El extracto acuoso de Phyllanthus orbicularis K protege al ADN plasmídico del daño inducido por las radiaciones ultravioletas / Phyllanthus orbicularis K aqueous extract protect plasmid DNA against the damage induced by ultraviolet radiation

Objetivo: Evaluar el efecto protector del extracto acuoso de Phyllanthus orbicularis, K ante el daño inducido por las radiaciones UVB y UVC. Material y métodos: Se empleó el ensayo con ADN plasmídico (pBluescript SK II) libre de célula. Se evaluó la capacidad del extracto acuoso de P. orbicularis de inducir roturas de cadenas en el plásmido, a las concentraciones 0,0001-2,0 mg/ml. Se realizaron estudios de protección del extracto frente a las radiaciones UVB y UVC a las concentraciones de 0,1-2,0 mg/ml. Se cuantificó la transmitancia del extracto frente ambos tipos de radiaciones. Resultados: Ninguna de las concentraciones evaluadas resultó genotóxica en 30 min de exposición. Las concentraciones ≥ 1 mg/ml de P. Obicularis sí indujeron roturas de cadenas a tiempos mayores de evaluación. El extracto de P. Orbicularis protegió al ADN frente a las radiaciones UVB y UVC a concentraciones ≥ 0,1 mg/ml y 0,5 mg/ml respectivamente. Conclusiones: En nuestras condiciones experimentales, el extracto acuoso de P. Orbicularis proteg eal ADN frente al daño inducido por las radiaciones UV (AU)

Aim: The aim of this work was to evaluate the protective effect of aqueous extract of Phyllanthus Orbicularis, K from the damage induced by UV radiation. Material and methods: The plasmid-based non cellular system was used. The extract capacity to induce DNA strand breaks was evaluated at 0.0001- 2.0 mg/ml concentrations. The protective effect of extract against UVB and UVC radiation was evaluated at 0.1- 2.0 mg/ml concentrations. The transmittance of extract was measured for both UV radiations. Results: The P. orbicularis aqueous extract was not genotoxic even after 30 min of exposure. Concentrations ≥ 1 mg/ml of extract induced strand breaks at major times of exposition. P. Orbicularis extract protected DNA against UVB and UVC radiation at concentrations ≥ 0.1 mg/ml and 0.5 mg/ml respectively. Conclusions: In our experimental conditions, P. Orbicularis aqueous extract protects DNA from damage induced by UV radiation (AU)

Elimination of free-living amoebae in fresh water with pulsed electric fields.

This study investigates the effects of pulsed electric fields on the inactivation of trophozoite form of Naegleria lovaniensis Ar9M-1 in batch and flow processes, systematically examining the lethal effect of field strength, pulse duration, number of pulses, and pulse frequency. Our results show that amoebae eradication is modulated by pulse parameters, composition of the pulsing medium, and physiological state of the cells. Cell survival is not related to the energy delivered to the cell suspension during the electrical treatment. For a given energy a strong field applied for a short cumulative pulse duration affects viability more than a weak field with a long cumulative pulsation. We also determine the optimal electrical conditions to obtain an inactivation rate higher than 95% while using the least energy. Flow processes allow to treat large-scale volumes. Our results show that the most efficient flow process for amoeba eradication requires a field parallel to the flow. Pulsed electric fields are a new and attractive method for inactivating amoebae in large volumes of fresh water.

Recent biotechnological developments of electropulsation. A prospective review.

During the last 25 years, basic research has improved our knowledge on the molecular mechanisms triggered at the membrane level by electric pulses. Applied aspects may now be used under safe conditions. Electropulsation is known as a very efficient tool for obtaining gene transfer in many species to produce genetically modified organisms (GMO). This is routinely used for industrial purposes to transfer exogenous activities in bacteria, yeasts and plants. The method is simple and of a low cost. But electropulsation is not limited to this application for biotechnological purposes. It is known that the field-associated membrane alterations can be irreversible. The pulsed species cannot recover after the treatment. Their viability is strongly affected. This appears as a very promising technology for the eradication of pathogenic microorganisms. Recent developments are proposed for sterilization purposes. New flow technologies of field generation allow the treatment of large volumes of solution. When high flow rates are used, microorganisms are submitted both to a hydromechanical and to an electrical stress. The synergy of the two effects may be present when suitable pulsing conditions are chosen. Several examples for the treatment of domestic water and in the food industry are described. Walled microorganisms are affected not only at the membrane level. We observed that alterations are present on the cell wall. A very promising technology is the associated controlled leakage of the cytoplasmic soluble proteins. Large dimeric proteins such as beta-galactosidases can be extracted at a high yield. High volumes can be treated by using a flow process. Extraction of proteins is obtained with many systems including mammalian cells.

The generation of reactive-oxygen species associated with long-lasting pulse-induced electropermeabilisation of mammalian cells is based on a non-destructive alteration of the plasma membrane.

Chinese hamster ovary (CHO) cells in suspension were subjected to pulsed electric fields suitable for electrically mediated gene transfer (pulse duration longer than 1 ms). Using the chemiluminescence probe lucigenin, we showed that a generation of reactive-oxygen species (oxidative jump) was present when the cells were electropermeabilised using millisecond pulses. The oxidative jump yield was controlled by the extent of alterations allowing permeabilisation within the electrically affected cell area, but showed a saturating dependence on the pulse duration over 1 ms. Cell electropulsation induced reversible and irreversible alterations of the membrane assembly. The oxidative stress was only present when the membrane permeabilisation was reversible. Irreversible electrical membrane disruption inhibited the oxidative jump. The oxidative jump was not a simple feedback effect of membrane electropermeabilisation. It strongly controlled long-term cell survival. This had to be associated with the cell-damaging action of reactive-oxygen species. However, for millisecond-cumulated pulse duration, an accumulation of a large number of short pulses (microsecond) was extremely lethal for cells, while no correlation with an increased oxidative jump was found. Cell responses, such as the production of free radicals, were present during electropermeabilisation of living cells and controlled partially the long-term behaviour of the pulsed cell.

Chinese hamster ovary cells sensitivity to localized electrical stresses.

Application of an external electric field on a cell suspension induces an alteration in the membrane structure giving free access to the cell cytoplasm. Under mild pulsation conditions, permeabilization is a reversible process which weakly affects cell viability while drastic electrical conditions lead to cell death. The field pulse must be considered as a complex stress applied on the cell assembly. This study is a systematic investigation of the stress effects of field strength, pulse duration and number of pulses, at given joule energy. The loss in cell viability is not related to the energy delivered to the system. At a given joule energy, a strong field during a short cumulated pulse duration affects more viability than using a weak field associated with a long cumulated pulsation. At a given field strength and for a given cumulated pulse duration an accumulation of short pulses is also observed to be very damaging for cells. A control by the delay between the pulses suggests a memory effect. The field effect appears also to be vectorial in line with the known asymmetry of the membrane organization. These results suggest that processes at a cellular level are involved, either an activation of cell death or damage in cellular functions.

The interchain disulfide bond between TCR alpha beta heterodimers on human T cells is not required for TCR-CD3 membrane expression and signal transduction.

In the present paper, it was attempted to define the amino acids or regions on TCR beta molecules that determine the TCR alpha-TCR beta interaction. Sequence studies on HBP-ALL variant cells with an intrinsic deficiency in TCR alpha beta dimer formation elucidated a conserved amino acid motif in the TCR-C beta beta-strand E, = Y(C)(L)(S)SRLR(V)(S)(A); this motif seems to represent one interaction area for the TCR alpha-TCR beta interaction. In addition, amino acids in the connecting peptide may be shaped in a precise structure (by the interactions with CD3 molecules?) involved in TCR alpha-TCR beta dimerization. This result was supported by the finding that the interchain disulfide bond between TCR alpha and beta chains is not required for membrane expression or transmembrane signal transduction of TCR alpha beta-CD3 complexes. Finally, comparative results from two membrane TCR-CD3-negative Jurkat variants R4.9 and E6.E12 suggest that TCR-C beta exon 1- and 2-encoded amino acids are important for the TCR beta-CD3 gamma epsilon association.