Non-irradiated Bystander Fibroblasts Attenuate Damage to Irradiated Cancer Cells

Introduction and aim: Radiation-induced bystander effect is described as a different type of responses, displayed by non-irradiated neighbouring cells, induced by signals transmitted from irradiated cells. These responses in bystander cells include genetic damage (SCE, micronuclei, genomic instability), apoptosis induction and other non-necessarily detrimental effects. Bystander effect might bear some implications for coexisting normal cells non-targeted by cancer radiotherapy. However, it is possible, that bystander effect can act in opposite direction, and non-irradiated cells can in some way influence the response of targeted cells. Our experiments in vitro were aimed at evaluating this concept. Materials and methods: Lung Lewis carcinoma cells (LLC), growing in monolayer in 6-well plates, were irradiated with 2 or 4 Gy dose of X-rays (using a 6 MeV accelerator suited for therapeutic purposes). After irradiation, the cells were co-cultured with non-irradiated NIH3T3 mice fibroblasts, the latter growing (in monolayer) in special inserts. Such system allowed separation of the two kinds of cells, with medium freely circulating through the separation membrane (pore size 0.4 µm). Thus, species released by irradiated cells could be transmitted to non-irradiated neighbours and vice versa. Results and discussion: The bystander effects, caused by irradiated cancer cells, which were observed in non-targeted fibroblasts included dose-dependent elevation of micronucleus and apoptosis frequency indicating that irradiated cancer cells can induce damage in normal fibroblast cells. However, irradiated LLC cancer cells co-incubated with fibroblasts presented lower levels of this type of cytogenetic damage and apoptosis in comparison with LLC cells incubated after irradiation without fibroblasts on inserts or without inserts at all. Our current studies attempt to search for agents and signalling responsible for observed bilateral bystander effects. Supported by the Polish Ministry of Science and High Education, grant no N406 101/31/387

Dose-rate Effects are Dependent on Cell Lines

Background: A number of studies, including our own, have indicated that low dose rate of ionizing radiation might be (within some range) more effective in killing cells than high dose rate. This phenomenon is defined as inverse dose-rate effect and is mediated mainly by post radiation oxidative reactions, as radiation peroxidative damage increases with decreased dose rate. Knowledge of biological dose rate-dependent effects might have influence upon the extension of therapeutic maneuvers and limitation of normal tissue reaction. Aim: The aim of the present study was to evaluate the dose-rate effects on lymphoblastic cells since this type of cells is a target in total body irradiation. Materials and methods: Lymphoblastic K562 cells were exposed to 2 Gy X-ray dose (using a 6 MeV accelerator for therapeutic purposes), at high dose rate (2Gy/min), low dose rate (0.07 Gy/min) and at high dose rate, but total dose was given as short separated pulses delivered during the same time as low dose rate (28 min 34 sec). Cells were suspended in RPMI medium supplemented with serum and irradiated in culture flasks. After irradiation cells were incubated for 0.5 60 h and micronuclei, apoptotic, necrotic and mitotic cells were scored under fluorescent microscope. Activity of superoxide dismutase isoenzymes (MnSOD and CuZuSOD), and glutathione peroxidise as well as end-product of lipid peroxidation (malondialdehyde) were also measured. Results and discussion: Our results indicated that 2 Gy dose at high dose rate induced higher yield of micronuclei than the same dose at low dose rate or 2 Gy dose given in many pulses. At the same time high dose-rate was less effective in apoptosis induction in K562 cells than low dose-rate or pulsed dose. This effect may differ in different cell lines, since our study on murine carcinoma AT478 cells indicated opposite trend. Furthermore, there was no dose-rate effect in the activation of antioxidant enzymes in lymphoblastic cells, whereas inverse dose rate effect was observed for carcinoma cells. The observed differences may also result from the type of cell growth, K562 grow in suspension, but AT478 are adherent cells. The adherent cells may suffer higher damage from low dose rate during long time of irradiation, in part due to bystander effects. The ongoing study is aimed at evaluating the mechanisms responsible for observed differences and possible signalling activated at different dose rate irradiation. Supported by the Polish Ministry of Science and High Education, grant no N406 101/31/387

Spectroscopic and biological studies of a novel synthetic chlorin derivative with prospects for use in PDT

Photosensitizers with desirable combinations of chemical, photophysical and biological properties are essential for improving the efficacy of photodynamic therapy (PDT) against various cancers. Chlorins seem to be promising candidates for photodynamic therapy (PDT) owing to their photophysical properties. This paper reports spectroscopic and biological properties of a novel synthetic chlorin derivative. Cytotoxicity, phototoxicity as well as subcellular localization of the novel derivative was studied using Lewis lung carcinoma cultured cells (LLC). In the examined concentration range no significant cytotoxic effects were found but high phototoxicity was observed. Confocal laser scanning microscopy demonstrated that the compound, upon entering cells, was localized in the perinuclear cytoplasm of LLC cells. Using fluorescent microscopy we investigated the impact of PDT based on the novel compound upon cytoskeleton and DNA structure of LLC cells. Our results indicate that liposomes are effective in transferring the chlorin photosensitizer into the studied cells, leading to their high photosensitization, whereas the non-carrier delivery mode (i.e., DMSO) is rather useless for such purposes

Synthesis and characterization of quinoline-based thiosemicarbazones and correlation of cellular iron-binding efficacy to anti-tumor efficacy

Abstract not available.Maciej Serda, Danuta S. Kalinowski, Anna Mrozek-Wilczkiewicz, Robert Musiol, Agnieszka Szurko, Alicja Ratuszna, Namfon Pantarat, Zaklina Kovacevic, Angelica M. Merlot, Des R. Richardson, Jaroslaw Polansk

Specific Targeting of Melanotic Cells with Peptide Ligated Photosensitizers for Photodynamic Therapy

Abstract A strategy combining covalent conjugation of photosensitizers to a peptide ligand directed to the melanocortin 1 (MC1) receptor with the application of sequential LED light dosage at near-IR wavelengths was developed to achieve specific cytotoxicity to melanocytes and melanoma (MEL) with minimal collateral damage to surrounding cells such as keratinocytes (KER). The specific killing of melanotic cells by targeted photodynamic therapy (PDT) described in this study holds promise as a potentially effective adjuvant therapeutic method to control benign skin hyperpigmentation or superficial melanotic malignancy such as Lentigo Maligna Melanoma (LMM)