Radiolysis, Photolysis, Sonolysis and Sonoprotection of Cells

Division Of Clinical Sciences - Nci

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Abstract

Summary of work: Sonodynamic therapy is a promising new modality for cancer treatment based on the synergistic effects of cell killing by a combination of sonosensitizer and ultrasound. Ultrasound can penetrate deeply into tissue and can be focused in a small region of tumor to activate non-toxic molecules (e.g. porphyrins ) thus minimizing undesirable side effects. The experimental evidence suggests that sonosensitization is due to the chemical activation of sonosensitizers inside or in close vicinity of hot collapsing cavitation bubbles to form sensitizer-derived radicals either by direct pyrolysis of the sensitizer at the water-gas interface or due to the reactions of hydrogen atoms and hydroxyl radicals formed by the pyrolysis of water. The free radicals derived from the sonosensitizer (mostly carbon-centered) react with oxygen to form peroxyl and alkoxyl radicals. Unlike OH radicals and H atoms which are formed by pyrolysis inside cavitation bubbles, the reactivity of alkoxyl and peroxyl radicals with organic compounds in biological media is much lower and hence they have a higher probability of reaching critical cellular sites. Our recent studies have shown that the long chain ( C5-C8 ) n-alkyl glucopyranosides completely inhibit ultrasound induced cytolysis .(This protective effect has possible applications in HIFU ( High intensity focused ultrasound ) for tumor treatment and in ultrasound assisted drug delivery and gene therapy. n-Alkyl glucopyranosides with hexyl ( 5mM ), heptyl ( 3mM ), octyl( 2mM ) n-alkyl chains protected 100 % of HL-60 cells in vitro from 1.057 MHz ultrasound induced cytolysis under a range of conditions which resulted in 35% to 100% cytolysis in the absence of glucopyranosides. However the hydrophilic methyl-beta-D-glucopyranoside did not protect cells. The surface active n-alkyl glucopyranosides accumulate at the gas-liquid interface of cavitation bubbles(1). The OH radicals and H atoms formed in collapsing cavitation bubbles react by H-atom abstraction from either the n-alkyl chain or the glucose moiety of the n-alkylglucopyranosides. Owing to the high concentration of the long chain surfactants at the gas-liquid interface of cavitation bubbles , the initially formed carbon radicals on the alkyl chains are transferred to the glucose moieties to yield radicals which react with oxygen leading to the formation of hydrogen peroxide. Our recent measurements of the hydrogen peroxide yields at 614 kHz and 1.057 MHz from oxygen-saturated solutions of long chain ( hexyl , octyl ) glucopyranosides compared with methyl-beta-D-glucopyranoside are consistent with the proposed mechanism of sonoprotection. This sequence of events prevents sonodynamic cell killing by initiation of lipid peroxidation chain reactions in cellular membranes by peroxyl and/or alkoxyl radicals. The effect of ultrasound frequency (from 47 kHz to 1 MHz ) on the ability of a homologous series of n-alkylglucopyranosides to protect cells from ultrasound-induced cytolysis was investigated. Comparisons of the protective ability of this series of n-alkylglucopyranosides with our earlier studies of their accumulation at the gas/solution interface of cavitation bubbles show that the ability of these surfactants to accumulate at this gas/solution interface is governed by the dynamic absorption properties and not the equilibrium absorption properties of these surfactants(1). Therapeutic applications of ultrasound to drug activation, apoptosis induction, gene transfer and changes of gene expression were reviewed (4). The ultrasound-induced cytolysis of human leukemia (HL-60) cells is enhanced in the presence of micron-sized alumina particles (3) . This effect is due to an increase of acoustic cavitation activity resulting in a greater amount of sonochemical activity as measured by an increase in the hydroxyl radical yield. Influence of changing Pulse Repetition Frequency on the Chemical and Biological Effects induced by Low Intensity Ultrasound in-vitro (2)(collaboration with T. Kondo et al.) . The influence of changing the pulse repetition frequency (PRF) of 1 MHz ultrasound from 0.5 to 100 Hz on the chemical and biological effects of low intensity ultrasound was studied (2). A significant effect of PRF on OH radical formation, cell killing of U 937 cells and of apoptosis induction was observed. The lowest free radical formation and cell killing and the highest cell viability were found at 5 Hz (100 millisecond pulse duration.) In contrast, no correlation was found between sucrose hydrolysis (a reaction that does not occur at normal temperature and pressure) and PRF. To our knowledge this is the first report devoted to the impact of low PRF at low intensities on ultrasound-induced chemical and biological effects and the mechanisms involved. This study has introduced the role of ultrasound streaming (convection) and explored its importance in comparison to standing waves. Modulation control over ultrasound-mediated gene delivery. Evaluating the importance of standing waves ( with T. Kondo et al. ) Low modulation frequencies from 0.5 to 100 Hz have been shown to alter the characteristics of the ultrasound field producing solution agitation ( less than 5 Hz: region of ultrasound streaming prevalence ) or stagnancy ( 5 Hz;region of standing waves establishment . In this study, the same conditions were used to depict the changes in exogoneous DNA delivery in these regions. The luciferase expression data revealed that lower modulations were more capable of enhancing delivery at the expense of viability. On the contrary, the viability was conserved at higher modulations while delivery was found to be null. Cavitational activity and acoustic streaming were the effectors beyond the observed pattern and delivery enhancement was shown to be mediated mainly through sonoporation. To, promote transfection, the addition of calcium ions or of an echo contrast agent ( Levovist ) was proposed. Depending on the mechanism involved in each approach , differential enhancement was observed in both regions and at the interim zone (5 Hz). In both cases, the enhancement in the standing waves was significant reaching 16.0 and 3.3 folds, respectively. Therefore, it was concluded that although the establishment of standing waves is not the only prerequisite for high transfection rates, yet it is a key element in optimization when other factors such as proximity and cavitation are considered. 1. Sostaric, J.Z., Miyoshi,N., Cheng J.Y.&Riesz,P. Dynamic adsorption properties of n-alkyl glucopyranosides determine their ability to inhibit cytolysis mediated by acoustic cavitation. J. of Physical Chemistry B 112, 12703-12709, (2008) 2. Buldakov, M.A., Hassan, M.A., Zhao, Q.L., Feril, L.B., Kudo, N., Kondo, T., Litvyakov, N.V., Bolshakov, M.A., Rostov, V.V., Cherdyntseva, N.V. and Riesz, P. Influence of changing pulse repetition frequency on chemical and biological effects induced by low intensity ultrasound in-vivo. Ultrasonics Sonochemistry 16, 392-397 (2009) 3. Miyoshi, N., Tuziuti, T., Yasui K., Iida, Y., Shimizu, N., Riesz,P., &J.Z. Sostaric. Ultrasound-induced cytolysis of cancer cells is enhanced in the presence of micron-sized alumina particles. Ultrasonics Sonochemistry 15, 881-890 (2008) 4. Yoshida,T., Kondo, T., Ogawa, R., Zhao, Q., Hassan, M., Watanabe, A., Takasaki, I., Tabuchi, Y., Shoji, M., Kudo, N., Feril, L., Tachibana, K., Buldakov, M., Honda, T., Tsukada, K.&Riesz, P., Molecular therapy by ultrasound. The mechanism [summary truncated at 7800 characters]