Andreza R. Simioni

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 cells 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.

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.

Photodynamic therapy with aluminum-chloro-phtalocyanine induces necrosis and vascular damage in mice tongue tumors

In this paper we describe the efficacy of the liposomal-AlClPc (aluminum-chloro-phthalocyanine) formulation in PDT study against Ehrlich tumor cells proliferation in immunocompetent swiss mice tongue. Experiments were conduced in sixteen tumor induced mice that were divided in three control groups: (1) tumor without treatment; (2) tumor with 100J/cm(2) laser (670nm) irradiation; and (3) tumor with AlClPc peritumoral injection; and a PDT experimental group when tumors received AlClPc injection followed by tumor irradiation. Control groups present similar macroscopically and histological patterns after treatments, while PDT treatment induced 90% of Ehrlich tumor necrosis after 24h of one single application, showing the efficacy of liposome-AlClPc (aluminum-chloro-phthalocyanine) mediated PDT on the treatment of oral cancer.

In vitro photodynamic therapy on human oral keratinocytes using chloroaluminum-phthalocyanine

In this study, oral carcinoma cells were used to evaluate chloroaluminum-phthalocyanine encapsulated in liposomes as the photosensitizer agent in support of photodynamic therapy (PDT). The genotoxicity and cytotoxicity behavior of the encapsulated photosensitizer in both dark and under irradiation using the 670-nm laser were investigated with the classical trypan blue cell viability test, the acridine orange/ethidium bromide staining organelles test, micronucleus formation frequency, DNA fragmentation, and cell morphology. The cell morphology investigation was carried out using light and electronic microscopes. Our findings after PDT include reduction in cell viability (95%) associated with morphologic alterations. The neoplastic cell destruction was predominantly started by a necrotic process, according to the assay with acridine orange and ethidium bromide, and this was confirmed by electronic microscopy analysis. Neither the PDT agent nor laser irradiation alone showed cytotoxicity, genotoxicity, or even morphologic alterations. Our results reinforce the efficiency of light-irradiated chloroaluminum-phthalocyanine in inducing a positive effect of PDT.

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.

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

BackgroundRhodium (II) citrate (Rh2(H2cit)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 Rh2(H2cit)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 Rh2(H2cit)4 and SPIOs can represent a strategy to enhance the formers therapeutic action. In this work, we report the cytotoxicity of free rhodium (II) citrate (Rh2(H2cit)4) and rhodium (II) citrate-loaded maghemite nanoparticles or magnetoliposomes, used as drug delivery systems, on both normal and carcinoma breast cell cultures.ResultsTreatment with free Rh2(H2cit)4 induced cytotoxicity that was dependent on dose, time, and cell line. The IC50 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 Rh2(H2cit)4-loaded maghemite nanoparticles (Magh-Rh2(H2cit)4) and Rh2(H2cit)4-loaded magnetoliposomes (Lip-Magh-Rh2(H2cit)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 Rh2(H2cit)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 Rh2(H2cit)4 induces cell death by apoptosis.ConclusionsThe 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 Rh2(H2cit)4 treatment. Thus, magnetic nanoparticles represent an attractive platform as carriers in Rh2(H2cit)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.

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.

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

BackgroundThe 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.MethodsEhrlich 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.ResultsIntratumoral 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.ConclusionsPDT 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.

Preparation, Characterization and in vitro Toxicity Test of Magnetic Nanoparticle-Based Drug Delivery System to Hyperthermia of Biological Tissues

The goals of this study were first to prepare and second to test albumin-based beads containing maghemite nanoparticles dispersed on them. Incorporation of maghemite nanoparticles in the albumin-based template, allowing full control of the magnetic content, was accomplished by adding ionic magnetic fluid to the albumin suspension, following heat denaturation. The material evaluation was performed using Zeta potential measurements and scanning electron microscopy from one side and by investigating the cell activity on the other hand.

In vitro uptake 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 (IC50) were 22.5 (±0.60) and 12.5 (±0.26) μg/ml, for UA and UA-LIPO, respectively. The MICs of UA and UA-LIPO were 6.5 and 5.8 μg/mL, respectively. The MBC of UA-LIPO was twice as low (16 μg/mL) as that of UA (32 μg/mL). An improvement in the intracellular uptake of UA-LIPO was found (21.6 × 104 ± 28.3 × 102 c.p.s), in comparison with UA (9.5 × 104 ± 11.4 × 102 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.