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Potentiation of hyperthermia-induced haemolysis of human erythrocytes by photodynamic treatment. Evidence for the involvement of the anion transporter in this synergistic interaction.

ABSTRACT: Heat treatment of human erythrocytes led to increased passive cation permeability, followed by haemolysis. K+ leakage was linear up to a loss of about 80% in the temperature range 46-54 degrees C. Kinetic analysis of the results revealed an activation energy of 246 kJ/mol, implicating a transition in the membrane as critical step. Pretreatment of erythrocytes with 4,4'-di-isothiocyano-2,2'-stilbenedisulphonate, chymotrypsin or chlorpromazine caused a potentiation of subsequent heat-induced K+ leakage. Photodynamic treatment of erythrocytes with Photofrin II, eosin isothiocyanate or a porphyrin-Cu2+ complex as sensitizer also induced an increase in passive cation permeability, ultimately resulting in colloid osmotic haemolysis. The combination of photodynamic treatment immediately followed by hyperthermia had a synergistic effect on K+ leakage. Analysis of the results by the Arrhenius equation revealed that both the activation energy and the frequency factor of heat-induced K+ leakage were decreased significantly by preceding photodynamic treatment, suggesting that hyperthermia and photodynamic treatment have a common target for the induction of K+ leakage. Several lines of reasoning indicate that this common target is band 3. A model is thus proposed for the observed potentiation of hyperthermically induced K+ leakage by photodynamic treatment, in which photo-oxidation of band 3 results in increased sensitivity to subsequent thermal denaturation. These phenomena may be of more general significance, as photodynamic treatment and hyperthermia interacted synergistically with respect to K+ leakage with L929 fibroblasts also.

SUBMITTER: Prinsze C

PROVIDER: S-EPMC1151208 | biostudies-other | 1991 Jul

REPOSITORIES: biostudies-other

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Project description:Thermal inactivation of glyceraldehyde-3-phosphate dehydrogenase appeared to be caused by a conformational mechanism, without involvement of covalent reactions. On the other hand, photodynamic inactivation of the enzyme (induced by illumination in the presence of Photofrin II) was caused by photo-oxidation of the essential thiol group in the active centre. A short photodynamic treatment of the enzyme, leading to only a limited inactivation, caused a pronounced potentiation of subsequent thermal inactivation, as measured over the temperature range 40-50 degrees C. Analysis of the experimental results according to the Arrhenius equation revealed that both the activation energy of thermal inactivation and the frequency factor (the proportionality constant) were significantly decreased by the preceding photodynamic treatment. The experimental results indicate a mechanism in which limited photodynamic treatment induced a conformational change of the protein molecule. This conformational change did not contribute to photodynamic enzyme inhibition, but was responsible for the decreased frequency factor and activation energy of subsequent thermal inactivation of the enzyme. The opposing effects of decreased activation energy and decreased frequency factor resulted in potentiation of thermal inactivation of the enzyme over the temperature range 40-50 degrees C. With other proteins, different results were obtained. With amylase the combined photodynamic and thermal effects were not synergistic, but additive, and photodynamic treatment had no effect on the frequency factor and the activation energy of thermal inactivation. With respect to myoglobin denaturation, the photodynamic and thermal effects were antagonistic over the whole practically applicable temperature range. Limited photodynamic treatment protected the protein against heat-induced precipitation, concomitantly increasing both the frequency factor and the activation energy of the process. These results offer a model for one of the possible mechanisms of synergistic interaction between photodynamic therapy and hyperthermia in cancer treatment.

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Potentiation of hyperthermia-induced haemolysis of human erythrocytes by photodynamic treatment. Evidence for the involvement of the anion transporter in this synergistic interaction.

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