RU2433794C2 - Method of modelling stimulation of reparative osteogenesis with extract of fetal bone tissue cells - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: invention relates to medicine and can be applied for modelling stimulation of reparative osteogenesis. Osteosynthesis is performed. Stimulating preparation is introduced. As stimulating preparation used is dry extract of fetal bone tissue cells of animal, prepared from bones of calvarium, which is dissolved in sterile solvent immediately before introduction and after 14 days of fixation is introduced once in dose 1 mg per 1 kg of body weight into paraossal tissues of fracture region.

EFFECT: method makes it possible to prevent resorption in zone of fragment junction, reduce term of fracture consolidation.

1 ex, 10 dwg

Description

The invention relates to medicine, namely to orthopedics and traumatology, and veterinary medicine, can be used to stimulate the reparative processes of damaged bones, in particular with a significant degree of trauma to osteogenic tissues (bone marrow, periosteum) and damage to the vessels feeding the bone.

A known method of modeling stimulation of bone tissue regeneration, which consists in delimiting the region of a circular defect of a long tubular bone from the penetration of surrounding tissues by a film of dinitrocellulose, which is applied circularly with overlapping edges of the defect by 5-7 mm and tightly fixed with nylon threads, previously removing the periosteum from the ends of bone fragments (RF Application 2005100265/14. Method for modeling bone tissue regeneration stimulation / Shevtsov V.I., Dyachkov A.N., Ruchkina I.V., Russian State Institution The 1st Scientific Center “Reconstructive Traumatology and Orthopedics” named after Academician G.A. Ilizarov (RF); declared on January 11, 2005, published on June 20, 2006, Bull. No. 17).

However, the known method is used to stimulate the reparative regeneration of bone tissue when replacing circular defects.

A known method of stimulating the repair process of the bone, which consists in the fact that after the end of the distraction period, the distal and proximal parts of the bone reagent are brought together, compressing the connective tissue layer of the regenerate (Patent 2071740 of the Russian Federation, MKI6 A61B 17/56. Method of stimulating the repair process of the bone / V.I. . Shevtsov (RF), A.V. Popkov (RF). - No. 94013185/14; application. 13.04.94; publ. 20.01.97, Bull. 2).

However, the known method is intended to stimulate the formation of bone regenerate during distraction osteogenesis.

A known method of mechanical stimulation of delayed osteogenesis in bone fractures, which consists in the fact that bone fragments are fixed with a device for transosseous osteosynthesis with the ability to move and 3-5 days after surgery, metered distraction efforts are applied to bone fragments at a rate of 0.5 mm per day, for 4 days, then the fragments are fixed in the reached position for 3 days, after which they are compressed, precisely matching the fragments, orienting them relative to the longitudinal axis, and fixing to the joint patent (Patent 2354322 RF, IPC8 A61B 17/58. Method for mechanical stimulation of delayed osteogenesis in bone fractures / N. Kononovich (RU), E. Dyuryagin (RU), A. Dyachkov (RU), Federal State Institution “ RRC “WTO” named after Academician G. A. Ilizarov of Rosmedtehnologii ”(RU). - No. 20071364269/14; declared. 01.10.2007; publ. 10.05.2009, Bull. No. 13).

However, the known method assumes as a direct stimulating effect the violation of the immobility of bone fragments at the fixation stage, which can be achieved only with the help of a special fixing structure that allows you to create metered distraction and compression forces in the fracture zone.

Known casting composition for stimulating bone formation and bone fusion, containing calcium sulfate and viscous biopolymers, preventing the growth of unwanted connective tissues, as well as inducing the growth of blood vessels and the development of bone osteoblasts at an early stage (RF Patent No. 2296588 C2, IPC A61L 27/40, 27/44, 27/00, A61P 19/00 Filling composition for stimulating bone formation and bone fusion containing calcium sulfate and viscous biopolymers / KIM In-San, Cho Biung Tea., REGEN BYOTEK, INC. (KR) - No. 2005113276/15; claimed 30.09.2002., Published 10.04.2007, Bull . No. 10).

However, the known composition is used to fill the cavities of bone defects, and is also introduced into the cavity of the diastasis in order to stimulate osteogenesis after stretching of the bones, in particular the jaw, and was not used to stimulate osteogenesis during the healing of bone fractures.

A known method of modeling delayed osteogenesis, including violation of the integrity of bone tissue, fixation of bone fragments by the apparatus for transosseous osteosynthesis, in which dissected adjacent to the bone soft tissue. An osteotomy of the tibia of the experimental animal is carried out, the ends of the fragments are removed from the wound. The contents of the bone marrow cavity and the area of the nutrient artery are removed from each. Then the ends of the fragments are compared and fixed to fusion. Under unfavorable physiological conditions created, 30 days after osteosynthesis, pronounced osteoclastic resorption of the ends of bone fragments is determined. Complete bone fusion coalescence occurs at 60-75 days of stable fixation and is characterized by endochondral osteogenesis (RF Patent No. 2301457, MKI8 G09B 23/28. Modeling method for delayed osteogenesis / Kononovich N.A. (RU), Dyuryagin EB (RU), Dyachkov AH (RU), Federal State Budgetary Institution Scientific and Research Center “WTO” named after Academician G.A. Ilizarov of Roszdrav (RU) - No. 2005104529/14; announced on February 18, 2005; published on June 20, 2007, Bull. 17).

It is known that nonviable human fetuses obtained from women who artificially terminate pregnancy for social and medical reasons (Doskaliev Zh.A., Kayupov B.A., The role of fetal stem cells in the restoration of impaired functions of organs and tissues // serve as a source of fetal material). Annual All-Russian and International Conference “Stem Cells and the Prospect for Their Use in Health Care.” 2007. URL: http://www.stemcell.ru/news/phpnews/cell_news.php?nom_news=930 (accessed September 28, 2009)) .

Fetal organ preparations are known (transplants, groups of cells, cell extracts, hydrolysates and individual biomolecules), which are prepared from organs and tissues of healthy animals (http://www.regbiomed.com/journal/cabines/Cabines№14_2004.doc).

A known method of stimulating osteogenesis, including intramuscular injection of a suspension of allogeneic hydroxylappatite in an isotonic sodium chloride solution and providing acceleration of bone tissue formation at the site of demineralized bone matrix in the experiment (RF Patent No. 2219933, IPC A61K 33/00. Method of stimulation of osteogenesis / Volova L.T. , Podkovkin V.G., Vlasov M.Yu. - No. 2002119768/14; declared. 22.07.2002., Published. 27.12.2003).

There is a method of stimulating reparative processes in bone tissue after osteosynthesis, during which animals in the postoperative period are injected with SELPLEX at a dose of 5 mg per 1 kg of live weight 1 time per day for 14 days (RF Patent No. 2295962 C1, IPC A61K 33 / 04, A61P 19/00 Method of stimulation of reparative processes in bone tissue after osteosynthesis / Sakhno NV, Sadovnikov NV, Papazyan TT, Sadovnikova N.Yu., Chervanev VA, Troyanovskaya L .P., Federal State Educational Establishment of Higher Professional Education "Oryol State stressed agricultural university "(RF). - No. 2005128000/14; declared. 07.09.2005, published. 03.03.2007, Bull. No. 9).

However, the known method does not have a direct stimulating effect on reparative processes in the lesion area, does not prevent the occurrence of osteoclastic resorption of the ends of the fragments, and does not contribute to the development of intramembranous osteogenesis.

The objective of the present invention is to develop a method that has a direct stimulating effect on reparative processes in the lesion zone, prevents the occurrence of osteoclastic resorption of the ends of the fragments, promotes the development of intra-membranous osteogenesis and thereby reduces the time of fracture consolidation. The problem is solved in that in the method for modeling stimulation of reparative osteogenesis with an extract of fetal bone tissue cells, including osteosynthesis and administration of a stimulating drug, a dry extract of fetal bone tissue cells prepared from bones of the cranial vault, which is dissolved in sterile immediately before administration, is used as a stimulating drug solvent and injected once at a dose of 1 mg per 1 kg of body weight into the paraossal tissue of the fracture area.

It is advisable to use water for injection or an isotonic sodium chloride solution as a solvent.

The present invention is illustrated by a detailed description, the attached photos, radiographs and histotopograms, on which:

Figure 1 - view of the leg of the dog after violation of the integrity of the bone and the imposition of the transosseous fixation apparatus (hereinafter the apparatus);

Figure 2 - dry extract of cells of fetal bone tissue;

Figure 3 - a solution of a dry extract of cells of fetal bone tissue;

Figure 4 - injection of a solution of extract of fetal bone tissue in the paraossal tissue of the fracture area;

5 is a copy of an x-ray of the bones of the leg, 30 days of fixation, according to the invention;

6 is a copy of the x-ray of the bones of the leg, 45 days of fixation, according to the invention;

7 is a copy of the histotopogram of the regeneration of the diaphysis of the tibia of the dog after 45 days of limb fixation in the apparatus. The drug is stained according to Van Gieson. Magnification - 1.5x .;

Fig. 8 is a copy of a micrograph of the intermediate region of the bone fusion zone. Hematoxylin and eosin stain. Magnification - 250x;

Figure 9 is a copy of the micrograph of the periosteal region of the bone fusion zone. Hematoxylin and eosin stain. Magnification - 250x;

Figure 10 is a copy of a micrograph of red bone marrow in the bone marrow channel of the regenerate. Hematoxylin and eosin stain. The magnification is 630x.

The method is as follows.

A bone fracture, accompanied by significant damage to the osteogenic tissues and the vessels supplying the bone, is precisely repaired and stably fixed (Figure 1). After 14 days of fixation, reparative osteogenesis is stimulated with an extract of fetal bone tissue cells prepared from bones of the cranial vault. For this, a dry extract of cells of fetal bone tissue (Figure 2), prepared from the bones of the cranial vault, at a dose of 1 mg per 1 kg of body weight is dissolved in a sterile solvent in an amount necessary for small volume injection (Figure 3). As a solvent, water for injection or an isotonic sodium chloride solution is used. The resulting solution is administered once by injection into paraossal tissues at the junction of bone fragments (Figure 4). As a result, by 30 days after osteosynthesis, there are no signs of osteoclastic resorption of the ends of the fragments in the area of their junction (Fig. 5) and after 45 days of stable fixation, complete bone fusion ensured by the formation of the spongy bone, both in the intermediate zone and due to periosteal bone formations (Fig.6, 7). During this period, the fixation of the fracture is stopped.

An example of the method.

In an experimental animal (dog No. 4790), an open fracture of the lower leg bones was simulated, accompanied by significant damage to the periosteum, bone marrow and rupture of the vessels feeding the bone. For this, in the operating room of an anesthetized animal, after treating the surgical field with a 5% alcohol solution of iodine, the Ilizarov apparatus was placed on the right shin. An open transverse osteotomy was performed in the middle third of the tibia diaphysis with a Jigley saw. To enhance the degree of trauma to osteogenic tissues from the bone marrow cavities of the proximal and distal fragments, the contents were removed to a depth of at least 1 cm, after which the fragments were accurately compared. Knotted stitches were applied to the soft tissue. The surgical field was treated with 5% alcohol solution of iodine.

14 days after osteosynthesis, a solution of a dry extract of fetal bone tissue cells prepared from bones of the cranial vault was injected into a paraossal tissue at the junction of the fragments by a small volume at a dose of 1 mg per 1 kg of body weight, where a sterile isotonic solution was used as a solvent.

30 days after osteosynthesis, radiological contours of the ends of the fragments were fuzzy. The inter-fragment gap is not enlarged. It was covered by homogeneous shadows of medium intensity. Near the fracture line, shadows of periosteal layers were observed, not combining fragments among themselves.

By 45 days after osteosynthesis in the radiographs, the fracture line was poorly visualized. The contours of the ends of the fragments were blurred. The inter-fragment space was blocked by high-intensity homogeneous shadows. The shadows of periosteal layers united bone fragments among themselves in the area of their junction.

In a clinical examination, pathological mobility and soreness were absent. After that, the fixation by the Ilizarov apparatus was stopped.

By this time, bone fusion determined by the formation of a trabecular bone both in the intermediate zone and due to periosteal bone formations soldering the ends of fragments was determined on histotopograms. In the intermediate zone, osteons are oriented along the growth of vessels from the side of the periosteum and endosta. Bone beams that form osteons were close in structure to lamellar bone tissue (Fig. 8). The periosteal zone of the regenerate was represented from the medial surface of the trabecular mediastinal bone (Fig. 9), from the lateral surface — portions of the trabecular bone of the coarse-fibrous structure and small fragments of fibrous connective tissue. Endosteal callus in the fracture area is represented by a narrow strip of cancellous bone. The formation of a bone marrow cavity filled in the projection of the fracture with sections of the reticular tissue with hematopoietic elements and loose fibrous connective tissue, single bone trabeculae was noted. Closer to the endostomy, the formation of red bone marrow was determined (Figure 10), single and bony trabeculae and a red-yellow bone marrow with dilated sinusoids were noted above and below the fracture site.

The proposed method for modeling stimulation of reparative osteogenesis with an extract of cells of fetal bone tissue allows creating favorable conditions for reparative regeneration in case of bone fractures, which helps to prevent the occurrence of osteoclastic resorption in the junction of the fragments and contributes to the formation of intramembranous osteogenesis. This leads to a reduction in the terms of stable fixation of fractures by 1.5 times. In addition, the proposed method is easy and quick to perform.

The proposed method was developed at the Federal State Institution “Russian Scientific Center“ Reconstructive Traumatology and Orthopedics ”named after Acad. GA Ilizarova Rosmedtekhnologii ”and is intended for its use in clinical medical practice and in veterinary medicine.

Claims (1)

A method of modeling stimulation of reparative osteogenesis with an extract of fetal bone tissue cells, including osteosynthesis and administration of a stimulating preparation, characterized in that the dry extract of fetal bone tissue cells of an animal prepared from bones of the cranial vault, which is dissolved in a sterile solvent immediately before administration, is used as the stimulating preparation after 14 days of fixation, they are administered once at a dose of 1 mg per 1 kg of body weight into the paraossal tissue of the fracture area.

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