KR101521411B1 - Composition for the treatment of joint diseases including bone and cartilage and manufacturing method thereof - Google Patents

Abstract

The present invention relates to a composition for treatment of arthropathies affecting bones and cartilages, comprising a stromal vascular fraction (SVF), and extra cellular matrix (ECM). A pharmaceutical composition for arthropathies of the present invention enables bones to recover without a side effect such that surgical operations such as incision, can be minimized, thereby having advantages of no scar and fast recovery. Also, by adding platelet rich plasma (PRP), a calcium chloride solution (CaC12), and hyaluronic acid to the composition, a treatment for arthropathies affecting bones and cartilages can be effectively performed.

Description

TECHNICAL FIELD The present invention relates to a composition for treating joint diseases including bone and cartilage, and a method for manufacturing the same.

The present invention relates to a composition for the treatment of joint diseases including bone and cartilage including an SVM (stromal vascular fraction), an extra cellular matrix (ECM), and a composition for treating platelet rich plasma ), Calcium chloride aqueous solution (CaCl ₂ ), and hyaluronic acid (hyaluronic acid).

Arthritis is a multiple disease with the highest prevalence among diseases suffered by mankind. Of these, degenerative arthritis is a representative geriatric disease, with 70% to 80% of people aged 65 years or older being 75 years old or older. Currently, more than 10% of the total population in Korea has degenerative arthritis, and this prevalence is rapidly increasing due to the rapid aging of Korean society. It is also estimated that there are more than 500 million people with degenerative arthritis worldwide. In addition, as the average life span of human beings is extended and the duration of human societal activity is increased, it occupies an important position to be urgently overcome in terms of qualitative aspects of human life.

Degenerative arthritis is a disease in which the knee joints become worn out as they get older. This occurs when both the inner and the anterior joints of the knee are overused. The knee cartilage is composed of hyaline cartilage and fibrous cartilage. These cartilage tissues are surrounded by membranes at the ends of the bones and act as structural buffers. It prevents the pain that may occur and the wear of the bones.

Chondrocyte is the only cellular component in cartilage tissue. It plays a role of synthesizing, secreting and decomposing matrix of collagen and proteoglycan which is essential for maintaining cartilage tissue's normal function, so as to maintain the functional homeostasis of articular cartilage tissue It plays an essential role in giving.

In order for chondrocyte tissue to function normally, chondrocyte production and differentiation must be appropriately performed, the survival of the chondrocyte should be well protected, and the calcification of the chondrocyte that is present is continuously suppressed, Should be prevented. The most prevalent cause of degenerative arthritis is that the essential function of these articular cartilage cells is lost as a result of aging.

One of the key factors in inducing degenerative arthritis is the quantitative loss of cartilage and cartilage tissue in the cartilage, which is closely related to the differentiation and survival of chondrocytes in cartilage tissue. One of the key factors of degenerative arthritis induction is the hardening of cartilage tissue in the joint, which is directly related to the calcification of cartilage cells. As the degenerative arthritis progresses, the proteoglycan matrix of the cartilage tissue is mechanically abraded and cartilage tissue is destroyed, resulting in the appearance of the bone under the cartilage. Also, at this stage, the small protruding bones newly formed due to the calcification of cartilage cells are deposited on the periphery, causing the hardening of the cartilage tissue, thereby accelerating the loss of cartilage tissue due to abrasion. Such complex deformation of joint tissue structure causes inflammation and pain, and when symptoms become severe, it causes joint deformation and movement limitation.

In order to treat such degenerative joint disease, a procedure for removing the damaged tissue or removing the entire damaged joint is utilized, and in Published Patent Application No. 2011-0106855 (published September 29, 2011), a stem Suggesting a method of transplanting the cells and the bone substrate to the injured joint region through surgical operation. However, these procedures have the limitation that severe incisions or injuries can occur and the recovery is slow.

Published Patent No. 2011-0106855 (Released on September 29, 2011)

It is an object of the present invention to solve the problems of the prior art, and it is an object of the present invention to provide a blood vessel fraction and an extracellular matrix and further comprising platelet-rich plasma, aqueous calcium chloride solution and hyaluronic acid, And to provide a composition for treating a joint disease, including bone and cartilage, which can minimize surgical operation.

In order to achieve the above object, the present invention provides a composition for treating cartilage damage comprising degenerative arthritis, which comprises 8 parts by weight of a stromal vascular fraction (SVF) and an extra cellular matrix (ECM) Wherein the substrate blood vessel fraction is obtained by centrifugation after treating the adipose tissue with collagenase, and wherein the substrate blood vessel fraction and the extracellular matrix are derived from human beings.

The composition for treating cartilage damage including degenerative arthritis of the present invention may further comprise 1 to 10 parts by weight of platelet-rich plasma, and the platelet-rich plasma is derived from autologous blood.

The composition for treating cartilage damage including degenerative arthritis of the present invention may further comprise 0.1 to 0.4 parts by weight of 1 to 5 wt% of calcium chloride aqueous solution and 0.5 to 2 parts by weight of hyaluronic acid in the composition.

The method for preparing a composition for treating cartilage damage comprising degenerative arthritis according to the present invention comprises the steps of (a) mixing and stirring an adipose tissue obtained by liposuction with collagenase at a ratio of 1: 1 to 1: 2, Extracting a stromal blood vessel fraction containing stem cells after the reaction; (b) extracting and separating a stromal blood vessel fraction containing the stem cells, pulverizing the remaining fat extract, mixing the extracted blood vessel fraction again, and removing oil and tumescent solution ; (c) washing the fat extract from which the oil and the citrate solution have been removed to remove the enzyme; (d) centrifuging the fat extract after the washing step to obtain a substrate blood vessel fraction and an extracellular matrix; And (e) 5 to 10 parts by weight of the substrate blood vessel fraction and 0.1 to 10 parts by weight of an extracellular matrix. In the step (e), 1 to 10 parts by weight of platelet-rich plasma may be further added, and further 0.1 to 0.4 parts by weight of 1 to 5 wt% of calcium chloride aqueous solution and 0.5 to 2 parts by weight of hyaluronic acid may be further added have.

The composition for treating a joint disease including bone and cartilage of the present invention, more specifically, a composition for treating cartilage damage including degenerative arthritis, comprises a substrate blood vessel fraction and an extracellular matrix, and can regenerate bone and cartilage without side effects It is possible to minimize the surgical operation such as incision, so there is no scar and the recovery is fast. In addition, the composition may further include platelet-rich plasma, an aqueous solution of calcium chloride, and hyaluronic acid to effectively raise the bone regeneration effect.

Fig. 1 is an image of a fat extract obtained by centrifuging and concentrating adipose tissue obtained by liposuction.
FIG. 2 is an image obtained by separating the blood vessel fraction of the substrate by centrifuging the fat extract of FIG. 1; FIG.
FIG. 3 is an image for pulverizing the remaining fat extract after substrate blood fractionation.
FIG. 4 is an image in which the substrate blood vessel fraction is mixed again with the fat extract pulverized in FIG.
Figure 5 is an image showing substrate blood vessel fraction and extracellular matrix obtained from adipose tissue.
Figure 6 is a knee MRI image before and after a knee with a composition comprising a matrix vessel fraction and an extracellular matrix.
FIG. 7 is a knee MRI image of a composition comprising a matrix blood vessel fraction, an extracellular matrix and a platelet rich plasma before and after knee surgery.
Figure 8 is an image of the femoral head MRI before and after a composition comprising a substrate blood vessel fraction, an extracellular matrix, and a platelet-rich plasma in the femoral head.
FIG. 9 is a knee MRI image of the knee before and after a composition comprising a substrate blood vessel fraction, an extracellular matrix, a platelet rich plasma, an aqueous calcium chloride solution, and hyaluronic acid.
10 is an image of a femoral head MRI before and after a composition comprising a substrate blood vessel fraction, an extracellular matrix, a platelet rich plasma, an aqueous calcium chloride solution, and hyaluronic acid in the femoral head.

Hereinafter, preferred embodiments of the present invention will be described in detail with reference to the drawings. Prior to the description, terms and words used in the present specification and claims should not be construed as limited to ordinary or dictionary meanings and should be construed in accordance with the technical concept of the present invention.

Throughout this specification, when an element is referred to as "including" an element, it is understood that it may include other elements as well, without departing from the other elements unless specifically stated otherwise.

Each step may be performed differently than the order specified unless explicitly stated in the context of the specific order. That is, each of the steps may be performed in the same order as described, or may be performed substantially concurrently or in the reverse order.

Hereinafter, the composition for treating a joint disease including bone and cartilage of the present invention, i.e., a composition for treating cartilage damage including degenerative arthritis, and a method for preparing the same will be described in detail.

The present invention relates to a composition comprising a matrix blood vessel fraction and an extracellular matrix, which can regenerate bones and cartilage without side effects, minimizes surgical operations such as incision, And a composition for treating joint diseases including cartilage.

In the present specification, the substrate blood vessel fraction means a cell obtained by treating fat tissue with collagenase and then centrifuging, and includes cells including immune cells, adipose stem cells and the like. In this case, collagenase is an enzyme that promotes the hydrolysis of collagen. It decomposes collagen in adipose tissue to separate adipocyte stem cells. It can treat other enzymes that play the same or similar roles according to the needs of those skilled in the art It can be replaced by mechanical treatment.

The extracellular matrix is a complex aggregate of biomolecules that fill tissue or extracellular spaces. Its main components are composed of fibrous proteins such as collagen and elastin, and cell adhesive proteins. The substrate blood vessel fraction and the extracellular matrix may be derived from humans, but are not particularly limited thereto.

Preferably, the substrate blood vessel fraction and the extracellular matrix are contained in an amount of 5 to 10 parts by weight and 0.1 to 10 parts by weight, respectively, in the therapeutic composition according to the present invention, and the extracellular matrix is 0.1 to 5 parts by weight The content may be appropriately changed depending on the condition, purpose, sex, age, etc. of the patient in need of transplantation. However, when the content of the matrix blood vessel fraction and the extracellular matrix is less than the above range, the treatment efficiency is low If it is higher than the above range, the increase in treatment efficiency is insufficient, which is economically undesirable.

The present invention relates to a composition for treating a joint disease including bone and cartilage, which further comprises platelet-rich plasma in the therapeutic composition. The platelet-rich plasma may be derived from autologous blood, but is not particularly limited thereto.

 Platelet-rich plasma is contained in the pharmaceutical composition according to the present invention in an amount of 1 to 10 parts by weight. When the content of the platelet-rich plasma is out of the above range, the content ratio with the blood vessel fraction is not appropriate and the synergistic regeneration effect may be lowered.

The present invention relates to a composition for treating joint diseases including bone and cartilage, which further comprises hyaluronic acid and an aqueous solution of calcium chloride of 1 to 5 wt% in the therapeutic composition.

Hyaluronic acid is a biodegradable polymer approved for injection into the body from the Food and Drug Administration and is a substance derived from a living body. The hyaluronic acid contained in the composition of the present invention may be purified from body fluids or tissues of mammals including humans, obtained by culturing recombinant microorganisms or cells, and those obtained by chemical synthesis methods. The weight average molecular weight of the hyaluronic acid may range from about 200,000 to 400,000. The hyaluronic acid may be contained within a range in which the substrate blood vessel fraction and the like can be appropriately dispersed. For example, it is preferable that 0.5 to 2 parts by weight of the hyaluronic acid is contained in the therapeutic composition of the present invention.

The calcium chloride aqueous solution activates the platelet rich plasma to release various growth factors in the platelets to induce tissue regeneration. The concentration of the aqueous solution of calcium chloride contained in the therapeutic composition of the present invention is preferably 1 to 5 wt%, more preferably 3 wt%. It is preferable that the aqueous solution of calcium chloride is contained in the therapeutic composition of the present invention in an amount of 0.1 to 0.4 part by weight, and if it is contained in an amount of less than 0.1 part by weight, the effect of the aqueous calcium chloride solution is not exhibited. If the amount exceeds 0.4 part by weight, platelet- It is possible to inhibit the change to -like substance and to inhibit the role of support.

According to the present inventors' study, it was found that the above-mentioned therapeutic compositions showed the most excellent effects in the treatment of degenerative arthritis and osteonecrosis of the femoral head.

The therapeutic composition of the present invention is characterized in that (a) an adipose tissue obtained by liposuction is mixed and stirred with collagenase at a ratio of 1: 1 to 1: 2 to effect an enzyme reaction, Extracting a substrate blood vessel fraction; (b) extracting and separating a stromal blood vessel fraction containing the stem cells, pulverizing the remaining fat extract, mixing the extracted blood vessel fraction again, and removing oil and tumescent solution ; (c) washing the fat extract from which the oil and the citrate solution have been removed to remove the enzyme; (d) centrifuging the fat extract after the washing step to obtain a substrate blood vessel fraction and an extracellular matrix; And (e) mixing 5-10 parts by weight of the substrate blood vessel fraction with 0.1-10 parts by weight of an extracellular matrix.

In step (e), 1 to 10 parts by weight of platelet-rich plasma may be further added. In step (e), 0.1 to 0.4 parts by weight of 1 to 5 wt% of calcium chloride aqueous solution and 0.5 to 2 parts by weight of hyaluronic acid may be added. And may further include additional.

The therapeutic composition of the present invention can be administered parenterally such as injection at the time of clinical administration and can be used in the form of a general pharmaceutical preparation. That is, the therapeutic composition of the present invention can be administered in various forms of parenteral administration at the time of clinical administration. In the case of formulation, a diluent such as a filler, an extender, a binder, a wetting agent, a disintegrating agent and a surfactant, . ≪ / RTI >

The preparation may contain a sterilized aqueous solution, a non-aqueous solution, a suspending agent, an emulsion, a freeze-drying agent, and the like. Examples of the non-aqueous solvent and the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As the base of the suppository, witepsol, macrogol, tween 61, cacao paper, laurin, glycerol and gelatin can be used.

In the method of using the therapeutic composition of the present invention, factors affecting the amount to be administered include, but are not limited to, the dosage regimen, the frequency of administration, the specific disease undergoing therapy, the severity of the disease, And whether the cooperative therapy is in progress, and the age, height, weight, health, and physical condition of the individual undergoing treatment. Generally, as the body weight of a patient undergoing treatment increases, it is preferable to administer the above-described ratio of therapeutic composition preparation in a larger amount.

The therapeutic composition of the present invention can be administered in an amount effective to stimulate cartilage regeneration. For the treatment of meniscal damage or degenerative arthritis, each of the above components is divided into halves, for example, once into the anterior lateral of the knee and once into the anterior medial.

The ratio of the therapeutic composition of the present invention is based on the results of the treatment for the patients desired by the inventor's clinic.

[Experimental Example 1]

Substrate blood vessel fraction  And Extracellular matrix  Obtain

Lipid fractionation and extracellular matrix acquisition were performed.

First, the patient was transferred to the operating room and laid down flat. Then, the area to be aspirated into the lower abdomen was disinfected with a provodine-iodine, covered with a sterilized cloth, and then incised about 5 cm below the navel and about 0.5 cm on both sides. And the lower abdomen was localized anesthetized using a solution of thiamine (500 cc NS + 2% Lidocaine 40 cc + 0.5% Marcaine 20 cc + Epinephrine 1: 1000 0.5 cc). Then a fat tissue of about 120 cc was obtained using a 3.0 Hartmann tube. The adipose tissue was centrifuged at 1000 x g-force for 5 minutes. As a result, a concentrated fat extract of about 60 cc was obtained as shown in Fig.

The concentrated fat extract and collagenase, which is a digestive enzyme, were mixed at a ratio of 1: 1 and the enzyme reaction was carried out for 30 minutes while stirring at 37 ° C. Then, the substrate blood vessel fraction containing stem cells was centrifuged for 4 minutes at 300 × g- Separated as shown in FIG.

Subsequently, the fat extract remaining after the separation of the matrix blood vessels was ground using a homogenizer (manual or mechanical) as shown in FIG. 3, and then mixed again with the substrate blood vessel fraction as shown in FIG. 4, The solution was removed.

The enzyme was then removed by washing three times with 500 cc of D5SR (Dextrose 5% Lactated Ringer). After each wash procedure, the fat extract was centrifuged at 300 x g-force. After the final centrifugation, a stromal blood vessel fraction containing approximately 0.1 to 5 g of extracellular matrix and 8 g of stem cells was obtained as shown in Fig. 5

[Experimental Example 2]

Knee joint procedure

In order to inject a composition for treating a joint disease including bone and cartilage of the present invention into a knee joint, the patient was placed on the floor and the right knee was placed at an angle of 90 degrees. The knee injection site was disinfected with beta-diene, cleanly covered with sterilized cloth, and anesthetized with 2% lidocaine. Using a 18-gauge 1/2-inch needle, a composition for treating joint diseases, including bone and cartilage, was injected into the anterior-medial region of the knee. After injection, the patient was instructed to maintain a floating posture for 1 hour for cell attachment.

[Experimental Example 3]

Femur part  Surgery

In order to inject the composition for treating joint diseases including bone and cartilage of the present invention into the femur part, the patient was laid face up. After disinfection with beta-diene and cleaning with a sterile cloth, the thigh femur was anesthetized with 2% lidocaine. The composition was injected into the femur with a 16-gauge 3½ needle under the guidance of an ultrasound. The legs were then raised and allowed to remain for 1 hour for attachment of the patient's cells.

[Example 1]

SVF + ECM composition Arthritis of the knee  Cartilage regeneration therapy

The substrate blood vessel fraction and extracellular matrix obtained in Experimental Example 1 were made into three kinds of compositions (A, B, and C) at the same dosage as shown in Table 1 below, .

The functional rating index (FRI) and visual analogue scale (VAS) were used to assess the treatment effects of the subjects before and after the procedure.

The functional grade index measures the body problems that are detected during daily activities and classifies the degree by each item. In addition, visual analogue scale is one of the most commonly used methods for measuring pain intensity. In this study, the visual analogue scale measures pain scores from 0 to 10, indicating that the lower the number, the less pain the subject felt And the results are shown in Table 1 below.

No. SVF ECM Result measuring means Before the procedure after 2 weeks After 4 weeks A 8g 0.1 g FRI 28 28 27 VAS 8 8 7 B 8g 1g FRI 26 23 22 VAS 7 6 6 C 8g 5g FRI 22 16 10 VAS 6 3 One

According to the above Table 1, in the case of the composition A, the pain of the patient was alleviated by 10% or more after 4 weeks of the procedure, and in the case of the composition B, the pain of the patient was alleviated by 10% or more after 2 weeks of the procedure.

Compared with the compositions A and B, it was found that in the case of the composition C, the patient's pain was alleviated by 50% after 2 weeks and by 80% after 4 weeks.

Also FIG. 4, which is an MRI photograph before and 12 hours after the administration of the composition C, shows that the thickness of the vitreous cartilage and fibrocartilage inside the right knee significantly increased after 12 weeks compared to that before the injection.

[Comparative Example 1]

SVF + ECM composition Arthritis of the knee  Cartilage regeneration therapy

In order to compare with Example 1, a composition comprising 16 g of substrate blood vessel fraction obtained in the same manner as in Experimental Example 1 and 10 g of extracellular matrix was applied to the knee joint in the same manner as Experimental Example 2, The results were as shown in Table 2 and Table 3 below.

Result measuring means Before the procedure after 2 weeks After 4 weeks After 12 weeks FRI 38 22 17 14 VAS 8 5 3 One

Physiotherapy
- Scope of Activity Bend (deg) Bending VAS Stretch (deg) Stretch VAS Before the procedure 94 9 +3 1/10 Two weeks after the procedure 105 6 +3 1/10 Four weeks after the procedure 114 4 +4 0/10 After 12 weeks of procedure 129 One +5 0/10

According to the above Tables 2 and 3, the patient's pain was alleviated by more than 30% after 2 weeks of the procedure and the knee bending was similarly improved. After 4 weeks of treatment, the patient's pain was reduced by more than 60% and the range of activity was further improved.

Comparing the results with Example 1, the composition of Comparative Example 1, which contained 16 g of substrate blood vessel fraction and 10 g of extracellular matrix, showed that composition C containing 8 g of substrate blood vessel fraction and 5 g of extracellular matrix, And there is a problem in that the larger the amount of the composition is, the more the patient feels uncomfortable. Thus, the composition C of Example 1 is the most preferable from the viewpoint of efficiency.

[Example 2]

SVF + ECM + PRP  Composition Arthritis of the knee  Cartilage regeneration therapy

Platelet Separation Plasma Preparation

30 cc of autologous blood was drawn from an anticoagulant acid citrate dextrose (ACD) anticoagulant of 2.5 cc and centrifuged at 200 × g-force for 15 minutes. After the first centrifugation, the supernatant was subjected to a second centrifugation for 5 minutes at 1000 x g-force.

After the second centrifugation, the supernatant was discarded and 2 g of the double layered microparticles (leukocyte, platelet thin layer on the left side of the neglected blood surface) were mixed with 8 g of the substrate blood vessel fraction obtained according to Experimental Example 1 and 5 g of the extracellular matrix . Table 4 and Table 5 show the results of evaluating the functional class index and visual similarity scale in order to evaluate the degree of pain of the subjects after the treatment composition was applied to the knee joint in the same manner as Experimental Example 2. [

Result measuring means Before the procedure after 2 weeks After 4 weeks After 12 weeks FRI 37 20 16 13 VAS 8 5 4 One

Physiotherapy
- Scope of Activity Bend (deg) Bending VAS Stretch (deg) Stretch VAS Before the procedure 100 8 +3 1/10 Two weeks after the procedure 110 4 +4 0/10 Four weeks after the procedure 130 3 +5 0/10 After 12 weeks of procedure 130 2 +5 0/10

According to the above Tables 4 and 5, the pain of the patient was alleviated by more than 30% after 2 weeks of the procedure, and the bending of the knee was similarly improved. After 12 weeks of treatment, the patient's pain was reduced by more than 80% and the range of activity was further improved.

7, which is an MRI photograph before and 12 weeks after the procedure, the thickness of the inner meniscus of the knee and the cartilage of the vitreous was significantly increased compared with that before the procedure. This indicates that the composition of Example 1 The difference is greater than the difference shown.

[Example 3]

SVF + ECM + PRP  Composition femoral head Avascular necrosis Bone regeneration  cure

The same composition as in Example 2 was used for the treatment of avascular necrosis of femoral head in the same manner as Experimental Example 3. After evaluating the degree of pain of the subjects, the functional class index and the visual superiority scale were evaluated. As shown in Table 7.

Result measuring means Before the procedure after 2 weeks After 4 weeks After 12 weeks FRI 26 21 14 8 VAS 8 4 4 2

Physiotherapy
- Scope of Activity Bend (deg) Bending VAS Abduction (deg) Abduction VAS Civil war (deg) Civil War VAS Before the procedure 90 6 19 7 9 8 Two weeks after the procedure 110 3 35 4 15 4 Four weeks after the procedure 115 2 37 3 18 3 After 12 weeks of procedure 125 2 40 2 20 2

According to the above Tables 6 and 7, the pain of the patient was alleviated by more than 50% after 2 weeks of operation, the pain of the patient was improved by more than 70% after 12 weeks of operation, and the activity range was further improved.

8, which is an MRI photograph taken before and 12 weeks after the procedure, it can be seen that defects in the upper bony bone are significantly reduced after 12 weeks of operation compared to before the procedure. Thus, bone regeneration in the upper glenohumeral and enhancement of the bone matrix in the subcortical region of the femur ball significantly increased.

[Example 4]

SVF + ECM + PRP + CaCl ₂ + Hyaluronic  acid composition Arthritis of the knee  Cartilage regeneration therapy

8 g of substrate blood vessel fraction obtained in Experimental Examples 1 and 2, 5 g of extracellular matrix, 2 g of platelet-rich plasma, 0.2 g of 3 wt% calcium chloride aqueous solution and 1 g of hyaluronic acid were further mixed to prepare a therapeutic composition The results are shown in Table 8 and Table 9, respectively.

Result measuring means Before the procedure after 2 weeks After 4 weeks After 12 weeks FRI 37 20 16 13 VAS 8 4 3 One

Physiotherapy
- Scope of Activity Bend (deg) Bending VAS Stretch (deg) Stretch VAS Before the procedure 95 9 +3 1/10 Two weeks after the procedure 110 5 +4 0/10 Four weeks after the procedure 115 4 +4 0/10 After 12 weeks of procedure 130 One +5 0/10

According to Tables 8 and 9, after 2 weeks of treatment, the patient's pain was alleviated by 50% or more and the knee flexion was similarly improved. After 12 weeks of treatment, the patient's pain was reduced by 80% or more, and the range of activity was further improved.

9, which is an MRI photograph before and 12 weeks after the procedure, shows that the thickness of the meniscus and vitreous cartilage in the right knee was increased after 12 weeks of operation compared with before the procedure, .

[Comparative Example 2]

SVF + ECM + PRP + CaCl ₂ + Hyaluronic  acid composition Arthritis of the knee  Cartilage regeneration therapy

For comparison with Example 4 above, 4 g of substrate blood vessel fraction obtained according to Experimental Example 1 and 2, 2 g of extracellular matrix, 0.1 g of 3 wt% calcium chloride aqueous solution and 2 g of hyaluronic acid in 1 g of platelet- Were mixed with each other to prepare a therapeutic composition. The results are shown in Tables 10 and 11, respectively.

Result measuring means Before the procedure after 2 weeks After 4 weeks After 12 weeks FRI 38 38 36 24 VAS 7 7 7 4

Physiotherapy
- Scope of Activity Bend (deg) Bending VAS Stretch (deg) Stretch VAS Before the procedure 104 7 +4 1/10 Two weeks after the procedure 105 7 +4 1/10 Four weeks after the procedure 104 7 +4 0/10 After 12 weeks of procedure 120 4 +4 0/10

According to Tables 10 and 11 above, patient's pain was improved by more than 40% after 12 weeks of operation, and knee flexion improved somewhat after 4 weeks of treatment.

Comparing the above results with Example 4, it was found that the treatment effect was poor when a smaller amount of substrate blood vessel fraction, extracellular matrix, platelet rich plasma and aqueous calcium chloride solution were contained than the therapeutic composition of Example 4 of the present invention .

[Example 5]

SVF + ECM + PRP + CaCl ₂ + Hyaluronic  acid composition Femoral head Avascular necrosis Bone regeneration therapy

The same composition as in Example 4 was used for the treatment of osteonecrosis of femoral head in the same manner as Experimental Example 3. After evaluating the degree of pain of the subjects, the functional grade index and the visual superiority scale were evaluated. 13.

Result measuring means Before the procedure after 2 weeks After 4 weeks After 12 weeks FRI 38 21 18 11 VAS 8 5 4 One

Physiotherapy
- Scope of Activity Bend (deg) Bending VAS Abduction (deg) Abduction VAS Civil war (deg) Civil War VAS Before the procedure 92 8 20 7 8 8 Two weeks after the procedure 112 5 34 4 15 4 Four weeks after the procedure 115 4 35 4 17 4 After 12 weeks of procedure 125 2 40 One 19 2

According to the above Tables 12 and 13, the pain of the patient was alleviated by more than 30% after 2 weeks of the procedure, and the pain of the patient was improved by more than 80% after 12 weeks of the procedure.

FIG. 10, which is an MRI photograph before and 12 weeks after the procedure, shows that the defect of the bone in the upper zygote is significantly reduced after 12 weeks of treatment compared with that before the procedure. This shows that the composition of Example 3 is shown in FIG. 5 Which is larger than the difference.

The results of the experiments of Examples 1 to 5 as described above show that the composition comprising the substrate blood vessel fraction, extracellular matrix, platelet rich plasma, aqueous calcium chloride solution and hyaluronic acid of the present invention can be used for bone and osteoarthritis such as degenerative arthritis and femoral head avascular necrosis It is very effective in the treatment of joint diseases including cartilage.

The MRI results of the patients showed that the composition of the present invention significantly changed the bone matrix size in the knee cartilage and the femoral cortex and that the measured physiotherapy results, pain index and functional grade index were all improved I could.

Also, the results of the tests of Comparative Examples 1 and 2 show that the therapeutic effect is significantly lower when the amount of the composition of the present invention is less than that of the composition of the present invention, and the therapeutic effect is similar when a large amount is administered. However, as the dose increases, the patient tends to feel uncomfortable and is not economically advantageous.

Therefore, when a composition for treating a joint disease including a bone marrow fraction, an extracellular matrix, a platelet rich plasma, an aqueous solution of calcium chloride, and a bone and cartilage containing hyaluronic acid proposed in the present invention is used, It is possible to treat joint diseases including bone and cartilage.

Claims (10)

In 100 parts by weight of a composition for treating cartilage damage comprising degenerative arthritis,
8 parts by weight of a stromal vascular fraction (SVF) and 5 parts by weight of an extra cellular matrix (ECM)
The composition for treating cartilage damage comprising degenerative arthritis, wherein the matrix blood vessel fraction is obtained by centrifuging after treating the adipose tissue with collagenase. The method according to claim 1,
The composition for treating cartilage damage comprising degenerative arthritis, wherein the composition further comprises 1 to 10 parts by weight of platelet rich plasma (PRP). 3. The method of claim 2,
Wherein the platelet-rich plasma is derived from autologous blood. The method according to claim 1,
Wherein the composition further comprises 0.1 to 0.4 parts by weight of calcium chloride aqueous solution (CaCl2) of 1 to 5 wt% and 0.5 to 2 parts by weight of hyaluronic acid (hyaluronic acid). The method according to claim 1,
A composition for treating cartilage damage comprising degenerative arthritis, wherein the matrix blood vessel fraction and the extracellular matrix are derived from a human. (a) mixing and stirring the adipose tissue obtained by liposuction with collagenase at a ratio of 1: 1 to 1: 2 to conduct an enzymatic reaction, and then extracting a stromal blood vessel fraction containing stem cells ;
(b) extracting and separating a stromal blood vessel fraction containing the stem cells, pulverizing the remaining fat extract, mixing the extracted blood vessel fraction again, and removing oil and tumescent solution ;
(c) washing the fat extract from which the oil and the citrate solution have been removed to remove the enzyme;
(d) centrifuging the fat extract after the washing step to obtain a substrate blood vessel fraction and an extracellular matrix; And
(e) mixing 5-10 parts by weight of the substrate blood vessel fraction with 0.1-10 parts by weight of an extracellular matrix. The method according to claim 6,
The method for preparing a composition for treating cartilage damage comprising degenerative arthritis, wherein the composition further comprises 1 to 10 parts by weight of platelet-rich plasma in step (e). 8. The method of claim 7,
Wherein said platelet-rich plasma is derived from autologous blood. ≪ RTI ID = 0.0 > 11. < / RTI > The method according to claim 6,
The composition for treating cartilage damage comprising degenerative arthritis according to claim 1, wherein in step (e), 0.1 to 0.4 parts by weight of 1 to 5 wt% of calcium chloride aqueous solution and 0.5 to 2 parts by weight of hyaluronic acid are further mixed. Gt; The method according to claim 6,
Wherein the matrix blood vessel fraction and the extracellular matrix are derived from a human.

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