CN113029918A - Application of Ru360 in enhancing sensitivity of mouse embryonic fibroblasts to 1800MHz radio frequency electromagnetic field - Google Patents
Application of Ru360 in enhancing sensitivity of mouse embryonic fibroblasts to 1800MHz radio frequency electromagnetic field Download PDFInfo
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Abstract
The invention discloses application of Ru360 in enhancing sensitivity of mouse embryonic fibroblasts to a radio frequency electromagnetic field. The Ru360 is an oxygen-bridged binuclear ruthenium amine complex, is a mitochondrial calcium ion pump inhibitor, and experiments show that the 3 mu M Ru360 can remarkably enhance the DNA damage and cell activity inhibition effect of a 1800MHz radio frequency electromagnetic field on mouse embryonic fibroblasts, can remarkably enhance the sensitivity of cells to radio frequency electromagnetic radiation, and has the application prospects of radio frequency electromagnetic field sensitization and establishment of a radio frequency electromagnetic field sensitive cell model.
Description
Technical Field
The invention belongs to the field of biomedicine, and particularly relates to application of a mitochondrial calcium ion pump inhibitor Ru360 in enhancing the sensitivity of mouse embryonic fibroblasts to 1800MHz radio frequency electromagnetic field.
Background
With the development of scientific technology, more and more mobile phones are used, and people worry about the influence of mobile phone radiation on body health. At present, whether the radiation of the mobile phone is carcinogenic is one of the most concerned problems of the public, and whether the radiation of the mobile phone can cause the DNA in the cell to be damaged is important research content. Although there are many studies related to the present, the effect is not strong enough due to the low energy of the radio frequency electromagnetic field, which leads to inconsistent results between different studies. Therefore, finding a cell model sensitive to radio frequency electromagnetic fields can accelerate research progress in this field. However, no sensitive cell model has been found or reported.
Ru360 is an oxygen-bridged binuclear ruthenium amine complex, a selective mitochondrial calcium uptake inhibitor. At present, the effect of Ru360 in enhancing the sensitivity of cells to electromagnetic fields has not been reported.
The chemical structure of Ru360 is:
disclosure of Invention
In order to solve the technical problem, the invention provides application of a mitochondrial calcium ion pump inhibitor Ru360 in enhancing the sensitivity of cells to radio frequency electromagnetic fields.
The technical scheme adopted by the invention is as follows: application of mitochondrial calcium ion uptake inhibitor Ru360 in enhancing sensitivity of mouse embryonic fibroblasts to 1800MHz radio frequency electromagnetic field.
Further, the concentration of Ru360 in the culture medium was 3. mu.M.
Further, the Ru360 is commercially available high-purity Ru360, and the purity is more than or equal to 97%.
Furthermore, the invention also provides the DNA damage effect of the Ru 360-enhanced 1800MHz radio frequency electromagnetic field on mouse embryo fibroblast and the inhibition effect on cell viability.
The invention provides application of a mitochondrial calcium ion pump inhibitor Ru360 in enhancing the sensitivity of cells to radio frequency electromagnetic fields, provides a method for enhancing the radiation sensitivity of cells to radio frequency electromagnetic fields for cell induction electromagnetic radiation mechanisms, related researches and related fields, and has important economic and scientific research values.
Drawings
FIG. 1 is a diagram of a basic comet assay for detecting DNA damage in mouse embryonic fibroblasts, wherein,
pseudo exposure of 1-4.0W/kg 1800MHz RF electromagnetic field for 15 min,
exposing the 1800MHz radio frequency electromagnetic field of 2-4.0W/kg for 15 min,
3-15 minutes ahead of time Ru360 treatment +4.0W/kg 1800MHz RF electromagnetic field pseudo exposure for 15 minutes,
4-15 minutes ahead of time Ru360 treatment +4.0W/kg 1800MHz RF electromagnetic field exposure for 15 minutes.
FIG. 2 is a statistical histogram of changes in the level of DNA damage in mouse embryonic fibroblasts, wherein,
pseudo exposure of 1-4.0W/kg 1800MHz RF electromagnetic field for 15 min,
exposing the 1800MHz radio frequency electromagnetic field of 2-4.0W/kg for 15 min,
3-15 minutes ahead of time Ru360 treatment +4.0W/kg 1800MHz RF electromagnetic field pseudo exposure for 15 minutes,
4-15 minutes ahead of time Ru360 treatment +4.0W/kg 1800MHz RF electromagnetic field exposure for 15 minutes,
**P<0.01,***P<0.001。
fig. 3 cell viability was measured 1 hour after the end of the rf electromagnetic field exposure, wherein,
pseudo exposure of 1-4.0W/kg 1800MHz RF electromagnetic field for 15 min,
exposing the 1800MHz radio frequency electromagnetic field of 2-4.0W/kg for 15 min,
3-15 minutes ahead of time Ru360 treatment +4.0W/kg 1800MHz RF electromagnetic field pseudo exposure for 15 minutes,
4-15 minutes ahead of time Ru360 treatment +4.0W/kg 1800MHz RF electromagnetic field exposure for 15 minutes,
*P<0.01。
Detailed Description
The present invention will be described in further detail with reference to examples.
1. The material method comprises the following steps:
1) reagent α MEM culture medium (Hyclone, china); fetal bovine serum (FBS, hangzhou ilex bioengineering materials ltd, china); 0.25 wt.% pancreatin (containing 0.02wt. edta, Gibco, usa); cell Counting Kit-8(CCK8, Japan Dojindo, Japan); ru360(Sigma, China).
2) The instrument comprises the following steps: a radio frequency electromagnetic field irradiation system (sXc1800, IT' IS Foundation, Switzerland); multifunctional microplate readers (Spark, Tecan, switzerland); biosafety cabinets (HFsafe-1200LC, Heal Force, China); carbon dioxide cell incubator (Forma Steri-Cycle, the mo scientific, china); a comet electrophoresis tank (JY-SC10, Beijing Junyi, China); electrophoresis power supply (Bio-Rad, USA).
3) Culture of mouse embryonic fibroblasts were cultured in alpha MEM containing 10% fetal bovine serum at 37 ℃ and 5% CO2The solution was changed every two days in the environment of (1). And digesting and passaging when the cell growth density reaches 80% -90%.
4) Inoculating mouse embryo fibroblast to obtain well-grown cells, digesting to obtain cell suspension, and culturing at a ratio of 1 × 104The density of the culture dish was measured in a 35mm cell culture dish, and Ru360 (final concentration: 3. mu.M) was added after 12 hours of ordinary culture. After 15 minutes, the cells were placed in an electromagnetic radiation device and exposed to a 4.0W/kg1800MHz radio frequency electromagnetic field for 15 minutes.
5) Alkaline comet assay for detecting DNA damage, washing a single-sided frosted glass slide with double distilled water, drying, spreading a layer of 0.65% agarose gel on the frosted surface, flattening with a cover glass, and placing on ice for accelerating coagulation. The cells were digested with 0.25% trypsin, and the culture medium was added to make a cell suspension, which was placed on ice for future use. mu.L of the cell suspension was mixed with 75. mu.L of 0.65% low-melting agarose, spread on the first agarose gel, flattened with a coverslip, and placed on ice to promote coagulation. The coverslip was removed and the slide was immersed in a pre-cooled alkaline cell lysis solution (2.5M NaCl, 1% sodium N-lauroyl sarcosinate, 100mM disodium EDTA, 10mM Tris base, pH 10, 1% Triton X-100) and lysed at 4 ℃ for 1 hour. Subsequently, the cells were immersed in an enzymatic hydrolysate (0.5mg/mL DNase-free protease K, 2.5M NaCl, 1% sodium N-lauryl sarcosinate, 100mM disodium EDTA, 10mM Tris base, pH 10, 1% Triton X-100) and subjected to enzymatic hydrolysis at 37 ℃ for 2 hours. Before electrophoresis, the gel was uncoiled in a pre-cooled electrophoresis solution (300mM NaOH, 0.1% 8-hydroxyquinoline, 2% DMSO, 10mM tetrasodium EDTA, pH 13) at 4 ℃ for 20 minutes; and (4) electrophoresis. 300mA, 20V (0.4V/cm), and electrophoresis at 4 ℃ for 20 minutes. After the completion of electrophoresis, the Gel was neutralized for 5 minutes in a neutralization buffer (0.4M Tris, pH 7.5), stained with Gel red (1 μ g/mL), and sectioned, photographed by a fluorescent microscope and analyzed by CASP software.
6) Statistical analysis the experimental data are expressed in' X + -SD and statistically analyzed using Microsoft Excel software, and the comparison between the two groups was performed using t-test, and it was considered statistically different when P < 0.05.
2. As a result:
3 mu M Ru360 is added in advance for treating mouse embryo fibroblasts 15 minutes, after 1800MHz 4.0W/kg radio frequency electromagnetic field is continuously exposed for 15 minutes, the DNA damage level in the cells is detected through an alkaline comet assay, and the Ru360 can remarkably enhance the DNA damage of the radio frequency electromagnetic field to the mouse embryo fibroblasts (shown in a figure 1-2); meanwhile, by detecting the cell viability by the CCK8 method, the Ru360 can obviously enhance the cell viability inhibition effect of the radio frequency electromagnetic field on the mouse embryo fibroblast (shown in figure 3), and has obvious radio frequency electromagnetic radiation sensitization effect.
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are included in the scope of the present invention.
Claims (4)
- Use of Ru360 for enhancing the sensitivity of mouse embryonic fibroblasts to 1800MHz radio frequency electromagnetic fields.
- 2. The use according to claim 1, wherein the Ru360 is present in the culture medium at a concentration of 3 μ M.
- 3. Use according to claim 1, wherein Ru360 is used for enhancing the sensitivity of mouse embryonic fibroblasts to 1800MHz radio frequency electromagnetic radiation.
- 4. The use according to claim 1, wherein Ru360 is used for enhancing the DNA damaging effect of 1800MHz radio frequency electromagnetic field on mouse embryonic fibroblasts and the inhibitory effect on cell viability.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20170105963A1 (en) * | 2015-10-16 | 2017-04-20 | The Board Of Trustees Of The University Of Illinois | Small molecules that induce intrinsic pathways apoptosis |
| US20180291347A1 (en) * | 2010-10-08 | 2018-10-11 | Mesoblast International Sarl | Enhanced msc preparation |
-
2021
- 2021-03-10 CN CN202110258468.0A patent/CN113029918A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20180291347A1 (en) * | 2010-10-08 | 2018-10-11 | Mesoblast International Sarl | Enhanced msc preparation |
| US20170105963A1 (en) * | 2015-10-16 | 2017-04-20 | The Board Of Trustees Of The University Of Illinois | Small molecules that induce intrinsic pathways apoptosis |
Non-Patent Citations (4)
| Title |
|---|
| G J GARCÍA-RIVAS ET AL: "Ru360, a specific mitochondrial calcium uptake inhibitor, improves cardiac post-ischaemic functional recovery in rats in vivo", 《BRITISH JOURNAL OF PHARMACOLOGY》, vol. 149, no. 7, 9 October 2006 (2006-10-09) * |
| 丁国莲 等: "极低频电磁场对生殖细胞的影响及其机制研究的进展", 《国外医学（计划生育/生殖健康分册）》, vol. 26, no. 1, 31 December 2007 (2007-12-31), pages 12 * |
| 孙川: "电磁场对不同遗传背景细胞基因组稳定性的影响研究", 《中国博士学位论文全文数据库 医药卫生科技辑》, no. 06, 15 June 2017 (2017-06-15), pages 19 - 69 * |
| 赵文旭 等: "维拉帕米对HeLa细胞12C6+离子辐射敏感性的影响", 《医药卫生科技》, vol. 19, no. 3, 31 December 2012 (2012-12-31) * |
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