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

Abstract\ud \ud Background\ud 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.\ud \ud \ud Methods\ud 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.\ud \ud \ud Results\ud 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.\ud \ud \ud Conclusions\ud 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.We are grateful to the Sabin Institute/Sabin Laboratories for technical\ud support and to the Brazilian National Council for Technological and Scientific\ud Development (CNPq), the Foundation to Support Research in the Federal\ud District (FAPDF), the Coordination for Further Training of Graduate Staff\ud (CAPES), the Nanobiotechnology-Network CON-NANO (CAPES), INCTNanobiotecnologia\ud (MCTI, CNPq, CAPES), CNANO-UnB, the São Paulo\ud Research Foundation (FAPESP) #08/53719-4 ACT, and the DPP-University of\ud Brasília, for financial support

Original Article

The pancreas taken from the frog (Rana nigromaculata) was fixed in 1% OsO_4 and sliced into ultrathin sections for electron microscopic studies. The following observations were made: 1. A great \u27number of minute granules found in the cytoplasm of a pancreatic cell were called the microsomes, which were divided into two types, the C-microsome and S-microsome. 2. Electron microsopic studies of the ergastoplasm showed that it is composed of the microsome granules and A-substance. The microsomes were seen embedded in the A-substance which was either filamentous or membranous. The membranous structure, which was called the Am-membrane, was seen to form a sac, with a cavity of varying sizes, or to form a lamella. 3. The Am-membrane has close similarity to α-cytomembrane of Sjostrand, except that the latter is rough-surfaced. It was deduced that the Am-membrane, which is smooth-surfaced, might turn into the rough-surfaced α-cytomembrane. 4. There was the Golgi apparatus in the supranuclear region of a pancreatic cell. It consisted of the Golgi membrane, Golgi vacuole and. Golgi vesicle. 5. The mitochondria of a pancreatic cell appeared like long filaments, and some of them were seen to ramify. 6. The membrane of mitochondria, i. e. the limiting membrane, consisted of the Ammembrane. The mitochondria contained a lot of A-substances, as well as the C-microsomes and S-microsomes. When the mitochondria came into being, there appeared inside them chains of granules, which appeared like strips of beads, as the outgrowths of the A-substance and the microsome granules attached to the Am-membrane. They are the so-called cristae mitochondriales. 7. The secretory granules originate in the microsomes. They came into being when the microsomes gradually thickened and grew in size as various substances became adhered to them. Some of the secretory granules were covered with a membrane and appeared like what they have called the intracisternal granule of Palade.It seemed that this was a phenomenon attendant upon the dissolution and liqutefaction of the secretory granule. 8. Comparative studies were made of the ergastoplasm of the pancreatic cells from the frogs in hibernation, the frogs artificially hungered, the frogs which were given food after a certain period of fasting, the frogs to which pilocarpine was given subcutaneously, and the very young, immature frogs. The studies revealed that the ergastoplasm of the pancreatic cells greatly varied in form with the difference in nutritive condition and with different developmental stages of the cell. The change in form and structure occured as a result of transformation of the microsomes and A-substance. The ergastoplasm, even after it has come into being, might easily be inactivated if nutrition is defective. The ergastoplasm is concerned in the secretory mechanism, which is different from the secretory phenomenon of the secretory granules. It would seem that structurally the mitochondria have no direct relation to this mechanism

The translation elongation factor 1A in tumorigenesis, signal transduction and apoptosis: Review article

An increasing number of evidences suggest the involvement of the eukaryotic elongation factor 1A, a core component of the protein synthesis machinery, at the onset of cell transformation. In fact, eEF1A is shown to be up-regulated in cell death; moreover, it seems to be involved in the regulation of ubiquitin-mediated protein degradation. In addition, eEF1A undergoes several post-translational modifications, mainly phosphorylation and methylation, that generally influence the activity of the protein. This article summarizes the present knowledges on the several extra-translational roles of eEF1A also in order to understand as the protein synthesis regulatory mechanisms could offer tools for cancer intervention

Optical Sensors for Vapors, Liquids, and Biological Molecules Based on Porous Silicon Technology

The sensing of chemicals and biochemical molecules using several porous silicon optical microsensors, based both on single-layer interferometers and resonant-cavity-enhanced microstructures, is reported. The operation of both families of sensors is based on the variation of the average refractive index of the porous silicon region, due to the interaction with chemical substances either in vapor or liquid state, which results in marked shifts of the device reflectivity spectra. The well established single-layer configuration has been used to test a new chemical approach based on Si-C bonds for covalent immobilization of biological molecules, as probe, in a stable way on the porous silicon surface. Preliminary results on complementary oligonucleotide recognition, based on this technique, are also presented and discussed. Porous silicon optical microcavities, based on multi- layered resonating structures, have been used to detect chemical substances and, in particular, flammable and toxic organic solvents, and some hydrocarbons. The results put in evidence the high sensitivity, the reusability, and the low response time of the resonant-cavity-enhanced sensing technique. The possibility of operating at room temperature, of remote interrogation, and the absence of electrical contacts are further advantages character- izing the sensing technique

Modulatory Effect of Gliadin Peptide 10-mer on Epithelial Intestinal CACO-2 Cell Inflammatory Response.

Celiac Disease (CD) is a chronic inflammatory enteropathy, triggered in genetically susceptible individuals by dietary gluten. Gluten is able to elicit proliferation of specific T cells and secretion of inflammatory cytokines in the small intestine. In this study we investigated the possibility that p10-mer, a decapeptide from durum wheat (QQPQDAVQPF), which was previously shown to prevent the activation of celiac peripheral lymphocytes, may exert an inhibitory effect on peptic-tryptic digested gliadin (PT-Gly)-stimulated intestinal carcinoma CACO-2 cells. In these cells, incubated with PT-Gly or p31-43 α-gliadin derived peptide in the presence or in the absence of p10-mer, IRAK1 activation and NF-kB, ERK1/2 and p38 MAPK phosphorylation were measured by immunoblotting, Cyclooxigenase 2 (COX-2) activity by PGE-2 release assay, and production of cytokines in the cell supernatants by ELISA. Our results showed that pre-treatment of CACO-2 cells with p10-mer significantly inhibited IRAK1 activation and NF-kB, ERK1/2 and p38 MAPK phosphorylation, as well as COX-2 activity (i.e. PGE-2 release) and production of the IL-6 and IL-8 pro-inflammatory cytokines, induced by gliadin peptides. These findings demonstrate the inhibitory effect of the p10-mer peptide on inflammatory response in CACO-2 cells. The results of the present study show that this p10-mer peptide can modulate "in vitro" the inflammatory response induced by gliadin peptides, allowing to move towards new therapeutic strategies. Turning off the inflammatory response, may in fact represent a key target in the immunotherapy of celiac disease

Modulatory Effect of Gliadin Peptide 10-mer on Epithelial Intestinal CACO-2 Cell Inflammatory Response

Celiac Disease (CD) is a chronic inflammatory enteropathy, triggered in genetically susceptible individuals by dietary gluten. Gluten is able to elicit proliferation of specific T cells and secretion of inflammatory cytokines in the small intestine. In this study we investigated the possibility that p10-mer, a decapeptide from durum wheat (QQPQDAVQPF), which was previously shown to prevent the activation of celiac peripheral lymphocytes, may exert an inhibitory effect on peptic-tryptic digested gliadin (PT-Gly)-stimulated intestinal carcinoma CACO-2 cells. In these cells, incubated with PT-Gly or p31-43 α-gliadin derived peptide in the presence or in the absence of p10-mer, IRAK1 activation and NF-kB, ERK1/2 and p38 MAPK phosphorylation were measured by immunoblotting, Cyclooxigenase 2 (COX-2) activity by PGE-2 release assay, and production of cytokines in the cell supernatants by ELISA. Our results showed that pre-treatment of CACO-2 cells with p10-mer significantly inhibited IRAK1 activation and NF-kB, ERK1/2 and p38 MAPK phosphorylation, as well as COX-2 activity (i.e. PGE-2 release) and production of the IL-6 and IL-8 pro-inflammatory cytokines, induced by gliadin peptides. These findings demonstrate the inhibitory effect of the p10-mer peptide on inflammatory response in CACO-2 cells. The results of the present study show that this p10-mer peptide can modulate "in vitro" the inflammatory response induced by gliadin peptides, allowing to move towards new therapeutic strategies. Turning off the inflammatory response, may in fact represent a key target in the immunotherapy of celiac disease

Effect of gliadin peptide 10-mer on p31–43 activity and entrance in CACO-2 cells.

<p>(<b>A</b>) CACO-2/TC7 cells, either unstimulated or stimulated with p31–43 (50 µg/ml), p10-mer (50 µg/ml) + p31–43 (50 µg/ml), were analyzed by Western blot for ERK phosphorylation and NF-kB activation. (<b>i</b>) Phosphorylated levels of ERK were analyzed in whole cell extracts by Western blot with anti-phospho-ERK1/2 antibodies; for control, the blotted membranes were stripped and reprobed with anti-ERK1/2 antibodies. Bound antibodies were visualized with HRP-conjugated IgG and immunoreactivity was assessed by ECL. (<b>ii</b>) NF-kB activation was analyzed in whole cell extracts by Western blot with anti-phospho-NF-kB p65 Ser antibodies; for control, the blotted membranes were stripped and reprobed with anti-NF-kB p65 antibodies. Bound antibodies were visualized with HRP-conjugated IgG and immunoreactivity was assessed by ECL. Densitometric analysis was performed using ImageJ version 1.46 software, peaks were reproduced by reading the Western Blot bands. One example representative of 3 experiments. (<b>B</b>) CACO-2/TC7 cells, either unstimulated or stimulated with biotinylated p31–43 (50 µg/ml), p10-mer (50 µg/ml) + biotinylated p31–43 (50 µg/ml), were immunostained with streptavidin-AlexaFluor and analyzed by a BD FACSCalibur flow cytometer. (<b>C</b>) CACO-2/TC7 cells, either unstimulated or stimulated with biotinylated p31–43 (50 µg/ml), p10-mer (50 µg/ml) + biotinylated p31–43 (50 µg/ml), were immunostained with streptavidin-AlexaFluor. The images were acquired using an Olympus U RFL fluorescence microscope.</p