Radioprotective agent

 

(57) Abstract:

The invention relates to medicine, namely to radiology. The invention consists in that the radiation protection means includes vitamins E and C in doses of 10 and 20 mg/kg of the Invention provides finding medicines to protect the body from exposure to low (non-lethal) doses of radiation has a number of important practical properties, in particular the universal nature of the radioprotective effect, low toxicity, efficacy of oral application. table 2.

The invention relates to medicine, namely to radiology, and can be used for the prevention and early treatment of radiation injuries of man.

In modern conditions of low doses (non-lethal) radiation exposure become one of the main factors destabilizing the genome of mammals. The possibility of radiation injuries of the body in a single exposure at doses not exceeding 1-2 Gr, and the danger of kumulirana cytogenetic effects of radiation during prolonged and repeated (fractionated) low-dose exposure to ionizing radiation determine the when which can lead to lesions of small doses of radiation, be extremely diverse, one of the most important requirements for these funds is the universality of the latter, i.e.:

- the ability to break the cytogenetic effects of various types of low doses of radiation effects (short-term and prolonged, single and repeated);

- demonstration of the protective effect at the level of both somatic and germ cells;

the presence of both preventive and curative radioprotective effect.

Obligatory requirements to the drugs designed for wide application, are also the effectiveness of the oral route of administration and good tolerance with repeated use, suitability for industrial production.

Known substances that have a radioprotective effect at low doses of irradiation. So, serosoderjaschei radioprotectors reduce the yield of chromosome aberrations in bone marrow cells of mice irradiated at a dose of 1 Gy [1] , but the protective effect of these compounds was observed only at short-term exposure and prophylactic use of drugs.

Connection indene series have preventive antimutagenic AK is n, related to the number of quinoid compounds, partially protects germ cells from small doses of radiation [3] . But the development of indene compounds and melanin stopped at the stage of laboratory tests, and standard dosage forms of these substances have not been created yet.

Also known drug Riboxin, which reduces the number of aberrant mitoses in cells of the bone marrow, maintains a pool of precursor cells in the bone marrow and its germinative epithelium at low doses of irradiation. But Riboxin also has a kind of "radiomimetic" effect, increasing the number of chromosomal abnormalities in myelokaryocytes non-irradiated animals [4] .

Have also been studied as a means of protection from low dose exposure to drugs among vitamins - vitamin E (-tocopherol acetate) and vitamin C (ascorbic acid). But current data do not allow an unambiguous conclusion about the effects of these vitamins on the radioresistance of biological objects. Along with radioprotective effect [5, 6] in the experiments was sometimes observed lack of radioprotective action of these vitamins [7, 8] . In experiments with lethal irradiation vitamins E and C could have and rapture, including vitamins E, C and A. Oral administration of this formulation reduced the yield of chromosome aberrations in bone marrow cells of rats subjected to x-ray irradiation at a dose of 3.5 Gy [10] . This recipe is considered as a prototype.

Analysis of the formulation of the prototype showed that, first, it is multi-component and one component (vitamin A) is strongly toxic properties [11] . Secondly, the protective effect of the formulation was registered only at the level of somatic cells. Thirdly, the effect of the formulation appeared at the dose of radiation, significantly exceeding the upper limit of "small" doses of ionizing radiation and coming on the biological effect on the body to discharge minimum lethal dose. Finally, the effectiveness of the formulation in the short-term x-ray irradiation cannot serve as a proof of its efficacy in other types of radiation effects.

In view of the above, the aim of the invention was to find drugs to protect the body from low doses of radiation treatment that meets the requirements listed above (the universal character of the protective effect, the effectiveness of oral Annot the surrounding vitamins E and C in the dose of 10 mg/kg and 20 mg/kg Known application of formulations of similar composition in combination with lecithin as antimutagen when exposed to toxic substances [12] , as additional funds to the chemotherapy of tuberculosis [13] , in combination with vitamin F (linetolom) as a means to treat nervous conditions [14] . The combination of vitamins E and C (in different doses and weights) is a member of many complex vitamin preparations General therapeutic application of domestic (Dekleva, Aerovit, Undevit, Hendevit, Kvadevit etc. ) [15] and foreign (Microvit, stresstabs and others ) [16] production.

The possibility of achieving the goal of the invention is proved by the following examples.

Example 1.

The experiments were performed on 75 outbred white rats-males weighing 180-200 g total Animals were exposed to gamma irradiation on the installation IGUR at a dose of 2 Gy in the following modes:

a) Single irradiation with a dose rate of 1.51 G/min

b) Fractionated irradiation fractions of 0.25 Gy with an interval between the effects of 1 day. The dose of 0.75 Gy/min

Criterion damaging effects of radiation and the protective effect of the drugs was selected number of cells with chromosoma, from each animal was analyzed 100 metaphases.

The methodology of the study of chromosomal aberrations in cells, bone marrow

Chromosomal aberrations were studied myelokaryocytes by the method of Ford and Hamerton [17] . The basis of this method is the registration of structural damage to chromosomes, which occur at different stages of the mitotic cell cycle and in its development reach metaphase.

Preparation of drugs bone marrow was performed at 24 h after irradiation according to the method described by A. M. Malashenko, and N. Zolotareva [18] , and B. N. Orlov and co-workers [19] . To do this for 2 h before slaughter animals were injected intraperitoneally with 0.2 ml of 0.025 % solution of colchicine. Rats were scored by the method of cervical dislocation, quickly identified the tibia and washed bone marrow warmed to 37oC environment N 199 in the centrifuge tube with 10 ml of the same medium. Tubes suspension was centrifuged at 150 g for 6 minutes the Supernatant was decanted, and the precipitate was added 8 ml of warmed (to 37oC) 0,56% solution of potassium chloride. The cells thoroughly resuspendable and was placed in a thermostat at 37oC for 20 min, then was again subjected to centrifugation. The supernatant fluid is on retainer, consisting of glacial acetic acid and methanol, taken in the ratio 1: 3. The mixture was prepared before fixation and kept in the refrigerator. The tubes were closed tubes, the contents were mixed by shaking and put in the fridge for 10-15 minutes and Then the suspension is again centrifuged and add 5-6 ml of fresh fixative. This procedure was repeated 2-3 times.

Before preparing drugs, the suspension was centrifuged at 150 g for 6 minutes the Supernatant was aspirated, and to the precipitate was added 1 ml of fresh fixative and resuspendable Pasteur pipette. Then on low fat, wet and chilled slides put 8-10 drops of the suspension and the latch burned in the burner flame. The glass was dried in the air.

The color of the preparations was carried out with a solution Azur-eosin according to Romanovsky-Institute for 40 minutes the Paint was prepared immediately before use by mixing 10 ml of distilled water, 5 ml of 0.1 % solution of Azura, 2 ml of 0.1 % solution of eosin and 0.25 ml of 5% sodium bicarbonate. After this, the slides were washed in running water and dried in air.

The analysis of the drugs was performed on a microscope with immersion object (increase HH,25), which were selected by nerastas is at least 100 metaphases, only option experience 500 cells. Take into account the percentage of cells with chromosome aberrations.

The results are shown in table. 1.

Experiments have shown that individual vitamins do not have radioprotective action according to selected criteria. The same recipe used once for 1 h before irradiation and after irradiation, almost 2 times reduces the yield of chromosome aberrations in bone marrow cells after single and fractionated exposure.

Example 2.

The experiments were performed on 60 outbred mice-males weighing 18-22, the Results are presented in table. 2.

Total one-time acute irradiation was carried out with gamma rays at a dose of 2 Gy apparatus IGUR-1 two radiation sources (cesium-137) at dose rate of 1.51 G/min, and prolonged exposure (in the same dose range) - using apparatus AGO at dose rate of 0,0083 Gr/min

Fractionated irradiation was performed with fractions of 0.25 and 0.5 Gy with daily interval between them. Used machine IGUR-1 with two sources of radiation at the dose of 0.75 Gy/min

Recipe (vitamins E+C) was administered by mouth 1 hour before irradiation (when fractionated, Obluch reciprocal translocations (RT)

Under the concept of "reciprocal translocations understand mutual exchanges of areas of non-homologous chromosomes.

Cytological analysis of the frequency of RT in stem spermatogonia investigated 45-60 days after exposure (i.e. poststelle period) in the spermatocyte stage diakinesis-metaphase of the first meiotic division permanent air-dried preparations prepared according to the method proposed by Evans, Ford and co-workers [20] .

Males were scored by displacement of the cervical vertebrae. Testes were removed, freed from the shells, crushed 2.2% solution of sodium citrate. Oprawki tissues and membranes of seminiferous tubules was besieged by centrifugation for 30 s at 100 g. The supernatant was subjected to repeated centrifugation for 5 min. the Precipitate resuspendable in 1% solution of sodium citrate to hypotonic treatment within 10 minutes After the second centrifugation the cells were fixed in a mixture of methanol and acetic acid (3: 1) with the addition of 2-3 drops of chloroform. After the double shift lock 2 drops of cell suspension in fixative was applied on a glass slide and dried under a lamp. The preparations were stained with acetic acid lactoria.

Statistical processing of data was performed using criterion Student.

The experiments showed that at different variants of low dose irradiation formula (vitamins E+C) has a pronounced protective effect on the criterion of frequency of reciprocal translocations in spermatogonia irradiated mice at all variants of radiation exposure.

Used in the formula doses of vitamin E (10 mg/kg) and vitamin C (20 mg/kg) do not exceed the average therapeutic dose for a person with multiple daily oral application [6, 21, 22] .

In experiments recipes the e was well tolerated recipe and at 20-fold increase doses of its constituent vitamins, taken in a weight ratio of 1: 2.

The invention meets the criterion of "novelty", as the first recipe, made up of vitamins E and C, taken in a weight ratio of 1: 2, used to protect the body at the same time at the level of somatic and germ cells from low doses of radiation effects.

The invention meets the criterion of "inventive step", since on the basis of known information sources of information the ability to use the specified recipe as a universal therapeutic and preventive effects tools at low doses of irradiation did not seem obvious.

According to the invention, the criterion of suitability for industrial applications" proved by the examples and the fact that both components of the formulation are released by the medical industry. Biological compatibility of vitamins E and C, used orally in a specified weight ratio, shown in the clinic [14] .

Thus, the data presented indicate the achievement of the objectives of the invention.

The possibility of a substantial extension of the scope of vitamins E and C. the proposed formulation can be found Mi substances, in diagnostic and therapeutic use of ionizing radiation sources in outpatient and clinical practice, in cases of occupational low dose irradiation of people. The recipe can be used as a personnel radioprotective agent in the first aid kits and special kits designed to provide medical assistance to the population in case of emergencies.

The components of the formulation are physiologically acceptable substance, chemically and pharmacologically compatible with each other official drugs [23, 24] .

References

1. Krasil'nikov, I. I. , Antosiewicz A. E. , Deyev S. P. , A. Myasoedov F. a Method of preventing biological effects of radiation due to small (sub-lethal) doses of radiation. Proc. Dokl. Vsesojuz. proc. "Radiobiological consequences of the accident at the Chernobyl NPP", 30 Oct. -1 November 1991 , Minsk, 1991, S. 63.

2. Mosse I. B. the Problem of chemical protection in radiation genetics. Minsk. Science and technology, 1974. - 152 C.

3. Mosse I. B. , I. Lyakh P. , S. Plotnikov I. , Kostrov L. N. To the question of the prevention of late effects of irradiation. Proc. Dokl. IV vsesojuz. proc. "Chemistry, pharmacology and mechanismsschemes medical radiation protection of participants of liquidation of consequences of large-scale radiation accidents. In Proc. : Military medicine. Problems of prevention, diagnosis, treatment of extreme conditions. M : VI, 1994, S. 76-85.

5. Tiunov L. A. , Vasiliev, A. , Wallenstein E. A. Radioprotective funds. M.-L. : Nauka, 1964, S. 38-40.

6. Alekseenko K. K. Effect of vitamin C on the peripheral blood during radiation therapy for breast cancer. Proc. Dokl. IX vsesojuz. scient. proc. Restorative and compensatory processes by radiation injuries", of L. , Oct. 1986 L. , 1986, S. 166-167.

7. Umegaki K. , Ikon T. , Ichikawa T. Effect of vitamin E on chromosomal damage in bone marrow cells of mice having received low dose of x-ray irradiation/Int. J. Vitamin and Nutrition Research, 1994, v. 64, No. 4, p. 249-252.

8. Eldor A. , Vlodavsky I. , E. Riklis , Fuks Z. Recovery of prostacyclin capacity of irradiated endothelial cells and the protective effects of vitamin C/Prostaglandins, 1987, v. 34, N 2, p. 241-255.

9. Srinivasan V. , Weiss J. F. Radioprotection by vitamin E: injectable vitamin E administered alone or with WR-3689 enhances survival of irradiated mice/Int. J. Radiat. Oncology. Biol. , Phys. , 1992, v. 23, N 4, p. 841-845.

10. Aliyev A. A. Antimutagenic properties of some vitamins in terms of induction of chromosome aberrations by ionizing radiation. VINITI, 15.07.83, N 3969 - Dup 83 (CIT. by RJ Radiobiology, 1983, N 10, 10.70.56 DEPT, S. 9).

11. Mashkovsky M. D. Retinol. Vitamin A. Medicines. T. 2. Kharkiv, Torching", 1997, S. 70-72.

12. Minnunni>13. Shcherbakova, I. C. Clinical value of the system state of free radical oxidation and antioxidant defense in patients with infiltrative pulmonary tuberculosis//Collection of abstracts of research and development, Vol. 8, h 1, 1993, N 1-2, S. 204.

14. Kovalev, Y. C. , Kondakov B. C. About the combined use of antioxidants in clinic of neurosises//bul. the experts. Biol. and the honey. , 1994, T. 117, No. 2, S. 209.

15. Mashkovsky M. D. Medicines. T. 2. Kharkiv, Torching", 1997, 590 S.

16. On The Main Page. Drugs in Russia. M , Attraversare, 1995, 1168 S.

17. Ford, C. E. , J. L. Hamerton Chromosomes of five rodent species //Nature. - 1956. - Vol. 31. - P. 247-254.

18. Malashenko A. M. determination of the mutagenicity of chemical compounds (genetic screening) in laboratory mice (meth. instructions). M. , 1977, 12 S.

19. Orlov, B. N. , G. A. Chudnovsky, E. P. Kryukov. The study of chromosome sets mammals. Met. the leadership. M. : Nauka, 1976, 15 S.

20. Evans, E. P. , Brecon G. , Ford, C. C. An air drying method for meiotic preparations from mammalian tests. -Cytogenetics, 1964. vol. 2, p. 289-294.

21. Andrusenko M. P. , Wild, C. I. , Zharkova N. B. , Kostrikina, I. E. the Use of antioxidants for the correction of adverse musculoskeletal effects of pharmacotherapy, Byull. the experts. Biol. and the honey. , 1994, so hasinai therapy in elderly patients//bul. the experts. Biol. and the honey. , 1994, T. 117, No. 2, S. 224.

23. Mashkovsky M. D. Tocopherol acetate. Medicinal product. So 2, S. 95-96.

24. Mashkovsky M. D. ascorbic Acid. Vitamin C. Medicines. So 2, S. 88-89.

Therapeutic and prophylactic to protect the body from low doses of radiation effects, including vitamin E in an amount of 10 mg/kg) and vitamin C (20 mg/kg

 

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