Study of the antibacterial and cytotoxic activity of chitosan and its derivatives chemically modified with phthalic anhydride and ethylenediamine.

The insertion of hydrophobic and hydrophilic chains in the chitosan molecule can improve its antibacterial activity, expanding its range of application in several areas of medical-pharmaceutical sciences. Thus, this work aimed to increase the antibacterial activity of chitosan through the modification reaction with phthalic anhydride (QF) and subsequent reaction with ethylenediamine (QFE). The chitosan and derivatives obtained were characterized by elemental analysis, 13C Nuclear Magnetic Resonance (13C NMR), X-Ray Diffraction (XRD), Fourier Transform Infrared Spectroscopy (FTIR) and Thermogravimetric Analysis (TG), where it was possible to prove the chemical modification. Both materials showed a greater antibacterial inhibitory effect against Gram-positive bacteria, Staphylococcus aureus, emphasizing antibacterial activity against Gram-negative bacteria, Escherichia coli, with values above 70 % of the inhibitory effect, which is a promising result. Assays with human fibroblast cells by the [3-(4,5-dimethylthiazolyl)-2,5-diphenyl tetrazolium (MTT)] bromide reduction test did not indicate toxicity in the materials. Thus, the derived materials showed promise for biomedical applications since they combined excellent antibacterial activity against gram-positive and gram-negative strains and did not show cytotoxicity.

Encapsulation of nor-ß-lapachone into poly(d,l)-lactide-co-glycolide (PLGA) microcapsules: full characterization, computational details and cytotoxic activity against human cancer cell lines.

In this work, we characterize nor-ß-lapachone-loaded (NßL-loaded) microcapsules prepared using an emulsification/solvent extraction technique. Features such as surface morphology, particle size distribution, zeta potential, optical absorption, Raman and Fourier transform infrared spectra, thermal analysis data, drug encapsulation efficiency, drug release kinetics and in vitro cytotoxicity were studied. Spherical microcapsules with a size of 1.03 ± 0.46 µm were produced with an encapsulation efficiency of approximately 19%. Quantum DFT calculations were also performed to estimate typical interaction energies between a single nor-ß-lapachone molecule and the surface of the microparticles. The NßL-loaded PLGA microcapsules exhibited a pronounced initial burst release. After the in vitro treatment with NßL-loaded microcapsules, a clear phagocytosis of the spheres was observed in a few minutes. The cytotoxic activity against a set of cancer cell lines was investigated.

Are salty liquid food flavorings in vitro antitumor substances?

The objective of this study was to evaluate the antiproliferative, cytotoxic and genotoxic potential of salty liquid synthetic flavorings of Butter, Cheddar Cheese and Onion. The antiproliferative potential (2.9-1500 µg/mL) was assessed by MTT assay after 72h using the human tumor lines SF-295 (glioblastoma), OVCAR-8 (ovarian), HCT-116 (colon) and HL-60 (promyelocytic leukemia) and primary cultures of murine Sarcoma 180 (S180) and peripheral blood mononuclear cells (PBMC). Allium cepa bulbs were exposed to growing respective doses (1 mL and 2 mL). Only Butter and Cheddar flavorings revealed cytotoxic activity on cancer cells, with IC50 values ranging from 125.4 µg/mL (Cheddar - HCT-116) to 402.6 µg/mL (Butter - OVCAR-8). Butter flavoring was the most cytotoxic on PBMC (136.3 µg/mL) and increased cell division rate in relation to the mitotic index but did not cause cellular aberrations. Onion and Cheddar flavorings reduced the mitotic index after 24h and 48h exposure, but only Onion flavoring resulted in cellular aberrations and mitotic spindle abnormalities, such as anaphase and telophase bridges, micronucleated cells, conchicine-metaphases and amplifications. So, Butter, Onion and/or Cheddar flavorings caused significant changes in the division of meristematic cells of A. cepa and presented cytotoxic action even on decontrolled proliferating human tumor cells.

Controlled Release of Nor-ß-lapachone by PLGA Microparticles: A Strategy for Improving Cytotoxicity against Prostate Cancer Cells.

Prostate cancer is one of the most common malignant tumors in males and it has become a major worldwide public health problem. This study characterizes the encapsulation of Nor-ß-lapachone (NßL) in poly(d,l-lactide-co-glycolide) (PLGA) microcapsules and evaluates the cytotoxicity of the resulting drug-loaded system against metastatic prostate cancer cells. The microcapsules presented appropriate morphological features and the presence of drug molecules in the microcapsules was confirmed by different methods. Spherical microcapsules with a size range of 1.03 ± 0.46 µm were produced with an encapsulation efficiency of approximately 19%. Classical molecular dynamics calculations provided an estimate of the typical adsorption energies of NßL on PLGA. Finally, the cytotoxic activity of NßL against PC3M human prostate cancer cells was demonstrated to be significantly enhanced when delivered by PLGA microcapsules in comparison with the free drug.

Molecular characterization of the Corynebacterium pseudotuberculosis hsp60-hsp10 operon, and evaluation of the immune response and protective efficacy induced by hsp60 DNA vaccination in mice.

BACKGROUND: Heat shock proteins (HSPs) are important candidates for the development of vaccines because they are usually able to promote both humoral and cellular immune responses in mammals. We identified and characterized the hsp60-hsp10 bicistronic operon of the animal pathogen Corynebacterium pseudotuberculosis, a Gram-positive bacterium of the class Actinobacteria, which causes caseous lymphadenitis (CLA) in small ruminants. FINDINGS: To construct the DNA vaccine, the hsp60 gene of C. pseudotuberculosis was cloned in a mammalian expression vector. BALB/c mice were immunized by intramuscular injection with the recombinant plasmid (pVAX1/hsp60). CONCLUSION: This vaccination induced significant anti-hsp60 IgG, IgG1 and IgG2a isotype production. However, immunization with this DNA vaccine did not confer protective immunity.

Evidence for reductive genome evolution and lateral acquisition of virulence functions in two Corynebacterium pseudotuberculosis strains.

BACKGROUND: Corynebacterium pseudotuberculosis, a gram-positive, facultative intracellular pathogen, is the etiologic agent of the disease known as caseous lymphadenitis (CL). CL mainly affects small ruminants, such as goats and sheep; it also causes infections in humans, though rarely. This species is distributed worldwide, but it has the most serious economic impact in Oceania, Africa and South America. Although C. pseudotuberculosis causes major health and productivity problems for livestock, little is known about the molecular basis of its pathogenicity. METHODOLOGY AND FINDINGS: We characterized two C. pseudotuberculosis genomes (Cp1002, isolated from goats; and CpC231, isolated from sheep). Analysis of the predicted genomes showed high similarity in genomic architecture, gene content and genetic order. When C. pseudotuberculosis was compared with other Corynebacterium species, it became evident that this pathogenic species has lost numerous genes, resulting in one of the smallest genomes in the genus. Other differences that could be part of the adaptation to pathogenicity include a lower GC content, of about 52%, and a reduced gene repertoire. The C. pseudotuberculosis genome also includes seven putative pathogenicity islands, which contain several classical virulence factors, including genes for fimbrial subunits, adhesion factors, iron uptake and secreted toxins. Additionally, all of the virulence factors in the islands have characteristics that indicate horizontal transfer. CONCLUSIONS: These particular genome characteristics of C. pseudotuberculosis, as well as its acquired virulence factors in pathogenicity islands, provide evidence of its lifestyle and of the pathogenicity pathways used by this pathogen in the infection process. All genomes cited in this study are available in the NCBI Genbank database (http://www.ncbi.nlm.nih.gov/genbank/) under accession numbers CP001809 and CP001829.

Actin-interacting and flagellar proteins in Leishmania spp.: Bioinformatics predictions to functional assignments in phagosome formation.

Several motile processes are responsible for the movement of proteins into and within the flagellar membrane, but little is known about the process by which specific proteins (either actin-associated or not) are targeted to protozoan flagellar membranes. Actin is a major cytoskeleton protein, while polymerization and depolymerization of parasite actin and actin-interacting proteins (AIPs) during both processes of motility and host cell entry might be key events for successful infection. For a better understanding the eukaryotic flagellar dynamics, we have surveyed genomes, transcriptomes and proteomes of pathogenic Leishmania spp. to identify pertinent genes/proteins and to build in silico models to properly address their putative roles in trypanosomatid virulence. In a search for AIPs involved in flagellar activities, we applied computational biology and proteomic tools to infer from the biological meaning of coronins and Arp2/3, two important elements in phagosome formation after parasite phagocytosis by macrophages. Results presented here provide the first report of Leishmania coronin and Arp2/3 as flagellar proteins that also might be involved in phagosome formation through actin polymerization within the flagellar environment. This is an issue worthy of further in vitro examination that remains now as a direct, positive bioinformatics-derived inference to be presented.

Encapsulation and release characteristics of DCTN/PLGA microspheres.

In the present study, poly (D,L-lactic-co-glycolic)-acid microspheres containing trans-Dehydrocrotonin (DCTN) were prepared by the double emulsion method. The hypoglycemic activity of DCTN-loaded microspheres was monitored in normal glycemic mice after administration of a daily dose of DCTN (50 mg kg(-1) body weight) for 7 days. Spherical microspheres with two populations of particles with 3.20 +/- 0.10 and 7.60 +/- 0.70 microm mean diameter size microm were observed. The encapsulation efficiency of DCTN was 85.5 +/- 3.9%. The in vitro kinetic profile of DCTN from PLGA-microspheres was initially fast (burst effect of 19.4% at 2 h). Such a burst step was maintained until achieving 35.7+/-2.0% at 7h, followed by a gradual release of DCTN attaining a maximum drug release at 55.7 +/- 2.6% within 30 h. DCTN was able to reduce glucose levels (14.3%) of normal glycemic animals and this effect was improved by its encapsulation into microspheres (26.8%). The optimum glucose levels in the blood of animals treated with DCTN suspension and DCTN-loaded microspheres were 119.21 +/- 19.75 mg dL(-1) at day 5 and 103.08 +/- 18.88 mg dL(-1) at day 7, respectively. DCTN-loaded microspheres are thus offered as a potential delivery system for the treatment of metabolic diseases characterized by hyperglycemia.

Actin-interacting and flagellar proteins in Leishmania spp: bioinformatics predictions to functional assignments in phagosome formation

Several motile processes are responsible for the movement of proteins into and within the flagellar membrane, but little is known about the process by which specific proteins (either actin-associated or not) are targeted to protozoan flagellar membranes. Actin is a major cytoskeleton protein, while polymerization and depolymerization of parasite actin and actin-interacting proteins (AIPs) during both processes of motility and host cell entry might be key events for successful infection. For a better understanding the eukaryotic flagellar dynamics, we have surveyed genomes, transcriptomes and proteomes of pathogenic Leishmania spp. to identify pertinent genes/proteins and to build in silico models to properly address their putative roles in trypanosomatid virulence. In a search for AIPs involved in flagellar activities, we applied computational biology and proteomic tools to infer from the biological meaning of coronins and Arp2/3, two important elements in phagosome formation after parasite phagocytosis by macrophages. Results presented here provide the first report of Leishmania coronin and Arp2/3 as flagellar proteins that also might be involved in phagosome formation through actin polymerization within the flagellar environment. This is an issue worthy of further in vitro examination that remains now as a direct, positive bioinformatics-derived inference to be presented.