Method and transplant for treatment of liver failure

FIELD: medicine.

SUBSTANCE: inventions relate to medicine, namely to cell transplantology, and deal with method and transplant for treatment of liver failure. For this purpose autologous progenitor cells of bone marrow are isolated and cultivated in vitro. Also realised is sampling of autologous liver cells. After that, immobilisation of autologous liver cells and progenitor cells of bone marrow on carrier - biodegradable biocompatible three-dimensional matrix is performed. After that, transplantation of carrier with cells is performed by its introduction into mesentery of small intestine. Transplant includes biodegradable biocompatible three-dimensional porous matrix with pore size 2-500 mcm and total porosity 50-97%, ensuring total concentration of liver cells and progenitor bone marrow cells 2×106-15×106 cells per 1 cm3 of matrix and ratio of progenitor bone marrow cells to liver cells from 1:1 to 1:4. Total volume of matrix constitutes not less than 0.05 cm3, its smallest linear size being not less than 0.2 mm.

EFFECT: inventions make it possible to improve results of liver failure treatment due to prolongation of terms of hepatocyte survival and activisation of their proliferation, creation of conditions for growing of vessels into creates carcass, diffusion of nutrients, oxygen and factors of tissue differentiation, make it possible to avoid application of immunosuppressive therapy.

6 cl, 6 dwg

 

The invention relates to medicine, transplantation, namely, cell transplantation, and can be used in the correction and treatment of liver failure. The proposed method and the graft can be used in specialized departments involved in the treatment and correction of liver failure.

There is a method of treatment of liver failure, based on the use of suspensions of isolated xenogeneic hepatocytes in extrakorporale system supporting the liver, through which perpusilla the blood of the recipient liver (Shumakov V.I. and co-authors. Essays on physiological problems of Transplantology and artificial organs // Tula. - Reprints. - 1998. - s-362).

Isolated hepatocytes in such systems, as xenogenic capable in the body of the patient only for a short time to carry out detoxification and synthetic function. At the same time, the ability of suspensions of isolated hepatocytes to allocate regulatory peptides, stimulates regeneration processes in the diseased liver recipient, extended to 0.5-8 hours (Shumakov V.I. and other Treatment of severe liver failure perfusion of the patient's blood through a suspension of cryopreserved hepatocytes. // Surgery. - 1990. No. 2. - C.113-116).

Know of any other way cured the I liver failure, using microfragments liver tissue, which helped to prolong the metabolic activity of hepatocytes contained in microfragments to 24-34 and even up to 48 hours (Solovyev V.V., Onishchenko N.A., Akatov B.C., Lezhnev EI Functional activity of hepatocytes in the slices of liver in vitro: dependence on size of the fragments and the length of their cultivation. // Bul. The experts. Biol. and Honey., - 1997, - №10, - s-408). Term increase in metabolic activity of hepatocytes in the composition of microfragments is due to the saving of intercellular contacts and the natural extracellular matrix, as well as due to the preservation of the spatial structure of the liver tissue, resulting in optimized conditions for the functioning of all cell types, including hepatocytes.

Also known is a method of treatment of hepatic failure involving for prolonging the time of survival of isolated hepatocytes in perfusion bioreactors use of isolated hepatocytes with Mironosetsky (using, for example, cytodex-3 with a ratio of 1.6 g of micronesias 1×109cells), which are pre-coated with collagen (Dimetriou A.A., Rozga J., L. Podesta Early clinical experience with a hybrid bioartificial liver // Scan. J. gastroenterol. - 1995. - 30. - Suppl.208. - p.111-117; Chen, S., Eguchi S. Watanabe Hepatic support strategies// Transplant. Proc. - 1996. - 28, No. 4. - R-2038).

As a prototype of the and selected known method of treatment of liver failure, using isolated hepatocytes for auxiliary support of the liver and, consequently, fill detoxification and bioregulatory functions of the damaged liver (Van de Kerkhove M.P., R. Hoekstra, Chamuleau R.A., van Gulik TM Clinical application of bioartificial liver support systems. // Ann. Surg. - 2004. - Vol.240. - P.216-230).

The disadvantages of known methods, including prototype, predpolagaemyh the use of extracorporeal bioreactor with microfragments liver tissue or isolated hepatocytes include:

- the necessity of using special grinders to obtain fragments of a given size, which increases the risk of infection from donor material;

- the necessity of using perfusion systems and inclusion in the perfusion circuit additional components for oxygenation and detoxification of the blood (plasma)coming to microfragments liver tissue, which increases the economic cost method.

- the need of the use of bioreactors microfragments in a mixture with particles of a porous biocompatible carrier to prevent adhesion of microfragments and ensure the effective mass transfer;

- the use of particles of porous media does not create conditions to prevent delayed death of hepatocytes in microfragments, because with the increase date the cultivation of microfragments due to the low rate of proliferation of hepatocytes begins to form a monolayer of quickly attached and proliferating stromal cells, enveloping local liver tissue and impairing their functioning;

- the necessity of using xenogenic material that increases the risk of transfer of dangerous infections and contributes to excessive activation of the immune system of the recipient with the reduction of the period of operation of this material;

- early termination (1-2 days) metabolic and regulatory functions of hepatocytes;

- the need to regularly exercise sessions connected perfusion systems "bioisostere liver".

A device for the treatment of liver failure in vitro method of connection of isolated hepatocytes (Dimetriou A.A., Rozga J., L. Podesta Early clinical experience with a hybrid bioartificial liver // Scan. J. gastroenterol. - 1995. - 30. - Suppl.208, - p.111-117). Also known is a device designed for the treatment of liver failure, which can increase the period of viability of isolated hepatocytes (Shumakov V.I. and co-authors. Essays on physiological problems of Transplantology and artificial organs // Tula. - Reprints. - 1998. - s-362).

As a prototype of the proposed transplant for the treatment of liver failure is selected intracorporeal transplant (bimodal) "auxiliary liver" (Uyama, S., Kaufman, P., Kneser U., Vacanti J., Rodriges X. Hepatocyte transplantation using biodegradable matrices in ascorbic acid-deficient rats: comparsion with heterotropically ransplanted liver grafts // Transplantation. - 2001. - Vol.7. - P.1-7).

The disadvantages of the known devices, including prototype, predpolagaemyh using bimodule with isolated hepatocytes include:

- severe inflammatory reaction to the graft;

- the death of a large number of isolated hepatocytes;

- low density attaching cells of the liver;

- high cost matrix carrier.

The objective of the invention is to develop a method that improves the efficiency of correction and treatment of liver failure due to prolongation of the terms of the survival of isolated hepatocytes and their implementation functional and organ-specific activity by transplant long-term functioning intracorporeal graft type "auxiliary liver".

Technical result achieved in the implementation of the invention, consists in the correction and treatment of liver failure by providing isolated hepatocytes detoxification function, prolongation of the terms of their survival by activation of proliferation through joint intracorporeal injection progenitor cells in the bone marrow, create a frame for placement in the space of associations formed cells, creating conditions for germination in this framework vessels, diffusion of nutrients, KIS is Orada and factors of tissue differentiation of the blood vessels of the mesentery and blood vessels, sprouted in the frame, to the immobilized liver cells and bone marrow cells; and in the prevention of complications associated with blood clots and cellular infiltration at the site of transplantation of the graft with the cellular material.

The advantages of the proposed method and graft for the treatment of liver failure, allowing essentially to create intracorporeal system auxiliary liver for long-term maintenance of functional status of isolated hepatocytes are:

- no need to use complex perfusion systems;

- creation using a 3-dimensional matrix of culture conditions for cell proliferation and the formation of a "tissue-like" structure;

- creation of adequate conditions for diffusion oxygenated interstitial fluid and sprouting of blood vessels through the matrix that prolong adequate conditions of life planted cells;

- the use of autologous cells, which reduce the degree of activation of the immune system, allow the system to provide in the body long bioregulatory effects.

In the proposed invention does not use tissue and/or cellular material of human embryos. Used cellular material adult donors.

Co-cultivation of liver cells and casinogaming before transplantation to further increase the functional activity of hepatocytes by releasing growth factors and cytokines, secreted by bone marrow cells and creates conditions for the differentiation of bone marrow cells in the endothelial cells to form blood vessels and their germination in the matrix. This contributes to a more rapid integration of the graft into the circulatory system after transplantation and maintain metabolism and vital functions of cells by delivery of the newly educated and sprouted vessels oxygen; creation of adequate conditions for diffusion oxygenated interstitial fluid that prolong adequate conditions of life planted cells. The use of anticoagulants and antiplatelet agents immediately after transplantation inhibits the cellular response to the transplant as foreign body and prevents the formation of thrombi at the site of transplantation.

The essence of the invention is as follows.

For the treatment of liver failure use of isolated liver cells. Moreover, pre-exercise intake of autologous progenitor cells in the bone marrow. Carry out culturing them in vitro. Then taking autologous liver cells. Perform landing freshly isolated liver cells and cultured bone marrow cells on three-dimensional biocompatible biodegradable porous matrix. The pore size of the matrix varies from 2 to 500 μm. The total porosity of the matrix of the leaves 50-97%. The concentration of total liver cells and bone marrow is 2×106- 15×106cells on 1 cm3matrix. The ratio of bone marrow cells to liver cells from 1:1 to 1:4. And the total amount of matrix is not less than 0.05 cm3. The smallest linear dimension of the matrix should be not less than 0.2 mm carried out the co-cultivation of autologous liver cells and bone marrow. Transplantation matrix with cells of the liver and bone marrow produce in the mesentery of the small intestine.

Cultivation of bone marrow cells is carried out for, e.g., 7 days.

In the particular case after the landing of the liver cells and bone marrow in the matrix prior to transplantation carry out the co-cultivation of autologous liver cells and bone marrow cells, for example, in 2-3 days.

In the particular case immediately after transplantation prescribe anticoagulants in the prevention dose. Dose: heparin 2500 ME every 12 hours for 7-10 days under the control of the blood coagulation system and antiplatelet agents in the prevention dose, e.g trental based 45 mg/m2body surface area every 12 hours for 30 to 90 days.

The proposed transplant for the treatment of liver failure includes a three-dimensional biocompatible biodegradable porous matrix and planted on him autologous cells in the bone MH is and after in vitro cultivation and freshly isolated autologous liver cells. The total volume of the matrix is not less than 0.05 cm3and the smallest linear dimension is not less than 0.2 mm pore Size of the matrix 2-500 μm, the total porosity 50-97%. Concentration of liver cells and bone marrow 2×106-15×106cells on 1 cm3matrix and the ratio of bone marrow cells to liver cells from 1:1 to 1:4.

The proposed method and the transplant allow prolonged corrected acute or chronic liver failure due to prolongation of the functioning of isolated hepatocytes. Transplantation is not allogeneic, and autologous cells eliminates complications associated with rejection reaction. The proposed group of inventions allows the following.

1. To create conditions for the attachment of isolated liver cells and bone marrow cells to a biocompatible matrix, stimulates their proliferation, which is supported by diffusion in the matrix of nutrients, oxygen and factors of tissue differentiation of blood flowing to the matrix of the mesenteric vessels.

2. To create a structure based on biocompatible biodegradable and porous matrices that mimic the spatial structure for attaching cells of the donor liver and progenitor cells from the bone marrow, as well as conditions for germination in this framework vessels feeding the attached cells.

<> 3. To create conditions that prevent cell infiltration of the graft and its supporting cells in a viable state through the use of anticoagulants and antiplatelet agents.

The method is as follows and includes several stages:

1) Isolation of autologous progenitor cells in the bone marrow.

The allocation of progenitor cells from the bone marrow carried out according to traditional methods (Shumakov V.I., Onishchenko N.A., Krasheninnikov M.E. Ter-Minassian and other Bone marrow as a source of mesenchymal cells to restore the treatment of damaged organs. //The Bulletin of transplantation and the suit. bodies 2002, 4, p.3-6; Shumakov V.I., Onishchenko N.A. et al. Biological reserves bone marrow cells and correction of organ dysfunction // Moscow, Laurel. 2009, - p.61-67). For 4-7 days before the main surgical intervention under local anesthesia dotted line in the iliac bone of the patient and take the bone marrow in the amount of 40-150 ml in a sterile container with Hanks solution containing 200 μg/ml gentamicin; and 10.0 μg/ml insulin; 0.25 μm of dexamethasone; 250 u/ml of heparin. Cell suspension centrifuged 5 min at 1500 rpm and the precipitate cells resuspended in lyse solution (114 mm NH4Cl; 7.5 mm KHCO3; 100 μm EDTA), at a ratio of 1:4 from the original volume of aspirate for 5-10 min and centrifuged 3 min the ri 1500 rpm at room temperature. Hamalsarany supernatant completely removed by suction. Achieve complete lysis of red blood cells, for which the procedure lizirovania spend three times with subsequent laundering of the cells by centrifugation. Cellular precipitate free of erythroid and platelet forms resuspended in the growth environment.

2) Cultivation of autologous progenitor cells from bone marrow in vitro.

Culturing progenitor cells in the bone marrow carried out according to traditional methods (Shumakov V.I., Onishchenko N.A., Krasheninnikov M.E. Ter-Minassian and other Bone marrow as a source of mesenchymal cells to restore the treatment of damaged organs. // The Bulletin of transplantation and the suit. bodies 2002, 4, p.3-6; Shumakov V.I., Onishchenko N.A. et al. Biological reserves bone marrow cells and correction of organ dysfunction // Moscow, Laurel. 2009, - p.77-100). Partially purified progenitor cells in the bone marrow were seeded for cultivation in Petri dishes (d=60 mm, the number of 1.5-2.0 million cells/ml were Cultured at 37°C in CO2the incubator atmosphere of 5% CO2and 95% humidity for 7 days with one change of medium on the third day. A week later, the culture of bone marrow cells contained up to 10% adherent to plastic spread fibroblast-like and macrophage cells and up to 90% of free-floating in suspension, not rounded CIDP is spivshihsya cells (hematopoietic cells). Neprecejusies to the plasticity of progenitor cells in the bone marrow was collected and then used for immobilization on porous matrix with the cells of the liver.

3) Isolation of autologous liver cells.

Produce resection of 2-4×2-4×1-2 cm of the liver tissue in patients with liver failure to obtain liver cells. The allocation of autologous cells of the liver are carried out by the traditional method (Fontaine M, Schloo B, Jenkins R, Uyama S, Hansen L, Vacanti J.P. Human hepatocyte isolation and transplantation into an athymic rat, using prevascularized cell polymer constructs. // Pediatr. Surg. - 1995. - vol.30(1). - P.56-60; Hang H, Shi X, Gu G, Wu Y, Ding Y. A simple isolation and cryopreservation method for adult human hepatocytes // Int J Artif Organs. 2009 Oct; 32(10). - P.720-7; Lehec S.C, Hughes RD, Mitry R.R, graver carving M.A, Verma A, Wade J.J, if you A. Experience of microbiological screening of human hepatocytes for clinical transplantation. // Cell Transplant. 2009; 18(8). - P.941-947; O. Seglen Preparation of isolated rat liver cells. // Methods. Cell. Biol. 1976. - vol.13. - P.29-83).

The allocation of autologous liver cells produce of the resected portion of the liver by 3 times washing piece of liver from the blood and grinding it to a cold (t=4°C) in a Petri dish, with 3 times washing the resulting suspension buffer solution without calcium [1000 ml of distilled water, 8.3 g of NaCl, 0.5 g KCl, 2.38 g HEPES, pH of 7.4, 37°C] for 7 minutes. After this, small pieces of liver are incubated 3 times with a solution of collagenase [1000 ml of distilled water, 8.3 g of NaCl, 0.5 g KCl, 0.7 g of CaCl2, 2.38 g HEPES, 7.5 mg trypsin inhibitor and 500 mg collagenase Type V-S (> 125 CDU/mg), pH 7.3; 37°C] for 6-8 minutes, followed by replacing the enzyme solution using centrifugation at 500 rpm for 1 minute at t=37°C. the resulting material was transferred to a sieve with 200 µm and filtered washing with culture medium William's E with 10% fetal bovine serum, after which a suspension of single cells and small aggregates are transferred to a centrifuge tube and centrifuged at 500 rpm at 4°C for 1 minute. The supernatant is removed, the residue resuspended in the same fresh medium and centrifuged again. The procedure was repeated 3 times. Cell viability assessed by the method of dyeing Trifanova blue. Get get the number of cells in the range from 3.0×108to 4.0×108hepatocytes 12-15 g of liver tissue. The cell suspension should contain: hepatocytes, neverthemore liver cells, which determine when light microscopy. The separation of parenchymal and nepriskonowennij cells do not perform. A suspension of liver cells concentrate in 1-2 ml of physiological solution.

4) Fit (immobilization) autologous liver cells and autologous bone marrow cells to the media.

Landing (immobilization) of cellular material is carried out according to traditional methods (D.J. Mooney, Sano K., R.M. Kaufmann, Majahod K., Schloo Century, Vacanti JP, Langer R. Long-term engraftment of hepatocytes transplanted on biodegradable polymersponges // J.Biomed. Mater. Res. - 1997. - Vol.5. - P.413-420).

Autologous liver cells and autologous progenitor cells in the bone marrow resuspendable in the growth medium [William's E with the substitution of arginine to ornithine, with the addition of fetal bovine serum, growth factor hepatocytes, epidermal growth factor, β-subunit of the cholera toxin, dexamethasone, ethanolamine, sodium Selenite, glucagon, insulin, insulin-like growth factor-I, ascorbic acid, linolevoi and linelevel fatty acids] in a concentration of 2.0 to 4.0×106liver cells/ml and 2,0-4,0×106progenitor bone marrow cells/ml Total cell suspension was applied to the matrix at a concentration of 2×106-15×106on 1 cm3on pre-saturated environment matrix.

As the matrix can be used, for example, the biomaterial-based polyoxometalate (3-oxybutyrate) - Elastopor. Polyoxometalate - homopolymer synthesized different types of prokaryotic cells. They are the substrate of endogenous respiration and maintain cell viability in non-optimal environmental conditions (Sevastianov V.I., Perova NV, Dovzhik I.A., Titushkin I.A., German E.A., Belomestnykh SM, shyshatskyy E.I., Volova T.G. biomedical properties of polyoxyalkylated - biodegradable bacterial polymers // advanced materials, 200, No. 5. - P.46-55; Sevastianov V.I., Perova NV, Shishatskaya E.I., Kalacheva G.S., Volova T.G., Production of purified polyhydroxyalkanoates (PHAs) for applications in contact with blood. J. Biomater. Sci. Polymer Edn., 2003, V.14(10). - P.1029-1042; T. Volova, E. Shishatskaya, Sevastianov, V., Efremov, S., Mogilnaya O., Results of biomedical investigations of PHB and PHB/PHV fibers, Biochem. Eng. J., 2003, No. 3736. - P.1-9).

Elastopor currently approved for medical use. The main component Elastopor is a biodegradable bacterial copolymer of β-oxybutyrate and β-oxovalerate (FOB-with-STAND) with the inclusion of oxovalerate 15÷30 mol.% (Mw=295-360 KDa, crystallinity 50-60%), obtained from the Institute of Biophysics SB RAS, Krasnoyarsk, Russia. Part Elastopor also includes high-molecular hydrophilic plasticizer (AIV), which increases the hydrophilicity and elasticity of the material.

Physico-chemical, mechanical and technological properties Elastopor make this biopolymer is very attractive to develop time frames for hybrid bioisosteric bodies (Sevastianov V.I., Egorov V.A., German E.A., Perova NV, Onishchenko N.A. a Biodegradable biopolymer material Elastopor for cell transplantation //advanced materials, 2004, №3. - P.35-41; Sevastianov V.I., German E.A., Volkov YEAR, Markovtsev MG three-Dimensional porous matrices for stem-cell-based biodegradable bacterial copolymer Bioplastics"// advanced materials, 2007, №6. - P.5-1).

Used matrices Elastopor had the following dimensions and parameters: diameter of the porous sponge - 10±2 mm; thickness of 1.2±0.5 mm; weight - 6,0-12,0 mg Porosity is not less than 95±2%. The size of the macropores is 300±100 microns.

5) co-cultivation of autologous progenitor cells in the bone marrow and autologous liver cells in VITRO.

In the particular case of co-cultivation was performed autologous liver cells and progenitor cells from bone marrow according to the standard technique (P.M. Kaufmann, K. Sano, Uyama, S., Breuer C.K., Organ G.M., Schloo B.L., Kluth D., Vacanti J.P. Evaluation of Methods of Hepatotrophic Stimulation in Rat Heterotopic Hepatocyte Transplantation Using Polymers // J. of Pediatric Surgeru. - 1999. - Vol.34. - P.1118-1123; Uyama, S., Kaufmann P.M., Takeda T., J.P. Vacanti Delivery of whole liver equivalent hepatocyte mass using polymer devices and hepatotrophic stimulation // Transplantation. - 1993. - Vol.55. - P.932-935).

This allowed to increase the number of viable liver cells and progenitor cells from the bone marrow, capable of performing specific functions: detoxification and synthetic functional activity of hepatocytes, and created conditions for the differentiation of progenitor cells from the bone marrow into the endothelial cells to form blood vessels and their germination in the matrix.

The matrix with cells were placed in a sterile chamber for incubation in vitro in the growth medium [William's E with the substitution of arginine to ornithine, with the addition of fetal bovine serum, growth factor hepatocytes, epidermal growth factor, β-is jedinica the cholera toxin, dexamethasone, ethanolamine, sodium Selenite, glucagon, insulin, insulin-like growth factor-1, ascorbic acid, linolevoi and linelevel fatty acids] for 2-3 days.

6) Transplantation of media, with immobilizerturning by autologous liver cells and progenitor cells in the bone marrow in the body.

Transplantation of matrices with immobilizerturning by autologous liver cells and autologous progenitor cells in the bone marrow provide patients with liver failure in the mesentery of the small intestine for 3-4 days after resection.

7) Use of anticoagulants and antiagregantov.

After transplantation of matrices with immobilizerturning by autologous liver cells and autologous progenitor cells in the bone marrow of patients prescribed anticoagulants in the prevention dose, for example: heparin 5,000 IU every 12 hours for 7-10 days under the control of the blood coagulation system and antiplatelet agents, for example, trental based 45 mg/m2body surface area every 12 hours for 30 to 90 days.

The proposed transplant for the treatment of liver failure consists of a three-dimensional biocompatible biodegradable porous matrix and planted him autologous progenitor cells from the bone marrow and autologous liver cells.

For the proof in is the possibility of achieving the stated purpose and achievement of the technical result here is the following data.

An example of the proposed method using the proposed transplant experiment.

Modeling of chronic liver insufficiency in animals (rats) were carried out according to recognized and adequate model (Fischer A. Physiology and experimental pathology of the liver // Afisher. - Budapest, 1961, - 230 S.; Kalashikov IVAN Effect of transplantation of cells of the thymus, bone marrow and spleen on regenerative processes in the diseased liver, Byull. the experimental. Biol. and med., - 1979, No. 10. - S-480; Kalashikov IVAN General and local changes in the body in experimental liver injury and regeneration // abstract. Prof. Diss. - 1982, Kazan. - 41 S.) by introducing a 60% solution of CCl4the first administration of 0.5 ml per 100 g mass, subsequent to 0.3 ml per 100 g body weight. Course introduction 6 weeks with a frequency of 2 introduction in a week.

Modeling of acute liver failure in animals (dogs) was carried out by resection 45-50% of the liver tissue, which is also a recognized and adequate model of acute liver failure (Demetriou, A., Reisher A., Sanchez J. Et al. Transplantation of microcarrier-attached hepatocytes into 90% partially hepatomized rats // Hepatology. - 1988. - Vol.8. - P.1006-1009; Chikoti S. p., Plekhanov A.N. Liver failure // in the book. Liver failure. Irkutsk. - 2002. - 260 S.; G.K. Michalopoulos Liver regeneration: Molecular mechanisms of growth control // FASEB J. - 1990. - Vol.l04. - P.16; Bowling WM, Kennedy SC, Cat SR, Duncan JR, Gao C, Flye MW. Portal branch occlusion safely facilitates in vivo retroviral vector transduction of rat liver. // Hum. Gen. Ther / - 1996, - vol.7(17), - p.2113-2121).

1) was Carried out according to traditional methods, the selection of autologous progenitor cells in the bone marrow.

Received cellular aspirate of bone marrow channel belieberzone bones (rats) or humerus (dogs) by lavage of a phosphate buffer solution containing 50 u/ml heparin and 0.25 mg/l of gentamicin using 18G needle mounted on the syringe. Cell suspension was centrifuged, the precipitate cells resuspendable in lytic solution at room temperature for 3 min and centrifuged for 3-5 min at 1500 rpm Hamalsarany supernatant was completely removed by aspiration, and the cell precipitate containing progenitor cells in the bone marrow, resuspendable in the growth environment. The interphase with mononuclear cells were collected from the surface of erythroid sediment and resuspendable in lytic solution, at a ratio of 1:4, for 5-8 min and centrifuged 5 min at 1500 rpm at room temperature. Hamalsarany supernatant was completely removed. Achieved complete lysis of red blood cells, for which the procedure lizirovania conducted twice or three times with the subsequent laundering of the cells by centrifugation. Cellular precipitate free of erythroid and platelet the forms, the number of 60-150×106cells were combined with sediment cells derived from plasma, and then resuspendable in the growth environment to stimulate cell growth.

2) was Carried out following the conventional method of culturing progenitor cells in the bone marrow.

Partially purified progenitor cells in the bone marrow were sown for cultivation in the amount of 1.5-2.0 million cells/ml were Cultured at 37°C in CO2the incubator atmosphere of 5% CO2and 95% humidity for 7 days with one change of medium on the third day.

3) Isolation of autologous liver cells was carried out according to traditional methods.

The allocation of autologous liver cells from resected area of the liver (4×4×2 cm in dogs and 1×1×0.5 cm in rats) was performed by 3 times washing with blood and grinding it out in the cold (t=4°C), with 3 times washing the resulting suspension buffer solution without calcium within 7 minutes. After this, small pieces of liver were preincubator 3 times with a solution of collagenase for 6-8 minutes, followed by replacing the enzyme solution using centrifugation at 500 rpm for 1 minute at t=37°C. the resulting material was transferred to a sieve with meshes of 200 μm and was filtered washing with culture medium William's E with 10% fetal bovine serum, after which the cell suspension was centrifuged at 500 rpm is Ying at 4°C for 1 minute. The supernatant was removed, the residue resuspendable in the same fresh medium and again centrifuged. The procedure was repeated 3 times. Cell viability, just in the range from 83 to 95% was estimated by the method of dyeing Trifanova blue. The number of cells ranged from 3.0×108to 4.0×108hepatocytes on 15 g of liver tissue. The cell suspension contained: hepatocytes ~95÷98%, and ~5÷2% nepriskonowennij liver cells, which were identified by light microscopy. Division parenhimatosny and neprekonatelnych cells was not performed. A suspension of liver cells was concentrated to 1-2 ml of physiological solution

4) Landing (immobilization) autologous liver cells and progenitor cells from the bone marrow to the matrix is also carried out according to traditional methods.

Autologous liver cells and progenitor cells in the bone marrow resuspendable in the growth medium at a concentration of 2.0 to 4.0×106liver cells/ml and 2,0-4,0×106progenitor bone marrow cells/ml Total cell suspension was applied to the matrix at a concentration of 2×106-15×106on 1 cm3on the pre-saturated environment matrix.

5) In the particular case of co-cultivation was performed autologous liver cells and progenitor cells from bone marrow according to the usual method.

It was possible to increase the number of viable liver cells and progenitor cells from the bone marrow, able to perform specific functions: detoxification and synthetic functional activity of hepatocytes, and to create conditions for the differentiation of progenitor cells from the bone marrow into the endothelial cells to form blood vessels and their germination in the matrix.

The matrix with cells were placed in a sterile chamber for incubation in vitro in the growth medium for 2-3 days.

As the matrix was used, for example, the biomaterial-based polyoxometalate (3-oxybutyrate) Elastopor. Elastopor has a molecular weight of 295-360 KDa synthesized by the Institute of Biophysics SB RAS (Krasnoyarsk), and is currently approved for medical use.

Used matrices Elastopor had the following dimensions and parameters: diameter of the porous sponge - 10±2 mm; thickness of 1.2±0.5 mm; weight - 6,0-12,0 mg Porosity is not less than 95±2%. The size of the macropores is 300±100 microns.

6) Transplantation of matrices with immobilizerturning by autologous liver cells and progenitor cells in the bone marrow in the body of animals (dogs) was carried out for 3-4 days after modeling liver failure by resection of 50% of the liver tissue.

Before transplantation, animals (dogs) were narcoticyou: a mixture of drugs was carried out by intramuscular injection of a mixture of products: Calypsol (5% of 10 mg/kg Droperidol (1.5 mg/kg), Atropine (0.1% to 0.02 mg/who), Diphenhydramine 1% to 0.5 ml and then, in the operating room was performed anaesthesia induction by intravenous injection of 1% Propofol at the rate of 7 mg/kg. the Operation was performed under endotracheal anesthesia. Maintenance of anesthesia was performed with 1% Propofol 0.5 mg/kg/h and Kalipsola 2 mg/kg In rats was carried out by anesthesia allowance per 100 g body weight, were injected intramuscularly: Calypsol 10% - 2 ml, Xylazine 2% - 1 ml Atropine of 0.2 ml.

Total cellular material containing autologous liver cells and bone marrow at a concentration of 2×106-15×106on cm3that is immobilized on the matrix Elastopor, transplanted in the mesentery of the small intestine of animals.

The abdominal cavity is sutured in layers tightly. For the prevention of infectious complications intraoperatively animals were intravenously administered drove Ofloxacin 40 ml; Metronidazole - 30 ml; Ditilin 5% to 20 mg/kg/h of the monitored indicators of biochemistry, coagulation, blood count at 1, 3. 9, 14, 18 days after the operation.

7) Use of anticoagulants and antiagregantov.

In the particular case to prevent cell infiltration of the graft by the red blood cells immediately after transplantation matrices with immobilizerturning by autologous liver cells and autologous progenitor cells in the bone marrow daily was prescribed anticoagulants in the pros is fakticheskoi dose for example: heparin 2500 ME every 12 hours for 7-10 days under the control of the blood coagulation system and antiplatelet agents in the prevention dose, for example: trental based 45 mg/m2body surface area every 12 hours for 30 to 90 days.

Altogether, we conducted 25 experiments, with correction of acute liver failure under the proposed method was carried out in 5 experiments and chronic liver failure in 20 experiments.

Figure 1 shows the dynamics of growth and reduction of acute liver failure in the control, where: 1 - AST, 2 - ALT 3 - GGT, 4 - ALP, 5 - LDH. The maximum increase in indicators of liver failure detected 1 day after modeling liver failure. Studies have shown that after resection of 50% of liver ALT levels compared with the outcome of increased 6 times, a AST 15 times, which indicates a significant functional impairment of liver cells. The level of alkaline phosphatase after resection has increased in 5 times. Then a gradual return of these indicators to normal by 18 days after modeling liver failure.

Figure 2 shows the dynamics of growth and reduction of acute liver failure in the experiment after transplantation matrix with planted at him autologin the mi progenitor cells in the bone marrow and autologous cells of the liver under the proposed method, where 6 - AST 7 - ALT, 8 - GGT, 9 - ALP, 10 - LDH.

From the presented data it follows that until the transplantation of autologous liver cells and progenitor cells from the bone marrow, planted on the matrix, the dynamics of biochemical indicators of liver failure was similar with the control group. However, since transplantation (3 days after modeling liver failure) autologous liver cells and progenitor cells from the bone marrow, planted on the matrix, indicated more rapid decline of biochemical indicators of liver failure. Alkaline phosphatase was higher than the original level only 1.5 times in comparison with 1 day after resection decreased 3.5 times. Especially bright lower level indicators of liver failure seen on day 5 after transplantation of autologous liver cells and progenitor cells from the bone marrow, planted on the matrix, these figures decreased almost 2 times compared to the control. Moreover, these figures are normalized to 14 days after the modeling of acute liver failure in the study group and only 18 days after surgery in the control group.

Figure 3 presents the group viable autologous hepatocytes 90 days after their transplantation into the body of EC the pilot of the animal on the proposed method. Color: hematoxylin - eosin. The magnification ×400 (where 11 - viable hepatocytes).

Figure 4 presents these histological examination after 180 days after the transplantation of autologous liver cells and progenitor cells from the bone marrow, planted on the matrix. In histological preparations of the animals that had not received anticoagulant and antiplatelet agents identified expressed lymphoid reaction (12), fibrotic changes (13) and giant cells of foreign bodies (14) (as inflammatory reaction on the substrate). Coloring PAS. Magnification ×400.

Figure 5 animals who received anticoagulants and antiplatelet agents, revealed significantly lower cell response. In the preparation of meet up groups of viable hepatocytes (15) and isolated mononuclear cells (16). Coloring PAS. Magnification ×400.

Figure 6 after 90 days in a drug identified the newly formed vessel (17). Coloring PAS. The magnification ×400.

In all cases, was achieved the desired result, and it was received and adequately compensated liver failure. The method can improve the results of treatment of liver failure by providing isolated autologous hepatocytes detoxification and bioregulatory functions, extension of the terms of their survival by activation of proliferation at the expense of sovmestnog the intracorporeal injection of autologous progenitor cells in the bone marrow, create a frame for placement in the space of associations formed cells, creating conditions for germination in this framework vessels, diffusion of nutrients, oxygen and factors of tissue differentiation of the blood vessels of the mesentery, sprouted in the frame, to the immobilized liver cells and bone marrow cells; provides for prevention of complications associated with the use of graft cells. In addition, the use of autologous liver cells and progenitor cells from the bone marrow avoids the use of immunosuppressive therapy, and long-term viability allows you to maintain homeostasis.

Thus, the above data clearly indicate that correction of liver failure is much faster and more appropriately in the application of the proposed method of correction of liver failure. In addition, our proven long-term survival of liver cells and the appearance of newly formed vessels in the graft after 90 and 180 days after transplantation animal matrix with autologous cells of the liver and autologous progenitor cells in the bone marrow.

Given the above, the extrapolation of experimental results to the clinic legitimate, the proposed method and the graft, providing prolonged the e zhiznedeyatelnosti isolated hepatocytes, can be used for the correction of liver failure.

1. A method for the treatment of liver failure, including the use of isolated liver cells, wherein the pre-exercise intake of autologous progenitor cells in the bone marrow and their cultivation in vitro, then taking autologous liver cells, perform landing freshly isolated autologous liver cells and cultured autologous progenitor bone marrow cells on three-dimensional biocompatible biodegradable porous matrix with a pore size of 2-500 μm and a total porosity of 50-97%, providing a total concentration of liver cells and progenitor cells from the bone marrow 2·106-15·106cells on 1 cm3matrix and the ratio of the progenitor cells from the bone marrow to the liver cells from 1:1 to 1:4, and the total amount of the matrix must be not less than 0.05 cm3and its the smallest linear dimension should be not less than 0.2 mm, and transplanted matrix with the cells of the liver and bone marrow in the mesentery of the small intestine.

2. The method according to claim 1, characterized in that after landing autologous liver cells and progenitor cells from the bone marrow to the matrix prior to transplantation produce their co-cultivation.

3. The method according to claim 1, characterized in that after transplantation of autologous CL is current and liver progenitor bone marrow cells on the matrix prescribe anticoagulants and antiplatelet agents in the prevention dose.

4. The method according to claim 1, characterized in that the culturing progenitor cells in the bone marrow is carried out for 7 days.

5. The method according to claim 2, characterized in that the co-cultivation of autologous liver cells and progenitor cells from the bone marrow is carried out in 2-3 days.

6. The transplant for the treatment of liver failure, including a three-dimensional biocompatible biodegradable porous matrix having a total volume of not less than 0.05 cm3and the smallest linear dimension of at least 0.2 mm, a pore size of 2-500 μm, the total porosity 50-97%, and planted him autologous progenitor bone marrow cells after in vitro cultivation and freshly isolated autologous liver cells, and the concentration of liver cells and bone marrow 2·106-15·106cells on 1 cm3matrix and the ratio of bone marrow cells to liver cells from 1:1 to 1:4.



 

Same patents:

FIELD: medicine.

SUBSTANCE: group of inventions relates to medicine and can be applied for treatment of liver failure. Transplant includes three-dimensional biocompatible biodegradable porous matrix, which has total volume not less than 0.05 cm3 and the smallest linear size not less than 0.2 mm, pore size 2-500 mcm, total porosity 50-97%, and implanted on it allogenic progenitor cells of bone marrow and allogenic liver cells, concentration of liver and bone marrow cells being 2×106-15×106 cells per 1 cm3 of matrix and ratio of bone marrow cells to liver cells being from 1:1 to 1:4. In realisation of method of treating liver failure transplant is placed into mesentery of small intestine.

EFFECT: group of inventions makes it possible to increase term of cell survival, activate their proliferation.

6 cl, 7 dwg

FIELD: medicine.

SUBSTANCE: group of inventions relates to medicine and can be applied for treatment of liver failure. Transplant includes heterogenic biocompatible biodegradable gel, which has total volume not less than 0.1 ml and the smallest linear size not less than 0.2 mm, pore size 30-500 mcm, total porosity 50-98%, implanted on it autologous progenitor cells of bone marrow after their cultivation in vitro and cultivated autologous liver cells, concentration of liver and bone marrow cells being 2×106-15×106 cells per of 1 cm3 of heterogeneous gel and ratio of bone marrow cells to liver cells being from 1:1 to 1:4. In method of treating liver failure transplant is placed into parenchyma of liver and/or mesentery of small intestine.

EFFECT: group of inventions makes it possible to increase term of cell survival, activate their proliferation.

4 cl, 6 dwg

FIELD: medicine.

SUBSTANCE: group of inventions relates to medicine and can be applied for treatment of liver failure. Transplant for treatment of liver failure includes heterogenic biocompatible biodegradable gel, which has total volume not less than 0.1 ml and the smallest linear size not less than 0.2 mm, pore size 30-500 mcm, total porosity 50-98%, implanted on it autologous progenitor cells of bone marrow after their cultivation in vitro and cultivated autologous liver cells, concentration of liver and bone marrow cells being 2×106-15×106 cells per 1 cm3 of heterogeneous gel and ratio of bone marrow cells to liver cells being from 1:1 to 1:4. In realisation of method of treating liver failure transplant is placed into parenchyma of liver and/or mesentery of small intestine.

EFFECT: group of inventions makes it possible to increase term of cell survival, activate their proliferation.

4 cl

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to chemical-pharmaceutical industry, namely to application of an extract of an elevated part of scabiose centaury (Centaurea scabiosa L.) as an antioxidant hepatoprotector. A method of preparing the antioxidant hepatoprotector consists in grinding the elevated part of scabiose centaury (Centaurea scabiosa L.) to particle size 2-4 mm followed by three-fold extraction in 70% ethanol at temperature 75-80°C for 1-1.5 hours in the raw material to extractant relation 1:15-1:20, evaporation to dryness of the prepared extract.

EFFECT: application of the extract of the elevated part of scabiose centaury (Centaurea scabiosa L) allows extending the range of herbal antioxidant hepatoprotectors, promotes normalising the antioxidant activity in hepatic tissue, regression of the main biochemical values in hepatitis.

2 cl, 20 ex, 9 tbl

FIELD: medicine.

SUBSTANCE: invention relates to medicine, namely, to efferent therapy, and can be applied in treatment of patients with hepatic failure after performed cardio-surgical operation. For this purpose realised is veno-venous hemofiltration with application of standard bicarbonate solution. If clinical and laboratory signs of hepatic failure increase following sessions of hemofiltration are carried out during 5-8 hours with application of dialytic solution based on standard bicarbonate solution with 2% content of albumen. After session of hemofiltration with such solution, laboratory control of basic functional hepatic indices is performed. In case of necessity additional sessions with application of dialytic solution which contains 2% albumen are carried out.

EFFECT: method makes it possible to increase efficiency of small-flow veno-venous hemofiltration due to increase of clearance of endotoxins, associated with blood plasma proteins.

2 cl, 2 ex

FIELD: medicine.

SUBSTANCE: invention relates to medicine, namely to abdominal surgery, hepatology and methods of detoxication, and can be applied for treatment of liver abscess of different etiology. For this purpose after sanitisation of purulent nidus cavity of liver abscess is processed with 0.25% solution of derinate, then blood sampling is carried out in quantity 150-200 ml of blood, after centrifugation of which erythrocytes are returned to patient, and leucocytes are extra-corporeally processed with 0.5% solution of glutoxim in dose 1 ml and diluted in 50-100 ml of 0.9% solution of NaCl with following intravenous drop introduction, and from the following day into cavity introduced is 0.25% solution of derinate in dose 1.5 ml one time per day during 5 days.

EFFECT: method makes it possible to reduce treatment duration and reduce number of complications due to improvement of indices of cellular link activity, connected with increase of quantitative indices of T-helpers and T-suppressors at the background of increase of phagocytic activity of neutrophils, as well as reduction of erythrodieresis processes due to normalisation of APFC content and humoral link of immunity.

2 tbl

FIELD: chemistry.

SUBSTANCE: invention relates to novel compounds of formula I, which are HSP90 (heat-shock proteins) inhibitors and can be used to prepare a medicinal agent for treating tumorous diseases affected by HSP90 inhibition. In formula I R1 denotes Hal, H, OA or A, R2, R3 each independently denotes -O-(X)s-Q, -NHCO-(X)s-Q, -CONH-(X)s-Q, -NH(CO)NH-(X)s-Q, -NH(CO)O-(X)s-Q, -NHSOr(X)s-Q, NHCOA, Hal, Het or H, where, if R2=H, then R3≠H, or if R3=H, then R2≠H, R4 denotes H, R5 denotes H, Hal, A, OA, (CH2)nCOOH, (CH2)nCOOA, O(CH2)oCONH2, NHCOOA, NHCO(CH2)nNH2, NHCONHA or O(CH2)oHet1, A denotes a straight or branched alkyl containing 1-10 carbon atoms, in which 1-5 hydrogen atoms may be substituted with F, Cl and/or Br, X denotes a straight or branched C1-C10 alkylene which is unsubstituted or substituted once, twice or thrice by A, O A, OH, Hal, CN, COOH, COOA, CONH2, NH2, NHCOA, NHCOOA, Q denotes H, Ar or Het, Ar denotes phenyl which is unsubstituted or substituted once, twice or thrice with A, OA, OH, NO2, Hal, CN, (CH2)nCOOH, (CH2)nCOOA and/or tetrazole, Het denotes a cyclic saturated or aromatic 5-6-member heterocycle containing 1-2 N and/or O atoms, optionally condensed with a benzene ring which may be substituted once, twice or thrice with A, OA, OH and/or =O (carbonyl oxygen), Het1 denotes a monocyclic saturated, unsaturated or aromatic heterocycle containing 1-2 N and/or O atoms, which may be mono- or disubstituted with A, OA, OH, Hal and/or =O (carbonyl oxygen), Hal denotes F, Cl, Br or I, n equals , 1, 2, 3 or 4, o equals 1, 2 or 3, s equals 0, 1 or 2.

EFFECT: high efficiency of using said derivatives.

4 cl, 4 dwg, 1 tbl, 29 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to drugs, and concerns a therapeutic agent for treating hepatitis containing 2-amino-2-[4-3-benzyloxyphenylthio)-2-chlorophenyl]ethyl-1,3-propanediole or its pharmaceutically acceptable salt, or hydrate as an active ingredient. Also, disclosed is a method of treating hepatitis by introducing to a patient an effective amount of the therapeutic agent 2-amino-2-[4-3-benzyloxyphenylthio)-2-chlorophenyl]ethyl-1,3-propanediole or its pharmaceutically acceptable salt, or hydrate as an active ingredient.

EFFECT: therapeutic agent shows a stronger effect in treating hepatitis as compared to the other analogues.

4 cl, 3 dwg, 1 tbl, 2 ex

FIELD: medicine.

SUBSTANCE: invention relates to medicine and is intended for treatment of gastroesophageal reflux disease (GERD) in patients with chronic opisthorchosis. PH-monitoring of upper parts of GIT and dehelmintisation are carried out. Type of reflux is determined. Preparations of ursodeoxycholic acid (UDCA) are administered. In case of mixed type of GER with prevalence of acid component dose is 250 mg/day for 2 weeks. In case of mixed type of GER with prevalence of alkali component dose is 500 mg/day before going to bed for 4 weeks. In isolated alkali version dose is 10 mg/kg in two intakes: morning and evening. In case of tgenpH>7 from 16.54 to 27.6%; NpH>7 from 27 to 31, and GER tgenpH>7 from 27.6 to 48.8%; NpH>7 from 31 to 35 - for 4 weeks. In case of tgenpH>7 from 16.54 to 27.6%; NpH>7 from 27 to 31, and GER tgenpH>7 from 27.6 to 48.8%; NpH>7 from 31 to 35 and in case of GER tgenpH>7 from 48.8 to 74.4%, NpH>7 from 36 to 42 - for 8 weeks. In case of tgenpH>7 from 48.8 to 74.4%; NpH>7 from 36 to 42, and GER tgenpH>7 from 16.54 to 74.4%; NpH>7 from 27 to 42 -for 12 weeks.

EFFECT: method makes it possible to reduce motor-tonic disturbances of biliary tract, reduce number of recurrences, enhance treatment efficiency.

4 cl, 5 ex

FIELD: medicine.

SUBSTANCE: invention relates to field of pharmacy, in particular to phytotherapy, and can be applied for treatment of drug hepatitis. Medication represents herbal mixture, including 29 herbs: herbal part of woodland strawberry, herbal part of common dandelion, inflorescences of pineapple weed, leaves of stinging nettle, inflorescences of tansy, inflorescences of pot marigold, fruits of common juniper, peppermint leaves, herbal part of St John's wort. Medication additionally contains: lime tree flowers, herbal part of heartsease, herbal part of Centaurium, flax seeds, dill seeds, burdock root, herbal part of common chicory, herbal part of common knotgrass, collective fruits of common hop, herbal part of cornflower, leaves of cowberries, leaves of Arctostaphylus, corn stigma, herbal part of immortelle, root and rhizome of Acorus, herbal part of Menyanthes, garden angelica root, herbal part of yellow sweet clover, Inula root, herbal part of common horsetail.

EFFECT: mixture reduces terms of hepatitis treatment, as well as improves clinico-laboratory indices due to normalisation of liver function tests and functional state of liver.

4 ex, 2 tbl

FIELD: medicine.

SUBSTANCE: invention refers to medicine, particularly to percutaneous active substance delivery. A horny layer punching apparatus comprises an element provided with micro flanges having a skin contact surface and a variety of horny layer punching micro flanges on said surface. The apparatus is designed to enable the element provided with the micro flanges moving transversely in relation to a skin surface when contacting said skin.

EFFECT: ensured percutaneous drug delivery and penetration.

5 cl, 15 dwg

FIELD: medicine.

SUBSTANCE: invention refers to medical equipment, namely to biomechanical material injection, and more specifically to a device for high-viscosity material injection in a portion of a patient's body. The device for high-viscosity material injection in a cannula comprises a container, a pressing unit in liquid junction with the container, an overload valve for pressure balance in the device and a material moving element. The container is inextensible and has an outlet hole used for connection with the cannula for viscous material transportation. The pressing unit limits a fluid flow path through which an incompressible fluid is transferred. The material moving element cuts the fluid flow path and forms a section for incompressible fluid reception from its one side and a section for viscous material reception from the opposite side. The section for incompressible fluid reception is connected with the fluid flow path of the pressing unit for applying pressure. The section for viscous material reception is connected with the outlet hole of the container. The material moving element is movable under pressure of the actuating incompressible fluid to displace the viscous material from the section for viscous material reception of the container into the cannula. The viscous material injection device contains the inextensible body, material moving element, pressing unit and overload valve for pressure balance in the device. The material moving element divides the inextensible body on a first cavity having a first volume designed for incompressible fluid reception, and a second cavity having a second volume designed for viscous material reception. The material moving element is movable to change the first and second volumes in inverse proportion. The pressing unit for transferring the incompressible fluid into the first cavity which moves the material moving element for increasing the first volume and decreasing the second volume, thereby injecting the viscous material from the inextensible body. The container for the device for viscous material injection in patient's tissue comprises a proximal end, a distal end whereon an outlet hole for cannula connection and a flexible and inextensible package integrated in the container and designed for viscous material reception are provided. The package has a hole connected with the distal end of the container and connected with its outlet hole. The container is designed for receiving pressurised incompressible fluid surrounding the package, and thereby compressing the package and displacing the viscous material contained therein through the outlet hole of the container. The device for viscous material injection in patient's tissue comprises the inextensible container, the pressing unit in liquid junction with the inextensible container and the material moving element. The container has a proximal end with an inlet hole, a distal end with an outlet hole designed for connection with the cannula. The pressing unit is designed to generate pressurisation of the incompressible fluid. The pressing unit comprises a body and a power piston. The body has a fluid inlet, a fluid outlet and a fluid flow path created in between, at least one return valve in the flow path controlling fluid flow. The fluid outlet is designed in liquid junction with the container. The power piston is coupled with the body, by liquid junction with the flow path. The power piston is designed to provide a mechanical advantage by pressurising the fluid flow path when the incompressible fluid is transferred between the first and second positions. When movable to the second position, the power piston creates suction force transferring the incompressible fluid through the inlet. The return valve prevents reverse flow through the outlet back to the body. If returned to the first position, the power piston creates pressure transferring the incompressible fluid through the return valve from the body through the output and into the inextensible container. The material moving element divides the flow path between the inextensible container and pressing unit actuating the section of incompressible fluid reception and the section for viscous material reception. The material moving element is movable under pressure of the incompressible fluid inside the section of incompressible fluid reception to the outlet of the inextensible container, thereby displacing the viscous material from the section for viscous material reception into the cannula. In the second version of the device for viscous material injection into the cannula, the material moving element comprises a membrane elastically stretched under pressure of the actuating incompressible fluid.

EFFECT: invention provides mechanical advantage in viscous material injection.

49 cl, 14 dwg

FIELD: medicine.

SUBSTANCE: group of inventions refers to biotechnology. A method involves microcapsule supply in a liquid flow onto an output of a microchannel plate 2. The diametre of its channels is less than the size of microcapsules which are kept on the input of the microchannel plate 2 after liquid drainage therethrough. A microneedle lattice 3 is moved to pierce a microcapsule enclosure and to introduce a liquid substance. Thereafter, the filled microcapsules are removed by changing the liquid flow direction through the microchannel plate 2 by a reverse one. The device comprises a microchannel plate 2, a channel 1 with a microcapsule containing liquid, a microneedle lattice 3 and a liquid substance container 4. The lattice 3 is coaxial to the microchannel plate 2. The microneedles are connected with the liquid substance container 4. The channel 1 comprises a microchannel plate 2, a microneedle lattice 3, a piston assembly 5 and valves 8, 9 for maintaining single-direction microcapsule containing liquid motion through the microchannel plate 2. The lattice 3 is movable towards the microchannel plate 2 by a distance of microcapsule enclosure piercing.

EFFECT: inventions provide a continuous process of liquid substance introduction simultaneously in a great number of microcapsules.

2 cl, 4 dwg

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely to occupational pathology, and can be used for pneumoconiosis prevention. That is ensured by the introduction of Aqua Maris nasal spray in each nasal passage by 4-6 injections for at least 6-8 times in a working shift. In addition, Bronchipret is administered by 20 drops in 1/2 glass of water once before a working shift. The therapeutic course is 2.5-3 months twice a year.

EFFECT: higher effectiveness of pneumoconiosis prevention ensured by more evident preventional effect on the upper and lower air passages.

6 tbl, 5 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine, namely to parodontics, and can be used for treating periodontal abscess. That is ensured by antiseptic preparation of the oral cavity, anaesthesia, opening of abscess by blunt dissection, drug-induced management. Thereafter, a dental core in number 1 to 3 containing metronidazole, a dry Echnacea extract, and copolymer of styrene and maleic anhydride as a base in the ratio, wt % 10:10:80 is inserted in the periodontal pocket.

EFFECT: invention allows to eliminate inflammation in single dosage and to avoid undesired gum retraction and systemic administration of antibiotics.

2 ex

FIELD: medicine.

SUBSTANCE: invention refers to medicine, particularly to obstetrics, and concerns a method of birth complications prevention in women with contracted pelvis. For this purpose, 3-4 days prior to an estimated delivery date, oestradiol dipropionate 1.0 ml is introduced intramuscularly, while 10% calcium gluconate 10.0 ml and 5% ascorbic acid 4.0 ml are introduced intravenously. Prepidil gel is introduced in the cervical canal.

EFFECT: invention provides facilitated advancement of a foetus head through the generative passages and, as consequence, reduced duration of delivery, decreased caesarian section and forceps operation cases, lower perinatal mortality and birth trauma rate.

2 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: there is claimed application of concentrated (from 30 mg/mg to 150 mg/ml) methotraxate solution for preparation of medication for subcutaneous introduction for therapy of inflammation-associated autoimmune diseases. Invention also relates to ready for application syringe, carpula, containing said pharmaceutic composition in form of solution, and handle-injector, which contains said carpula and/or ready for application syringe - all contain from 30 mg/mg to 150 mg/ml methotrexate. It is demonstrated that increase of solution concentration to 50 mg/ml reduces volume of daily introduced liquid to 0.6 ml.

EFFECT: there is ensured easier tolerance by patients, which has positive impact on their trust.

32 cl, 2 ex

FIELD: medicine.

SUBSTANCE: group of inventions refers to medical equipment and aims at injection of an active pharmaceutical component. All versions of said device contain retainers for prevention of medicine loss before injection. The first and second versions of the device comprise a case wherein there is a sectioned needle travelling along the central advance axis (X-X). The medicine is supplied to the needle. In the first and third versions of the device, the retainers are placed on the walkway of the needle. The needle is deformed with respect to the central advance axis (X-X) in the injection device when it passes through the retainers. In the second and fourth versions of the device, the retainers comprise an elastic flange ledge which when in rest seals the needle and is taken back when the needle advances.

EFFECT: reduced risk of implant fall-out during storage and transportation of the injection device to an injection place.

50 cl, 4 dwg

FIELD: medicine.

SUBSTANCE: invention refers to medicine and shall find application in making acrylic resin. A container comprises the first tubular member which forms the first solid phase storage chamber with the first open-ended side wall, and a bottom face. The first tubular member forms a longitudinal axis. The second tubular member with the second side wall, the first and second end walls forms the second liquid phase storage chamber. The first and second chambers form a hermetically sealed space. The second tubular member forms a piston which slides in the first tubular member to interact with the compound. The bottom face of the first tubular member has a compound passage to deliver thereof from the container. The piston accommodates filter blocked through recesses to connect the first and second chambers. A barrier comprises one membrane broken by a breaking device, a membrane connected to the bottom face of the first chamber. An external feeder for compound delivery from the container and direct implantation aseptically is connected through the broken membrane to the first chamber.

EFFECT: reduced risk of medium contamination at the stages of compound preparation and implantation.

15 cl, 9 dwg

FIELD: medicine.

SUBSTANCE: invention relates to medical equipment, namely to devices for injection, in particular device for intramuscular or subcutaneous injection of active pharmaceutical component in solid or semi-solid state, usually called implant. Device for injecting implant in the process of reverse injection operation into the subject skin includes hollow main body, to which attached is hollow needle into which implant is introduced, additional body, located coaxially inside main body and surrounding needle and rod with piston which can slide coaxially inside hollow needle. Included are means which allow rod with piston to preserve its position with respect to needle unchanged, when device for injection is pressed to the subject's skin and when additional body is pulled inside main body and which allows rod with piston to enter hollow needle for holding implant at the required depth in the subject's skin while hollow needle in withdrawn from the subject's skin, in which process additional body leaves main body. Device includes means for springy withdrawal of additional body from main body. Resilient means of reverse movement which control withdrawal of additional body from main body are released automatically bringing additional body into initial position, in which it surrounds hollow needle again.

EFFECT: invention considerably simplifies operation of implant injection.

29 cl, 32 dwg

FIELD: medicine.

SUBSTANCE: there is described composition, which includes thermoplastic polymer, speed modifying agent and biologically active agent, which is suitable as implant for medication delivery with slow release into human or animal organism and which can be introduced in organism in liquid form.

EFFECT: characteristics of release and biodegradation of polymeric system with slow release are considerably improved.

2 tbl, 2 ex, 9 cl

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