Combined Effect of Neutron Radiation and Curcumin on Breast Cancer Cells Cytotoxicity in 3D Culture Medium

Introduction: Introduction: Chemotherapy, biotherapy, and radiotherapy play a limited but important role in treating breast cancer. For more efficient treatment, combination therapy could be an appropriate option. In this study, radiotherapy using neutron radiation emitted from a 241Am-Be neutron source, as well as biotherapy using curcumin (80 µM) was combined to investigate the efficiency of treatment towards MCF-7 breast cancer in a three dimensional (3D) culture medium. Methods: Methods: MTT, neutral red uptake assay, nitric oxide, glutathione assay, catalase, cytochrome c, comet assay, and caspase-3 were used to determine the effect of neutron radiation and also neutron and curcumin combination on the viability of cancer cells. Results: Results: The results of cytotoxicity test showed that neutron irradiation with or without curcumin at 5, 10, 15, and 20 h reduced the survival of tumor cells. Moreover, the rate of apoptosis due to the neutron effect at different irradiation times enhanced with the increasing time. Conclusion: Conclusion: Due to the significant anticancer effect of curcumin in 3D culture, using this molecule before or after neutron therapy is recommended.

Effects of Americium-241 and humic substances on Photobacterium phosphoreum: bioluminescence and diffuse reflectance FTIR spectroscopic studies.

The integral bioluminescence (BL) intensity of live Photobacterium phosphoreum cells (strain 1883 IBSO), sampled at the stationary growth stage (20 h), was monitored for further 300 h in the absence (control) and presence of (241)Am (an α-emitting radionuclide of a high specific activity) in the growth medium. The activity concentration of (241)Am was 2 kBq l(-1); [(241)Am]=6.5×10(-11) M. Parallel experiments were also performed with water-soluble humic substances (HS, 2.5 mg l(-1); containing over 70% potassium humate) added to the culture medium as a possible detoxifying agent. The BL spectra of all the bacterial samples were very similar (λ(max)=481±3 nm; FWHM=83±3 nm) showing that (241)Am (also with HS) influenced the bacterial BL system at stages prior to the formation of electronically excited states. The HS added per se virtually did not influence the integral BL intensity. In the presence of (241)Am, BL was initially activated but inhibited after 180 h, while the system (241)Am+HS showed an effective activation of BL up to 300 h which slowly decreased with time. Diffuse reflectance infrared Fourier transform (DRIFT) spectroscopy, applied to dry cell biomass sampled at the stationary growth phase, was used to control possible metabolic responses of the bacteria to the α-radioactivity stress (observed earlier for other bacteria under other stresses). The DRIFT spectra were all very similar showing a low content of intracellular poly-3-hydroxybutyrate (at the level of a few percent of dry biomass) and no or negligible spectroscopic changes in the presence of (241)Am and/or HS. This assumes the α-radioactivity effect to be transmitted by live cells mainly to the bacterial BL enzyme system, with negligible structural or compositional changes in cellular macrocomponents at the stationary growth phase.

Comparison of RBE values of high-LET α-particles for the induction of DNA-DSBs, chromosome aberrations and cell reproductive death.

BACKGROUND: Various types of radiation effects in mammalian cells have been studied with the aim to predict the radiosensitivity of tumours and normal tissues, e.g. DNA double strand breaks (DSB), chromosome aberrations and cell reproductive inactivation. However, variation in correlations with clinical results has reduced general application. An additional type of information is required for the increasing application of high-LET radiation in cancer therapy: the Relative Biological Effectiveness (RBE) for effects in tumours and normal tissues. Relevant information on RBE values might be derived from studies on cells in culture. METHODS: To evaluate relationships between DNA-DSB, chromosome aberrations and the clinically most relevant effect of cell reproductive death, for ionizing radiations of different LET, dose-effect relationships were determined for the induction of these effects in cultured SW-1573 cells irradiated with gamma-rays from a Cs-137 source or with α-particles from an Am-241 source. RBE values were derived for these effects. Ionizing radiation induced foci (IRIF) of DNA repair related proteins, indicative of DSB, were assessed by counting gamma-H2AX foci. Chromosome aberration frequencies were determined by scoring fragments and translocations using premature chromosome condensation. Cell survival was measured by colony formation assay. Analysis of dose-effect relations was based on the linear-quadratic model. RESULTS: Our results show that, although both investigated radiation types induce similar numbers of IRIF per absorbed dose, only a small fraction of the DSB induced by the low-LET gamma-rays result in chromosome rearrangements and cell reproductive death, while this fraction is considerably enhanced for the high-LET alpha-radiation. Calculated RBE values derived for the linear components of dose-effect relations for gamma-H2AX foci, cell reproductive death, chromosome fragments and colour junctions are 1.0 ± 0.3, 14.7 ± 5.1, 15.3 ± 5.9 and 13.3 ± 6.0 respectively. CONCLUSIONS: These results indicate that RBE values for IRIF (DNA-DSB) induction provide little valid information on other biologically-relevant end points in cells exposed to high-LET radiations. Furthermore, the RBE values for the induction of the two types of chromosome aberrations are similar to those established for cell reproductive death. This suggests that assays of these aberrations might yield relevant information on the biological effectiveness in high-LET radiotherapy.

Lauriston S. Taylor Lecture: the quest for therapeutic actinide chelators.

All of the actinides are radioactive. Taken into the body, they damage and induce cancer in bone and liver, and in the lungs if inhaled, and U(VI) is a chemical kidney poison. Containment of radionuclides is fundamental to radiation protection, but if it is breached accidentally or deliberately, decontamination of exposed persons is needed to reduce the consequences of radionuclide intake. The only known way to reduce the health risks of internally deposited actinides is to accelerate their excretion with chelating agents. Ethylendiaminetetraacetic acid (EDTA) and diethylenetriaminepentaacetic acid (DTPA) were introduced in the 1950's. DTPA is now clinically accepted, but its oral activity is low, it must be injected as a Ca(II) or Zn(II) chelate to avoid toxicity, and it is structurally unsuitable for chelating U(VI) or Np(V). Actinide penetration into the mammalian iron transport and storage systems suggested that actinide ions would form stable complexes with the Fe(III)-binding units found in potent selective natural iron chelators (siderophores). Testing of that biomimetic approach began in the late 1970's with the design, production, and assessment for in vivo Pu(IV) chelation of synthetic multidentate ligands based on the backbone structures and Fe(III)-binding groups of siderophores. New efficacious actinide chelators have emerged from that program, in particular, octadentate 3,4,3-LI(1,2-HOPO) and tetradentate 5-LIO(Me-3,2-HOPO) have potential for clinical acceptance. Both are much more effective than CaNa3-DTPA for decorporation of Pu(IV), Am(III), U(VI), and Np(IV,V), they are orally active, and toxicity is acceptably low at effective dosage.

Effect of americium-241 alpha-particles on the dose-response of chromosome aberrations in human lymphocytes analysed by fluorescence in situ hybridization.

PURPOSE: To evaluate by the fluorescent in-situ hybridization (FISH) technique the dose-response and intercellular distribution of alpha-particle-induced chromosome aberrations. In particular, the validity of using the yield of characteristic types of chromosome abnormalities in stable cells as quantitative indicators for retrospective dose reconstruction has been evaluated. MATERIAL AND METHODS: Monolayers of human peripheral lymphocytes were exposed at doses from 0.02 to 1 Gy to alpha-particles emitted from a source of americium-241. The most probable energy of the alpha-particles entering the cells was 2.7 MeV. FISH painting was performed using DNA probes for chromosomes 2, 4 and 8 in combination with a pan-centromeric probe. In complete first-division cells, identified by harlequin staining, aberrations involving painted target chromosomal material were recorded as well as aberrations involving only unpainted chromosomal material. RESULTS: In total, the percentage of complex aberrations was about 35% and no dose dependence was observed. When complex-type exchanges were reduced to simple base types, the different cell distributions were clearly over-dispersed, and the linear coefficients of the dose-effect curves for translocations were significantly higher than for dicentrics. For past dose reconstruction, only a few complex aberrations were in stable cells. The linear coefficient obtained for transmissible aberrations in stable cells was more than seven times lower than that obtained in all analysed cells, i.e. including unstable cells. CONCLUSION: FISH-based analysis of complex rearrangements allows discrimination between partial-body exposures to low-linear energy transfer radiation and high-linear energy transfer exposures. In assessing past or chronic exposure to alpha-particles, the use of a dose-effect curve obtained by FISH-based translocation data, which had not excluded data determined in unstable cells, would underestimate the dose. Insertions are ineffective biomarkers because their frequency is too low.

Apoptosis of rat kidney cells after 241-americium administration.

Tumors induction by americium is well known but there are no data on the biological effects of this radionucleide at subcellular level. In order to study the possible ultrastructural lesions induced by this element, a group of rats were injected with 241-Americium-citrate (9 kBq), once a week for five weeks and sacrificed 7 days after the last injection. We describe the alterations observed in the cortex kidney using cytochemical (TUNEL reaction) and histochemical (PAS staining) methods for light microscopy as well as electron microscopy techniques. Various types of lesions were detected: condensation of nuclear chromatine, fragmentation of the nuclei, swollen mitochondria, disappearance of mitochondrial crests and skrinking of the cytoplasm. This study clearly demonstrated the induction of apoptosis by americium in rat cortex kidney cells.

[The significance of modifying factors in evaluating the effect of radionuclides]. / Znachenie modifitsiruiushchikh faktorov pri otsenke vozdeistviia radionuklidov.

The authors provide the data obtained in experimental animals, pertaining to the influence of certain modifying factors on metabolism and biological effects induced by radioactive isotopes of iodine, by transuranium elements and beryllium. Out of all the modifying factors investigated, the form of the introduced compound and age turned out the most significant ones.

[Alkaline phosphatase activity of the blood serum in dogs exposed jointly to external gamma irradiation and alpha radiation]. / Aktivnost' shchelochnoi fosfatazy v syvorotke krovi sobak pri sovmestnom vozdeistvii vneshnego gamma-oblucheniia i al'fa-izlucheniia.

Alkaline phosphatase activity in dog blood serum was studied for two years following separate and combined exposure to gamma radiation (6.45 to 51.6 mC/kg) and inhaled submicron 239Pu oxide, containing 25 per cent of 241Am, in chronically effective amounts (approximately 7-10 kBq/kg). Alkaline phosphatase activity was of an undulatory character, and the significance of changes depended upon the kind and the level of radiation as well as the time lapsed from the start of the exposure. With the combined exposure to gamma and alpha radiation in the doses used, no enhancement of the effect was noted as compared with the action of each factor applied separately.

Influence of pregnancy and lactation on maternal deposition and perinatal uptake of 241Am in the rat.

To investigate the metabolism of 241Am as affected by pregnancy and lactation, female rats were injected with 5 muCi of 241Am intravenously while nulliparous, pregnant or lactating. The females and subsequent litters were killed at various times after injection to determine 241Am distribution and retention. The temporal relationship between injection and pregnancy influenced the tissue retention of 241Am in both dams and progeny. The half-time of 241Am in livers of pregnant or lactating rats was more than twice that of nulliparous rats, although the initial uptake was approximately 50% of the activity in all groups. Both spleen and femur accumulated more 241Am at 7-10 weeks after injection than at earlier times, but this increase could not be related to pregnancy. Approximately 10 times more 241Am was transferred to offspring from dams that were lactating when exposed than was transferred via the placenta when exposure occurred late in gestation. Furthermore, more 241Am was transmitted to the progeny via milk if exposure of the dam occurred during lactation rather than during pregnancy.

Radiosensitivity of vertebral bone marrow CFU-S surviving in mice internally contaminated with 239Pu or 241Am.

The radiosensitivity of pluripotent hemopoietic stem cells was studied in ICR "Swiss" mice (28 g/mouse) given i.v. 198.6 kBq 239Pu/kg as citrate complex or 208.6 kBq 241 Am/kg as nitrate at the age of 10 weeks. The bone marrow cells were examined at the early and late phases of radionuclide contamination. To obtain data for survival curves and D0 of stem cells the CFU-S assay was used and the donor vertebral marrow cells were exposed to the complementary X-irradiation either early after injection to the heavily irradiated recipients or to the "in vitro" irradiation given before the transplantation. To determine the iron uptake in splenic erythroid progeny the recipients given marrow cells unexposed to the X-rays received 37 kBq 59Fe 6 h before they were killed and the relative activity per colony was calculated. The radiation effect of the used actinides on the bone marrow cells resulted in decreased cellularity and seriously altered both relative and absolute CFU-S numbers. The radiosensitivity of CFU-S increased in all intervals examined (D0 from 0.60 to 0.86 Gy, in controls 0.97 to 1.06 Gy) and was more expressed when the CFU-S were exposed to the X-rays immediately after the bone marrow cell transplantation to the heavily irradiated hosts. The stem cell pool appeared, especially at older age, to be affected also in its ability to produce erythrocytic progeny.

Relative effectiveness of 241Am vs 226Ra approached by haemopoietic stem cell studies in various bone marrow sites of contaminated mice.

Haemopoietic stem cell assays allow investigation of local radiation damage at any time after contamination. These techniques were used to determine the relative effectiveness of 241Am and 226Ra in inducing damage in various cortical and trabecular bone marrow sites. Male Balb/c mice were injected with either 230 or 660 kBq 226RaCl2/kg body weight or with one of three doses of monomeric 241Am citrate at 138, 552 or 768 kBq 241Am/kg body weight. At 7 time intervals after injection (from 4 hr to 100 days) the colony-forming capacity of pluripotent stem cells (= CFU-s) and macrophage-granulocyte committed stem cells (CFU-c) was assayed in sternal marrow, marrow of lumbar vertebrae, of the trabecular distal end of the femur and of the femoral shaft (in which were distinguished axial marrow and marrow situated peripherally near the cortical bone). 226Ra retention and 241Am retention were measured in these bone sites. Skeletal doses and doses to the bone marrow sites were calculated. On the base of injected activity, 241Am was more effective than 226Ra in reducing the number of CFU-s and CFU-c but the effectiveness varied in the various bone marrow sites. CFU-s and CFU-c response is considered in relation to the retained radioactivity in the surrounding bone matrix and in relation to the mean marrow dose.