Traditional Chinese medicine preparation for treating knee osteoarthritis as well as preparation method and application thereof

Abstract

The invention provides a traditional Chinese medicine preparation for treating knee osteoarthritis and a preparation method and application thereof, and belongs to the technical field of stem cell preparations. The traditional Chinese medicine preparation comprises the following components: prepared rehmannia root, astragalus membranaceus, angelica sinensis, achyranthes bidentata, salvia miltiorrhiza, ligusticum wallichii, divaricate saposhnikovia root, fried spina date seed, earthworm, caulis spatholobi, safflower carthamus, lycopodium clavatum, rhizoma drynariae, radix paeoniae alba, paniculate swallowwort root and radix angelicae pubescentis. The traditional Chinese medicine preparation and the stem cells are combined for use, the unique advantages of the traditional Chinese medicine preparation in the aspect of treating knee osteoarthritis are exerted, and meanwhile the traditional Chinese medicine preparation can effectively promote differentiation and proliferation of cartilage cells. When being combined with stem cells, the composition can effectively promote the stem cells to be differentiated into cartilage cells, so as to achieve a treatment effect on knee osteoarthritis.

Description

Technical Field

[0001] The present invention belongs to the technical field of stem cell preparations, and in particular relates to a traditional Chinese medicine preparation for treating knee osteoarthritis, and a preparation method and application thereof. Background Art

[0002] Knee osteoarthritis is a chronic joint disease characterized by cartilage degeneration and destruction, as well as bone hyperplasia. It is more common among middle-aged and elderly individuals, and its prevalence is increasing annually with the aging population. Statistics show that the prevalence of knee osteoarthritis exceeds 50% among people over 65 years old, severely impacting knee joint function and quality of life, placing a heavy burden on patients, their families, and society.

[0003] Currently, there are various clinical treatments for knee osteoarthritis, including medication, surgery, and physical therapy, but all have limitations. Commonly used medications for knee osteoarthritis include nonsteroidal anti-inflammatory drugs (NSAIDs) (such as ibuprofen and diclofenac sodium), which primarily relieve pain and inflammation. However, long-term use may cause side effects such as gastrointestinal discomfort and liver and kidney damage, and cannot prevent the progression of the disease. While hyaluronic acid intra-articular injection therapy can improve joint lubrication and relieve symptoms to a certain extent, the efficacy is short-lived and requires multiple injections, which is inconvenient for patients. For patients with severe disease, surgical treatment such as arthroscopic surgery, osteotomy, and total joint replacement may be required. Although surgical treatment can effectively improve joint function, it is invasive, expensive, and carries risks such as infection and prosthesis loosening, making it not suitable for all patients. Physical therapy, such as hot compresses, massage, and rehabilitation training, can be used as an adjunct treatment to help relieve symptoms and improve joint mobility, but its efficacy is limited when used alone, making it difficult to achieve the desired therapeutic effect. Therefore, finding a safe, effective treatment method that can fundamentally improve the condition is of great clinical significance for the treatment of knee osteoarthritis. Summary of the Invention

[0004] In light of this, the present invention aims to provide a composition for treating knee osteoarthritis, its preparation method, and its use. This invention combines a traditional Chinese medicine preparation with stem cells. This preparation not only promotes the differentiation of stem cells into chondrocytes, but also promotes the differentiation and proliferation of the chondrocytes themselves, achieving a therapeutic effect on knee osteoarthritis.

[0005] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0006] The invention provides a traditional Chinese medicine preparation for treating knee osteoarthritis. The traditional Chinese medicine preparation comprises the following components in parts by weight: 10-15 parts of prepared rehmannia root, 10-20 parts of astragalus root, 10-15 parts of angelica root, 10-15 parts of achyranthes bidentata, 10-20 parts of salvia miltiorrhiza, 10-15 parts of chuanxiong rhizome, 10-15 parts of siler, 10-15 parts of stir-fried spiny jujube seeds, 5-15 parts of earthworm, 10-20 parts of millettia reticulata, 10-20 parts of safflower, 10-20 parts of striata herb, 10-20 parts of drynaria rhizome, 5-15 parts of white peony root, 5-15 parts of cynanchum wilfordii and 5-15 parts of angelica dahurica.

[0007] The present invention provides a method for preparing the traditional Chinese medicine preparation, comprising the following steps:

[0008] (2.1) mixing the components of the traditional Chinese medicine preparation to obtain a mixture;

[0009] (2.2) The mixture is mixed with water, decocted 1 to 3 times, filtered, the filtrate is collected, the filtrates are combined, concentrated, dried, and sterilized to obtain a traditional Chinese medicine preparation.

[0010] Preferably, the mass ratio of the mixture to water is 1:3-7.

[0011] Preferably, the decocting time is 15 to 40 minutes per time, and the decocting temperature is 90 to 100°C.

[0012] Preferably, the concentration is reduced-pressure concentration, the concentration temperature is 35-55° C., and the volume after concentration is 2 / 5-3 / 5 of the original volume.

[0013] Preferably, the sterilization is performed by radiation sterilization using cobalt 60, and the sterilization time is 20 to 28 hours.

[0014] The present invention also provides the use of the traditional Chinese medicine preparation or the traditional Chinese medicine preparation obtained according to the preparation method in preparing a drug for treating knee osteoarthritis, wherein the traditional Chinese medicine preparation is used in combination with stem cells.

[0015] Preferably, when the Chinese medicine preparation is used in combination with stem cells, the dosage of the Chinese medicine preparation is 1 to 3 g / day, and the concentration of the stem cells is 1 to 5×10 5 / mL, the dosage is 0.5-2mL / week.

[0016] Preferably, the stem cells are bone marrow mesenchymal stem cells.

[0017] Preferably, the dosage form of the traditional Chinese medicine preparation is a powder, and the dosage form of the stem cells is an injection.

[0018] Compared with existing technologies, the present invention has the following beneficial effects: By combining a traditional Chinese medicine preparation with stem cells, the present invention leverages the unique advantages of traditional Chinese medicine preparations in treating knee osteoarthritis while effectively promoting the differentiation and proliferation of chondrocytes. Combined with stem cells, it effectively promotes the differentiation of stem cells into chondrocytes, achieving a therapeutic effect on knee osteoarthritis. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 The effect of different concentrations of Chinese herbal medicine preparations on chondrocyte proliferation;

[0020] Figure 2 To investigate the effects of different Chinese herbal medicines on the differentiation of bone marrow mesenchymal stem cells into chondrocytes;

[0021] Figure 3 The effects of different treatment groups on bone tissue imaging in KOA rats;

[0022] Figure 4 The effects of different treatment groups on the pathological changes of articular cartilage in KOA rats. DETAILED DESCRIPTION

[0023] The present invention provides a traditional Chinese medicine preparation for treating knee osteoarthritis, comprising the following components in parts by weight:

[0024] 10-15 parts of Rehmannia root, preferably 11-14 parts, more preferably 13 parts;

[0025] 10-20 parts of Astragalus, preferably 13-17 parts, more preferably 15 parts;

[0026] 10-15 parts of Chinese Angelica sinensis, preferably 11-14 parts, more preferably 13 parts;

[0027] 10-15 parts of Achyranthes bidentata, preferably 11-14 parts, more preferably 13 parts;

[0028] 10-20 parts of Salvia miltiorrhiza, preferably 12-18 parts, more preferably 15 parts;

[0029] 10-15 parts of Chuanxiong, preferably 11-14 parts, more preferably 13 parts;

[0030] 10-15 parts of windproof, preferably 11-14 parts, more preferably 12 parts;

[0031] 10-15 parts of stir-fried Chinese date seeds, preferably 11-14 parts, more preferably 13 parts;

[0032] 5-15 parts of earthworms, preferably 7-13 parts, more preferably 10 parts;

[0033] 10-20 parts of Millettia reticulata, preferably 12-18 parts, more preferably 15 parts;

[0034] 10-20 parts of safflower, preferably 12-18 parts, more preferably 16 parts;

[0035] 10-20 parts of Herba Lycopodii Herba, preferably 12-18 parts, more preferably 15 parts;

[0036] 10-20 parts of Drynaria rhizome, preferably 12-18 parts, more preferably 16 parts;

[0037] 5-15 parts of white peony root, preferably 8-12 parts, more preferably 10 parts;

[0038] 5-15 parts of Cynanchum wilfordii, preferably 11-14 parts, more preferably 12 parts;

[0039] 5 to 15 parts of Angelica dahurica, preferably 11 to 14 parts, more preferably 13 parts.

[0040] In the present invention, there is no particular limitation on the source of the raw materials, and conventional commercially available products can be used. In the present invention, Rehmannia root is sweet and slightly warm in nature, enters the liver and kidney meridians, and contains ingredients such as catalpol, rehmannia root, and polysaccharides, and has the effects of enhancing immune function, improving hematopoietic function, anti-oxidation, and regulating endocrine function.

[0041] Astragalus is sweet and slightly warm in nature, and enters the spleen and lung meridians. It has the effects of replenishing qi and raising yang, strengthening the exterior and stopping sweating, promoting diuresis and reducing swelling, promoting fluid production and nourishing blood, relieving stagnation and numbness, expelling toxins and pus, and healing sores and promoting tissue regeneration.

[0042] Angelica sinensis is sweet, pungent and warm in nature, and enters the liver, heart and spleen meridians. It has the functions of nourishing blood, activating blood circulation, regulating menstruation, relieving pain and moistening the intestines.

[0043] Achyranthes bidentata is bitter, sweet, sour and flat in nature and enters the liver and kidney meridians. It has the effects of promoting blood circulation, tonifying the liver and kidneys, strengthening tendons and bones, promoting diuresis and relieving stranguria, and guiding blood (fire) downward.

[0044] Danshen is bitter and slightly cold in nature, and enters the heart and liver meridians. It has the effects of promoting blood circulation and removing blood stasis, relieving pain through menstruation, cooling blood and eliminating carbuncle, and relieving restlessness and calming the mind.

[0045] Chuanxiong is pungent and warm in nature, and enters the liver, gallbladder, and pericardium meridians. It has the functions of activating blood circulation and promoting qi, and is good at "going up to the head, going down to the sea of ​​blood, and relieving depression in the middle";

[0046] Saposhnikovia root is pungent, sweet, and slightly warm in nature. It enters the bladder, liver, and spleen meridians and has the effects of dispelling wind and relieving exterior symptoms, removing dampness and relieving pain, and stopping spasms.

[0047] Stir-fried Chinese jujube seeds are sweet, sour and flat in nature, and enter the liver, gallbladder and heart meridians. They have the effects of nourishing the liver and heart, astringing yin blood, astringing sweat and promoting body fluid.

[0048] Earthworms are salty and cold in nature, and enter the liver, spleen, and bladder meridians. They are good at moving around, can promote bowel movements, and have the effects of clearing heat, unblocking meridians, relieving asthma, and promoting urination.

[0049] Millettia reticulata is bitter, sweet and warm in nature and enters the liver and kidney meridians. It has the effects of promoting blood circulation, unblocking collaterals, and replenishing blood. It is suitable for the symptoms of "blood deficiency and stasis" or "blood stasis and deficiency".

[0050] Safflower is pungent and warm in nature, and enters the heart and liver meridians. It has the effects of promoting blood circulation, relieving menstruation, dispersing blood stasis and relieving pain.

[0051] The herb is pungent, bitter and warm in nature and enters the liver, spleen and kidney meridians. It has the effects of promoting blood circulation, dispelling wind and dampness, reducing swelling and relieving pain.

[0052] Drynaria fortunei is bitter and warm in nature, and enters the liver and kidney meridians. It has the effects of nourishing the kidneys and strengthening bones, promoting blood circulation and healing wounds, and relieving pain and stopping bleeding.

[0053] White peony root is sweet, sour, and slightly cold in nature. It enters the liver and spleen meridians and has the effects of nourishing blood and regulating menstruation, restraining yin and stopping sweating, softening the liver and relieving pain, and calming liver yang.

[0054] Cynanchum wilfordii is pungent and warm in nature, and enters the liver and stomach meridians. It has the effects of dispelling wind and dampness, dredging meridians and relieving pain, promoting blood circulation and detoxification, reducing swelling and itching, and relieving cough and asthma.

[0055] Duhuo tastes pungent, bitter and slightly warm, and enters the kidney and bladder meridians. It has the effects of dispelling wind and dampness, relieving numbness and relieving pain.

[0056] The present invention provides a method for preparing the traditional Chinese medicine preparation, comprising the following steps:

[0057] (2.1) mixing the components of the traditional Chinese medicine preparation to obtain a mixture;

[0058] (2.2) The mixture is mixed with water, decocted 1 to 3 times, filtered, the filtrate is collected, the filtrates are combined, concentrated, dried, and sterilized to obtain a traditional Chinese medicine preparation.

[0059] In the present invention, the components of the traditional Chinese medicine preparation are mixed to obtain a mixture; in the present invention, there is no limitation on the mixing method, and the components of the traditional Chinese medicine preparation can be mixed uniformly in a conventional manner.

[0060] In the present invention, the mixture is mixed with water, decocted 1 to 3 times, filtered, the filtrate is taken, the filtrates are combined, concentrated, dried, and sterilized to obtain a traditional Chinese medicine preparation; the mass ratio of the mixture to water is 1:3 to 7, preferably 1:4 to 6, and more preferably 1:5; the decocting time is 15 to 40 minutes per time, preferably 20 to 35 minutes, and more preferably 30 minutes, and the decocting temperature is 90 to 100°C; 4 layers of gauze are preferably used for filtration; the concentration is reduced pressure concentration, the concentration temperature is 35 to 55°C, preferably 40 to 50°C, and more preferably 45°C, and the volume after concentration is 2 / 5 to 3 / 5 of the original volume, preferably 9 / 20 to 11 / 20, and more preferably 1 / 2; the drying is preferably carried out by freeze drying; the sterilization is carried out by cobalt 60 radiation sterilization, and the sterilization time is 20 to 28 hours, preferably 22 to 26 hours, and more preferably 24 hours.

[0061] The present invention also provides the use of the traditional Chinese medicine preparation or the traditional Chinese medicine preparation obtained according to the preparation method in preparing a drug for treating knee osteoarthritis, wherein the traditional Chinese medicine preparation is used in combination with stem cells.

[0062] In the present invention, when the Chinese medicine preparation is used in combination with stem cells, the dosage of the Chinese medicine preparation is 1 to 3 g / day, and the concentration of the stem cells is 1 to 5×10 5 / mL, the usage is 0.5-2mL / week; the stem cells are bone marrow mesenchymal stem cells; the dosage form of the traditional Chinese medicine preparation is powder, and the dosage form of the stem cells is injection.

[0063] The technical solutions provided by the present invention are described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0064] Example 1

[0065] Weigh 130g of Rehmannia glutinosa, 150g of Astragalus membranaceus, 130g of Angelica sinensis, 130g of Achyranthes bidentata, 130g of Salvia miltiorrhiza, 130kg of Chuanxiong, 120g of Saposhnikovia divaricata, 130g of stir-fried Semen Ziziphi Spinosae, 100g of Earthworm, 150g of Millettia reticulata, 160g of Carthamus tinctorius, 150g of Herba Lycopodii, 160g of Rhizoma Drynariae, 100g of White Peony Root, 120g of Cynanchum chinense, and 130g of Angelica dahurica, mix well, add 10.15kg of water, boil over high heat, simmer for 30min over low heat, filter through 4 layers of gauze, take the filtrate for standby use, add 10.15kg of water to the filter residue and decoct again in the same way, filter through 4 layers of gauze, combine the two filtrates, concentrate under reduced pressure at 45°C to 1 / 2 of the original volume, freeze-dry, obtain powder, and sterilize by radiation with cobalt 60 for 24h to obtain a traditional Chinese medicine preparation.

[0066] Example 2

[0067] Weigh 100 g of Rehmannia root, 100 g of Astragalus root, 100 g of Angelica sinensis, 100 g of Achyranthes bidentata, 100 g of Salvia miltiorrhiza, 100 g of Chuanxiong, 100 g of Saposhnikovia divaricata, 100 g of stir-fried Semen Ziziphi spinulosae, 50 g of Earthworm, 100 g of Millettia reticulata, 100 g of Carthamus tinctorius, 100 g of Herba Lycopodii herba, 100 g of Drynaria fortunei, 50 g of Paeonia lactiflora, 50 g of Cynanchum chinense, and 50 g of Angelica dahurica, mix well, add 4.2 kg of water, boil over high heat, simmer for 20 min over low heat, filter through 4 layers of gauze, take the filtrate for later use, add 4.2 kg of water to the filter residue and decoct again in the same way, filter through 4 layers of gauze, combine the two filtrates, concentrate under reduced pressure at 45 ° C to 3 / 5 of the original volume, freeze-dry, obtain a powder, and sterilize by cobalt 60 radiation for 20 h to obtain a traditional Chinese medicine preparation.

[0068] Example 3

[0069] Weigh 150g of Rehmannia glutinosa, 200g of Astragalus membranaceus, 150g of Angelica sinensis, 150g of Achyranthes bidentata, 200g of Salvia miltiorrhiza, 150g of Ligusticum chuanxiong, 150g of Saposhnikovia divaricata, 150g of stir-fried Semen Ziziphi spinulosae, 150g of Pheretima wormwood, 200g of Millettia reticulata, 200g of Carthamus tinctorius, 200g of Herba Lycopodii herba, 200g of Rhizoma Drynariae, 150g of Paeonia lactiflora, 150g of Cynanchum chinense, and 150g of Angelica dahurica, mix well, add 18.9kg of water, boil over high heat, simmer for 40min, filter through 4 layers of gauze, take the filtrate for standby use, add 18.9kg of water to the filter residue and decoct again in the same way, filter through 4 layers of gauze, combine the two filtrates, concentrate under reduced pressure at 45°C to 2 / 5 of the original volume, freeze-dry, obtain powder, and perform radiation sterilization with cobalt 60 for 28h to obtain a traditional Chinese medicine preparation.

[0070] Comparative Example 1

[0071] The Chinese medicine preparation was prepared in the same manner as in Example 1, except that Saposhnikovia divaricata was replaced with Asarum sieboldii.

[0072] Comparative Example 2

[0073] The Chinese medicine preparation was prepared in the same manner as in Example 1, except that safflower was removed and the missing amount was supplemented with Achyranthes bidentata.

[0074] Experimental Example 1

[0075] The Chinese medicine preparation obtained in Example 1 was used as a sample, and sterile water was used to prepare solutions with concentrations of 0.2 μg / mL, 10 μg / mL, 25 μg / mL, 50 μg / mL, and 100 μg / mL under aseptic conditions. 4100 μL of Chinese medicine preparations of different concentrations were added to each well of the 96-well plate and incubated in a constant temperature incubator at 37°C and 5% CO2. After the cells adhered to the wall, 100 μL of Chinese medicine preparations of different concentrations were added. The wells without Chinese medicine preparations were used as controls. After incubation at 37°C and 5% CO2 for 24 hours, 10 μL of CCK-8 solution was added to the 96-well plate and incubated in a constant temperature incubator at 37°C and 5% CO2 for 4 hours. The absorbance value at a wavelength of 450 nm was detected by a microplate reader. The results are shown in the table. Figure 1 .

[0076] Depend on Figure 1 It can be seen that the traditional Chinese medicine preparation obtained by the present invention can significantly promote the proliferation of chondrocytes, and the promotion effect is the best when the concentration is 50 μg / mL.

[0077] Experimental Example 2

[0078] The traditional Chinese medicine preparations obtained in the manner of Example 1 and Comparative Examples 1 and 2 were used as samples. Sterile water was used under aseptic conditions to prepare solutions with a concentration of 50 μg / mL, which were used as induction solutions. 5 Cells were seeded in 6-well plates and 100 μL of different induction solutions were added to each well. Wells without induction solution were used as controls and covered with coverslips for cell climbing. The cells were cultured in a 37°C, 5% CO2 incubator for 24 hours and then digested and harvested using 0.25% trypsin. After 48 hours of digestion, the cells were washed 1-2 times with PBS and resuspended in serum-free medium to adjust the cell density to 1×10 5 / mL. Take 200mL of cell suspension and add it to the Transwell chamber. Add 600μL of culture medium containing 10% FBS to the lower chamber of the 24-well plate and culture at 37℃ for 24h. Remove the Transwell chamber and add 4% tissue fixative to fix it. Observe and take pictures under a microscope. The results are shown in the figure. Figure 2 .

[0079] Depend on Figure 2 As can be seen, the cells in the control group were long, spindle-shaped, flattened, with long cell bodies and protrusions, showing the original morphology of bone marrow mesenchymal stem cells. The cells using Example 1 as the induction solution increased in volume, became polygonal, and had multiple protrusions. The cells using Comparative Examples 1 and 2 as the induction solution showed little change in volume, and the cell bodies remained spindle-shaped with fewer protrusions. This indicates that the traditional Chinese medicine preparation obtained by the method of Example 1 of the present application can effectively promote the differentiation of bone marrow mesenchymal stem cells into chondrocytes.

[0080] Experimental Example 3

[0081] Fifty-four healthy SPF-grade 8-week-old male SD rats (purchased from Hubei Provincial Center for Disease Control and Prevention) with a body weight of 200-225 g were selected. After 1 week of adaptation, 6 of them were not treated (control group), and the remaining 48 rats were used to prepare a rat model of knee osteoarthritis according to the modified Hulth method (Rao Qi, Guan Junhang, Chen Dong. Modified Hulth method combined with glutaraldehyde injection to establish a rat knee osteoarthritis model [J]. Journal of Clinical Orthopaedics, 2022, 25(03): 440-443.). The rats with successful model construction were randomly divided into 8 groups, each with 6 rats. Groups 1 to 3 were gavaged with 1 g / day of the traditional Chinese medicine preparations obtained in Example 1 and Comparative Examples 1 and 2, and groups 4 were intraperitoneally injected with 0.1 mL of a concentration of 1×10 6 The bone marrow mesenchymal stem cells were injected once a week. Groups 5 to 7 were gavaged with 1 g / day of the traditional Chinese medicine preparations obtained in Example 1 and Comparative Examples 1 and 2, and injected intraperitoneally with 0.1 mL of a concentration of 1×10 6 Bone marrow mesenchymal stem cells were injected into the 8 groups without any treatment (model group). After 3 consecutive weeks of treatment, all rats were anesthetized and killed. The knee joint conditions and tissue pathology of the rats in each group were tested. Figure 3 and Figure 4 .

[0082] Micro-CT was used to examine the knee joints of the rats in each group. Anteroposterior and lateral radiographs of the knee joints of the rats in each group were used to observe joint damage. All specimens were fixed and then scanned with micro-CT. Scanning parameters were as follows: tube current 200 μA, voltage 85 kV, full-body scan, resolution 10.160993 μm, exposure time 384 ms, and scan angle 180 degrees. A phantom (Phantom; provided by the equipment manufacturer) was calibrated under the same conditions. Raw images were obtained after scanning. Three-dimensional reconstruction software NRecon (software version V1.7.4.2, Bruker, Germany) was used to reconstruct selected regions of the raw images. Reconstructed images were previewed before reconstruction. To reduce image artifacts and achieve optimal reconstruction results, reconstruction parameters were adjusted to the following values: smoothing = 5 (smoothing), beam-hardening = 8 (beam intensity), and ring artifacts = 25%. After setting the parameters, the image reconstruction process was initiated by editing and selecting the designated folder. CTAnalyser (software version 1.20.3.0, Bruker, Germany) was used to analyze the region of interest (ROI). Unified parameters were set and the software calculated parameters such as total tissue volume (TV), bone volume (BV), volume ratio (BV / TV), bone area (BS), trabecular number (Tb.N), and trabecular thickness (Tb.th). The results are shown in Figure 3. Figure 3 .

[0083] Histopathological examination method: The articular cartilage tissue of each group of rats was separated and fixed in 4% paraformaldehyde solution for 3 days, decalcified with 10% EDTA for 30 days, and then embedded in paraffin after dehydration. After dewaxing, gradient ethanol immersion, staining, differentiation, anti-blueing, eosin staining, ethanol dehydration, transparency, and sealing, the rat tissue was observed under an optical microscope for HE staining and photographed. Paraffin sections were prepared according to the instructions, stained with safranin fast green, observed under a microscope and photographed. The results are shown in the table. Figure 4 .

[0084] Depend on Figure 3 It can be seen that compared with the model group, the group using Example 1 in combination with stem cells can significantly promote the increase in bone volume and thickening of trabeculae in KOA rats. However, whether it is the Chinese medicine preparation obtained in Example 1 alone or the stem cell group alone, although it can promote the increase in bone volume and thickening of trabeculae in KOA rats, the effect is not obvious. Compared with the model group, only the comparative example 1 and comparative example 2 groups were used. There was no significant difference in the bone volume or trabeculae of KOA rats. However, the comparative example 1 combined with stem cells and the comparative example 2 combined with stem cells groups showed that the bone volume and trabeculae of KOA rats were similar to those of the stem cell group alone.

[0085] Depend on Figure 4 It can be seen that the surface of the knee cartilage in the blank control group was smooth and flat, with cells arranged neatly and no inflammatory cells. The cartilage edge of the rats in the model group was severely damaged, the cells were disordered, and a large number of inflammatory cells infiltrated. Compared with the model group, the damage to the cartilage tissue edge of the rats in the Example 1 group and the stem cell group was alleviated, the cells were arranged more neatly, and the infiltration of inflammatory cells was reduced. This shows that only the Chinese medicine preparation or stem cells obtained by using Example 1 of the present invention have a certain repair effect on the cartilage of KOA rats. Compared with the model group, the damage to the cartilage tissue edge of the rats in the Example 1 combined with stem cell group was significantly alleviated, the cells were arranged neatly, and no inflammatory cell infiltration was seen, indicating that the use of Example 1 of the present application in combination with stem cells can significantly repair the cartilage of KOA rats. However, the difference between the groups using only Comparative Example 1 or Comparative Example 2 and the model group was not much, indicating that only using Comparative Example 1 or Comparative Example 2 can only maintain the condition of knee osteoarthritis from worsening, but cannot achieve a good therapeutic effect. The results of the groups using Comparative Examples 1 and 2 in combination with stem cells are comparable to those of the group using only stem cells. It can be seen from this that the combined use of the traditional Chinese medicine preparation of the present invention and bone stem cells has a significant effect on the treatment of knee osteoarthritis.

[0086] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. A Chinese medicine preparation for treating knee osteoarthritis, characterized in that: The traditional Chinese medicine preparation comprises the following components in parts by weight: 10-15 parts of Rehmannia root, 10-20 parts of Astragalus root, 10-15 parts of Chinese Angelica sinensis, 10-15 parts of Achyranthes bidentata, 10-20 parts of Salvia miltiorrhiza, 10-15 parts of Chuanxiong, 10-15 parts of Saposhnikovia divaricata, 10-15 parts of stir-fried Ziziphus jujuba seeds, 5-15 parts of earthworms, 10-20 parts of Millettia reticulata, 10-20 parts of Carthamus tinctorius, 10-20 parts of Lycopodiella ciliata, 10-20 parts of Drynaria rhizome, 5-15 parts of White Peony Root, 5-15 parts of Cynanchum chinense, and 5-15 parts of Angelica dahurica.

2. The method for preparing the Chinese medicine preparation according to claim 1, wherein The following steps are involved: (2.1) mixing the components of the traditional Chinese medicine preparation according to claim 1 to obtain a mixture; (2.2) The mixture is mixed with water, decocted 1 to 3 times, filtered, the filtrate is collected, the filtrates are combined, concentrated, dried, and sterilized to obtain a traditional Chinese medicine preparation.

3. The preparation method according to claim 2, characterized in that The mass ratio of the mixture to water is 1:3-7.

4. The preparation method according to claim 2, characterized in that The decocting time is 15 to 40 minutes per time, and the decocting temperature is 90 to 100° C.

5. The preparation method according to claim 2, characterized in that The concentration is performed under reduced pressure, the concentration temperature is 35-55° C., and the volume after concentration is 2 / 5-3 / 5 of the original volume.

6. The preparation method according to claim 2, characterized in that The sterilization is performed by radiation sterilization using cobalt 60, and the sterilization time is 20 to 28 hours.

7. Use of the traditional Chinese medicine preparation according to claim 1 or the traditional Chinese medicine preparation obtained by the preparation method according to any one of claims 2 to 6 in preparing a drug for treating knee osteoarthritis, characterized in that: Combine traditional Chinese medicine preparations with stem cells.

8. The use according to claim 7, characterized in that When the Chinese medicine preparation is used in combination with stem cells, the dosage of the Chinese medicine preparation is 1 to 3 g / day, and the concentration of the stem cells is 1 to 5×10 5 / mL, the dosage is 0.5-2mL / week.

9. The use according to claim 7, characterized in that The stem cells are bone marrow mesenchymal stem cells.

10. The use according to claim 7, characterized in that The dosage form of the traditional Chinese medicine preparation is powder, and the dosage form of the stem cells is injection.

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