Exosomes of a compound preparation for treating mural thrombus

Abstract

The present invention provides an application of a compound preparation exosome in a medicament for treating mural thrombus, belonging to the field of biomedicine. Specifically, it includes the extraction and preparation method of the compound preparation exosome and its application. The extraction of the compound preparation exosome includes the following steps: Weigh and mix 17 traditional Chinese medicine compounds, namely Ligusticum chuanxiong, Prunus persica, Angelica sinensis, Carthamus tinctorius, Panax notoginseng (Tianqi), Salvia miltiorrhiza, Lonicera japonica, Paeonia lactiflora, Scutellaria baicalensis, Sappanwood, Eupatorium japonicum, Olibanum, Myrrha, Tabanus, Hirudo, Mylabris and Strychnos nux-vomica, according to weight fractions, and obtain the mixed exosomes of the 17 traditional Chinese medicine compounds through exosome concentration and size exclusion chromatography column separation. Mix the mixed exosomes with hyaluronic acid to prepare a dressing. The dressing prepared by the present invention is a novel dressing of a traditional Chinese medicine preparation mixed exosome, which can effectively solve the problems in the current treatment process of mural thrombus. Therefore, the composite dressing involved in the present invention has wide application value in the treatment of mural thrombus.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technology, and in particular to application of a compound preparation of exosomes in a drug for treating mural thrombosis. Background Art

[0002] Paramural thrombus (also called mural thrombus) refers to the situation where a thrombus is attached to the wall of a blood vessel. The formation of this type of thrombus is usually related to damage to the vascular endothelium. Platelets adhere to the damaged vascular endothelium and activate the coagulation pathway, leading to the formation of a thrombus. Depending on its nature, the time of formation, and the location, paramural thrombus may be at risk of detachment, thereby causing embolism. According to the search results, if the paramural thrombus is not formed for a long time and has a loose texture, it may detach and move along the blood flow to other parts, such as the lungs (causing pulmonary embolism) or the brain (causing cerebral infarction). If the thrombus becomes stable over time and adheres firmly to the blood vessel wall, the risk of detachment will be reduced.

[0003] At present, in addition to oral medication, TCM may also combine external treatments such as Chinese medicine fumigation, external application, TCM targeted drug therapy, and lifestyle adjustments, including appropriate exercise, acupuncture and moxibustion, etc. However, there are no reports on the use of exosomes extracted from Chinese medicine compound preparations in the form of patches to treat paramural thrombosis. Summary of the invention

[0004] The purpose of the present invention is to provide a compound preparation of exosomes for use in a drug for treating adventitious thrombosis. The dressing prepared by the present invention is a new type of dressing of a Chinese medicine preparation mixed with exosomes, which can effectively solve the problems in the current treatment of adventitious thrombosis.

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

[0006] The invention discloses application of a compound preparation of exosomes in a medicine for treating mural thrombosis.

[0007] Furthermore, the raw materials of the compound preparation include: Chuanxiong, peach kernel, angelica, safflower, Panax notoginseng (Pansenosideae), Salvia miltiorrhiza, honeysuckle, red peony root, scutellaria baicalensis, Sappan wood, Zedoaria, frankincense, myrrh, horsefly, leech, blister beetle and Strychnos nux vomica.

[0008] Furthermore, the weight of the raw materials of the compound preparation is: 5g of Chuanxiong, 5g of peach kernel, 5g of angelica, 5g of safflower, 5g of Panax notoginseng, 5g of Salvia miltiorrhiza, 5g of honeysuckle, 5g of red peony root, 5g of Scutellaria baicalensis, 10g of Sappan wood, 10g of Zedoaria, 10g of frankincense, 5g of myrrh, 5g of horsefly, 5g of leech, 5g of Cantharides, and 5g of Strychnos nux vomica.

[0009] Furthermore, the method for preparing the compound preparation exosomes comprises the following steps:

[0010] Step 1: Exosome extraction: 17 Chinese medicines in the compound preparation were weighed according to weight fraction, mixed, crushed into particles of about 3-5 mm, then soaked in 2000 mL of warm water at 37°C for 2 h, crushed with a beater for 30 min, and the operation of crushing for 1 min and stopping for 1 min was repeated to obtain a suspension; the obtained suspension was placed at 4°C for 2 h, then centrifuged at 4000 rpm, the precipitate was discarded, and the supernatant A was collected;

[0011] Step 2: Exosome concentration: Use ultrafiltration stick to concentrate exosomes;

[0012] (1) The inlet and outlet of an ultrafiltration rod with a pore size of 200 nm are connected to the supernatant A collected in step 1 through a pipeline, and the supernatant is circulated by a pump. At the same time, the first filtrate B is collected at the filtrate receiving port until the residual volume of supernatant A reaches 30 mL, and then the collection is stopped;

[0013] (2) The collected first filtrate B is concentrated using an ultrafiltration stick with a pore size of 30 nm according to the process described in (1) above, until the residual volume of the first filtrate B reaches 30 mL, and the operation is stopped to obtain the residual liquid, which is the concentrated liquid C;

[0014] Step 3 Exosome separation: Take an exclusion chromatography column tube, pump the concentrated solution C obtained in step 2 into the chromatography column at a flow rate of 2 mL / min, and continue to pump purified water at this flow rate after all the liquid enters the chromatography column, and receive the liquid flowing out of the chromatography column, 30 mL for each fraction; a total of 8 fractions are collected, and fractions 3, 4, and 5 are exosome fractions, which are combined to obtain exosomes of the Chinese medicine compound preparation.

[0015] Furthermore, the medicine also contains pharmaceutically acceptable excipients.

[0016] Furthermore, the dosage form of the drug is a dressing.

[0017] The present invention also provides a method for preparing exosomes of a Chinese medicine compound preparation, characterized in that the method specifically comprises the following steps:

[0018] Step 1: Exosome extraction: 17 Chinese medicines in the compound preparation were weighed according to weight fraction, mixed, crushed into particles of about 3-5 mm, then soaked in 2000 mL of warm water at 37°C for 2 h, crushed with a beater for 30 min, and the operation of crushing for 1 min and stopping for 1 min was repeated to obtain a suspension; the obtained suspension was placed at 4°C for 2 h, then centrifuged at 4000 rpm, the precipitate was discarded, and the supernatant A was collected;

[0019] Step 2: Exosome concentration: Use ultrafiltration stick to concentrate exosomes;

[0020] (1) The inlet and outlet of an ultrafiltration rod with a pore size of 200 nm are connected to the supernatant A collected in step 1 through a pipeline, and the supernatant is circulated by a pump. At the same time, the first filtrate B is collected at the filtrate receiving port until the residual volume of supernatant A reaches 30 mL, and then the collection is stopped;

[0021] (2) The collected first filtrate B is concentrated using an ultrafiltration stick with a pore size of 30 nm according to the process described in (1) above, until the residual volume of the first filtrate B reaches 30 mL, and the operation is stopped to obtain the residual liquid, which is the concentrated liquid C;

[0022] Step 3 Exosome separation: Take an exclusion chromatography column tube, pump the concentrated solution C obtained in step 2 into the chromatography column at a flow rate of 2 mL / min, and continue to pump purified water at this flow rate after all the liquid enters the chromatography column, and receive the liquid flowing out of the chromatography column, 30 mL for each fraction; a total of 8 fractions are collected, and fractions 3, 4, and 5 are exosome fractions, which are combined to obtain exosomes of the Chinese medicine compound preparation.

[0023] Furthermore, the raw materials and weight fractions of the Chinese medicine compound preparation described in step 1 are: 5g of Chuanxiong, 5g of peach kernel, 5g of angelica, 5g of safflower, 5g of Panax notoginseng, 5g of Salvia miltiorrhiza, 5g of honeysuckle, 5g of red peony root, 5g of Scutellaria baicalensis, 10g of Sappan wood, 10g of Zedoaria, 10g of frankincense, 5g of myrrh, 5g of horsefly, 5g of leech, 5g of Cantharides, and 5g of Strychnos nux vomica.

[0024] Furthermore, the exosomes of the traditional Chinese medicine compound preparation prepared by the above-mentioned preparation method are used in drugs for treating mural thrombosis.

[0025] The formula of each Chinese medicine in the Chinese medicine compound preparation of the present invention is as follows:

[0026] Chuanxiong: It has anticoagulant and antiplatelet aggregation effects and can promote blood circulation.

[0027] Peach kernel: Rich in fatty oil and aromatic acid, it effectively inhibits platelet aggregation and dissolves blood clots.

[0028] Angelica: A blood-replenishing Chinese medicine, rich in volatile oils and flavonoids, which inhibit platelet aggregation.

[0029] Safflower: Contains ingredients such as anthocyanins and volatile oils, which can inhibit platelet aggregation.

[0030] Panax notoginseng (Tianqi): Contains ingredients such as notoginseng saponins and notoginseng glycosides, which effectively inhibit platelet aggregation.

[0031] Danshen: Contains ingredients such as tanshinone and tanshinone, which inhibit platelet aggregation.

[0032] Honeysuckle: Contains flavonoids and volatile oils, which can inhibit platelet aggregation.

[0033] Red Peony Root: Contains ingredients such as paeoniflorin and peony root, which effectively inhibit platelet aggregation.

[0034] Scutellaria baicalensis: It is rich in ingredients such as baicalin, which can inhibit platelet aggregation.

[0035] Sophora japonica, zelan, frankincense, and myrrh: have the effect of promoting blood circulation and removing blood stasis, and help to promote blood circulation.

[0036] Horseflies, leeches, and blister beetles: have the effect of breaking up blood and removing blood stasis.

[0037] Strychnos nux vomica: has the effect of dispersing lumps and reducing swelling.

[0038] Compared with the prior art, the beneficial effects of the present invention are as follows.

[0039] Exosomes from traditional Chinese medicine are extracellular matrix vesicles that can carry the genetic information of their own cells. As a carrier, they deliver the cell information of traditional Chinese medicine to defective cells in the human body and exert their effects. Based on this characteristic, the exosomes from traditional Chinese medicine compound that can treat thrombosis are extracted and made into dressings. They are easy to use and act directly on the thrombosis site with obvious effects. The side effects of oral drugs and the trauma of surgical operations are reduced, greatly reducing the cost of treating diseases for patients. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 The extracted exosomes.

[0041] Figure 2 This is the result of exosome detection.

[0042] Figure 3 Electron micrograph of exosomes.

[0043] Figure 4 For exosome dressing.

[0044] Figure 5 This is a comparison of the effects before and after medication for Case 1. A is the ultrasound test result before medication; B is the ultrasound test result after medication.

[0045] Figure 6 This is a comparison of the effects before and after medication for case 2. A is the ultrasound test result before medication; B is the ultrasound test result after medication. DETAILED DESCRIPTION

[0046] The present invention is further described in detail below with specific embodiments, but it should not be understood that the scope of the above subject matter of the present invention is limited to the following embodiments, and all technologies realized based on the content of the present invention belong to the scope of the present invention.

[0047] Unless otherwise specified, all reagents and materials used in the present invention are commercially available.

[0048] Example 1. Preparation of exosomes from Chinese medicine compound preparations.

[0049] The raw materials and weight fractions of the Chinese medicine compound preparation are: 5g of Chuanxiong, 5g of peach kernel, 5g of Angelica sinensis, 5g of safflower, 5g of Panax notoginseng (Tianqi), 5g of Salvia miltiorrhiza, 5g of honeysuckle, 5g of red peony root, 5g of Scutellaria baicalensis, 10g of Sappan wood, 10g of Zedoaria, 10g of frankincense, 5g of myrrh, 5g of horsefly, 5g of leech, 5g of Cantharides, and 5g of Strychnos nux vomica.

[0050] Step 1: Exosome extraction: The above 17 Chinese medicines were weighed by weight, mixed, crushed into particles of about 3-5 mm, and then soaked in 2000 mL of warm water at 37°C for 2 hours, and crushed with a beater, repeating the operation of crushing for 1 minute and stopping for 1 minute for a total of 30 minutes. The resulting suspension was placed at 4°C for 2 hours, then centrifuged at 4000 rpm, the precipitate was discarded, and the supernatant A was collected.

[0051] Step 2: Exosome concentration: The exosomes were concentrated using an ultrafiltration rod (filter rod length 300 mm, diameter 40 mm). The filter rod had an inlet and an outlet, as well as a filtrate receiving port.

[0052] (1) Connect the inlet and outlet of an ultrafiltration rod with a pore size of 200 nm to the supernatant A collected in step 1 through pipes, and circulate the supernatant with a pump. At the same time, collect the first filtrate B at the filtrate receiving port until the residual volume of supernatant A reaches 30 mL, then stop collecting.

[0053] (2) The collected first filtrate B is concentrated using an ultrafiltration rod with a pore size of 30 nm according to the process described in (1) above, until the residual volume of the first filtrate B reaches 30 mL. The operation is stopped and the residual liquid obtained is the concentrated liquid C.

[0054] Step 3 Exosome separation: Take a 25×200mm (inner diameter×length) size exclusion chromatography column tube and fill it with Sephadex chromatographic filler suspended in water. The filler should be evenly filled in the chromatographic column tube to ensure that there are no bubbles and faults. The inlet of the chromatographic column is connected to a peristaltic pump. Pump the concentrated solution C obtained in step 2 into the chromatographic column at a flow rate of 2mL / min. After all the concentrated solution C enters the chromatographic column, continue to pump purified water at this flow rate to receive the liquid flowing out of the chromatographic column, 30mL for each fraction. A total of 8 fractions were collected, and fractions 3, 4, and 5 were exosome fractions. The exosomes of the traditional Chinese medicine compound preparation were obtained by merging (such as Figure 1 as shown).

[0055] The collected exosomes were tested for particle size and electron microscopy images (e.g. Figure 2~3 as shown).

[0056] Example 2: Preparation of exosome patch of traditional Chinese medicine compound preparation.

[0057] According to the steps described in Example 1, exosomes of the traditional Chinese medicine compound preparation were prepared.

[0058] The obtained exosomes were mixed with hyaluronic acid (1:1, v / v) to prepare a dressing (such as Figure 4 ), stored in a refrigerator at 4°C, the maximum storage time is 180 days.

[0059] Example 3: Treatment of mural thrombus with exosomes from a Chinese herbal compound preparation.

[0060] Clinical usage: Apply a small amount of exosome dressing to the affected area each time, use it once every 24 hours, and determine the area to be applied to the skin according to the actual surface area of ​​the thrombus until it is healed.

[0061] Case 1: Wang XX, male, 61 years old, Chinese medicine doctor.

[0062] Chief complaint: Intermittent dizziness, tinnitus, and blurred vision for more than a year, which has worsened recently.

[0063] Physical examination: Ultrasonic examination of the carotid artery confirmed that a strong echo was visible on the posterior wall of the left common carotid artery, with a length of approximately 0.59 cm. Two strong echo plaques were visible in the left common carotid artery, with the larger diameter of approximately 0.11 cm.

[0064] Diagnosis: mural thrombosis of the left common carotid artery.

[0065] Medication records and treatment effects ( Figure 5 ):

[0066] The Chinese herbal compound exosome dressing of the present invention was applied once a day, and a course of treatment was seven days. After seven days of application, ultrasound imaging was performed again, and a strong echo was seen on the posterior wall of the left common carotid artery, with a length of about 0.35 cm, and two strong echo plaques were seen in the left common carotid artery, with the largest diameter of about 0.09 cm.

[0067] Case 2: Jiang XX, male, 69 years old.

[0068] Chief complaint: Eight months after right carotid artery stripping, the patient felt dizzy and had intermittent tinnitus, so he came to see a doctor.

[0069] Physical examination: The diameters of both common carotid arteries were symmetrical, with thickening of the left intima-media with a thickness of 1.1 mm and that of the right intima-media with a thickness of 1.4 mm. A 26.8×1.7 mm hypoechoic plaque was seen on the proximal inner wall of the left common carotid artery and a 17.5×1.5 mm hypoechoic plaque was seen on the proximal outer wall. A 24.2×1.6 mm equal-echoic plaque was seen on the proximal inner wall of the right common carotid artery, a 40.6×2.5 mm mixed-echoic plaque was seen on the middle inner wall, and a 35.5×2.1 mm hypoechoic plaque was seen on the distal outer wall. The blood flow velocity and spectral morphology were normal. The diameters of the bifurcation of the bilateral carotid arteries were symmetrical, and the intima and media were not thick. A 12.6×1.9mm mixed echo plaque was seen on the inner wall of the left carotid bifurcation, and a 9.5×1.7mm equal echo plaque was seen on the outer wall. The blood flow velocity and spectral morphology were normal. A 17.2×2.1mm mixed echo plaque was seen on the inner wall of the right carotid bifurcation, resulting in localized stenosis of the lumen. The original diameter was 8.2mm, the residual diameter was 4.1mm, the diameter stenosis rate was 50%, the blood flow velocity was 132 / 53cm / s, the frequency window was filled, and the sound frequency was rough. The diameters of the bilateral internal carotid arteries were symmetrical. The intima-media thickening of the left carotid bulb was 1.3 mm thick. An 8.4×1.8 mm iso-echoic plaque was visible on the outer wall. The blood flow velocity was 78 / 35 cm / s and the spectrum was normal. The intima-media thickening of the right carotid bulb was 1.2 mm thick. A 14.7×1.5 mm hypo-echoic plaque was visible on the inner wall, leading to localized stenosis of the lumen. The original diameter was 3.8 mm, the residual diameter was 1.0 mm, the diameter stenosis rate was 74%, the blood flow velocity was 525 / 264 cm / s, the frequency window was filled, and the sound frequency was rough. The blood flow velocity and spectrum morphology of the left external carotid artery were normal. A 23.1×2.8 mm mixed echo plaque was visible on the outer wall of the right carotid bifurcation extending to the beginning of the external carotid artery, leading to localized stenosis of the lumen. The original diameter was 5.5 mm, the residual diameter was 2.2 mm, the diameter stenosis rate was 58%, the blood flow velocity was 251 / 61 cm / s, the frequency window was filled, and the sound frequency was rough.

[0070] Diagnosis: 8 months after right carotid artery stripping, the intima-media thickening and plaque formation (single) of the carotid bulb are present, with restenosis of 70-99%; the intima-media thickening and plaque formation (multiple) of the bilateral common carotid arteries; plaque formation (multiple) at the left carotid bifurcation; plaque formation (single) at the right carotid bifurcation, with stenosis of 50-69%; the intima-media thickening and plaque formation (single) of the left carotid bulb; plaque formation (single) of the right external carotid artery, with stenosis of 50-69%.

[0071] Medication records and treatment effects ( Figure 6 ):

[0072] The Chinese medicine compound exosome dressing of the present invention was applied once a day, and a course of treatment was seven days. After continuous application for 10 days, ultrasound examination was performed again: the inner diameter of the left blood vessel was normal, the stenosis rate near the right common carotid artery bifurcation was about 56%, the intima on both sides was thickened and not smooth, the color blood flow filling was acceptable, and multiple strong echo plaques were visible in both common carotid arteries, the larger one on the right was about 0.27×0.10cm in size, and the larger one on the left was about 0.29×0.06cm in size, and the blood flow velocity of the bilateral common carotid arteries was slightly reduced.

[0073] Case 3: Qi XX, male, 74 years old.

[0074] Chief complaint: On April 19, 2024, a physical examination revealed sclerosis-like changes in bilateral carotid arteries with plaque formation, and the patient came for treatment.

[0075] Physical examination: right common carotid artery inner diameter 0.56cm, intima-media thickness 0.8mm, peak velocity 0.65m / s. Left common carotid artery inner diameter 0.56cm, intima-media thickness 0.8mm, peak velocity 0.73m / s. Right internal carotid artery inner diameter 0.40cm, peak velocity 0.43m / s. Left internal carotid artery inner diameter 0.42cm, peak velocity 0.50m / s. Right vertebral artery inner diameter 0.38cm, peak velocity 0.25m / s. Left vertebral artery inner diameter 0.35cm, peak velocity 0.28m / s. The echo of the bilateral common carotid artery and internal carotid artery was rough, mixed echo plaques were seen in the right carotid artery, mainly with strong echoes, the largest of which was about 2.3×1.9mm, mixed echo plaques were seen in the left carotid artery, mainly with strong echoes, the largest of which was about 5.2×2.6mm, the local femoral stenosis was about <50%, the color blood flow filling was good, and the maximum blood flow peak velocity was within the normal range. The echo of the bilateral vertebral artery wall was rough, the color blood flow filling was good, and the maximum blood flow peak velocity was within the required range.

[0076] Diagnosis: Bilateral carotid artery sclerosis-like changes and plaque formation (stenosis rate <50%); bilateral vertebral artery sclerosis-like changes.

[0077] Medication records and treatment effects:

[0078] The Chinese medicine compound exosome dressing of the present invention was applied once a day, and a course of treatment was seven days. After 10 days of application, ultrasound imaging was performed again. The bilateral carotid arteries had normal vascular diameters, thickened intima, about 0.19 cm thick, not smooth, and good color blood flow filling. Multiple strong echo plaques were visible in both common carotid arteries, the larger one on the right was about 0.21×0.04 cm in size, and the larger one on the left was about 0.37×0.12 cm in size. The blood flow velocity of the bilateral common carotid arteries was normal.

[0079] In summary, the dressing prepared by the present invention is a new type of dressing that is a mixture of Chinese medicine preparations and exosomes. It is easy to use, directly acts on the thrombus site, and has obvious effects. It reduces the side effects of oral drugs and the trauma of surgical operations, and greatly reduces the cost of treating diseases for patients. It can effectively solve the problems in the current treatment of mural thrombosis and has the clinical significance of Chinese medicine.

[0080] The above description is only a preferred embodiment of the present invention and is not intended to limit the scope of the present invention. For those skilled in the art, the present invention may have various modifications and variations. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. Use of a compound preparation of exosomes in the preparation of a drug for treating mural thrombus, wherein the raw materials of the compound preparation include: Chuanxiong, peach kernel, Chinese angelica, safflower, Panax notoginseng, salvia miltiorrhiza, honeysuckle, red peony root, scutellaria baicalensis, sappan wood, zedoaria, frankincense, myrrh, horseflies, leeches, blister beetles and strychnos nux vomica; The weight of the raw materials of the compound preparation is: 5g of Chuanxiong, 5g of peach kernel, 5g of angelica, 5g of safflower, 5g of Panax notoginseng, 5g of Salvia miltiorrhiza, 5g of honeysuckle, 5g of red peony root, 5g of scutellaria, 10g of Sappan wood, 10g of Zedoaria, 10g of frankincense, 5g of myrrh, 5g of horsefly, 5g of leech, 5g of mylabris, and 5g of Strychnos nux vomica. The method for preparing the compound preparation exosomes comprises the following steps: Step 1: Exosome extraction: 17 Chinese medicines in the compound preparation were weighed according to weight fraction, mixed, crushed into 3-5 mm particles, then soaked in 2000 mL warm water at 37°C for 2 h, crushed with a beater for 30 min, and the operation of crushing for 1 min and stopping for 1 min was repeated to obtain a suspension; the obtained suspension was placed at 4°C for 2 h, then centrifuged at 4000 rpm, the precipitate was discarded, and the supernatant A was collected; Step 2: Exosome concentration: Use ultrafiltration stick to concentrate exosomes; (1) The inlet and outlet of an ultrafiltration rod with a pore size of 200 nm are connected to the supernatant A collected in step 1 through a pipeline, and the supernatant is circulated by a pump. At the same time, the first filtrate B is collected at the filtrate receiving port until the residual volume of supernatant A reaches 30 mL, and then the collection is stopped; (2) The collected first filtrate B is concentrated using an ultrafiltration stick with a pore size of 30 nm according to the steps described in (1) above, until the residual volume of the first filtrate B reaches 30 mL, and the operation is stopped to obtain the residual liquid, i.e., the concentrated liquid C; Step 3 Exosome separation: Take an exclusion chromatography column tube, pump the concentrated solution C obtained in step 2 into the chromatography column at a flow rate of 2 mL / min, and continue to pump purified water at this flow rate after all the liquid enters the chromatography column, and receive the liquid flowing out of the chromatography column, 30 mL for each fraction; a total of 8 fractions are collected, and fractions 3, 4, and 5 are exosome fractions, which are combined to obtain exosomes of the Chinese medicine compound preparation.

2. The use according to claim 1, characterized in that The drug further comprises pharmaceutically acceptable excipients.

3. The use according to claim 1, characterized in that The dosage form of the medicine is a dressing.

4. A method for preparing exosomes from a Chinese medicine compound preparation, characterized in that: The method specifically comprises the following steps: Step 1: Extraction of exosomes: Weigh the 17 Chinese medicines in the compound preparation according to weight fraction, mix them, crush them into 3-5 mm particles, and then soak them in 2000 mL warm water at 37°C for 2 hours, crush them with a beater for 30 minutes, repeat the operation of crushing for 1 minute and stopping for 1 minute to obtain a suspension; place the obtained suspension at 4°C for 2 hours, and then centrifuge it at 4000 rpm, discard the precipitate, and collect the supernatant A; the raw materials and weight fractions of the Chinese medicine compound preparation are: 5g of Chuanxiong, 5g of peach kernel, 5g of angelica, 5g of safflower, 5g of Panax notoginseng, 5g of Salvia miltiorrhiza, 5g of honeysuckle, 5g of red peony root, 5g of scutellaria, 10g of Sappanwood, 10g of Zedoariae, 10g of frankincense, 5g of myrrh, 5g of horseflies, 5g of leeches, 5g of mylabris, and 5g of Strychnos nux vomica; Step 2: Exosome concentration: Use ultrafiltration stick to concentrate exosomes; (1) The inlet and outlet of an ultrafiltration rod with a pore size of 200 nm are connected to the supernatant A collected in step 1 through a pipeline, and the supernatant is circulated by a pump. At the same time, the first filtrate B is collected at the filtrate receiving port until the residual volume of supernatant A reaches 30 mL, and then the collection is stopped; (2) The collected first filtrate B is concentrated using an ultrafiltration stick with a pore size of 30 nm according to the steps described in (1) above, until the residual volume of the first filtrate B reaches 30 mL, and the operation is stopped to obtain the residual liquid, which is the concentrated liquid C; Step 3 Exosome separation: Take an exclusion chromatography column tube, pump the concentrated solution C obtained in step 2 into the chromatography column at a flow rate of 2 mL / min, and continue to pump purified water at this flow rate after all the liquid enters the chromatography column, and receive the liquid flowing out of the chromatography column, 30 mL for each fraction; a total of 8 fractions are collected, and fractions 3, 4, and 5 are exosome fractions, which are combined to obtain exosomes of the Chinese medicine compound preparation.

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