotherapy. Nevertheless, few studies have been performed to explore this potential. Superparamagnetic particles of iron oxide (SPIOs) represent an attractive platform as carriers in drug delivery systems (DDS) because they can present greater specificity to tumor cells than normal cells. Thus, the association between Rh(2)(H(2)cit)(4) and SPIOs can represent a strategy to enhance the former's therapeutic action. In this work, we report the cytotoxicity of free rhodium (II) citrate (Rh(2)(H(2)cit)(4)) and rhodium (II) citrate-loaded maghemite nanoparticles or magnetoliposomes, used as drug delivery systems, on both normal and carcinoma breast cell cultures. RESULTS: Treatment with free Rh(2)(H(2)cit)(4) induced cytotoxicity that was dependent on dose, time, and cell line. The IC(50) values showed that this effect was more intense on breast normal cells (MCF-10A) than on breast carcinoma cells (MCF-7 and 4T1). However, the treatment with 50 µM Rh(2)(H(2)cit)(4)-loaded maghemite nanoparticles (Mag(h)-Rh(2)(H(2)cit)(4)) and Rh(2)(H(2)cit)(4)-loaded magnetoliposomes (Lip-Magh-Rh(2)(H(2)cit)(4)) induced a higher cytotoxicity on MCF-7 and 4T1 than on MCF-10A (p < 0.05). These treatments enhanced cytotoxicity up to 4.6 times. These cytotoxic effects, induced by free Rh(2)(H(2)cit)(4), were evidenced by morphological alterations such as nuclear fragmentation, membrane blebbing and phosphatidylserine exposure, reduction of actin filaments, mitochondrial condensation and an increase in number of vacuoles, suggesting that Rh(2)(H(2)cit)(4) induces cell death by apoptosis. CONCLUSIONS: The treatment with rhodium (II) citrate-loaded maghemite nanoparticles and magnetoliposomes induced more specific cytotoxicity on breast carcinoma cells than on breast normal cells, which is the opposite of the results observed with free Rh(2)(H(2)cit)(4) treatment. Thus, magnetic nanoparticles represent an attractive platform as carriers in Rh(2)(H(2)cit)(4) delivery systems, since they can act preferentially in tumor cells. Therefore, these nanopaticulate systems may be explored as a potential tool for chemotherapy drug development.

Impact of photodynamic therapy on inflammatory cells during human chronic periodontitis.

The aim of this study was to evaluate the effects of the photodynamic therapy (PDT) on the inflammatory infiltrate and on the collagen network organization in human advanced chronic periodontitis. Two different drug delivery systems (DDS) were tested (liposomes and nanoemulsions) to determine if the effects of PDT could differ according to the DDS used. Sixteen patients presenting two teeth with chronic advanced periodontitis and important tooth mobility with clinical indication of extraction were included in the group liposomes (group L, n=8) or in the group nanoemulsions (group N, n=8) in order to compare the effects of each DDS. Seven days before extractions one tooth of each patient was treated with PDT using phthalocyanine derivatives as photosensitizers and the contralateral tooth was taken as control. In group L the density of gingival collagen fibers (66±19%) was significantly increased (p<0.02) when compared to controls (35±21%). Concerning the antigen-presenting cells, PDT had differential effects depending on the drug delivery system; the number of macrophages was significantly decreased (p<0.05) in group L while the number of Langerhans cells was significantly decreased in group N (p<0.02). These findings demonstrate that PDT presents an impact on gingival inflammatory phenomenon during chronic periodontitis and leads to a specific decrease of antigen-presenting cells populations according to the drug delivery system used.

Poly(lactic acid-glycolic acid) nanoparticles markedly improve immunological protection provided by peptide P10 against murine paracoccidioidomycosis.

BACKGROUND AND PURPOSE: The present study reports on the preparation and testing of a sustained delivery system for the immunomodulatory peptide P10 aimed at reducing the in vivo degradation of the peptide and the amount required to elicit a protective immune response against paracoccidioidomycosis. EXPERIMENTAL APPROACH: BALB/c mice were infected with the yeast Paracoccidioides brasiliensis to mimic the chronic form of paracoccidioidomycosis. The animals were treated daily with sulfamethoxazole/trimethoprim alone or combined with peptide P10, either emulsified in Freund's adjuvant or entrapped in poly(lactic acid-glycolic acid) (PLGA) nanoparticles at different concentrations (1 microg, 5 microg, 10 microg, 20 microg or 40 microg.50 microL(-1)). Therapeutic efficacy was assessed as fungal burden in tissues and the immune response by quantitative determination of cytokines. KEY RESULTS: Animals given combined chemotherapy and P10 nanotherapy presented a marked reduction of fungal load in the lungs, compared with the non-treated animals. After 30 days of treatment, P10 entrapped within PLGA (1 microg.50 microL(-1)) was more effective than 'free' P10 emulsified in Freund's adjuvant (20 microg.50 microL(-1)), as an adjuvant to chemotherapy. After treatment for 90 days, the higher doses of P10 entrapped within PLGA (5 or 10 microg.50 microL(-1)) were most effective. Treatment with P10 emulsified in Freund's adjuvant (20 microg.50 microL(-1)) or P10 entrapped within PLGA (1 microg.50 microL(-1)) were accompanied by high levels of interferon-gamma in lung. CONCLUSIONS AND IMPLICATIONS: Combination of sulfamethoxazole/trimethoprim with the P10 peptide entrapped within PLGA demonstrated increased therapeutic efficacy against paracoccidioidomycosis. P10 incorporation into PLGA nanoparticles dramatically reduced the peptide amount necessary to elicit a protective effect.

In vitrouptake and antimycobacterial activity of liposomal usnic acid formulation.

The cellular uptake and antimycobacterial activity of usnic acid (UA) and usnic acid-loaded liposomes (UA-LIPOs) were assessed on J774 macrophages. The minimal inhibitory concentration (MIC) and the minimal bactericidal concentration (MBC) of UA and UA-LIPO against Mycobacterium tuberculosis were determined. Concentrations required to inhibit 50% of cell proliferation (IC(50)) were 22.5 (+/-0.60) and 12.5 (+/-0.26) microg/ml, for UA and UA-LIPO, respectively. The MICs of UA and UA-LIPO were 6.5 and 5.8 microg/mL, respectively. The MBC of UA-LIPO was twice as low (16 microg/mL) as that of UA (32 microg/mL). An improvement in the intracellular uptake of UA-LIPO was found (21.6 x 10(4) +/- 28.3 x 10(2) c.p.s), in comparison with UA (9.5 x 10(4) +/- 11.4 x 10(2) c.p.s). In addition, UA-LIPO remains much longer inside macrophages (30 hours). All data obtained from the encapsulation of usnic acid into liposomes as a drug delivery system (DDS) indicate a strong interaction between UA-liposomes and J774 macrophages, thereby facilitating UA penetration into cells. Considering such a process as ruling the Mycobacterium-transfection by magrophages, we could state that associating UA with this DDS leads to an improvement in its antimycobacterial activity.

Amphotericin B in poly(lactic-co-glycolic acid) (PLGA) and dimercaptosuccinic acid (DMSA) nanoparticles against paracoccidioidomycosis.

OBJECTIVES: The present study reports on the preparation and testing of a desoxycholate amphotericin B (D-AMB) sustained delivery system based on poly(lactic-co-glycolic acid) (PLGA) and dimercaptosuccinic acid (DMSA) polymeric blends (Nano-D-AMB) aimed at reducing the number of AMB administrations required to treat mycosis. METHODS: BALB/c mice were infected with the yeast Paracoccidioides brasiliensis intravenously to mimic the chronic form of paracoccidioidomycosis. At 30 days post-infection, the animals were treated with Nano-D-AMB [6 mg/kg of encapsulated D-AMB, intraperitoneally (ip), interval of 72 h] or D-AMB (2 mg/kg, ip, interval of 24 h). Drug efficacy was investigated by the fungal burden recovery from tissues. Toxicity was assessed by renal and hepatic biochemical parameters, physical appearance of the animals and haematological investigation. The control groups used were non-infected and the infected mice mock treated with PBS. RESULTS: Nano-D-AMB presented results comparable to free D-AMB, with a marked antifungal efficacy. The Nano-D-AMB-treated group presented lower loss of body weight and absence of stress sign (piloerection and hypotrichosis) observed after D-AMB treatment. No renal [blood urea nitrogen (BUN), creatinine] or hepatic (pyruvic and oxalacetic glutamic transaminases) biochemical abnormalities were found. The micronucleus assay showed no significant differences in both the micronucleus frequency and percentage of polychromatic erythrocytes for Nano-D-AMB, indicating the absence of genotoxicity and cytotoxic effects. CONCLUSIONS: The D-AMB-coated PLGA-DMSA nanoparticle showed antifungal efficacy, fewer undesirable effects and a favourable extended dosing interval. Nano-D-AMB comprises an AMB formulation able to lessen the number of drug administrations. Further studies would elucidate whether Nano-D-AMB would be useful to treat systemic fungal infections such as paracoccidioidomycosis, candidiasis, aspergillosis and cryptococcosis.

Photophysical and photobiological studies of a silicon tribenzonaphthoporphyrazinato incorporated into liposomes for photodynamic therapy use.

Nanostructured drug delivery systems (NDDS), such as liposomes, represent a growing area in biomedical research. These microheterogeneous media can be used in many biological systems to provide appropriate drug levels with a specific biodistribution. The photophysical properties of a silicon derivative of tribenzonaphthoporphyrazinato (Si-tri-PcNc) incorporated into liposome were studied by steady-state techniques, time-resolved fluorescence and laser flash photolysis. All the spectroscopy measurements performed allowed us to conclude that Si-tri-PcNc in liposome is a promising NDDS for PDT. The in vitro experiments with liposomal NDDS showed that the system is not cytotoxic in darkness, but exhibits a substantial phototoxicity at 1 microM of photosensitizer concentration and 10.0 J/cm2 of light. These conditions are sufficient to kill about 80% of the cells.

Photoinduced electron-transfer processes based on novel bipyridine-Ru(II) complex: properties of cis-[Ru(2,2'-bipyridine)2(5,6-bis(3-amidopyridine)-7-oxanorbornene)](PF6)2 and cis-[Ru(2,2'-bipyridine)2(3-aminopyridine)2](PF6)2 complexes.

This paper presents the synthesis, MO calculations, and photochemical and photophysical properties of cis-[Ru(bpy)2(3Amdpy2oxaNBE)](PF6)2 (2), where bpy is 2,2'-bipyridine and 3Amdpy2oxaNBE is the novel 5,6-bis(3-amidopyridine)-7-oxanorbornene chelate-ligand (1). Complex 2 is considered in relation to the cis-[Ru(bpy)2(3Amnpy)2](PF6)2 (3) analogous complex, where 3Amnpy is 3-aminopyridine. Complexes 2 and 3 exhibit absorptions near 350 nm and in the 420-500 nm region attributable to a contribution from MLCT transitions (dpi-->bpy and dpi-->L; L=3Amdpy2oxaNBE or 3Amnpy). Whereas complex 3 is photochemically reactive, complex 2 shows luminescence either at 77 K or at room temperature in fluid solution. The emission of 2 assignable as an MLCT (Ru-->bpy) emission is characterized by a long lifetime at room temperature (650 ns in CH3CN and 509 ns in H2O). It is independent of lambdairr, but it is temperature dependent; i.e., it increases as the temperature is lowered. Considering the chelate ring of 1 contributes to the stability of the complex 2 under continuous light irradiation, the difference in the primary photoprocesses of 3 (loss of 3Amnpy) and 2 (luminescence) may be caused by a lowering of the lowest excited state from 3 to 2. The surface crossing to the lowest MC state value of 987 cm-1 (similar to that of [Ru(bpy)3]2+) will be prevented in the case of complex 2, and as a result, efficient 3Amdpy moiety loss cannot occur. The electronic depopulation of the {Ru(bpy)2} unit and population of a bpy* orbital upon excitation are evident by comparing the photophysical properties with those of a [Ru(bpy)3]2+ related complex. Moreover, a reduction of a bpy ligand in the MLCT excited state is indicated by time-resolved spectra that show features typical of bpy*-. The photocatalytic property of 2 is spectroscopically demonstrated by oxidative quenching using either methylviologen2+ or [RuCl(NH3)5]+2 electron-acceptor ions.

Putative role of heparan sulfate proteoglycan expression and shedding on the proliferation and survival of cells after photodynamic therapy.

INTRODUCTION: Photodynamic therapy is based on the selective retention of a photosensitizer by highly proliferating cells and its activation with light at the appropriate wavelength. This combination generates reactive oxygen species that ultimately kill the cells. Some cells, however, may survive photodynamic therapy and the interaction of these cells with the extracellular matrix has profound effect in tumor biology. The knowledge of photodynamic therapy action on the extracellular matrix has not been fully explored. It has been focused mainly on integrins, matrix metalloproteinases and on growth factors and immunological mediators. Other important molecules involved in the regulation of many cell processes are the glycosaminoglycans, polymers of disaccharide units, present on the cell surface and in the extracellular matrix. In most cases, the glycosaminoglycans occur as proteoglycans. AIMS: The purpose of the present investigation is to evaluate heparan sulfate proteoglycan expression and shedding, and its relation to the survival of the remaining cells, after a liposomal-AlClPc based photodynamic treatment. MATERIALS: A wild-type endothelial cell derived from rabbit aorta and its counterpart transfected with EJ-ras oncogene were used. RESULTS: Both cell lines presented augmented heparan sulfate proteoglycan syndecan-4 mRNA expression, augmented synthesis of heparan sulfate chains and increased shedding. Also, the formation of stress fibers on the border of the cells and the arrest in G(1) phase of the cell cycle was observed. CONCLUSIONS: These results show that surviving cells after photodynamic therapy exhibit changes in their morphology and cell processes that differ from that of non-treated cells, and these changes are probably hindering the cells from resuming normal proliferation.

Synthesis and photochemical properties of a bis(n-Hexanoyl)silicon tribenzonaphthoporphyrazate.

OBJECTIVE: The aim of the present work is to describe the synthesis and photochemical studies of a silicon tribenzonaphthoporphyrazate (Si-tri-PcNc) derivative to be used in photomedicine. BACKGROUND DATA: Properties of the photosensitizer localization in tumor tissue and photochemical generation of reactive oxygen species are combined with precise delivery of laser-generated light to induce a procedure offering effective local tumoricidal activity. This technique is called photodynamic therapy (PDT). METHODS: This new phthalocyanine (Pc) compound obtained from 2,3-dicyanonaphthalene and phthalonitrile is analyzed and chemically characterized by (1)H-NMR, elemental analyses, infrared (IR), ultraviolet and visibile absorption spectroscopy (UV-Vis) and fluorescence spectra. RESULTS: The spectroscopic analysis of this compound in EtOH showed an electronic spectrum with characteristic bands in the Soret and Q region. Steady-state and time-resolved spectral analyses revealed a fluorescence quantum yield of phi(f) = 0.24, in agreement with other Pc compounds. This compound also shows a triplet excited lifetime of 0.40 microsec in air-saturated solution and a transient with the maximum absorption located at 610 nm, 120 nm shifted to the red if compared to the excited triplet state observed with other Pc compounds. CONCLUSION: Based on our results, we can assume that Si-tri-PcNc induces a higher singlet oxygen production under photoactivation, which is in agreement with the expected photobiological behavior. This result could lead to the testing and eventual use of this new generation of dyes in PDT.