Paris polyphylla exosome, preparation method thereof and application of paris polyphylla exosome in treatment of psoriasis

Abstract

The invention belongs to the technical field of biomedicine, and particularly relates to a paris polyphylla exosome, a preparation method thereof and application of the paris polyphylla exosome in treatment of psoriasis, in the method, fresh paris polyphylla rhizomes are cleaned and washed with a buffering agent, the washed paris polyphylla rhizomes and the buffering agent are mixed according to the mass ratio of 1: (1.5-2.5), after mixing, crushing and filtering are conducted, and the paris polyphylla exosome is obtained; the preparation method comprises the following steps: extracting paris polyphylla from paris polyphylla to obtain a paris polyphylla crude extract for later use, carrying out differential centrifugation on the paris polyphylla crude extract, taking supernate and concentrating to obtain an exosome crude extract, sequentially adding the exosome crude extract into cane sugar with the concentration of 15%, 30%, 45% and 60%, and carrying out cane sugar gradient purification to obtain the paris polyphylla exosome. The paris polyphylla exosome is in the shape of a cup-shaped circle and has a complete double-layer membrane structure, the protein concentration is 3.5-4.5 ug / ml, the particle size is 100-200 nm, and the paris polyphylla exosome is beneficial to improvement of psoriasis, is good in curative effect and fast in effect taking, and has high economical efficiency and safety.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and particularly relates to Paris polyphylla exosomes, a preparation method thereof, and an application in the treatment of psoriasis. Background Art

[0002] Psoriasis is a chronic, recurrent, inflammatory, systemic skin disease mediated by the immune system. Clinically, it is mainly manifested as erythemas of different sizes throughout the body covered with a large amount of white scales. The course of the disease is protracted, easy to relapse, and difficult to cure. In addition, psoriasis patients often suffer from complications such as cardiovascular diseases, metabolic syndrome, cancer, and depression. These complications synergistically increase the physical and psychological burdens of patients and also bring a huge social burden. According to epidemiological surveys, there are more than 60 million psoriasis patients globally, and the incidence of psoriasis in China is also increasing year by year. At present, the main clinical treatment methods for psoriasis are topical medications, systemic treatment, and physical therapy, etc. However, they have adverse reactions such as inducing potential infections and are expensive, and the indications need to be strictly grasped when using them. Therefore, it is particularly important to urgently find an economical, safe, and effective drug for the treatment of psoriasis. Summary of the Invention

[0003] In view of this, the present invention provides a Paris polyphylla exosome, a preparation method thereof, and an application in the treatment of psoriasis, so as to solve the technical problems of existing topical medications, systemic treatment, and physical therapy, which have adverse reactions such as inducing potential infections and are expensive.

[0004] To achieve the above object, the present application adopts the following scheme: A method for preparing Paris polyphylla exosomes, comprising the following steps: S10. Wash the fresh rhizome of Paris polyphylla clean, and wash it with a buffer. After washing, mix it with the buffer according to a mass ratio of 1:(1.5 to 2.5), and after mixing, break and filter it to obtain a crude extract of Paris polyphylla for standby; S20. Perform differential centrifugation on the crude extract of Paris polyphylla, take the supernatant, and concentrate the supernatant to obtain a crude exosome extract; S30. Perform sucrose gradient purification on the crude exosome extract to obtain Paris polyphylla exosomes. The Paris polyphylla exosomes are in a cup-shaped circular shape and have a complete bilayer membrane structure, with a protein concentration of 3.5 ug / ml to 4.5 ug / ml and a particle size of 100 nm to 200 nm. The purification gradient is 15%, 30%, 45%, and 60%.

[0005] Preferably, in the step S10, the buffer is phosphate buffered saline, and the concentration of the phosphate buffered saline is PBS1×.

[0006] Preferably, S20 specifically includes the following steps: S21. Centrifuge the crude extract of Paris polyphylla Smith at 1000 g for 10 min, discard the precipitate, and take the supernatant. S22. Centrifuge the supernatant obtained in S21 at 2000 g for 30 min, discard the precipitate, and take the supernatant. S23. Put the supernatant obtained in S22 into an ultrafiltration tube, centrifuge at 3500 r for 30 min for concentration to obtain a concentrated solution. S24. Centrifuge the concentrated solution at 10000 g for 1 h, take the supernatant, filter the supernatant under 0.22 um aqueous phase, take the filtrate, centrifuge the filtrate at 100000 g for 1 h, take the precipitate, and add a buffer to the precipitate for resuspension to obtain a crude extract of exosomes.

[0007] Preferably, S30 is specifically: sequentially add the crude extract of exosomes to sucrose with concentrations of 15%, 30%, 45%, and 60%, centrifuge at 150000 g for 2 h, aspirate the liquid of the corresponding gradient band, centrifuge the gradient band liquid at 150000 g for 1 h to remove sucrose, take the precipitate, add an appropriate amount of buffer to the precipitate for resuspension, and filter under 0.22 um aqueous phase to obtain Paris polyphylla Smith exosomes.

[0008] A pharmaceutical composition comprising Paris polyphylla Smith exosomes prepared by the above method for preparing Paris polyphylla Smith exosomes.

[0009] Preferably, it further comprises a pharmaceutically acceptable carrier.

[0010] Use of Paris polyphylla Smith exosomes prepared by the above method for preparing Paris polyphylla Smith exosomes in the medicament for treating psoriasis.

[0011] In the above-mentioned Paris polyphylla exosomes, their preparation method and application in treating psoriasis, through steps such as washing, disruption, differential centrifugation and sucrose gradient purification, exosomes are effectively extracted and purified from the rhizome of Paris polyphylla. During the preparation process, through reasonable operating conditions and parameter settings, the biological activity and function of exosomes are maintained as much as possible. The obtained Paris polyphylla exosomes have a specific morphology (cup-shaped circle), structure (intact bilayer membrane structure) and protein concentration (3.5 ug / ml to 4.5 ug / ml), as well as a particle size range (100 nm to 200 nm), thus obtaining Paris polyphylla exosomes with relatively high purity, reducing the influence of impurities on subsequent experiments and applications. During the preparation process, the biological activity of exosomes is effectively maintained, providing strong support for subsequent functional research and applications. After treating psoriasis mice with the obtained Paris polyphylla exosomes for a period of time, it can significantly reduce the infiltration of inflammatory cells under the skin of psoriasis mice, indicating that Paris polyphylla exosomes can reduce the inflammatory response of psoriasis, and can also inhibit the angiogenesis of psoriasis mice, thereby helping to improve the vasodilation manifestation of psoriasis, and can also improve the inflammatory factors in psoriasis mice, indicating that Paris polyphylla exosomes can treat psoriasis by regulating the immune inflammatory response, and also has the effect of inhibiting epidermal thickening. Therefore, Paris polyphylla exosomes help to improve psoriasis, and the curative effect is close to that of methotrexate. It can be seen that it has a fast onset, good curative effect and less dosage.

[0012] Since plant cells are easy to culture, large-scale production can be achieved, so that the extraction cost and production cost of the Paris polyphylla exosomes provided by the present invention are relatively low. Moreover, the Paris polyphylla exosomes provided by the present invention have a stable lipid bilayer structure, which can protect their contents from being damaged by the external environment. This enables them to maintain their biological activity during storage and transportation, thereby reducing the storage and transportation costs and having high economy.

[0013] In addition, Paris polyphylla exosomes are derived from plant cells and have great differences from human cells, so their immunogenicity is relatively low. This means that when using Paris polyphylla exosomes to treat psoriasis, the risk of triggering a human immune response is relatively low. At the same time, the toxicity of Paris polyphylla exosomes is also relatively low, which enables them to reduce the harm to the human body during the treatment of psoriasis, thereby improving the safety of treatment. Brief Description of the Drawings

[0014] Figure 1 It is a schematic diagram of the preparation of Paris polyphylla exosomes in the present invention.

[0015] Figure 2 It is a morphological diagram of the transmission electron microscope characterization of Paris polyphylla exosomes in the present invention.

[0016] Figure 3Representative back skin photos of mice in each group on the 7th day in the present invention.

[0017] Figure 4 Graph showing the total PASI score results of mice in the present invention.

[0018] Figure 5 Graph showing the histopathological observation of skin tissues of mice in each group in the present invention.

[0019] Figure 6 Graph showing the expression results of IL-17A in mice in each group in the present invention. Detailed implementation manners

[0020] To make the objectives and technical solutions of the present invention clearer, the content of the present invention will be further described in detail through experiments. However, it should not be understood that the scope of the above-mentioned subject matter of the present invention is limited to the following experiments only. Any corresponding replacement or modification made according to the common general knowledge and customary means in the art without departing from the above-mentioned technical solutions of the present invention is included in the present invention.

[0021] Please refer to Figure 1 and Figure 2 , in a specific embodiment, the preparation method of Paris polyphylla exosomes is as follows: Step 1: Select fresh and pest-free rhizomes of Paris polyphylla as the starting material, wash the soil, impurities, etc. on the surface of the rhizomes with an appropriate amount of clean water. Subsequently, further wash the rhizomes with a buffer (such as phosphate buffer PBS) to remove surface microorganisms, residual pesticides, etc.

[0022] Step 2: Mix the washed rhizomes with the buffer in a mass ratio of 1:(1.5 to 2.5), preferably 1:2. The buffer is preferably phosphate buffered saline (PBS1×). Use an appropriate crushing device (such as a grinder, an ultrasonic crusher, etc.) to crush the mixture to release the components inside the cells. After crushing, filter to remove large particle impurities, and obtain the crude extract of Paris polyphylla for standby.

[0023] Step 3: Perform differential centrifugation on the crude extract of Paris polyphylla to separate components of different sizes and densities. In this embodiment, by adjusting the centrifugation speed and centrifugation time, impurities such as cell debris and macromolecular substances can be gradually removed. Finally, obtain the supernatant and concentrate the supernatant to increase the concentration of exosomes, and obtain the crude exosome extract.

[0024] Step 4: Purify the crude exosome extract by sucrose gradient, specifically: Prepare sucrose solutions with concentration gradients of 15%, 30%, 45%, and 60%. Add the crude exosome extract to the above sucrose gradients in sequence, and then centrifuge at 150,000 g for 2 h. During this process, the exosomes will layer in the sucrose gradient according to density and size. After centrifugation, aspirate the liquid corresponding to the gradient band, which is rich in exosomes. Then, centrifuge again at 150,000 g for 1 h to remove sucrose and other impurities. After taking the precipitate, add an appropriate amount of buffer to resuspend it, so that the exosomes are dispersed in the buffer. Finally, filter under 0.22 um aqueous phase to remove possible residual impurities and microorganisms. After filtration, the collected liquid is the Paris polyphylla exosomes.

[0025] The purpose of this preparation method is to extract and purify exosomes with specific morphology, structure, and protein concentration from fresh Paris polyphylla rhizomes. Through steps such as washing, disruption, differential centrifugation, and sucrose gradient purification, this method effectively extracts and purifies exosomes from Paris polyphylla rhizomes. During the preparation process, through reasonable operating conditions and parameter settings, the biological activity and function of exosomes are maintained as much as possible. The obtained Paris polyphylla exosomes have a specific morphology (cup-shaped circle), structure (intact bilayer membrane structure), and protein concentration (3.5 ug / ml to 4.5 ug / ml), as well as a particle size range (100 nm to 200 nm), obtaining Paris polyphylla exosomes with higher purity, reducing the influence of impurities on subsequent experiments and applications. During the preparation process, the biological activity of exosomes is effectively maintained, providing strong support for subsequent functional research and applications.

[0026] Through steps such as washing and disruption, intracellular components including exosomes are effectively extracted from Paris polyphylla rhizomes. Through steps such as differential centrifugation and sucrose gradient purification, impurities are removed to improve the purity and concentration of exosomes. The obtained Paris polyphylla exosomes have a specific morphology (cup-shaped circle), structure (intact bilayer membrane structure), and protein concentration (3.5 ug / ml to 4.5 ug / ml), as well as a particle size range (100 nm to 200 nm), providing a reliable material basis for subsequent research and applications. Through this preparation method, Paris polyphylla exosomes with higher purity can be obtained, reducing the influence of impurities on subsequent experiments and applications. During the preparation process, through reasonable operating conditions and parameter settings, the biological activity and function of exosomes are maintained as much as possible. The obtained Paris polyphylla exosomes have broad application prospects in the fields of biology, medicine, etc., such as being used as drug carriers for disease treatment and as biomarkers for disease diagnosis. At the same time, it also provides strong support for in-depth research on the bioactive components and action mechanisms of Paris polyphylla.

[0027] Among them, the specific steps of S20 include: centrifuging the crude extract of Paris polyphylla Smith at 1000 g for 10 min to remove large particulate impurities and cell debris in the crude extract. These impurities usually have a relatively high density and will precipitate to the bottom during centrifugation, obtaining a supernatant which mainly contains intracellular soluble components and small particulate matter. Centrifuge the obtained supernatant at 2000 g for 30 min to further remove medium-sized particles and impurities in the supernatant, so as to improve the purity of exosomes in subsequent steps, and obtain the supernatant again. At this time, the supernatant mainly contains exosomes and other small molecular components. Put the supernatant obtained after secondary centrifugation into an ultrafiltration tube and centrifuge it at 3500 r (usually corresponding to a certain centrifugal force, but expressed as the rotational speed here, and it needs to be adjusted according to the specifications of the ultrafiltration tube and the model of the centrifuge in actual operation) for 30 min. Through ultrafiltration, small molecular substances and solvents in the supernatant are removed, and the exosomes are concentrated to obtain a concentrated solution, in which the concentration of exosomes is significantly increased. Centrifuge the concentrated solution at 10000 g for 1 h, take the supernatant, then filter the supernatant under 0.22 um aqueous phase, take the filtrate, and finally centrifuge the filtrate at 100000 g for 1 h, take the precipitate. Through deep centrifugation and filtration, impurities and small particles in the supernatant are further removed, and at the same time the exosomes precipitate. The 0.22 um filtration step can remove most microorganisms and cell debris to ensure the purity of exosomes. Next, add an appropriate amount of buffer to the precipitate for resuspension to obtain a crude extract of exosomes. The purpose of this step is to disperse the exosomes in the buffer for subsequent processing and analysis. Through the treatment of this step (S20), a crude extract of exosomes with relatively high purity can be obtained.

[0028] The present invention also provides a pharmaceutical composition, comprising the Paris polyphylla Smith exosomes prepared by the above preparation method.

[0029] The core component of the pharmaceutical composition provided by the present invention is the Paris polyphylla Smith exosomes prepared by the above method. This pharmaceutical composition not only contains the active ingredients of Paris polyphylla Smith, but also utilizes the unique delivery mechanism of exosomes, aiming to improve the bioavailability, targeting and efficacy of the drug. This pharmaceutical composition has broad application prospects in the fields of skin diseases and inflammatory diseases, etc., and provides new means for the treatment of these diseases by regulating the immune system, inhibiting inflammatory reactions, etc.

[0030] Furthermore, it further comprises a pharmaceutically acceptable carrier.

[0031] Pharmaceutically acceptable carriers are used to combine with Paris polyphylla exosomes to form stable solid or liquid dosage forms. This combination can not only protect the active ingredients of Paris polyphylla exosomes, but also improve the bioavailability of the drug and patient compliance. For example, Paris polyphylla exosomes can be mixed with appropriate fillers such as lactose and mannitol, and appropriate solvents such as water or ethanol can be added to make dosage forms such as tablets, capsules or oral liquids.

[0032] Use of Paris polyphylla exosomes prepared by the above-mentioned preparation method of Paris polyphylla exosomes in the treatment of psoriasis drugs.

[0033] It is proved by the following examples that after treating psoriasis mice with the obtained Paris polyphylla exosomes for a period of time, it can significantly reduce the infiltration of inflammatory cells under the skin of psoriasis mice, indicating that Paris polyphylla exosomes can reduce the inflammatory reaction of psoriasis, and can also inhibit the angiogenesis of psoriasis mice, thereby helping to improve the vasodilation performance of psoriasis, and can also improve the inflammatory factors in psoriasis mice, indicating that Paris polyphylla exosomes can treat psoriasis by regulating the immune inflammatory reaction, and also has the effect of inhibiting epidermal thickening. Therefore, Paris polyphylla exosomes help to improve psoriasis, and the curative effect is close to that of methotrexate. It can be seen that it has a fast onset, good curative effect and less dosage.

[0034] Since plant cells are easy to culture, large-scale production can be achieved, so that the extraction cost and production cost of Paris polyphylla exosomes provided by the present invention are relatively low. Moreover, the Paris polyphylla exosomes provided by the present invention have a stable lipid bilayer structure, which can protect their contents from being damaged by the external environment, which enables them to maintain their biological activity during storage and transportation, thereby reducing the storage and transportation costs and having high economy.

[0035] In addition, Paris polyphylla exosomes are derived from plant cells and have great differences from human cells, so their immunogenicity is relatively low, which means that when using Paris polyphylla exosomes to treat psoriasis, the risk of triggering human immune response is relatively low. At the same time, the toxicity of Paris polyphylla exosomes is also relatively low, which enables it to reduce the harm to the human body during the treatment of psoriasis, thereby improving the safety of treatment.

[0036] The following specific experimental examples are used to further illustrate the technical solutions and effects of the present invention. It should be noted that the inventors have carried out relevant pharmacodynamic experimental studies to prove the efficacy of Paris polyphylla exosomes in the treatment of psoriasis. It should be noted that the following experimental studies are all carried out on the basis of proving the drug safety, and the drug administration doses in the experimental studies are all within the safe dose range. The drugs selected in the following pharmacodynamic tests are the drugs obtained from the representative formulations of the present invention and their preparation methods; the inventors have also carried out pharmacodynamic experiments on other drugs, and the experimental results show the same or similar effects, but due to space limitations, they are not listed one by one here.

[0037] 1. Drug materials Methotrexate (from the market); 5% IMQ ointment (from the market); Paris polyphylla exosomes prepared by the Paris polyphylla exosome preparation method provided by the present invention (prepared by the laboratory of Ningxia Medical University).

[0038] 2. Related technical solutions 2.1 Collection of experimental animals SPF-grade male BALB / c mice with a body weight of 18 g to 22 g are selected, provided by the Experimental Animal Center of Ningxia Medical University, and are freely given water and fed a fixed amount of mouse feed in an animal house with ventilation, dryness, a room temperature of 22°C to 25°C, and a humidity of 50% to 70%.

[0039] 2.2 Grouping of experimental animals Take 28 SPF-grade BALB / c mice that have passed the quarantine, and randomly divide them into 4 groups, namely the normal group, the model group, the methotrexate group, and the Paris polyphylla exosome group.

[0040] 2.3 Model establishment After 3 days of adaptive feeding, the SPF-grade male BALB / c mice in the above-mentioned model group, methotrexate group, and Paris polyphylla exosome group are used to establish a classical IMQ-induced psoriasis-like skin lesion model. One day before the experiment, the backs of the mice are subjected to relevant hair removal treatments. First, the hair on the backs of the mice is basically removed using a pet hair clipper, and then Veet hair removal cream is used to thoroughly remove the remaining back fluff, obtaining a 2*3 cm smooth back skin of the mice. The treated mice are normally fed for 2 days to eliminate the possible damage to the back skin of the mice caused by the hair clipper and hair removal cream; The mice in the normal group are smeared with 62.5 mg of white petrolatum on the back skin every day, and 0.2 mL of normal saline is given by gavage 1 hour later, for 7 consecutive days; The mice in the model group were topically applied with 62.5 mg of 5% (mass fraction) IMQ ointment (imiquimod) daily. One hour later, they were intragastrically administered 0.2 mL of normal saline for 7 consecutive days. The mice in the methotrexate group were topically applied with 62.5 mg of 5% (mass fraction) IMQ ointment daily. One hour later, they were intragastrically administered 1 mg / kg of methotrexate for 7 consecutive days. The mice in the Paris polyphylla Smith exosome group were topically applied with 62.5 mg of 5% (mass fraction) IMQ ointment daily. One hour later, they were intragastrically administered 5 mg / kg of Paris polyphylla Smith exosomes for 7 consecutive days. Before each administration to the mice in each of the above groups, photographic records were taken. After the last administration, as Figure 3 shown, the mice were sacrificed by cervical dislocation.

[0041] 2.4 PASI score The visual observation method was used to observe whether characteristic changes of psoriasis such as erythema, skin lesion hypertrophy, and scales appeared on the backs of the mice in each group. The evaluation was carried out according to the internationally accepted evaluation method for the area and severity of psoriasis skin lesions (PASI). The evaluation results are as Figure 4 shown. The specific judgment criteria are shown in Table 1: Table 1 PASI scoring table

[0042] It was Figure 3 and Figure 4 reflected that compared with the normal group, the mice in the model group showed obvious erythema, scales, and thickness, indicating successful modeling; compared with the model group, the symptoms of skin erythema, scales, and thickness in the mice of the Paris polyphylla Smith exosome group were significantly improved (see Figure 3 ), and the total PASI score was significantly reduced (see Figure 4 ), almost on a par with the total score of the methotrexate group, indicating that its efficacy was close to that of methotrexate. This result shows that Paris polyphylla Smith exosomes can effectively relieve the skin lesion symptoms of psoriasis in mice.

[0043] 2.5 H&E staining The pathological changes of the back skin of the mice in each group were analyzed by H&E staining. The results are as Figure 5 shown.

[0044] It was Figure 5 found that compared with the normal group mice, the epidermal thickness of the model group mice was significantly increased, the subcutaneous inflammatory cell infiltration was significantly increased, and the number of newly formed capillaries increased. Compared with the model group, the epidermal thickness of the mice in the Paris polyphylla Smith exosome group was significantly reduced, the subcutaneous inflammatory cell infiltration was significantly reduced, and the number of newly formed capillaries decreased.

[0045] The epidermal thickness of mice in the Paris polyphylla Smith exosome group was significantly reduced, indicating that Paris polyphylla Smith exosomes may have the effect of inhibiting epidermal thickening, thus contributing to the improvement of the skin pathological manifestations of psoriasis; the infiltration of subcutaneous inflammatory cells in mice in the Paris polyphylla Smith exosome group was significantly reduced, indicating that Paris polyphylla Smith exosomes may have an anti-inflammatory effect and can reduce the inflammatory response of psoriasis; the number of newly formed capillaries in mice in the Paris polyphylla Smith exosome group was reduced, indicating that Paris polyphylla Smith exosomes may have the effect of inhibiting angiogenesis, thus contributing to the improvement of the vascular abnormalities in psoriasis.

[0046] 2.6 Immune inflammatory factors The levels of IL-17A in the sera of mice in each of the above groups were detected by ELISA, and the detection results are as Figure 6 shown.

[0047] It is Figure 6 reflected that, compared with the model group, the expression of IL-17A in the Paris polyphylla Smith exosome group was significantly decreased (p < 0.01), indicating that Paris polyphylla Smith exosomes may treat psoriasis by regulating the immune inflammatory response.

[0048] 3. Experimental results Paris polyphylla Smith exosomes can significantly reduce the infiltration of subcutaneous inflammatory cells, indicating that they can reduce the inflammatory response of psoriasis; they can also inhibit angiogenesis, thus contributing to the improvement of the vascular dilation in psoriasis; and they can also improve inflammatory factors, treat psoriasis by regulating the immune inflammatory response, and have the effect of inhibiting epidermal thickening; considering the above points, Paris polyphylla Smith exosomes contribute to the improvement of psoriasis.

[0049] As described above, it is only the specific implementation manners of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art within the technical scope disclosed by the present invention can easily think of various equivalent modifications or substitutions, and these modifications or substitutions should be covered within the protection scope of the present invention.

Claims

1. A preparation method of Paris polyphylla exosomes, characterized in that, It includes the following steps: S10. Clean the fresh Paris polyphylla rhizome, and wash it with a buffer. After washing, mix it with the buffer according to a mass ratio of 1:(1.5 to 2.5), break and filter the mixture after mixing to obtain the crude extract of Paris polyphylla for standby; S20. Perform differential centrifugation on the crude extract of Paris polyphylla, take the supernatant, and concentrate the supernatant to obtain the crude exosome extract; S30. Add the crude exosome extract to sucrose with concentrations of 15%, 30%, 45%, and 60% in sequence, centrifuge at 150,000 g for 2 h, aspirate the liquid of the corresponding gradient band, centrifuge the gradient band liquid at 150,000 g for 1 h to remove sucrose, take the precipitate, add an appropriate amount of buffer to the precipitate for resuspension, and filter it under 0.22 um aqueous phase to obtain the exosomes of Paris polyphylla. The exosomes of Paris polyphylla are in a cup-shaped circular form and have a complete double-layer membrane structure, with a protein concentration of 3.5 ug / ml to 4.5 ug / ml and a particle size of 100 nm to 200 nm.

2. The preparation method of Paris vietnamensis exosomes according to claim 1, characterized in that, In the step S10, the buffer is phosphate buffered saline, and the concentration of the phosphate buffered saline is PBS1×.

3. The preparation method of Paris vietnamensis exosomes according to claim 1, characterized in that, The step S20 specifically includes the following steps: S21. Centrifuge the crude extract of Paris polyphylla at 1000 g for 10 min, discard the precipitate, and take the supernatant; S22. Centrifuge the supernatant taken in S21 at 2000 g for 30 min, discard the precipitate, and take the supernatant; S23. Put the supernatant taken in S22 into an ultrafiltration tube and centrifuge it at 3500 r for 30 min for concentration to obtain a concentrated solution; S24. Centrifuge the concentrated solution at 10,000 g for 1 h, take the supernatant, filter the supernatant under 0.22 um aqueous phase, take the filtrate, centrifuge the filtrate at 100,000 g for 1 h, take the precipitate, and add a buffer to the precipitate for resuspension to obtain the crude exosome extract.

4. A pharmaceutical composition, characterized in that, It includes the exosomes of Paris polyphylla prepared by the method for preparing exosomes of Paris polyphylla according to any one of claims 1 to 3.

5. The pharmaceutical composition according to claim 4, wherein, It also includes a pharmaceutically acceptable carrier.

6. Use of the exosomes of Paris polyphylla prepared by the method for preparing exosomes of Paris polyphylla according to any one of claims 1 to 3 for the treatment of psoriasis.

Images