Effects of preparation conditions of poly(lactide-co-glycolide) nanoparticles loaded with amphiphilic porphyrins and their photoactivities.

Three porphyrins, (5,10,15,20-tetra(3-hydroxyphenyl)porphyrin, 5-hexyl-10,20-bis(3-hydroxyphenyl)-porphyrin and 5-hexyl-10,15,20-tris(3-hydroxyphenyl)porphyrin), with different amphiphilicities and equal singlet oxygen quantum yields in ethanol, were encapsulated into 50:50 poly(lactide-co-glycolide), nanoparticles prepared by the emulsion/evaporation technique. A 22 factorial design was utilized to evaluate the influence of the porphyrin/polymer mass ratio and the percentage of ethanol in the aqueous phase on the size and zeta potential of the nanoparticles. Increasing both the amount of ethanol and the porphyrin/polymer ratio decreases the size and increases zeta potential for the photosensitizers studied, except for 5-hexyl-10,15,20-tri(3-hydroxyphenyl)porphyrin. Entrapment efficiency depended on the individual m-hydroxyphenylporphyrin and ranged from 69 to 97%. After 1.5 h incubation with m-hydroxyphenylporphyrin-loaded nanoparticles the percentages of intracellular uptake were the same for all porphyrins since the molecules are confined in the nanoparticles, hampering the interaction of the amphiphilic photosensitizers with the cellular membrane. All encapsulated porphyrins caused the same decrease of cell viability and always localized in the perinuclear region of the cells. Results show that these m-hydroxyphenylporphyrins, although with different amphiphilicities, have equal photodynamic efficacies.

Effects of preparation conditions on the characteristics of poly(lactide-co-glycolide) nanospheres loaded with chloro(5,10,15,20-tetraphenylporphyrinato)indium(III).

A 2(4-1) fractional factorial design was utilized to evaluate the influence of four preparation conditions on six characteristics of poly(lactide-co-glycolide) nanospheres loaded with chloro(5,10,15,20-tetraphenylporphyrinato)indium(III). Ethanol in the aqueous phase and the stirring rate were the factors that most influenced the nanosphere characteristics. An increase in these factors caused a decrease in nanosphere size, recovery yield and residual chloroform and an increase in the percent of residual poly(vinyl alcohol). The synergic interaction between these two factors caused an increase in the percent residual chloroform. The entrapment efficiency was increased by an increase of ethanol in the aqueous phase or an increase in the percent poly(vinyl alcohol), but an overall decrease was obtained due to a synergic interaction between these factors. The stirring rate was the only parameter that caused an increase of the zeta potential. Evolutionary operations were then carried out based on the results from the fractional factorial design and nanospheres were obtained with sizes smaller than 200 nm.

In vitro photodynamic activity of chloro(5,10,15,20-tetraphenylporphyrinato)indium(III) loaded-poly(lactide-co-glycolide) nanoparticles in LNCaP prostate tumour cells.

In(III)-meso-tetraphenylporphyrin (InTPP) was encapsulated into nanoparticles (smaller than 200 nm) of poly(d,l-lactide-co-glycolide) (PLGA) using the emulsification-evaporation technique. The photodynamic efficacy of InTPP-loaded nanoparticles and its cellular uptake was investigated with LNCaP prostate tumour cells, in comparison with the free InTPP. The effects of incubation time (1-3h), drug concentration (1.8-7.7 micromol/L) and incident light dose (15-45 J/cm(2)) with both encapsulated and free InTPP were studied. The type of cell death induced by the photochemical process using both encapsulated and free InTPP was also investigated. Cell viability was reduced more significantly with increasing values of these effects for InTPP-loaded nanoparticles than with the free drug. The cellular death induced by both encapsulated and free InTPP was preponderantly apoptotic. Confocal laser scanning microscopy data showed that the InTPP-loaded nanoparticles, as well free InTPP, were localized in the cells, and always in the perinuclear region. Encapsulated InTPP was measured by the intensity of fluorescence intensity of cell extracts and was three times more internalized into the cells than was the free InTPP. Electron paramagnetic resonance experiments corroborated the participation of singlet oxygen in the photocytotoxic effect of nanoparticles loaded with InTPP.

Evaluation of the performance of different plastics used to seal nylon cDNA arrays / Avaliação da performance de diferentes plásticos usados para selar arranjos de cDNA em náilon

cDNA arrays are a powerful tool for discovering gene expression patterns. Nylon arrays have the advantage that they can be re-used several times. A key issue in high throughput gene expression analysis is sensitivity. In the case of nylon arrays, signal detection can be affected by the plastic bags used to keep membranes humid. In this study, we evaluated the effect of five types of plastics on the radioactive transmittance, number of genes with a signal above the background, and data variability. A polyethylene plastic bag 69 μm thick had a strong shielding effect that blocked 68.7 percent of the radioactive signal. The shielding effect on transmittance decreased the number of detected genes and increased the data variability. Other plastics which were thinner gave better results. Although plastics made from polyvinylidene chloride, polyvinyl chloride (both 13 μm thick) and polyethylene (29 and 7 μm thick) showed different levels of transmittance, they all gave similarly good performances. Polyvinylidene chloride and polyethylene 29 mm thick were the plastics of choice because of their easy handling. For other types of plastics, it is advisable to run a simple check on their performance in order to obtain the maximum information from nylon cDNA arrays.

Os arranjos de cDNA são uma poderosa ferramenta para o estudo de padrões de expressão gênica. Os arranjos em membranas de náilon apresentam ainda a vantagem de poderem ser reutilizados diversas vezes. Porém, um ponto bastante delicado em estudos de expressão gênica em larga escala é a sensibilidade. No caso de arranjos em membranas de náilon, a detecção dos sinais pode ser afetada pelo envoltório plástico utilizado para manter as membranas úmidas. Nesse estudo, nós avaliamos os efeitos de cinco tipos de plásticos na transmissão radioativa detectada, no número de genes com sinal acima da emissão de fundo e na variabilidade dos dados. O plástico produzido com polietileno com 69 μm de espessura apresentou uma forte interferência na emissão radioativa, bloqueando 68.7 por cento do sinal detectado. Este bloqueio na transmitância diminuiu o numero de genes detectados e aumentou a variabilidade dos dados. Outros plásticos mais finos tiveram resultados melhores. Apesar de plásticos feitos de cloreto de polivinilideno e cloreto de polivinila (ambos com 13 μm de espessura) e polietileno (29 e 7 μm de espessura) terem diferentes níveis de transmitância, todos apresentaram performances semelhantes nos testes realizados. Cloreto de polivinilideno e polietileno com 29 μm de espessura foram os plásticos escolhidos devido à facilidade de manuseio. Para outros tipos de plásticos, é recomendável realizar um teste de suas performances antes de utilizá-los para envolver membranas de náilon, de forma a obter o máximo de informação dos experimentos com arranjos de cDNA.

Cadmium accumulation in sunflower plants influenced by arbuscular mycorrhiza.

In order to investigate the cadmium (Cd) accumulation patterns and possible alleviation of Cd stress by mycorrhization, sunflower plants (Helianthus annuus L.) were grown in the presence or absence of Cd (20 micromol L(-1)) and inoculated or not inoculated with the arbuscular mycorrhizal fungus (AMF) Glomus intraradices. No visual symptoms of Cd phytotoxicity were observed; nevertheless, in non-mycorrhizal plants the presence of Cd decreased plant growth. The addition of Cd had no significant effect on either mycorrhizal colonization or the amount of extra-radical mycelia that was produced by the AMF. Cd accumulated mainly in roots; only 22% of the total Cd absorbed was translocated to the shoots, where it accumulated to an average of 228 mg Cd kg(-1). Although the shoot-to-root ratio of Cd was similar in both the AMF inoculated and non-inoculated plants, the total absorbed Cd was 23% higher in mycorrhizal plants. Cd concentration in AMF extra-radical mycelium was 728 microg g(-1) dry weight. Despite the greater absorption of Cd, mycorrhizal plants showed higher photosynthetic pigment concentrations and shoot P contents. Cd also influenced mineral nutrition, leading to decreased Ca and Cu shoot concentrations; N, Fe and Cu shoot contents; and increased S and K shoot concentrations. Cd induced guaiacol peroxidase activity in roots in both mycorrhizal and non-mycorrhizal plants, but this increase was much more accentuated in non-mycorrhizal roots. In conclusion, sunflower plants associated with G. intraradices were less sensitive to Cd stress than non-mycorrhizal plants. Mycorrhizal sunflowers showed enhanced Cd accumulation and some tolerance to excessive Cd concentrations in plant tissues.

Involvement of mitochondria in apoptosis of cancer cells induced by photodynamic therapy

A terapia fotodinâmica (TFD) é um promissor tratamento clínico que emprega a combinação de luz visível, oxigênio e uma substância fotossensibilizadora para tratar doenças de caráter oncológico. Esse tipo de tratamento induz a morte celular programada (apoptose) em tecido tumoral. No entanto, o mecanismo de indução de apoptose por TFD não está bem caracterizado. Muitos fotossensibilizadores possuem como alvo principal a mitocôndria. O tratamento com TFD causa danos nessa organela e induz a apoptose através da liberação de citocromo c, fator de indução de apoptose, Smac/DIABLO e certas pró-caspases. A liberação dessas proteínas, a partir da mitocôndria, é regulada por uma família de proteínas pró e antiapoptóticas, conhecida como família BCL-2. A proposta dessa minirrevisão é apresentar algumas publicações recentes que focalizam o envolvimento da mitocôndria em apoptose de células cancerígenas induzida por TFD.