Saratov JOURNAL of Medical and Scientific Research

Osipov A.N.

State Scientific Research Center n.a. A.I. Burnasyan — Burnasyan Federal Medical Biophysical Center of Federal Medical Biological Agency, head of laboratory, Doctor of Biological Sciences

Effectiveness of autologous cell products derived from adipose tissue for the treatment of severe local radiation injuries

Year: 2014, volume 10 Issue: №4 Pages: 838-844
Heading: radiation medicine Article type: Original article
Authors: Eremin P.S., Pigaleva N.A., Murzabekov M.B., Lebedev V.G., Lazareva N.L., Eremin I.I., Pulin A.A., Osipov A.N., Bushmanov A.Yu., Kotenko K.V.
Organization: State Scientific Research Center n.a. A.I. Burnasyan — Federal Medical Biophysical Center of Federal Medical Biological Agency
Summary:

Aim: to investigate the dynamics of ulcer surface healing depending on type of radiation ulcer and cell product in murine experimental model of severe local radiation injuries after exposure to X-rays. Material and Methods. Mature Wistar rats were used for experiments (males, weight 180-200 grams). Standard model of severe local radiation injuries was used: X-ray irradiation of animals on a modified RAP100-10 device. Isolation of the stromal-vascular fraction carried out by enzymatic treatment of adipose tissue. Autologous cells transplantation performed on day 20 after irradiation in the case of acute radiation injury model (early radiation ulcers) and day 120 in the case of long-term effects of radiation injury model (chronic radiation ulcers). As a positive control allogeneic MMSC derived from rat bone marrow were used. Results. Application of autologous cell products derived from adipose tissue in animal model of severe local radiation injuries was investigated for the first time. It was shown that usage of stromal vascular fraction of adipose tissue for the treatment of early radiation injuries not only leads to better improvement (as compared with the use of MMSCs), but also pronounced therapeutic effect could be observed at an earlier time. On the model of long-term effects of radiation injuries therapeutic effect was observed for all cell products. Conclusion. Cell-based products derived from adipose tissue are promising material for future research and clinical application for treatment of local radiation injuriesK) Described method allows choosing treatment strategy basing on patient's individual characteristics and the type of local radiation injuries.

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Characteristics of changes in the number of yH2AX and Rad51 protein foci in human skin fibroblasts after prolonged exposure to low-dose rate X-ray radiation

Year: 2014, volume 10 Issue: №4 Pages: 739-743
Heading: Genetics Article type: Original article
Authors: Ozerov I.V., Eremin P.S., Osipov A.N., Eremin I.I., Tsvetkova A.D., Guseva S.S., Ivanova K.Yu., Gavrilenko 0.I., Pustovalova M.V., Smetanina N.M., Grekhova A.K., Lazareva N.L., Pullin A.A., Maksimova О.A., Gordeev A.V., Bushmanov A.Yu., Kotenko K.V.
Organization: Lomonosov Moscow State University, State Scientific Research Center n.a. A.I. Burnasyan — Federal Medical Biophysical Center of Federal Medical Biological Agency
Summary:

Aim: to compare the repair process of DNA double-strand breaks in mammalian cells after acute versus prolonged exposure to X-ray irradiation with different dose rates. Material and methods. Studies were performed on primary human fibroblasts isolated from skin biopsies of healthy volunteers (women, 29 and 30 years). Cells were irradiated using an X-ray machine RUB RUST-M1 (JSC "Ruselectronics", Moscow, Russia) at 37°C temperature with a dose rate of 400 mGy/min (200 kV, 2*2.4 mA, a filter of 1.5mm AI) or 4 mGy/min (50 kV, 2*0.4 mA, a filter of 1.5 mm AI). Immuno-cytochemical protein staining was utilized for yH2AX and Rad51 foci analysis. Results. Phosphorylated histone H2AX (yH2AX) and the key protein of homologous recombination Rad51 foci formation and disappearance kinetics were investigated simultaneously in primary human dermal fibroblasts after acute and prolonged exposure to X-ray radiation at a same dose. It was shown that the relative yield of yH2AX foci per dose reduces with decrease in dose rate, while the relative yield of Rad51 foci conversely increases. Conclusion. Our findings suggest the fundamental differences in the ratio of non-homologous end joining and homologous recombination DNA repair in acute versus prolonged irradiated cells.

Effect of incubation in sodium chloride hypertonic solutions on y-radiation and hydrogen peroxide induced DNA damage in human peripheral blood lymphocytes

Year: 2014, volume 10 Issue: №4 Pages: 735-739
Heading: Genetics Article type: Original article
Authors: Vorobyeva N.Yu., Arkhangelskaya E.Yu., Bushmanov A.Yu., Osipov A.N.
Organization: State Scientific Research Center n.a. A.I. Burnasyan — Federal Medical Biophysical Center of Federal Medical Biological Agency
Summary:

Aim: to study the effect incubation in NaCI hypertonic solutions on the yield of DNA single-strand breaks (SSB) and alkali-labile sites (ALS) induced by y-radiation and hydrogen peroxide in human peripheral blood lymphocytes in vitro. Material and methods. The lymphocytes isolated from the peripheral blood of healthy women donors aged 21-26 years were used for the study. Cell irradiation at doses of 1, 2, 3 and 5 Gy was performed using the "AGAT P1" unit (Russia, y-radiation source — 60Co, a dose rate of 1 Gy/min) at 4°C. In case of hydrogen peroxide exposure (25, 50 and 100 uM) the cells were incubated for 15 minutes at 4°C. Analysis of the DNA SSB and ALS was performed using the alkaline DNA comet assay. Results. Incubation of cells in 0.35 M NaCI solution (non-histone proteins dissociation) increases the yield of DNA SSB and ALS induced by v-radiation and hydrogen peroxide at ~ 1.3 times. Increasing the NaCI concentration to 0.6 M (linker histone H1 dissociation) leading to almost double increase in the DNA damage yield (~ 1.7-1.8 times). Further increasing the NaCI concentration leading to disruption of the structure of nucleosomes (1.2 M) and almost complete chromatin deproteinization (2 M) increased the yield of DNA damage not such pronounced as expected (~ 2.2-fold at 2 M concentration). Conclusion. The results showed that incubation in NaCI hypertonic solutions significantly increases the sensitivity of lymphocytes to DNA damaging action of free radicals.

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Comparative study of the amount of DNA single-strand breaks and alkali-labile sites in human peripheral blood lymphocytes exposed to 365 nm UV versus X-ray radiation

Year: 2013, volume 9 Issue: №4 Pages: 791-795
Heading: Genetics Article type: Original article
Authors: Smetanina N.M., Pustovalova М.V., Bushmanov A.Yu., Osipov A.N.
Organization: State Scientific Research Center n.a. A.I. Burnasyan — Federal Medical Biophysical Center of Federal Medical Biological Agency
Summary:

Aim: to compare the amount of DNA single-strand breaks (SSB) and alkali-labile sites (ALS) induced by DNA 365 nm ultraviolet (UV) versus X-rays radiation in human peripheral blood lymphocytes at in vitro conditions. Material and methods. Blood lymphocytes were isolated in a Ficoll-verografin density gradient. Irradiation of cells was performed using BLX-365 UV-unit (wavelength 365 ± 10nm)and RUSTM1 biological X-ray unit. Analysis of DNA SSB and ALS was carried out using comet assay at alkaline conditions. Results. For the first time it was demonstrated that the amount of SSB and ALS induced by 365 nm UV radiation at a dose of 10 kJ/m2 in human lymphocytes corresponds to the effect of X-rays at a dose of- 1 Gy. Conclusion. Exposure of human peripheral blood lymphocytes to 365 nm UV radiation at a dose of 1 kJ/m2 induces - 100 DNA SSB per cell.

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Induction and reparation of double-strand DNA breaks in V79 cells continuously exposed to low dose-rate Y-radiation

Year: 2013, volume 9 Issue: №4 Pages: 787-791
Heading: Genetics Article type: Original article
Authors: Ozerov I.V., Bushmanov A.Yu., Anchishkina N.A., Guryev D.V., Pustovalova M.V., Smetanina N.M., Arkhangelskaya E.Yu., Vorobyova N.Y., Osipov A.N.
Organization: State Scientific Research Center n.a. A.I. Burnasyan — Federal Medical Biophysical Center of Federal Medical Biological Agency
Summary:

Aim: to study the patterns of changes in the number of DNA double-strand breaks (DSB) in mammalian cells continuously exposed to low dose-rate y- radiation. Material and methods. Chinese hamster lung fibroblasts (V79) were used in this study. The y- irradiation of cells at a dose rate of 0.1 mGy/min was performed using the «Gamma-Panorama» unit (Cs-137). The fluorescence immunoassay of the phosphorylated H2AX-histone (y-H2AX) foci was used to investigate the DNA DSBs formation. Frequency of apoptotic cells was evaluated using «DNA halo» assay. 5 (6) — chloromethyl-2,7-dichlorodihydrofluorescein diacetate was used to estimate the reactive oxygen species (ROS) production. Results, it was showed that continuous low dose-rate irradiation of Chinese hamster V79 cells induces an increase of the y-H2AX foci number and ROS production rate at the early stages of exposure time (6-24 h, doses 3.6-14.4 cGy), while increasing exposition time and, therefore, the radiation dose (48-72 h, 28.8-43.2 cGy) caused a decrease in these endpoints to almost the control level. There was observed no significant changes in the frequency of apoptotic cells. Conclusion. It is assumed that the processes causing the DSB amount changes in mammalian cells continuously exposed to low dose-rate y-radiation are associated with the development of oxidative stress and subsequent activation of cellular antioxidant defense systems.

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