Traditional Chinese medicine composition for crymodynia in department of traumatology as well as preparation method and application of traditional Chinese medicine composition
By developing a cold pain patch for the trauma department containing a variety of Chinese medicine ingredients, the problem of pain in KOA patients is difficult to control in cold environments, and significant pain relief and inflammation relief effects are achieved, avoiding the side effects of existing drugs.
Patent Information
- Application Number
- CN202510066698.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-01-16
- Publication Date
- 2025-03-11
AI Technical Summary
The prior art is difficult to effectively control pain in patients with knee osteoarthritis (KOA), especially in cold environments, and existing drugs have obvious side effects.
Develop a Chinese medicine composition for cold pain in the trauma, including Bauhinia peel, Viagra, Salvia miltiorrhiza, Turmeric, Forsythia, Clove, Cinnamon and Unisex, to make it into patch form, and relieve pain in KOA patients through the effects of warming the meridians and promoting blood circulation, removing cold and removing dampness.
The cold pain patch of this injury department significantly alleviated synovial inflammation and cold pain sensitivity in KOA mice, increased the cold pain sensitivity threshold, lowered the content of serum inflammatory factors and pain media, and improved the symptoms of KOA patients, especially the symptoms of cold pain sensitivity.
Smart Images
Figure CN119587669A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of traditional Chinese medicine, and in particular to a traditional Chinese medicine composition for traumatic cold pain, and a preparation method and application thereof. Background Art
[0002] Osteoarthritis is an age-related degenerative joint disease. The latest epidemiological survey results show that about 600 million people worldwide are affected by OA, among which knee osteoarthritis (KOA) is the most common. KOA is highly disabling, and the typical clinical symptoms are recurrent chronic cyclical pain. In the terminal stage of the disease, patients often have to undergo knee replacement surgery due to loss of joint function, which brings a heavy medical burden to society. KOA patients have a lower pain threshold to cold stimulation and will feel more intense pain in a cold environment. Controlling pain and relieving symptoms are the primary issues that need to be urgently addressed in the clinical treatment of KOA.
[0003] Chronic inflammatory stimulation may be an important factor leading to the progression of KOA. Previous reports have pointed out that KOA animal models show lower doses of inflammation re-triggering or more severe inflammatory responses and pain behaviors in repeated inflammatory stimulation. Circulating monocytes play an important role in the progression of KOA. Circulating monocytes complete pro-inflammatory differentiation during the process of being recruited to the synovium of KOA, and this process plays a key role in the generation of KOA pain. High mobility group box 1 protein (HMGB1) is a classic immune triggering molecule. Many studies have reported that HMGB1 can enhance the activity of chemokine CXC motif ligand 12 (CXCL12) and bind to it to form a heterogeneous complex, enhancing the directional migration of inflammatory cells to the site of injury through the chemokine receptor CXCR4.
[0004] There is no specific drug for KOA at present. The first-line drugs for controlling KOA pain in clinical practice, such as non-steroidal anti-inflammatory drugs, have obvious gastrointestinal and cardiovascular side effects. Traditional Chinese medicine has significant advantages in treating chronic inflammatory diseases. KOA belongs to the category of "arthralgia" and "weakness" in traditional Chinese medicine theory. Its main pathogenesis can be summarized as deficiency of the root and excess of the superficial symptoms, which is mostly caused by insufficient vital energy, liver and kidney deficiency, and the invasion of external evils such as wind, cold and dampness into the body. Summary of the invention
[0005] Purpose of the invention: The technical problem to be solved by the present invention is to provide a traditional Chinese medicine composition for treating cold pain in traumatology and its preparation method and application in view of the deficiencies in the prior art.
[0006] In order to solve the above technical problems, the present invention discloses a Chinese medicine composition for treating cold pain in trauma, and its preparation method and application. The specific scheme is as follows:
[0007] A Chinese medicine composition for cold pain in traumatology, comprising the following components in parts by weight: 6-10 parts of Bauhinia bark, 6-10 parts of Vitex rotundifolia, 1-3 parts of Salvia miltiorrhiza, 1-3 parts of Curcuma longa, 1-2 parts of Forsythia suspensa, 1-3 parts of Cloves, 1-3 parts of Cinnamon bark and 1-3 parts of Herba Lepidium. Preferably, the composition comprises the following components in parts by weight: 8 parts of Bauhinia bark, 8 parts of Vitex rotundifolia, 2 parts of Salvia miltiorrhiza, 2 parts of Curcuma longa, 1 part of Forsythia suspensa, 2 parts of Cloves, 2 parts of Cinnamon bark and 2 parts of Herba Lepidium.
[0008] Wherein, the dosage form of the traditional Chinese medicine composition for treating cold pain in trauma is a patch.
[0009] In a second aspect, the present invention provides a traumatic cold pain patch, which comprises an adhesive and the traumatic cold pain Chinese medicine composition described in the first aspect.
[0010] Wherein, the adhesive is honey, preferably jujube honey.
[0011] The mass ratio of the adhesive to the traditional Chinese medicine composition for treating traumatic cold pain is 1 to 3:1 to 3. Preferably, the mass ratio is 2:1.
[0012] In a third aspect, the present invention provides a method for preparing the traumatic cold pain patch described in the second aspect, comprising the following steps: weighing a formulated amount of the traumatic cold pain Chinese medicine composition, grinding it into powder, mixing it with the adhesive, and stirring to obtain the traumatic cold pain patch.
[0013] Preferably, the adhesive is mixed after being refined, wherein the refining is performed at 145° C. for 3 hours, and the stirring is performed at 50 Hz for 15 minutes.
[0014] In a fourth aspect, the present invention provides use of the traumatology cold pain Chinese medicine composition described in the first aspect or the traumatology cold pain patch described in the second aspect in the preparation of a drug for treating osteoarthritis.
[0015] Wherein, the osteoarthritis is knee osteoarthritis.
[0016] Wherein, the knee osteoarthritis includes knee synovitis and / or knee cold pain sensitivity.
[0017] In the Chinese medicine composition of the present invention, cinnamon bark tastes pungent and sweet, has a very hot nature, replenishes fire and assists yang, draws fire back to the origin, dispels cold and relieves pain, and warms and unblocks the meridians; Bauhinia bark tastes bitter, has a flat nature, promotes blood circulation and detoxification, dispels wind and dampness, and the two are monarch drugs, warming the meridians and promoting blood circulation, dispelling cold and dampness. Cloves are pungent, warm in nature, tonify the kidney and assist yang, warm the middle and reduce adverse reactions; Vitex rotundifolia tastes pungent and bitter, has a slightly cold nature. Dispel wind and dampness, relax the meridians and relieve pain, and the two are minister drugs, helping cinnamon bark and Bauhinia bark to warm the meridians and promote blood circulation, dispel rheumatism, and eliminate arthralgia. Salvia miltiorrhiza tastes bitter, has a slightly cold nature, promotes blood circulation and removes blood stasis, and relieves pain by promoting menstruation; Forsythia suspensa tastes bitter, has a slightly cold nature, clears away heat and detoxifies, eliminates swelling and dissipates knots, and evacuates wind-heat. Wind-cold-damp evil obstructs the arthralgia for a long time, and there are different degrees of heat transformation and joint swelling. The two are adjuvant drugs, which promote blood circulation and remove blood stasis, and clear away heat and relieve swelling. Du Yiwei tastes sweet and bitter, is neutral in nature, and can promote blood circulation and stop bleeding, dispel wind and relieve pain; Turmeric tastes spicy and bitter, and is warm in nature. It can promote blood circulation and qi, dredge menstruation and relieve pain. The liver governs the tendons, and the knee is the seat of the tendons. Both herbs belong to the liver meridian as guiding herbs, leading the other herbs in the prescription directly to the affected area. The combination of herbs can warm the meridians and promote blood circulation, dissipate blood stasis and relieve pain, dispel wind and dampness, and clear away heat and reduce swelling.
[0018] Beneficial effects:
[0019] The invention discloses a traumatology cold pain Chinese medicine composition and a traumatology cold pain patch. After the traumatology cold pain patch is used to intervene in mice with knee osteoarthritis, the inflammation of the synovial membrane of the mice is alleviated, the serum inflammatory factors and pain mediators are reduced, the cold pain sensitivity threshold is upregulated, and the upregulation of the expression levels of cold pain sensitivity, fibrosis and nerve sprouting related indicators and training immunity related indicator proteins and genes is significantly reversed. Therefore, the traumatology cold pain patch can significantly improve the synovial inflammation and cold pain sensitivity of KOA mice. The "warming meridians and activating blood circulation" drugs are used in the prescription, and the clinical application has significant therapeutic effects, especially for the symptoms of cold pain sensitivity in KOA patients, the effect is more significant. BRIEF DESCRIPTION OF THE DRAWINGS
[0020] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments, and the above and / or other advantages of the present invention will become more clear.
[0021] Figure 1 The results of in vivo imaging of mice.
[0022] Figure 2 These are the results of the cold pain threshold measurement in mice.
[0023] Figure 3 The results of HE staining of mouse synovial tissue (400×).
[0024] Figure 4 It is the content of IL-1β, TNF-α, CGRP and Substance P in mouse serum. ## P < 0.01, compared with the Sham group; # P < 0.05, compared with the KOA group; ##P<0.01, compared with the KOA group.
[0025] Figure 5 is the expression level of TRPM8 and TRPA1 proteins and genes in mouse synovial tissue. Figure 5 A shows the representative protein bands and relative protein expressions of TRPM8 and TRPA1; Figure 5 B is the relative gene expression of TRPM8 and TRPA1. ## P < 0.01, compared with the Sham group; # P < 0.05, compared with the KOA group; ## P<0.01, compared with the KOA group.
[0026] Figure 6 The expression levels of Netrin1 and Collagen I in mouse synovial tissue and DCC protein and gene in DRG tissue. Figure 6 A is the representative protein bands and relative protein expressions of Netrin1, Collagen I and DRG; Figure 6 B is the relative gene expression of Netrin1, Collagen I and DRG. ## P < 0.01, compared with the Sham group; # P < 0.05, compared with the KOA group; ## P<0.01, compared with the KOA group.
[0027] Figure 7 is the expression level of HMGB1, CXCL12, and CXCR4 proteins and genes in the synovial tissue of each group of mice, Figure 7 A shows the representative protein bands and relative protein expressions of HMGB1, CXCL12, and CXCR4; Figure 7 B is the relative gene expression of HMGB1, CXCL12, and CXCR4, among which, ## P < 0.01, compared with the Sham group; # P < 0.05, compared with the KOA group; ## P<0.01, compared with the KOA group. DETAILED DESCRIPTION
[0028] The mice used in the following examples were 50 2-month-old SPF male C57 mice, weighing 20-25 g (Vital River Company). The room temperature of the animal room was maintained at 25±2°C, relative humidity 60±5%, and a light / dark cycle of 12 hours. The animal experiment procedures were approved by the Animal Care and Use Committee of Nanjing University of Chinese Medicine and were conducted under the guidelines of the National Institutes of Health for the protection and use of experimental animals.
[0029] The reagents and instruments used in the following examples are as follows: DiR fluorescent dye (Shanghai Yisheng Biotechnology Co., Ltd., batch number: 40757ES25); HMGB1 recombinant protein, HMGB1 inhibitor (Glycyrrhizin, GL) (MCE, USA, batch number: HY-P73104A, HY-N0184); Hematoxylin-eosin (HE) staining kit (Beijing Solebold Technology Co., Ltd., batch number: G1120); β-actin, TRPA1, CXCL12, CXCR4 antibodies (Wuhan Sanying Biotechnology Co., Ltd., catalog number: 21829-1-AP, 19124-1-AP, 17402-1-AP, 11073-2-AP); TRPM8, TNF-α, Collagen I, Netrin-1, DCC, HMGB1 antibodies (Affinity, USA, batch number: DF7966, AF7014, F7001, DF8579, DF3611, AF7020); IL-1β antibody (CST, USA, batch number: 31202); horseradish peroxidase-conjugated goat anti-rabbit secondary antibody, RIPA lysis buffer (APEx BIO, USA, batch number: K1223, K1020), protein loading buffer (Shanghai Biotech, batch number: P0015); RNA extraction kit, reverse transcription and PCR quantification kit (Nanjing Novogene Biotech Co., Ltd., batch number: R401-01, R223, Q331); IL-1β, TNF-α ELISA detection kit (Wuhan Yacoin Biotechnology Co., Ltd., batch number: KTE7005, KTE7015), Substance P, CGRP ELISA detection kit (Wuhan Fine Biotechnology Co., Ltd., batch number: EM1589, EM0922).
[0030] IVIS Spectrum animal live imager (Perkin Elmer, USA), RM2016 pathology slicer (Leica Instruments, Shanghai, China), ABI 7500 qRT-PCR system (Applied Biosystems, USA), Typhoon gel imaging system (GE, USA).
[0031] All the medicinal materials used in the following examples were obtained from Jiangsu Provincial Hospital of Traditional Chinese Medicine and met the requirements of the Chinese Pharmacopoeia.
[0032] Example 1 Preparation of Traumatology Cold Pain Patch
[0033] The trauma cold pain patch described in this embodiment is prepared according to the following steps:
[0034] (1) Grinding Bauhinia bark, Vitex rotundus, Salvia miltiorrhiza, Curcuma longa, Forsythia suspensa, Clove, Cinnamomum cassia bark and Lamiophlomis lappa into powder in a mass ratio of 8:8:2:2:1:2:2:2 to obtain Shangke Lengtong Plaster powder;
[0035] (2) refining the jujube honey at 145°C for 3 h;
[0036] (3) The powder of the Shangke Lengtong Patch prepared in step (1) and the jujube honey prepared in step (2) were added to the pot in a mass ratio of 1:2, and then stirred at 50 Hz for 15 min to prepare the Shangke Lengtong Patch ointment.
[0037] Example 2 Construction of knee osteoarthritis (KOA) model by destabilization of medial meniscus (DMM) and animal grouping
[0038] Fifty mice were randomly divided into: Sham group, KOA group, KOA+HMGB1 reinfusion group (HMGB1 group), KOA+Shangke cold pain patch group (cold pain patch group) and KOA+HMGB1 inhibitor group (GL group). The modeling methods of mice in each group are as follows:
[0039] After the mice were adaptively raised for 1 week, they were anesthetized with 3% sodium pentobarbital injected intraperitoneally, the knee joints were prepared and disinfected, the mice were incised from the medial side of the knee joint to the joint cavity, the medial meniscus tibial ligament was severed, and penicillin was injected intramuscularly for 3 days (80,000 units per day) to prevent infection, and the KOA model was constructed.
[0040] Sham group: only the skin at the knee joint of mice was incised and sutured;
[0041] KOA group: The constructed KOA model was not given any medication;
[0042] HMGB1 group: Ficoll density gradient centrifugation was used to extract mouse peripheral blood mononuclear cells, and CD14 magnetic beads were used to sort circulating monocytes. 50 ng / mL of HMGB1 recombinant protein was used in 1640 medium to intervene in circulating monocytes for 24 h (the amount of circulating monocytes added was 500,000 / well). 10 mL of fresh 1640 medium was replaced, and after resting for 48 h, 50 ng / mL of HMGB1 recombinant protein was used again to stimulate for 24 h to obtain 500,000 trained circulating monocytes, which were then returned to the tail vein of KOA mice.
[0043] Cold pain patch group: medication started on the 14th day after surgery, with a cold pain patch applied to the knee joint every day, fixed with tape, for a total of 28 days, with a dose of 0.1 g / cm per patch 2 ;
[0044] GL group: The patients started taking medication on the 14th day after surgery, and the HMGB1 inhibitor glycyrrhizin was intraperitoneally injected at a dose of 10 mg / kg per day for 28 days.
[0045] Example 3 Study on the efficacy of cold pain patch for trauma
[0046] The data results in this example are expressed as mean ± standard deviation (x ± s), and statistical analysis was performed using Graphpad Prism 8.0 software. One-way ANOVA or t-test was used for comparison between groups, and P < 0.05 was considered to be statistically significant.
[0047] 1. Animal live imaging
[0048] According to the method described in Example 2 for constructing the HMGB1 group, HMGB1 trained circulating monocytes were obtained.
[0049] 24 hours after the DiR dye-labeled HMGB1 trained circulating monocytes were injected back into the tail vein, the HMGB1 group of mice was obtained, and the recruitment sites in the mice were observed using IVIS Spectrum animal in vivo imaging.
[0050] In vivo imaging results showed that Figure 1 As shown, compared with the Sham group, circulating mononuclear cells labeled with DiR injected through the tail vein were enriched in the knee joints of mice in the KOA group and the HMGB1 group, and the signal intensity of circulating monocytes enriched in the knee joints after HMGB1 training was higher.
[0051] 2. Mouse cold plate paw lifting test
[0052] The cold pain sensitivity threshold of each group of mice was measured using an adjustable temperature metal plate system (35150-001, Ugo Basil SLR, Italy). The temperature of the thermostatic metal plate was set to 0°C. The time when the mice withdrew their paws, raised their paws, and licked their paws on the cold plate was observed and recorded on days 0, 7, 14, 21, and 28 after modeling.
[0053] The results of cold pain threshold test showed that Figure 2 As shown in the figure, compared with the Sham group, the cold stimulation perception threshold of the mice in the KOA group was decreased, and the decrease was more obvious in the HMGB1 group than in the KOA group (P < 0.05). Compared with the KOA group, the cold stimulation perception threshold of the mice in the cold pain patch group and the GL group was significantly increased (P < 0.05).
[0054] 3. Tissue sample extraction and HE staining
[0055] After the intervention, the mice in each group were blooded from their eyes and killed by dislocation of the neck. The skin of the knee joint was cut and the joint cavity was opened. The synovial tissue was scraped with a blade and stored in a -80°C refrigerator or 4% paraformaldehyde solution for hematoxylin-eosin (HE) staining to evaluate the level of synovial pathological changes. Specifically, the synovial tissue fixed with 4% paraformaldehyde was embedded in paraffin, sliced, and the synovial tissue was routinely stained with HE. In addition, the skin on the back was opened, the lumbar spine was found and removed, the excess muscle tissue was removed, and the spine was cut into two sections along the spinal canal. The DRG of the L3-L5 spine was taken out and stored in a -80°C refrigerator.
[0056] The results of HE staining of synovial tissue showed that Figure 3 As shown, the synovial cells in the Sham group were arranged in an orderly manner, without inflammatory cell infiltration; in the KOA group, the synovial cells proliferated in large numbers and were distributed disorderly, accompanied by significant inflammatory cell infiltration, and the synovial cell proliferation and inflammatory cell infiltration in the HMGB1 group were more obvious; compared with the KOA group, the synovial cell proliferation and inflammatory infiltration in the cold pain patch group and the GL group were significantly reduced.
[0057] 4.ELISA
[0058] The collected mouse serum was taken according to the instructions of the kit, and the OD value was measured at a wavelength of 450nm to detect the content of inflammatory factors IL-1β (proinflammatory factor interleukin-1β), TNF-α (tumor necrosis factor-α) and pain mediators CGRP (calcitonin gene-related peptide) and Substance P (substance P) in the serum.
[0059] ELISA results showed that Figure 4 As shown in the results, compared with those in the Sham group, the levels of inflammatory factors IL-1β, TNF-α and pain mediators CGRP and Substance P in the serum of the mice in the KOA group and HMGB1 group were increased, and the increase in the HMGB1 group was more obvious (P<0.05). Compared with those in the KOA group, the levels of the above indicators in the mice in the cold pain patch group and GL group were significantly decreased (P<0.05).
[0060] 5. Western blot and real-time PCR test
[0061] Recurrent chronic and intractable pain is a typical symptom of KOA and a difficult problem that urgently needs to be solved clinically. Many studies have shown that synovial inflammation is significantly correlated with KOA pain symptoms and pathological structural changes. However, in preclinical studies for the treatment of KOA pain, the efficacy of inhibitors and mRNA vaccines developed targeting the classic proinflammatory factors IL-1β and TNF-α is often unsatisfactory. Targeting a single tissue or individual proinflammatory factors does not seem to effectively inhibit KOA peripheral pain sensitivity, so there are certain limitations in analgesic treatment. The latest KOA diagnosis and treatment guidelines point out that KOA pain is a mixed pain that combines nociception with neuropathic physiology, which means that repeated cold stimulation may affect pain nerve conduction, leading to an increase in the body's perception of pain and amplifying the degree of KOA pain. Consistent with this view, previous studies by our research group have shown that TRPA1 and TRPM8, members of the transient receptor potentials (TRPs) ion channel family, act as cold nociceptors and are directly related to the cold pain sensitivity of KOA, which is characterized by "pain when exposed to cold." In the synovial fibrosis state of KOA, endothelial cells secrete axon guidance factor 1 (Netrin1) which binds to the neuronal surface receptor DCC to induce sensory nerve sprouting, increase the density of ascending conduction of pain signals, and cause cold pain sensitivity.
[0062] In this example, Western blot and Real-time PCR methods were used to detect the protein and gene expressions of cold pain sensitivity related indicators TRPA1 and TRPM8, fibrosis indicator Collagen I, HMGB1 / CXCL12 / CXCR4 signaling pathway related indicators, and nerve sprouting indicators Netrin-1 and DCC in synovial tissue or DRG tissue. The synovial tissue or DRG tissue extracted in "3. Tissue sample extraction" of this embodiment was taken out from -80°C, ground into powder with liquid nitrogen, added with RIPA lysis solution and continued to grind and lyse for 5 minutes, 12000rpm, 15min centrifugation, and the supernatant was taken. The protein concentration was determined by the BCA method, and the protein was boiled and denatured and loaded for electrophoresis and transfer. After blocking with a rapid blocking solution at room temperature for 20 minutes, the internal reference antibody β-actin (dilution ratio of 1:20000) and primary antibodies such as TRPA1, TRPM8, TNF-α, IL-1β, Collagen I, Netrin-1, DCC, HMGB1, CXCL12, and CXCR4 (dilution ratio of 1:1000) were added, and incubated at 4°C until the next day, the primary antibody was recovered and the corresponding secondary antibody was added, and the ECL color was exposed. Image Lab software analyzed the gray value of the target protein and calculated the relative expression.
[0063] After total RNA was extracted from synovial tissue or DRG by Trizol method and the concentration was determined, the reverse transcription system was calculated for reverse transcription, and PCR amplification was performed using SYBR fluorescence quantitative method. PCR data results were all based on β-actin as the internal reference, 2 -ΔΔCt The relative expression of each target gene was calculated and analyzed by the PCR primer sequence shown in Table 1.
[0064] Table 1 Primer sequences
[0065]
[0066] Firstly, the expression of TRPM8 and TRPA1 proteins and genes, which are related indicators of cold pain sensitivity in the synovial tissue of each group of mice, was detected.
[0067] WB and PCR results showed that Figure 5 As shown in the figure, compared with the Sham group, the protein and gene expression levels of TRPM8 and TRPA1 in the synovial tissue of mice in the KOA group and HMGB1 group were increased, and the increase in the HMGB1 group was more obvious (P < 0.05). Compared with the KOA group, the protein and gene expression levels of the above indexes in mice were significantly decreased after cold pain patch or GL intervention (P < 0.05).
[0068] This example also detected the expression of Netrin1, Collagen I in the synovial tissue and DCC protein and gene in the DRG tissue of each group of mice. The results of WB and PCR showed that Figure 6 As shown in the figure, compared with the Sham group, the expression levels of Netrin1, Collagen I in the synovial tissue and DCC protein and gene in the KOA and HMGB1 groups were increased, and the increase in the HMGB1 group was more obvious (P < 0.05). Compared with the KOA group, the expression levels of the above-mentioned index proteins and genes in the mice were significantly decreased after the intervention of cold pain patch or GL (P < 0.05).
[0069] Regarding the expression of HMGB1, CXCL12, and CXCR4 proteins and genes in the synovial tissues of each group of mice, WB and PCR results showed that Figure 7 As shown in the figure, compared with the Sham group, the protein and gene expression levels of HMGB1, CXCL12, and CXCR4 in the synovial tissue of mice in the KOA group and HMGB1 group were increased, and the increase in the HMGB1 group was more obvious (P < 0.05). Compared with the KOA group, the protein and gene expression levels of the above indexes in mice were significantly decreased after cold pain patch or GL intervention (P < 0.05).
[0070] In summary, animal in vivo imaging showed that circulating monocytes can be recruited in the knee joint after being infused back into KOA mice through the tail vein, and more circulating monocytes were recruited in the knee joint of mice in the HMGB1 group. In addition, compared with the Sham group, the mice in the KOA group and HMGB1 group showed inflammatory pathological changes in the synovium, increased expression of serum inflammatory factors and pain mediators, decreased cold pain threshold, and up-regulated protein and gene expression of cold pain sensitivity, fibrosis and nerve sprouting related indicators in synovial tissue and DRG tissue, as well as HMGB1 / CXCL12 / CXCR4 signaling pathway related indicators, and the above changes in the HMGB1 group were more significant than those in the KOA group (P < 0.05); after intervention with the trauma cold pain patch of the present invention, the synovial inflammation of mice was alleviated, the serum inflammatory factors and pain mediators were reduced, the cold pain threshold was increased, and the up-regulation of cold pain sensitivity, fibrosis and nerve sprouting related indicators and training immune related indicators protein and gene expression levels was significantly reversed (P < 0.05).
[0071] The traumatic cold pain patch described in the present invention is formulated based on the above pathogenesis combined with its many years of experience in treating KOA. It has a significant effect on the symptoms of KOA cold pain sensitivity, and the various medicines in the prescription work together to achieve the effect of "warming the meridians and activating blood circulation, dispelling cold and relieving pain". KOA cold pain sensitivity is closely related to synovial inflammation, synovial fibrosis and sensory nerve sprouting. This study showed that compared with the KOA group, the traumatic cold pain patch intervention can significantly improve the synovial inflammation of KOA mice, adjust the cold pain sensitivity threshold, reduce the levels of inflammatory factors IL-1β, TNF-α and pain mediators calcitonin gene related peptide (Calcitonin gene related peptide, CGRP), substance P in serum, and down-regulate the cold pain sensitivity related indicators TRPA1, TRPM8 and fibrosis and nerve sprout related indicators type I collagen (Collagen I), Netrin-1, DCC protein and mRNA expression. The above results suggest that the traumatic cold pain patch has a good intervention effect on KOA cold pain sensitivity.
[0072] Trained immunity is an emerging research field. Its mechanism is the epigenetic remodeling and cellular metabolic reprogramming that occur when innate immune cells respond to the initial stimulus. The transcriptional reserve after the stimulus is removed still exists, and it enters a long-term enhanced defense state, and then responds more quickly and strongly to the next stimulus. This mechanism is highly similar to the mechanism of chronic pain caused by repeated attacks of KOA. Circulating monocytes are the most common innate immune cells. They have a high degree of dynamic plasticity and can be mobilized quickly and in large quantities to the site of inflammation. Studies have shown that the increased inflammatory reactivity of monocytes and macrophages after trained immunity may play a core role in certain inflammatory diseases. This mechanism may make an important contribution to the generation of cold pain sensitivity in KOA. Repeated and intensified inflammatory stimulation lowers the threshold of nociceptors for pain perception, leading to KOA pain sensitization.
[0073] The latest evidence shows that HMGB1 mediates the training immunity of local macrophages in the liver through immune response gene protein 1, so that the animal model of liver ischemia-reperfusion injury presents a more active inflammatory response when facing liver surgery. HMGB1 is widely distributed in mammalian cells. Its nuclear-cytoplasmic translocation and subsequent extracellular release are key factors in causing local inflammatory responses and are closely related to neuropathic peripheral pain sensitivity. There is a large number of local recruitment of macrophages during the training immunity process, which is likely to depend on the recruitment effect of the HMGB1 / CXCL12 / CXCR4 signaling axis. Our results show that after the fluorescently labeled circulating monocytes were transfused into mice, the circulating monocytes were recruited in the knee joints of KOA mice, and the circulating monocytes trained by HMGB1 recruited more circulating monocytes in the knee joints of mice after transfusion. Compared with the KOA group, after the HMGB1 training of circulating monocytes, the serum levels of inflammatory factors and pain mediators increased, and the protein and mRNA expressions of cold pain sensitivity-related indicators and fibrosis and nerve sprouting-related indicators were upregulated, which were reversed after intervention with HMGB1 inhibitors. In addition, compared with the Sham group, the expression of HMGB1 / CXCL12 / CXCR4 signaling axis-related proteins and mRNA in the KOA group mice was significantly higher than that in the KOA group, and the expression levels of the above indicators in the KOA mice stimulated by HMGB1 were higher. After intervention with the cold pain patch or HMGB1 inhibitor, the expression of HMGB1 / CXCL12 / CXCR4 signaling axis-related proteins and mRNA was significantly downregulated.
[0074] In summary, this study not only disclosed a trauma cold pain patch, but also clarified the potential mechanism of the trauma cold pain patch in alleviating KOA cold pain sensitivity. Specifically, there is a circulating monocyte training immunity driven by the HMGB1 / CXCL12 / CXCR4 signaling axis in KOA, which causes repeated exacerbation of inflammation and sensory nerve sprouting. The trauma cold pain patch improves KOA cold pain sensitivity by inhibiting this process.
[0075] The present invention provides a cold pain patch for trauma and its preparation method and application ideas and methods. There are many methods and ways to implement the technical solution. 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 principle of the present invention. These improvements and modifications should also be regarded as the protection scope of the present invention. All components not specified in this embodiment can be implemented by existing technologies.
Claims
1. A Chinese medicine composition for treating cold pain in trauma, characterized in that: The invention comprises the following components in parts by weight: 6-10 parts of Bauhinia bark, 6-10 parts of Vitex rotundifolia, 1-3 parts of Salvia miltiorrhiza, 1-3 parts of Curcuma longa, 1-2 parts of Forsythia suspensa, 1-3 parts of Clove, 1-3 parts of Cinnamon bark and 1-3 parts of Lamiophlomis lappa.
2. The Chinese medicine composition for treating cold pain in trauma according to claim 1, characterized in that: The invention comprises the following components in parts by weight: 8 parts of Bauhinia bark, 8 parts of Vitex rotundifolia, 2 parts of Salvia miltiorrhiza, 2 parts of Curcuma longa, 1 part of Forsythia suspensa, 2 parts of Clove, 2 parts of Cinnamon bark and 2 parts of Lamiophloea lappa.
3. The Chinese medicine composition for treating cold pain in trauma according to claim 1 or 2, characterized in that: The dosage form of the Chinese medicine composition is a patch.
4. A cold pain patch for trauma, characterized in that: The traumatic cold pain patch comprises an adhesive and the traumatic cold pain Chinese medicine composition according to claim 1.
5. The cold pain patch for traumatology according to claim 4, characterized in that: The adhesive is honey, preferably jujube honey.
6. The cold pain patch for traumatology according to claim 4, characterized in that: The mass ratio of the adhesive to the traditional Chinese medicine composition for traumatic cold pain is 1-3:1-3.
7. The method for preparing the cold pain patch for traumatology according to any one of claims 4 to 6, characterized in that: The method comprises the following steps: weighing a prescribed amount of the traditional Chinese medicine composition for traumatology, cold pain, grinding it into powder, mixing it with the adhesive, and stirring it to obtain a traumatology, cold pain patch.
8. Use of the traditional Chinese medicine composition for treating cold pain according to claim 1 or 2 or the cold pain patch for treating cold pain according to any one of claims 4 to 6 in the preparation of a drug for treating osteoarthritis.
9. The use according to claim 8, characterized in that: The osteoarthritis is knee osteoarthritis.
10. The use according to claim 9, characterized in that: The knee osteoarthritis includes knee synovitis and / or knee cold pain sensitivity.