Cerebroprotective agent

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to medicine, namely pharmacology and may be used for cerebral protection in treating cerebral affections. That is ensured by using Zongorine as a cerebroprotective agent.

EFFECT: agent provides the cerebroprotective action and prevents the death in animals suffering experimental post-hypoxic encephalopathy due to the activation of cerebral neutral stem cells.

2 ex, 3 tbl

 

The invention relates to medicine, specifically to pharmacology, neurology and cell technology.

Many cerebroprotective means and methods of treatment and prevention of brain lesions [1].

The disadvantage of these tools is often low efficiency [2].

Problem solved by the present invention is an expanding Arsenal of highly effective cerebroprotective funds.

This object is achieved by the use of songaria as cerebroprotective tools.

New in the present invention is the use as cerebroprotective means zongoene.

Used the original tool Zagorin was developed and received NII pharmacology" SB RAMS (Tomsk) in conjunction with the National research Irkutsk State Technical University (Irkutsk, Russia) and was a 0,0005% aqueous solution of this alkaloid. Tangorin were extracted from plant material (plants of the family Ranunculaceae) in the form of free bases by standard extraction method [3].

Today cerebroprotective with the purpose of treatment and prevention of the development of the brain use a wide range of drug and non-drug therapies [1, 2]. While pharmacological dei is a journey of existing cerebroprotective, is primarily to protect or stimulate preserved cellular elements. However, this concept of pharmacological intervention in some cases is absolutely untenable. Available cerebroprotective are not often able not only to fully restore the morphological and functional state of the organ, but also to prevent the development of progredient course the nature of the pathologic process [1].

In this regard, of course, important to develop a brand new cerebroprotective funds with new mechanisms of action.

Received in recent years, information about the properties and laws of life multipotent precursor cells of the adult organism has opened the possibility of the development of a new direction in the treatment of many diseases using stem cell therapy. The most physiological and promising approach to solving problems of regenerative medicine is a pharmacological strategy for cell therapy based on the principle of stimulation of endogenous stem cells (SC) by imitation activities of natural regulatory systems functioning in the body [4].

In NII pharmacology" SB RAMS shown the principal possibility and high efficiency farmacologicas the OI strategies of cell therapy in various disease models including and pathological conditions of the CNS [4]. At the same time using the method of thin-layer chromatography of us in the alkaloid fraction of plants in the family Ranunculaceae was established content zingarina (C22H33NO3),

and revealed a pronounced stimulating effect in respect of stromal progenitor cells of the skin and the healing process of skin wounds using externally [5].

The influence of songaria on the processes of regeneration of the damaged CNS pathological process, and the possibility of cerebroprotective due to the activation of the regeneration mechanisms associated with the stem cells (SC) brain, when applied in vivo has not been studied. The experiment showed unexpected results.

The fact of the use zingarina with achieving a new technical result consists in obtaining expressed cerebroprotective effects for the specialist is not obvious,

The new properties are not derived explicitly from the prior art in this field and is not found in the patent and scientific literature.

The present invention can be used in medicine.

Based on the above, you should consider the claimed solution meets the criteria of "Novelty", "Inventive step", "Industrial primenimo the ü".

The experiments were carried out at 68 outbred mice. Animals obtained from the nursery of the Department of experimental biomedical modeling NII pharmacology" SB RAMS.

Studies were performed in accordance with good laboratory practice (GLP), the Order of healthcare of the Russian Federation No. 708n from 23.08.2010 "On approval of rules of good laboratory practice", the Federal Law of April 12, 2010№61-FZ "On circulation of medicines", "guidelines for preclinical testing of drugs" (Moscow, 2012),

Example 1

The proposed tool was obtained from the herb Aconite Baikal and iokasti high (plants of the family Ranunculaceae). The aboveground part of the plants collected during the flowering period in the Irkutsk region, were ground to a particle size less than 5 mm, were treated with sodium carbonate solution and was subjected to continuous extraction with chloroform for 5 days. The chloroform extract was evaporated to small volume and carefully was extracted with 5% sulfuric acid. The acid extract was podslushivaet with sodium carbonate to pH 9-10 and consistently were extracted first with ether and then with chloroform. The ether extract was evaporated to dryness, dissolved in a small amount of ether and was chromatographically on deactivated alumina in the system hexane-acetone (90→50%). Ester-soluble fraction was subjected to on abnoy extraction buffer solutions with increasing pH values. The ether solution remaining after extraction of the most alkaline buffer was evaporated to dryness and chromatographically on aluminium oxide in the system hexane-methanol to elution Sonoline occurring after the release natalina. The substance was dissolved in distilled water to a final concentration of 0.0005%.

Example 2

Pharmacological properties of the tools studied on the model of hypoxic lesions of the Central nervous system. Hypoxic exposure was a hypoxia harmonyum and was modeled using thermocamera [6]. Thermocamera was a glass vessel with a balanced volume of 500 ml (±<1.0%) and an airtight lid. Animals were placed in thermocamera, then its cap tightly closed. Mouse was there until the death of convulsive seizure or first agonal breaths, after which they were removed and given to razdelatsya". To obtain severe encephalopathy hypoxic exposure was simulated twice with an interval of 48 hours. When using thermocamera 500 ml in animals immediately after hypoxic injury occurs quickly (within 2-3 hours) recovery of all functions. Functional disorders of the Central nervous system caused by hypoxic exposure, was estimated by the change in orientation / exploratory behavior on the indoor field and security of passive avoidance reaction [7].

When this formulation techniques within 10-15 days after the last hypoxic mice subjected to its influence, is almost indistinguishable from normal animals, however, since 14-21 days they begin to manifest violation of mnestic activity, and by the end of the fourth week, there is partial destruction. Assessment of animals for safety passive avoidance reaction was performed at 48 hours, 7, 14, 21 and 28 days after running out of reflex. The death of animals was considered to 35 days of the experiment. The results were evaluated based on the proportion of animals with preserved reflex and the proportion of animals killed by the end of the fifth week after hypoxic injury. Orienting-exploratory behavior in the open field and the production of passive avoidance reaction was carried out 1 hour after re-hypoxic injury.

The experimental set-up the "open field" was a camera the size of 40×40×20 cm with a square floor and white walls [7]. Its floor, is divided into 16 squares, had in each square round hole with a diameter of 3 see the Top of the chamber was covered by an electric incandescent lamp of 100 watts, which is located at a height of 1 m from the floor. The mouse was placed in one of its corners and within 3 minutes, separately for the first and subsequent two minutes, recorded the number of moves from square to square (horizontal activity), the number is in stevani on the hind feet (vertical activity), the number of surveys holes (mink reflex), the number of washings (grooming) and the number of acts of defecation by the number of fecal balls (boles) and calculated the asymmetry factor behavior as the ratio of the amount of horizontal displacement to total physical activity, expressed in percent. The results of the first and subsequent two minutes of the test were evaluated separately and in total.

The technique of passive avoidance reaction based on the suppression of innate reflex preference dark space available in rodents [7]. The experimental setup consisted of a camera, consisting of two compartments: a large - lit and small - dark. The animal was placed in the bright compartment and soon (10-20 seconds), due to innate reflex preference dark space, passed into the small compartment, after which the door connecting the two compartments, overlap and on the floor of the dark compartment, consisting of parallel stripe electrodes, the applied electric current pulses with a duration of 50 Me, with a frequency of 5 Hz and an amplitude of 50 mA. After 10 seconds the door was opened and the animal was able to jump out in the bright compartment with a conventional floor. In the described procedure, the animals developed a conditioned reflex avoidance dark space. When checking the reproducibility of reflex animals were placed in a light the compartment in the corner opposite from the entrance to the dark compartment and watched for 3 minutes. We registered the time of the first entry into the dark compartment (latent time of call), the total time spent in the dark compartment. Developed the reflex was considered, if for all 3 minutes of observing the animal never visited dark Bay or latent time of sunset exceed 150 C. About the quality of the reflex was assessed by the proportion of animals with the presence of reflex. Additional indicators of the conditioned-reflex activity and behavioral status were the number of bowel movements, grooming, a number of surveys of the entrance to the dark compartment, the time spent in the light compartment, the number of entries into the dark compartment and the time spent in the dark compartment. Animals that after placing in the bright compartment remained motionless, and did not come close to the entrance to the dark compartment, the calculation results were not taken into account.

To study mechanisms of drug action by using the method kleinova of in vitro studied its effect on neural SC paraventrikulyarnoe brain areas of animals undergoing hypoxia [8].

To study cerebroprotective properties, the proposed tool was administered at a dose of 0.2 ml orally in the form of 0.0005% - solution of two schemes: 1) daily, starting 4 days before hypoxic exposure and 5 times for 1 h before hypoxia; 2) 1 time per day for 5 days after modeling pathological status is I. These schemes allow to evaluate cerebroprotective, antihypoxic and nootropic effect of pharmacological substances [9].

As the comparison drug used G-CSF (Neupogen, "Hoffman-1a Roche, Switzerland), which was administered subcutaneously at a dose of 125 mg/kg for similar schemes. This tool is the closest to the technical essence of the achieved result. It has a pronounced cerebroprotective action defined by the stimulation of regeneration mechanisms "deep pool"associated with SC [8], however, is quite toxic [4], which makes it unlikely that its use in effective modes in the clinic according to testimony.

Studies have shown (table 1, 2) that migrated hypoxia induced a decrease in orienting-exploratory behavior in the open field with a shift of asymmetry factor behavior. In the hypoxic group control noted an increase in latency time in a dark chamber for the generation of reflex that indicates a breach of the indicative reflex. The use of the proposed drug for both schemes had led to the normalization of the orienting-exploratory behavior in the open field, while G-CSF was effective only if 2 the scheme of injection (table 1).

Manifestations of posthypoxic encephalopathia animals hypoxic control at check-mnestic functions (passive avoidance reaction) began with 14 days, progressive increases to 28 days in this group mentioned the death of 65% of the animals. Application zingarina had a protective effect on animals. Had increased levels of reproducibility reflex and prevent death. The use of the reference preparation was accompanied by the development of therapeutic effects of its introduction after modeling pathological conditions (table 2)

Investigation of the mechanisms cerebroprotective this means revealed its significant stimulating effect in regional neural precursor cells paraventrikulyarnoe brain areas that were manifested in a significant increase in their number (table 3).

The results indicate a pronounced cerebroprotective activity songaria, coupled with activation of the realization of the growth potential of resident neural SC brain.

Table 1
Influence zingarina in comparison with G-CSF indices of orienting-exploratory behavior in offspring of male mice in the open field after suffering hypoxic injury
Group observations doseTotal motor activityHorizontal activityVertical activityMink reflexGroomingDefecationThe asymmetry factor
Control intact65,9±7,4*32,3±3,8*4,2±1,224,1±2,81,0±0,31,5±0,2*51±4*
Control hypoxic35,7±3,227,5±2,81,0±0,65,7±1,41,2±0,50,2±0,173±4
G-CSF under the first scheme37,8±4,328,4±1.90,9±0,36,2±1,10,4±0,20,1±0,171±5
G-CSF according to the second schema61,7±6,3*29,7±4.2* 3,5±1,3*19,7±3,8*0,9±0,40,3±0,353±3*
The inventive tool under the first scheme60,4±4,3*33,1±2,4*4,3±0,9*22,6±1,17*0,9±0,20,4±0,252±4*
The inventive tool according to the second schema64,3±4,2*31,6±3,4*4,2±0,5*24,7±0,6*1,0±0,41,1±0,3*51±3*
* - the difference is significant in comparison with hypoxic control at p<0,05

Table 2
Influence zingarina in comparison with G-CSF on the safety of the conditioned reflex of passive avoidance offspring of male mice after suffering hypoxic injury
Group observations doseShare zhivotnyh preserved reflex when checking through (in %) The proportion of animals that died by the end of 35 days, in %
48 hours7 days14 days21 days28 days
Control intact100*100*100*92100*0*
Control hypoxic838333171765
G-CSF under the first scheme808033181850*
G-CSF according to the second schema100*100*83*92*90*0*
The inventive tool under the first scheme100*100 100*100*100*0*
The inventive tool according to the second schema100*100100 *100100*0*
* - the difference is significant in comparison with hypoxic control at p<0,05

Table 3
Influence zingarina in comparison with G-CSF on the dynamics of the content of neural stem cells (NSC) in paraventrikulyarnoe brain areas outbred mice after modeling encephalopathy
The timeframe of the study dayThe NSC, 5×104the nuclears
5thControl intactby 5.87±0,39
Control hypoxic6,3±0,62
G-CSF under the first schemeto 5.93±0,36
G-CSF WTO is Oh scheme 12,64±0,63*
The inventive tool under the first scheme10,8±0,72*
The inventive tool according to the second schema15,9±0,52*
* - the difference is significant in comparison with hypoxic control at p<0,05

Cited literature:

1. Mashkovsky PPM Medicines: 15-ed. - M: PA "Publishing house New Wave", 2008. - 1206 S.

2. Alekseev, G.V., Gurvich A.M., Semchenko V.V. Postresuscitation encephalopathy (pathogenesis, clinical features, prevention and treatment). - Omsk; the Omsk regional printing house, 2003. - 152 C.

3. Pogodaev N.N., Gapova C., Vereshchagin A.L., Gorshkov A.G., Semenov A.A. study of the alkaloid composition of some species of Siberian aconito // Rast. resources, issue 2, 2000, S-84.

4. Digi A.M., Sushkov GN. Fundamental aspects of the prospects of using nano-tech modifiers functions of stem cells in regenerative medicine // Nanotechnology and health. - 2012. - Volume IV. No. 2 (11), - P.30-38.

5. Sushkov G.N., Krapivin V., Nesterov Y., Povetieva T.N., Zhdanov V., Suslov., Fomin TI, Udut E.V., Miroshnichenko L.A., Kamanina E.V., A.A. Semenov, S. Kravtsova, Digi A.M. regenerative Mechanisms of action of diterpene alkaloids of Aconite Baikal // BJ is. the experimental. Biol. and medicine, 2012. No. 6. - S-827.

6. Patent (RU) for the invention №2240604 "Method of modeling post-hypoxic encephalopathy and related disorders in the blood system", 2004 (publ. 20.11.2004,, bull. No. 32). Authors: Goldberg ED, Digi A.M., Sushkov G.N., Suslov N

7. Bureš I Buresova O., Houston J. P. Techniques and basic experiments for the study of brain and behavior./Per. s angl. Under the editorship of Professor A.S. Batueva). - M.: Higher school, 1991. - 398 S.

8. Patent (RU) for the invention №2284060 "experimental therapy encephalopathy", 2006, (publ. 20.09,2006, bull. No. 26). Authors: Goldberg ED, Digi A.M., Sushkov G.N., Zhdanov V., Suslov N

9. Manual on experimental (preclinical) study of new pharmacological substances, Ed. RU Khabriev. - 2 ed. - M.: JSC "Publishing house "Medicine", 2005. - P.54-69.

Application zingarina as cerebroprotective funds.



 

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FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmaceutics, namely using an aqueous-alcohol solution of Hymenocardia acida leaves for preparing a pharmaceutical composition for treating hypertension.

EFFECT: using the aqueous-alcohol solution of Hymenocardia acida leaves enables preparing the composition for treating hypertension which is well-tolerated by the patients if administered regularly and continuously.

10 cl, 4 dwg, 4 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to pharmaceutics, namely to an agent for preventing or relieving gastrointestinal diseases. Compositions for preventing or relieving gastrointestinal diseases consists of two or more herbal therapeutic agents specified in a group of: Bupleuri Radix, Coptidis Rhizoma and Glycyrrhizae Radix which represent an unprocessed extract in a solvent specified in a group consisting of water, alcohols C1-C4 and combinations thereof. The therapeutic agent for preventing or relieving a gastrointestinal disease containing the above composition and a pharmaceutically acceptable carrier or a pharmaceutically acceptable additive. A functionally therapeutic product for preventing or relieving the gastrointestinal disease containing the above composition consisting of two or more herbal unprocessed extracts of Bupleuri Radix, Coptidis Rhizoma and Glycyrrhizae Radix.

EFFECT: using the declared composition enables the more effective treatment and prevention of the gastrointestinal diseases.

8 cl, 6 tbl, 6 ex, 4 dwg

FIELD: medicine, oncology, amino acids.

SUBSTANCE: invention relates, in particular, to the development of an antitumor preparation based on natural substances. Invention relates to an amino acid preparation comprising at least one modified essential amino acid obtained by treatment of amino acid by ultraviolet radiation (UV) at wavelength 250-350 nm for 12-80 h at temperature 15-30oC or with ozone at temperature 15-25oC. The modified amino acid has no toxicity for health cells. Also, invention relates to a method for preparing such preparation. Invention provides the development of an antitumor preparation based on modified amino acids and expanded assortment of antitumor preparations being without cytotoxicity for normal cells.

EFFECT: valuable medicinal antitumor properties of preparation.

8 cl, 4 tbl, 2 dwg, 4 ex

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