Mouse Model for the Biodosimetry Using Quantification of mRNA for DNA-damage Induced Genes in Peripheral White Blood Cells

Biodosimetry by the measurement of chromosomal damage in lymphocyte has been established. Although the principle is useful to estimate high-dose exposure, expansion of the method to estimate low-dose exposure is difficult. We aimed to develop a biodosimetric method that is suitable for low-dose (around 100 mGy) radiation exosure, and focused on the quantification of RNA in white blood cells. For the future application of the methodology to human blood, basic knowledges were obtained using mouse model. We previously reported increase in the levels of mRNAs for DNA damage-induced (DDI) genes in peripheral blood from whole-body x-irradiated mice (JRR 51, 265-275, 2010). The DDI genes are known to be expressed in radiation-sensitive proliferating cell which is slightly contained in peripheral blood, and it is expected that the relative rates of the level of DDI gene expression per the proliferating cells would show dose-dependency that is necessary for biodosimetry. When bax and puma mRNAs were chose as indexes of DDI gene expression level and c-myc mRNA was used as an index of proliferating cell number, the values of bax/myc and puma/myc rates in peripheral blood were kept low level without significant change among all the mice. When blood were isolated from mice 2h after whole-body x-irradiation in the various doses from 25 to 500mGy, these values of relative rates were increased in exposed dose-dependent manner. Similar dose-dependent increase was also observed in blood samples isolated from 4h after irradiation, but all the values elevated as compared with blood isolated from 2h after irradiation. Thus, records of blood sampling time and predicted irradiation time are essential in this model. No significant change was observed in these values of relative rates among different irradiation experiments using any production batches of mice. This mouse model can be useful not only to study radiation effect in vivo but also to elucidate basic knowledge that is necessary to expand the biodosimetric methodology to human.15th International Congress of Radiation Researc

Hydrogel formulation of Cs-absorbent and laxative drugs for enhancement of decorporation rate of internally contaminated radiocesium

Clinical decorporation procedure using Radiogardase to decrease body-burden of radiocesium caused by to the internal contamination accidents has been established in the last century. We aimed to improve the decorporation treatment to minimize stress of patients as the current tendency. The effects of several drugs were estimated by the measurement of the fecal and urinary excretion rates of radiocesium using mouse model. First, we examined the effect of pharmaceutical modification of the drug formulation of radiocesium-absorbents, because water-insoluble form of absorbents such as Prussian blue is sharp-pointed large crystal and may irritate the mucosa of gastro-intestinal (GI)-tract physically. Crystals of ferrous ferricyanides in the sizes of less than 1 micrometer were embedded in polyvinyl alcohol hydrogel with globular shap in the diameters from 10 to 50 micrometer. They were lyophilized for storage and hydrated just before administration. Though the rude crystal is rapidly excreted within 3h after the intrastomach administration, its hydrogel preparation elongates the retention time in GI-tract up to 6h, suggesting mitigation of irritation in mucosa by the formulation. Simultaneously, the excretion rate of internal radiocesium was increased by the use of the hydrogel formulation. Next, the effects of clinical drugs on the cesium excretion were examined in mouse. No diuretic drugs accelerated urinary excretion rate of internal radiocesium. In contrast, several drugs which has mild laxative action enhance intestinal excretion rate of internal radiocesium. These data show that decorporation rate of intestinal radiocesium can be enhanced by the procedure focusing on internal excretion rate, e.g. the simultaneous use of pharmaceutically formulated absorbents and laxatives.15th International Congress of Radiation Researc

Modification of nitroxyl contrast agents with multiple spins on the molecules and its proton T1-relaxivities

Paramagnetic nitroxyl radicals have been proposed as a redox sensitive MR contrast agent. A time-sequence of the T1-weighted MR images at 4.7 T showed a readily visible enhancement of image intensity in mice due to paramagnetic nitroxyl contrast agent and then sequentially showed gradual decrement of the image enhancement level. The dose 1.5 umol/g b.w. of the carbamoyl-PROXYL, which was a nitroxyl contrast agent used in the previous report, was enough lower than its maximum tolerated dose (MTD) level for mice. However, the dose 1.5 umol/g b.w. is near to the MTD of TEMPOL for mice, which is reported as 275 mg/kg b.w. (1.6 umol/g b.w.). In addition the dose 1.5 umol/g b.w. is lager than dose levels of common medicines (< 1 umol/g b.w.). Although the dose of the nitroxyl contrast agent used in future clinical trial should be decreased, the relatively small T1-relaxivity of the nitroxyl radicals (= 0.2 mM-1s-1) makes difficult to detect low concentration levels. Higher magnetic field, such as 7 T, may increase the percentage of signal enhancement and the signal to noise ratio (SNR), while most clinical machines were worked at lower magnetic field, such as 3 T or 1.5 T. To obtain enough detectable T1-image enhancements quantitatively using lower dose, modification of T1-relaxivity of the nitroxyl based contrast agents was tested. In this paper, the several nitroxyl contrast agents which have different number of spins in the molecule were chemically synthesized. The relaxivity of those nitroxyl contrast agents were measured. The suitable concentration range gave a linear relation with the image enhancement was observed for each nitroxyl contrast agent. The availability of the multi-spin nitroxyl contrast agent for MR-based redox imaging was described. The T1-relaxivity of nitroxyl contrast agents were increased depending on the number of spins on the molecule. The suitable linear relation between image enhancement and concentration of the contrast agent was obtained for a nitroxyl contrast agent in this paper.SFRBM\u27s 13th Annual Meetin

Caffeic acid phenethyl ester and caffeic acid ethyl ester activate transcription of heme oxygenase-1 gene in RAW 264.7 mouse monocyte-macrophage cell line independently of redox-regulated pathway

Activation of the transcription of HO-1 gene by elements from foods was examined in a mouse monocyte-macrophage cell line RAW264.7 by measuring relative levels of HO-1 mRNA by real-time reverse transcription-polymerase chain reaction, and it was confirmed that caffeic acid phenethyl ester (CAPE), a component of propolis, had a distinguished potential to induce HO-1 gene expression in the macrophage tumor cell. When RAW cells were incubated with 2 mM of CAPE, the level of the mRNA of HO-1 gene was enhanced significantly even at 1 h, and reached a peak at 4 h leading to an drastic increase of 25 times, then was down-regulated at 6 h. A similar induction was observed in another mouse macrophage cell line, J774.1, but not in the human hepatocarcinoma cell line HepG2, indicating that the activation is common and specific to macrophages. Caffeic acid ethyl ester (CAEE) also induced transcription of HO-1 mRNA in RAW 264.7 cells, leading to 20 times activation. However, chlorogenic acid exhibited a little activation of HO-1 as 1.3 times although it is also an ester of caffeic acid. 2 mM of curcumin and ethyl ferulate increased HO-1 mRNA by 6.4 and 7.2 times, respectively. Resveratrol also elevated HO-1 mRNA 3.4 times, though at a 66 mM. When a non-phenolic antioxidant, cysteamine, was used, 10 mM of cysteamine was required to activate HO-1 gene as 2.3 times. To investigate whether this activation is dependent upon the reductive capacity of the chemicals, the one-electron oxidation potentials of these phenolic compounds were determined by the second-harmonic alternative current voltammetry. The E0ox values of CAPE (0.48), CAEE (0.49), and chlorogenic acid (0.48) determined from the intersection of the voltammograms were almost the same. However, the value of curcumin (0.42) was smaller than those three caffeic acid esters. Thus, the reductive capacity of curcumin was the most potent, and those of CAPE, CAEE, and chlorogenic acid were almost the same and were lower than that of curcumin. However, chlorogenic acid exhibited a very little activation of HO-1 gene, and curcumin was less potent than CAPE. From these results we suggest that the transcriptional activation of HO-1 gene by CAPE and CAEE is not dependent on a redox-regulated mechanism, but on a structure-specific interaction mediated by receptors.HO Conference 2005. 4th International Congress