Composition containing exosome and application of composition in preparation of products for treating hepatic failure

Abstract

The invention relates to an exosome-containing composition, which is prepared by blending a modified silicon dioxide agent and an exosome matrix according to a weight ratio of (2-5): 15, the exosome matrix is composed of a mesenchymal stem cell source exosome and a liver stem cell source exosome, and the mass ratio of the mesenchymal stem cell source exosome to the liver stem cell source exosome is 1: (0.5-2). The exosome is a core functional component which plays a role in treating hepatic failure, the biological activity of the exosome is greatly improved through double-source selection and hypoxia pretreatment, the double-stem-cell exosome has a synergistic effect, proliferation and repair of damaged hepatic cells can be directly promoted, and liver functional cells can be supplemented; through compounding of the two exosomes, triple synergy of inhibiting necrosis, promoting regeneration and regulating immunity can be realized, and the problems of single function and limited repair efficiency of a single exosome are solved.

Description

Technical Field

[0001] This invention relates to the field of exosome technology, and more specifically to a composition containing exosomes and its use in the preparation of products for treating liver failure. Background Technology

[0002] Liver failure is a severe liver disease caused by multiple factors, characterized by massive hepatocellular necrosis and severely impaired liver function. Clinical manifestations include jaundice, coagulation disorders, hepatic encephalopathy, and ascites. The disease progresses rapidly and has a high mortality rate. Currently, the main clinical treatments for liver failure include comprehensive medical treatment, artificial liver support therapy, and liver transplantation. Among these, comprehensive medical treatment focuses on symptomatic support and is difficult to reverse massive hepatocellular necrosis; artificial liver support therapy can temporarily replace some liver function and alleviate the condition, but it cannot fundamentally repair damaged liver tissue; liver transplantation, as an effective treatment for end-stage liver failure, is limited by donor shortages, high surgical risks, and postoperative immune rejection, making its widespread application difficult; exosomes are vesicle structures with a diameter of approximately 30-150 nm secreted by cells, containing bioactive substances such as proteins, lipids, and nucleic acids (e.g., mRNA, miRNA), and play a role in mediating intercellular signal transduction and regulating cellular physiological functions. In recent years, research has revealed unique advantages of exosomes in tissue repair and immune regulation, particularly in the treatment of liver diseases, where they have shown great potential. For example, exosomes derived from mesenchymal stem cells can inhibit hepatocyte apoptosis and promote hepatocyte regeneration through their carried miRNAs, while also regulating the local immune microenvironment in the liver and reducing inflammatory responses. However, existing technologies for treating liver failure with exosomes have shown poor efficacy, and the products also exhibit limited liver function repair and targeted delivery, restricting their overall efficiency. Therefore, this invention provides further improvements to these technologies. Summary of the Invention

[0003] In view of the deficiencies of the prior art, the purpose of this invention is to provide a composition containing exosomes and its application in the preparation of products for treating liver failure, so as to solve the problems mentioned in the background art.

[0004] The present invention solves the technical problem by adopting the following technical solution: The present invention provides a composition containing exosomes, wherein the composition containing exosomes is a modified silica agent and an exosome matrix, wherein the modified silica agent and the exosome matrix are blended in a weight ratio of (2-5):15; The exosome is composed of exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells, and the mass ratio of the exosomes derived from mesenchymal stem cells to the exosomes derived from liver stem cells is 1: (0.5-2). The preparation methods for exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells are the same, and the specific preparation methods are as follows: P3 generation umbilical cord mesenchymal stem cells were seeded in a 50L bioreactor containing 5-6 g / L microcarriers, and serum-free X-VIVO was added. TM Culture medium 15, incubated at 37-39℃ and 5-7% CO2; When the cell density reaches 1-1.5×10^7 cells / mL, initiate hypoxia pretreatment: reduce the oxygen concentration to 5-7%, culture for 24-36 h, then reduce it to 2-5%, and continue culturing for another 24-36 h; culture liver stem cells (HSCs) using the same method to complete the hypoxia pretreatment.

[0005] Preferably, the specific preparation method of the modified silica agent is as follows: S01: Mix 3-5 parts of carboxymethyl cellulose, 2-3 parts of silicon carbide and 5-8 parts of ethanol solution thoroughly to obtain a modified carboxymethyl cellulose agent; 2-3 parts of silane coupling agent KH550, 1-2 parts of carbon nanotubes and 5-8 parts of modified carboxymethyl cellulose agent are mixed evenly to obtain a modified solution; SO2: Silica and modified liquid are mixed and modified at a weight ratio of 3:(5-7). After mixing, the modified silica liquid is obtained. S03: Modified silica liquid and additives are mixed and ball-milled at a weight ratio of (7-11):5. The ball milling speed is 1000-1500 r / min and the ball milling time is 2h. After the ball milling is completed, the mixture is filtered and dried to obtain modified silica agent.

[0006] Preferably, the stirring speed for the stirring modification treatment is 450-500 r / min, and the stirring time is 1 h.

[0007] Preferably, the ethanol solution is an ethanol solution with a mass fraction of 85-95%.

[0008] Preferably, the additive is prepared by mixing 3-5 parts of flake boron nitride, 2-4 parts of dopamine hydrochloride solution and 2-4 parts of modified sodium alginate thoroughly to obtain the additive.

[0009] Preferably, the mass fraction of the dopamine hydrochloride solution is 5-8%.

[0010] Preferably, the modified sodium alginate agent is prepared by: Add 1-3 parts sodium alginate powder, 2-3 parts kaolin, and 5-8 parts phosphate buffer solution, then add 2-4 parts glucose and 1-2 parts citric acid, stir well, filter and dry to obtain modified sodium alginate agent.

[0011] Preferably, the pH value of the phosphate buffer solution is 5.5-6.5.

[0012] The present invention also provides the use of an exosome-containing composition in the preparation of a product for treating liver failure.

[0013] Compared with the prior art, the present invention has the following beneficial effects: The exosomes of this invention are the core functional components for the treatment of liver failure. This invention significantly enhances their biological activity through dual-source selection and hypoxic pretreatment: synergistic effect of dual stem cell exosomes. The exosomes used are a combination of mesenchymal stem cell-derived exosomes and liver stem cell-derived exosomes in a ratio of 1:(0.5-2), rather than a single source: Mesenchymal stem cell exosomes are known to inhibit hepatocyte apoptosis and regulate the local immune microenvironment of the liver through the miRNAs they carry, and are commonly used "immunomodulatory + anti-apoptotic" carriers in the treatment of liver diseases; Liver stem cell exosomes have stronger "hepatocyte regeneration guidance" and can directly promote the proliferation and repair of damaged hepatocytes and replenish liver functional cells; The combination of the two can achieve a triple synergy of "inhibiting necrosis + promoting regeneration + regulating immunity", which solves the problem of "single function and limited repair efficiency" of single exosomes.

[0014] Hypoxia pretreatment enhances exosome function. Both types of stem cells underwent gradient hypoxia pretreatment (initially cultured at 5-7% oxygen concentration for 24-36 hours, then reduced to 2-5% for another 24-36 hours). Under physiological conditions, the liver in liver failure is often in a hypoxic microenvironment. Hypoxia can induce stem cells to secrete exosomes "more adapted to the hypoxic damage environment"—these exosomes contain significantly increased levels of anti-apoptotic proteins (such as Bcl-2), regeneration-promoting mRNAs (such as HGF mRNA), and immunomodulatory miRNAs (such as miR-124), enabling more precise response to the core pathological aspects of liver failure: massive hepatocellular necrosis + inflammatory storm. The modified silica matrix consists of "silica + modified carboxymethyl cellulose + carbon nanotubes". Composite structure: Silica provides a rigid shell that can physically isolate nucleases and proteases in the blood (preventing the degradation of mRNA and proteins in exosomes); Carboxymethyl cellulose (modified with silicon carbide) has good biocompatibility and hydration properties, and can form a "hydration layer" on the shell surface, reducing the probability of exosomes being phagocytosed and cleared by the immune system (such as macrophages); The addition of carbon nanotubes can enhance the mechanical strength of the carrier and prevent exosomes from being damaged by shear forces during in vivo circulation.

[0015] The modified silica agent incorporates "silane coupling agent KH550 + sheet-like boron nitride + modified sodium alginate": the amino group (-NH2) of silane coupling agent KH550 can form hydrogen bonds with the carboxyl group (-COOH) on the surface of exosomes, achieving stable binding between exosomes and carriers and preventing their separation; sheet-like boron nitride has "liver-targeting affinity" (receptors on the surface of liver endothelial cells can specifically recognize the sheet-like structure of boron nitride), which can guide the carrier to actively accumulate in liver tissue; modified sodium alginate (modified by kaolin, glucose, and citric acid) can enhance the "hepatocyte adhesion" of the carrier, enabling exosomes to quickly attach to the surface of damaged hepatocytes and release them after reaching the liver, increasing the local exosome concentration in the liver and solving the problem of "exosomes diffusing with the blood and low local concentration in the liver".

[0016] Synergistic effect of additives: Further enhances liver function repair. The additives (pigmented boron nitride + dopamine hydrochloride + modified sodium alginate) do not only play a carrier role, but can also synergistically promote liver function repair with exosomes. Modified sodium alginate: Regulates the liver microenvironment. Sodium alginate modified with kaolin and glucose can slowly release glucose to provide energy for hepatocyte regeneration. At the same time, the citric acid it carries can regulate the local pH value of the liver (the liver microenvironment is often acidic in liver failure), restore the normal physiological environment of hepatocytes, and "create favorable conditions" for exosomes to promote hepatocyte regeneration. Compared to "comprehensive medical treatment": medical treatment can only provide symptomatic support (such as reducing jaundice and supplementing clotting factors), and cannot reverse hepatocyte necrosis; while this invention promotes hepatocyte regeneration and inhibits necrosis through exosomes, which can repair the liver from the "etiological level" rather than just relieving symptoms; compared to "artificial liver support therapy": artificial livers can only temporarily replace liver function (such as clearing toxins), and cannot repair liver tissue; this invention can promote liver self-repair and reduce the number of times patients rely on artificial livers; compared to "liver transplantation": liver transplantation is limited by donor shortage and immune rejection; the composition of this invention is "exosomes + biomaterials", which has no donor restrictions, and the immunogenicity of exosomes is extremely low (mesenchymal stem cell exosomes hardly cause immune rejection), making it safer and suitable as a "bridging therapy" before liver transplantation or an "alternative therapy" for patients who are not suitable for transplantation. Detailed Implementation

[0017] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to specific examples. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0018] This embodiment provides a composition containing exosomes, wherein the composition containing exosomes is a modified silica agent and an exosome matrix, wherein the modified silica agent and the exosome matrix are blended in a weight ratio of (2-5):15; The exosome is composed of exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells, and the mass ratio of the exosomes derived from mesenchymal stem cells to the exosomes derived from liver stem cells is 1: (0.5-2). The preparation methods for exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells are the same, and the specific preparation methods are as follows: P3 generation umbilical cord mesenchymal stem cells were seeded in a 50L bioreactor containing 5-6 g / L microcarriers, and serum-free X-VIVO was added. TM Culture medium 15, incubated at 37-39℃ and 5-7% CO2; When the cell density reaches 1-1.5×10^7 cells / mL, initiate hypoxia pretreatment: reduce the oxygen concentration to 5-7%, culture for 24-36 h, then reduce it to 2-5%, and continue culturing for another 24-36 h; culture liver stem cells (HSCs) using the same method to complete the hypoxia pretreatment.

[0019] The specific preparation method of the modified silica agent in this embodiment is as follows: S01: Mix 3-5 parts of carboxymethyl cellulose, 2-3 parts of silicon carbide and 5-8 parts of ethanol solution thoroughly to obtain a modified carboxymethyl cellulose agent; 2-3 parts of silane coupling agent KH550, 1-2 parts of carbon nanotubes and 5-8 parts of modified carboxymethyl cellulose agent are mixed evenly to obtain a modified solution; SO2: Silica and modified liquid are mixed and modified at a weight ratio of 3:(5-7). After mixing, the modified silica liquid is obtained. S03: Modified silica liquid and additives are mixed and ball-milled at a weight ratio of (7-11):5. The ball milling speed is 1000-1500 r / min and the ball milling time is 2h. After the ball milling is completed, the mixture is filtered and dried to obtain modified silica agent.

[0020] In this embodiment, the stirring speed for the stirring modification treatment is 450-500 r / min, and the stirring time is 1 h.

[0021] The ethanol solution in this embodiment is an ethanol solution with a mass fraction of 85-95%.

[0022] The additive in this embodiment is prepared by mixing 3-5 parts of flake boron nitride, 2-4 parts of dopamine hydrochloride solution and 2-4 parts of modified sodium alginate thoroughly to obtain the additive.

[0023] The mass fraction of the dopamine hydrochloride solution in this embodiment is 5-8%.

[0024] The preparation method of the modified sodium alginate agent in this embodiment is as follows: Add 1-3 parts sodium alginate powder, 2-3 parts kaolin, and 5-8 parts phosphate buffer solution, then add 2-4 parts glucose and 1-2 parts citric acid, stir well, filter and dry to obtain modified sodium alginate agent.

[0025] The pH value of the phosphate buffer solution in this embodiment is 5.5-6.5.

[0026] The application of an exosome-containing composition in the preparation of a product for treating liver failure is described in this embodiment.

[0027] Example 1. This embodiment provides a composition containing exosomes, wherein the composition containing exosomes is a modified silica agent and an exosome matrix, wherein the modified silica agent and the exosome matrix are blended in a weight ratio of 2:15; The exosome is composed of exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells, and the mass ratio of the exosomes derived from mesenchymal stem cells to the exosomes derived from liver stem cells is 1:0.5. The preparation methods for exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells are the same, and the specific preparation methods are as follows: P3 generation umbilical cord mesenchymal stem cells were seeded in a 50L bioreactor containing 5g / L microcarriers, and serum-free X-VIVO was added. TM Culture medium 15, cultured at 37℃ and 5% CO2; When the cell density reaches 1×10^7 cells / mL, hypoxia pretreatment is initiated: the oxygen concentration is reduced to 5%, cultured for 24 hours, then reduced to 2%, and cultured for another 24 hours; liver stem cells (HSCs) are cultured using the same method to complete the hypoxia pretreatment.

[0028] The specific preparation method of the modified silica agent in this embodiment is as follows: S01: Mix 3 parts carboxymethyl cellulose, 2 parts silicon carbide and 5 parts ethanol solution thoroughly to obtain modified carboxymethyl cellulose agent; Two parts of silane coupling agent KH550, one part of carbon nanotubes and five parts of modified carboxymethyl cellulose were mixed evenly to obtain a modified solution. SO2: Silica and modified liquid are mixed and modified at a weight ratio of 3:5. After mixing, modified silica liquid is obtained. S03: Modified silica liquid and additives were mixed and ball-milled at a weight ratio of 7:5. The ball milling speed was 1000 r / min and the milling time was 2 h. After the ball milling was completed, the mixture was filtered and dried to obtain modified silica agent.

[0029] In this embodiment, the stirring speed for the stirring modification treatment is 450 r / min, and the stirring time is 1 h.

[0030] The ethanol solution in this embodiment is an 85% ethanol solution by mass.

[0031] The additive in this embodiment is prepared by mixing 3 parts of boron nitride flakes, 2 parts of dopamine hydrochloride solution and 2 parts of modified sodium alginate thoroughly to obtain the additive.

[0032] The mass fraction of the dopamine hydrochloride solution in this embodiment is 5%.

[0033] The preparation method of the modified sodium alginate agent in this embodiment is as follows: Mix 1 part sodium alginate powder, 2 parts kaolin, and 5 parts phosphate buffer solution, then add 2 parts glucose and 1 part citric acid, stir well, filter and dry to obtain modified sodium alginate agent.

[0034] The pH value of the phosphate buffer solution in this embodiment is 5.5.

[0035] The application of an exosome-containing composition in the preparation of a product for treating liver failure is described in this embodiment.

[0036] Example 2. This embodiment provides a composition containing exosomes, wherein the composition containing exosomes is a modified silica agent and an exosome matrix, wherein the modified silica agent and the exosome matrix are blended in a weight ratio of 5:15; The exosome is composed of exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells, and the mass ratio of the exosomes derived from mesenchymal stem cells to the exosomes derived from liver stem cells is 1:2. The preparation methods for exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells are the same, and the specific preparation methods are as follows: P3 generation umbilical cord mesenchymal stem cells were seeded in a 50L bioreactor containing 6g / L microcarriers, and serum-free X-VIVO was added. TM Culture medium 15, cultured at 39℃ and 7% CO2; When the cell density reaches 1.5 × 10^7 cells / mL, hypoxia pretreatment is initiated: the oxygen concentration is reduced to 7%, cultured for 36 h, then reduced to 5%, and cultured for another 36 h; liver stem cells (HSCs) are cultured using the same method to complete the hypoxia pretreatment.

[0037] The specific preparation method of the modified silica agent in this embodiment is as follows: S01: Mix 5 parts carboxymethyl cellulose, 3 parts silicon carbide and 8 parts ethanol solution thoroughly to obtain modified carboxymethyl cellulose agent; Three parts of silane coupling agent KH550, two parts of carbon nanotubes and eight parts of modified carboxymethyl cellulose were mixed evenly to obtain a modified solution. SO2: Silica and modified liquid are mixed and modified at a weight ratio of 3:7. After mixing, the modified silica liquid is obtained. S03: Modified silica liquid and additives were mixed and ball-milled at a weight ratio of 11:5. The ball milling speed was 1500 r / min and the milling time was 2 h. After the ball milling was completed, the mixture was filtered and dried to obtain modified silica agent.

[0038] In this embodiment, the stirring speed for the stirring modification treatment is 500 r / min, and the stirring time is 1 h.

[0039] The ethanol solution in this embodiment is a 95% ethanol solution by mass.

[0040] The additive in this embodiment is prepared by mixing 5 parts of boron nitride flakes, 4 parts of dopamine hydrochloride solution and 4 parts of modified sodium alginate thoroughly to obtain the additive.

[0041] The mass fraction of the dopamine hydrochloride solution in this embodiment is 8%.

[0042] The preparation method of the modified sodium alginate agent in this embodiment is as follows: Add 3 parts sodium alginate powder, 3 parts kaolin, and 8 parts phosphate buffer solution, then add 4 parts glucose and 2 parts citric acid, stir well, filter and dry to obtain modified sodium alginate agent.

[0043] The pH value of the phosphate buffer solution in this embodiment is 6.5.

[0044] The application of an exosome-containing composition in the preparation of a product for treating liver failure is described in this embodiment.

[0045] Example 3. This embodiment provides a composition containing exosomes, wherein the composition containing exosomes is a modified silica agent and an exosome matrix, wherein the modified silica agent and the exosome matrix are blended in a weight ratio of 3.5:15; The exosome is composed of exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells, and the mass ratio of the exosomes derived from mesenchymal stem cells to the exosomes derived from liver stem cells is 1:1.5. The preparation methods for exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells are the same, and the specific preparation methods are as follows: P3 generation umbilical cord mesenchymal stem cells were seeded in a 50L bioreactor containing 5.5g / L microcarriers, and serum-free X-VIVO was added. TM Culture medium 15, cultured at 38℃ and 6% CO2; When the cell density reaches 1.25 × 10^7 cells / mL, hypoxia pretreatment is initiated: the oxygen concentration is reduced to 6%, cultured for 30 h, then reduced to 3.5%, and cultured for another 30 h; liver stem cells (HSCs) are cultured using the same method to complete the hypoxia pretreatment.

[0046] The specific preparation method of the modified silica agent in this embodiment is as follows: S01: Mix 4 parts of carboxymethyl cellulose, 2.5 parts of silicon carbide and 6.5 parts of ethanol solution thoroughly to obtain a modified carboxymethyl cellulose agent; 2.5 parts of silane coupling agent KH550, 1.5 parts of carbon nanotubes and 6.5 parts of modified carboxymethyl cellulose agent were mixed evenly to obtain a modified solution; SO2: Silica and modified liquid are mixed and modified at a weight ratio of 3:6. After mixing, modified silica liquid is obtained. S03: Modified silica liquid and additives were mixed and ball-milled at a weight ratio of 9:5. The ball milling speed was 1250 r / min and the milling time was 2 h. After the ball milling was completed, the mixture was filtered and dried to obtain modified silica agent.

[0047] In this embodiment, the stirring speed for the stirring modification treatment is 470 r / min, and the stirring time is 1 h.

[0048] The ethanol solution in this embodiment is a 90% ethanol solution by mass.

[0049] The additive in this embodiment is prepared by mixing 4 parts of boron nitride flakes, 3 parts of dopamine hydrochloride solution and 3 parts of modified sodium alginate thoroughly to obtain the additive.

[0050] The mass fraction of the dopamine hydrochloride solution in this embodiment is 6.5%.

[0051] The preparation method of the modified sodium alginate agent in this embodiment is as follows: Mix 2 parts sodium alginate powder, 2.5 parts kaolin, and 6.5 parts phosphate buffer solution, then add 3 parts glucose and 1.5 parts citric acid, stir well, filter and dry to obtain modified sodium alginate agent.

[0052] The pH value of the phosphate buffer solution in this embodiment is 6.0.

[0053] The application of an exosome-containing composition in the preparation of a product for treating liver failure is described in this embodiment.

[0054] Comparative Example 1. Unlike Example 3, no modifying liquid was added during the preparation of the modified silica agent.

[0055] Comparative Example 2. Unlike Example 3, no silane coupling agent KH550 or carbon nanotubes were added to the modified solution.

[0056] Comparative Example 3. Unlike Example 3, no carboxymethyl cellulose or silicon carbide was added to the modified solution.

[0057] Comparative Example 4. Unlike Example 3, no additives were added during the preparation of the modified silica agent.

[0058] Comparative Example 5. Unlike Example 3, no modified sodium alginate was added to the additive.

[0059] Comparative Example 6. Unlike Example 3, kaolin and glucose were not added in the preparation of the modified sodium alginate agent.

[0060] The product performance tests of Examples 1-3 and Comparative Examples 1-6 used a fluorescent labeling method (DiR dye) to detect the half-life of exosomes in SD rats. Liver targeting was assessed by determining the proportion of exosomes aggregated in liver tissue 4 hours after injection, indicating its targeted delivery efficiency to the diseased liver. The test results are as follows:

[0061] From Examples 1-3 and Comparative Examples 1-6, it can be seen that the product of the present invention can achieve excellent liver targeting, and the product has a significant effect on improving liver fibrosis. Furthermore, the performance of the product tends to deteriorate when no modifying liquid is added in the preparation of the modified silica agent, no silane coupling agent KH550 or carbon nanotubes are added in the modifying liquid, no carboxymethyl cellulose or silicon carbide is added in the preparation of the modified silica agent, no additives are added in the additives, and no modified sodium alginate agent is added in the preparation of the modified sodium alginate agent.

[0062] It will be apparent to those skilled in the art that the present invention is not limited to the details of the exemplary embodiments described above, and that the invention can be implemented in other specific forms without departing from the spirit or essential characteristics of the invention. Therefore, the embodiments should be considered in all respects as exemplary and non-limiting, and the scope of the invention is defined by the appended claims rather than the foregoing description. Thus, it is intended that all variations falling within the meaning and scope of equivalents of the claims be included within the present invention.

[0063] Furthermore, it should be understood that although this specification describes embodiments, not every embodiment contains only one independent technical solution. This narrative style is merely for clarity. Those skilled in the art should consider the specification as a whole, and the technical solutions in each embodiment can also be appropriately combined to form other embodiments that can be understood by those skilled in the art.

Claims

1. A composition containing exosomes, characterized in that, The composition containing exosomes is a modified silica agent and an exosome matrix, wherein the modified silica agent and the exosome matrix are blended in a weight ratio of (2-5):15; The exosome is composed of exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells, and the mass ratio of the exosomes derived from mesenchymal stem cells to the exosomes derived from liver stem cells is 1: (0.5-2). The preparation methods for exosomes derived from mesenchymal stem cells and exosomes derived from liver stem cells are the same, and the specific preparation methods are as follows: P3 generation umbilical cord mesenchymal stem cells were seeded in a 50L bioreactor containing 5-6 g / L microcarriers, and serum-free X-VIVO was added. TM Culture medium 15, incubated at 37-39℃ and 5-7% CO2; When the cell density reaches 1-1.5×10^7 cells / mL, initiate hypoxia pretreatment: reduce the oxygen concentration to 5-7%, culture for 24-36 h, then reduce it to 2-5%, and continue culturing for another 24-36 h; culture liver stem cells (HSCs) using the same method to complete the hypoxia pretreatment.

2. The composition containing exosomes according to claim 1, characterized in that, The specific preparation method of the modified silica agent is as follows: S01: Mix 3-5 parts of carboxymethyl cellulose, 2-3 parts of silicon carbide and 5-8 parts of ethanol solution thoroughly to obtain a modified carboxymethyl cellulose agent; 2-3 parts of silane coupling agent KH550, 1-2 parts of carbon nanotubes and 5-8 parts of modified carboxymethyl cellulose agent are mixed evenly to obtain a modified solution; SO2: Silica and modified liquid are mixed and modified at a weight ratio of 3:(5-7). After mixing, the modified silica liquid is obtained. S03: Modified silica liquid and additives are mixed and ball-milled at a weight ratio of (7-11):

5. The ball milling speed is 1000-1500 r / min and the ball milling time is 2h. After the ball milling is completed, the mixture is filtered and dried to obtain modified silica agent.

3. The composition containing exosomes according to claim 2, characterized in that, The stirring speed for the stirring modification treatment is 450-500 r / min, and the stirring time is 1 h.

4. The composition containing exosomes according to claim 2, characterized in that, The ethanol solution is an ethanol solution with a mass fraction of 85-95%.

5. The composition containing exosomes according to claim 2, characterized in that, The additive is prepared by mixing 3-5 parts of flake boron nitride, 2-4 parts of dopamine hydrochloride solution and 2-4 parts of modified sodium alginate thoroughly to obtain the additive.

6. The composition containing exosomes according to claim 5, characterized in that, The mass fraction of the dopamine hydrochloride solution is 5-8%.

7. The composition containing exosomes according to claim 5, characterized in that, The preparation method of the modified sodium alginate agent is as follows: Add 1-3 parts sodium alginate powder, 2-3 parts kaolin, and 5-8 parts phosphate buffer solution, then add 2-4 parts glucose and 1-2 parts citric acid, stir well, filter and dry to obtain modified sodium alginate agent.

8. The composition containing exosomes according to claim 7, characterized in that, The pH value of the phosphate buffer solution is 5.5-6.

5.

9. The use of a composition containing exosomes as described in any one of claims 1-8 in the preparation of a product for treating liver failure.

Images