RU2295964C1 - Method for treating ischemic tissue alterations - Google Patents

Abstract

FIELD: medicine, surgery.

SUBSTANCE: in area of the lesion one should form canals due to laser radiation at wave length of 0.97-1.064 mcm in uninterrupted or impulse modes, at power being 7-15 Wt. In intervals between canals one should fulfill implantation of medullary cells. As an implant one should apply a punctate of rubra medulla ossium of a patient's trabecular bones separated from mature erythrocytes, bony elements and fats. The method normalizes circulation in area of the lesion, increases viability of implanted cells and possibility for their differentiation up to endothelial cells of new blood vessels.

EFFECT: higher efficiency of therapy.

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Description

The invention relates to medicine and can be used to treat trophic disorders and stimulate reparative processes in tissues with their ischemic damage of a different nature and localization.

There is a method of implanting autologous bone marrow cells, which consists in introducing autologous bone marrow cells at points located on the ischemic muscle of the affected limb (Surgery 2001. Jul 130 (1) 44-54. The induction of angiogenesis by the implantation of autologous bone marrow cells. Hamano K., Li TS., Kobayashi T., Tanaka N., Kobayashi S., Matsuzaki M., Esato K.)

In a known technique, bone marrow cells are prepared with the removal of red blood cells.

The results of the study after the bone marrow cell implantation procedure showed that angiogenesis was significantly induced in ischemic muscle. Laser Doppler showed a significant increase in blood flow.

However, this method still does not provide sufficient treatment effectiveness.

A known method of stem cell implantation in an ischemic myocardium in an experiment, selected as the closest analogue, comprising injecting a stem cell suspension injection into the epimiocardial channels formed in the myocardial zones with a post-infarction scar laser in a continuous mode (see patent RU No. 2237440, IPC-7: A 61 B 17/00, published on 05/20/2004).

In the known method, stem cells are isolated in a biological protection box from the bone marrow of the rib removed from the experimental animal as a result of its subperiosteal resection of the rib.

In this work, it was noted that the use of stem cell transplantation to replace myocardial defects allows us to fix newly formed cardiomyocytes on the 9th day, which fill the infarct zone by 68%, significantly reducing dyskinesia sites.

According to the creators of the known method, the proposed method eliminates the diffuse spread of stem cells through tissues associated with injection, improves the accuracy of localization of implanted autologous stem cells and ensures minimal trauma, which facilitates the process of sampling material for histological examination and provides a more detailed assessment of stem interaction cells with surrounding myocardium and, as a result, allows you to solve the problems of treatment of myocardial infarction with the power of cell cardiomyoplasty.

However, the use of only bone marrow stem cells as an implant does not provide ischemic tissues with the whole set of biologically active substances contained in the bone marrow puncture with blood cells, which does not allow to effectively resolve the treatment of trophically altered tissues using a known method.

This is because the stem cells isolated in the known method are washed (isolated) from the components of the extracellular matrix, which in the bone marrow is a significant source of biologically active substances (growth factors, heparan sulfates, ferments, cytokines, etc.) and is a necessary source of normal the existence of stem cells.

In addition, the use of a resected rib of a biological object to extract stem cells from the bone marrow is too traumatic and not acceptable for humans. In this case, the selection of stem cells in a special biological protection box cannot but affect the cost of this treatment method.

And maintaining a suspension of stem cells directly into the channels made by the laser cannot provide adequate treatment for ischemic tissue changes, since the channel made using laser radiation using the parameters specified in the known method is surrounded by a tissue necrosis zone, which reduces the survival rate of the introduced stem cells, since they are devoid of normal microenvironment and nutrient medium.

Thus, the technical result, the solution of which this invention is directed, is to achieve a higher degree of effectiveness of the treatment of ischemic tissue changes.

The specified technical result is achieved by the fact that in the known method of treating ischemic tissue changes, including the implementation of the channels in the affected area, carried out using laser radiation and the subsequent implantation of bone marrow cells in this area, according to the invention in the spaces between the channels made by laser radiation with a length waves of 0.97-1.064 microns in continuous or pulsed modes, with a power of 7-15 W, carry out the implantation of punctate of the red bone marrow of the patient’s spongy bones, separated from mature eras Irocytes, bone elements and fats.

The authors claim that it’s the claimed combination of performing channels in the affected area using laser radiation and implantation of red bone marrow cells in the spaces between these channels, using spongy bones separated from mature red blood cells, bone elements and fats for implantation of punctured red bone marrow, allows you to achieve the highest, not achieved today by any of the known methods, the degree of effectiveness of the treatment of ischemic tissue changes.

According to the authors, this is due to the fact that the implementation of channels in the tissues of the affected area using laser radiation promotes the formation of a network of small vessels in the places around them, which successfully compensates for blood flow in this tissue and significantly accelerates the normalization of blood flow in the affected area, and implantation in the intervals between the channels of punctured red bone marrow of the patient’s spongy bones, separated from mature red blood cells, bone elements and fats, it contributes to the creation of sufficient the concentration of biologically active substances and enzymes necessary for the successful repair and growth of new blood vessels.

In addition, the bone marrow stem cells themselves, which are present in the inventive suspension, can differentiate from the endothelial cells of new blood vessels, including due to the presence of a normal microenvironment and a nutrient medium.

The suspension claimed by the authors contains growth factors, cytokines, proteolytic enzymes of the collagenase and plasminogen group, secreted by bone marrow cells, which are a powerful source of biologically active substances, unlike stem cells used in the closest analogue, washed from the extracellular matrix components.

The authors experimentally established that the best result of the treatment of tissue ischemia can be achieved with the claimed parameters of laser radiation.

The method is as follows.

With trophic changes in the tissues of the lower limb.

The implementation of the channels of ischemic tissue is as follows.

The formation of channels in the tissues of the affected limb is carried out using laser radiation with a wavelength of 0.97-1.064 microns in continuous and pulsed modes. The power of the radiation used is from 7 to 15 watts. A light guide with a diameter of 0.4-0.6 mm is used.

Using a source of high-intensity laser radiation, perforations (channels) are applied through the skin and soft tissues through 1-5 cm through the skin and soft tissues. There can be many options for applying perforations (channels):

- throughout the lower leg;

- throughout the thigh;

- throughout the lower leg and in the proximal and distal parts of the thigh;

- throughout the thigh and proximal and distal parts of the leg;

- from hip to foot, etc.

In this case, the canals can be performed both to the periosteum, and can be limited only by soft tissues.

When performing channels limited by soft tissues, the fiber is punctured through the injection needle into the tissue of the affected limb. The implementation of perforations (channels) to the periosteum is performed by a light guide through the skin, soft tissues. When the laser source is turned on, the fiber is inserted into the soft tissues of the affected limb through distances from 1 to 5 cm. The channels are formed around the circumference of the affected limb, with the exception of the projection zones of the neurovascular bundles. The upper limit of exposure passes above the site of occlusion of the main bloodstream.

Holes are applied from the outside and inside of the affected limb.

The total number of channels made both to the periosteum and limited to soft tissues depends on the extent and volume of trophic changes in the lower extremities.

Before making perforations (channels), the patient is sampled 10-100 ml of punctate from the ilium or sternum containing red bone marrow and venous blood.

By centrifugation under aseptic conditions, a fraction of mature red blood cells is isolated from the obtained punctate (at the bottom of the tube). At the same time, the suspension is washed (isolated, separated) from bone elements and fat.

Plasma and a fraction of immature cellular elements (middle fraction of cells) are collected in a syringe in a volume of 10-70 ml.

Injection (0.3-0.5 ml at each point of injection), after performing the channels with laser radiation, chipping tissue located in the spaces between the made channels.

The number of laser channels performed and the number of injections of the inventive suspension and their volume depends on the volume of ischemic tissue.

The clinical criteria are used as criteria for the effectiveness of the therapy: subjective sensation in the patient of warming of the feet and improvement of tactile sensitivity of the toes of the feet, increase in the distance of painless walking, reduction in pain, healing of necrotic defects.

With trophic changes in liver tissue.

Tests of the proposed method were carried out on biological objects - rats with a model of liver cirrhosis of ischemic origin.

In the liver tissues with laser radiation with a wavelength of 0.97-1.64 μm, a radiation power of 7-15 W in perforated and continuous modes perform perforations (channels). The number of holes made depends on the volume of trophic changes in the tissues of the biological object and on the volume of the affected organ.

Punctate of myeloid tissue and blood is taken from the iliac bones, which are washed by centrifugation (isolated, separated) from mature red blood cells, bone elements and fats. The resulting suspension is injected into the liver tissue of a biological object. Places of introduction of the suspension are located in the gaps between the previously made laser channels. The amount of suspension administered and the number of injections performed also depends on the volume of ischemic tissue and the volume of the affected organ.

With trophic changes in myocardial tissue.

Tests of the proposed method were carried out on biological objects - dogs with a model of coronary heart disease.

Perforation holes (channels) are made on the heart of a biological object by laser radiation with a wavelength of 0.97-1.064 microns in continuous or pulsed modes with a power of 7-15 watts. The number of channels depends on the extent of the lesion.

Bone marrow punctate is taken from the ileum. The resulting punctate is centrifuged to separate mature red blood cells, bone elements and fats. The resulting suspension is punctured into the heart of a biological object - a dog. Suspension injections are performed in the heart tissue located in the gaps between the previously made laser channels. The amount of suspension administered and the number of injections depends on the volume of ischemic tissue and the volume of the affected organ.

The results of the studies are evaluated by morphological studies of the treated organs.

The claimed method is confirmed by examples of specific performance.

Example No. 1. Patient X. 72 years. Diagnosis:

Obliterating atherosclerosis of the lower extremities. Occlusion of the right femoral artery, left popliteal artery, stenosis of the iliac arteries, chronic ischemia of the III degree according to Fontaine.

The patient underwent surgical treatment of the claimed method.

Using laser radiation with a wavelength of 0.97 in a pulsed mode with a radiation power of 10 W, a 0.6 mm optical fiber made 18-10 perforations (channels) respectively to the periosteum on the thigh, lower leg, and foot of the right and left limbs, respectively. Performed on 8 channels in the muscle tissue of the affected limbs.

A 100 ml punctate containing red bone marrow and venous blood was taken from the iliac crest. By aseptic centrifugation, a fraction of mature red blood cells was isolated (at the bottom of the tubes).

At the same time, the suspension is washed (separated) from bone elements and fat. 60 ml of plasma with an average cell fraction was collected into the syringe, which was injected into the trophic lesion zone between the perforations (laser channels) by repeated injections of 0.3-0.5 ml at each injection site.

The day after the operation, the patient had pain in his legs at rest, his feet became warmer, a week later the distance of painless walking increased 4 times (from 50 m to 200 m).

Example No. 2. The biological object is a rat. Model of liver cirrhosis of ischemic origin.

In the liver tissue of a biological object, 10 perforations (channels) were made by laser radiation with a wavelength of 0.97 μm in a continuous mode with a power of 7 W, a fiber with a diameter of 0.4 mm.

1 ml of bone marrow punctate was taken from the iliac bones of the biological object. By centrifugation under aseptic conditions, the fraction of mature red blood cells (at the bottom of the tube) was isolated from the obtained punctate. At the same time, the suspension is washed (separated) from bone elements and fat. Plasma was collected in the syringe with an average fraction of cells in a volume of 0.7 ml. The resulting suspension for 4 injections is introduced into the liver tissue in the gaps between the previously performed laser channels.

A week later, with a morphological study in the liver tissues against the background of cirrhotic changes, foci of liver tissue regeneration were determined.

Example No. 3. Biological object - a dog with a model of coronary heart disease.

On the working heart of a biological object, 10 perforations (channels) were made by laser radiation in a pulsed mode with a wavelength of 1.64 μm with a power of 15 W, a fiber diameter of 0.4 mm.

15 ml of bone marrow punctate were taken from the iliac bones of the biological object. Processing the received punctate according to examples No. 1 and No. 2. Received 7 ml of the claimed suspension, which by injection of 0.5 ml at each point was introduced into the gaps between the previously performed laser channels.

After 3 weeks, during a morphological study of the myocardium, an active process of neoangiogenesis was observed - a lot of newly formed capillaries, arteries and veins of small caliber, neoplasm of cardiomyocytes.

Claims (1)

A method for the treatment of ischemic tissue changes, including the implementation of the channels in the affected area, carried out using laser radiation and the subsequent implantation of bone marrow cells in this area, characterized in that in the spaces between the channels made by laser radiation with a wavelength of 0.97-1.064 μm in continuous or pulsed modes, with a power of 7-15 watts, the punctate of the red bone marrow of the patient’s spongy bones, separated from mature red blood cells, bone elements and fats, is implanted.