CN114432260A - Medicine-carrying sustained-release microsphere based on exosome and preparation method thereof - Google Patents
Medicine-carrying sustained-release microsphere based on exosome and preparation method thereof Download PDFInfo
- Publication number
- CN114432260A CN114432260A CN202011110573.1A CN202011110573A CN114432260A CN 114432260 A CN114432260 A CN 114432260A CN 202011110573 A CN202011110573 A CN 202011110573A CN 114432260 A CN114432260 A CN 114432260A
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- Prior art keywords
- exosome
- drug
- loaded
- sustained
- release microsphere
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Abstract
The invention relates to a medicine-carrying sustained-release microsphere based on exosome and a preparation method thereof. The drug-loaded sustained-release microsphere based on exosome comprises an inner core and a carrier material coated on the surface of the inner core; the inner core is an exosome loaded with a medicament; the carrier material is polylactic acid-glycolic acid copolymer and/or polylactic acid. The drug-loaded sustained-release microsphere has good biocompatibility, can not cause adverse effect on the physiological balance of the microenvironment of an organism in the degradation process, is beneficial to the healing of wounds when being applied to body surface wounds, can effectively load drugs and has good drug sustained-release effect.
Description
Technical Field
The invention relates to the technical field of pharmaceutical preparations, in particular to a drug-loaded sustained-release microsphere based on exosomes and a preparation method thereof.
Background
Exosomes are small membrane vesicles containing complex RNAs and proteins, which were first discovered in sheep reticulocytes in 1983 and were named "exosomes" by Johnstone in 1987. Various cells can secrete exosomes under normal and pathological states, mainly come from a multivesicular body formed by invagination of intracellular lysosome particles, and are released into extracellular matrix after fusion of outer membranes and cell membranes of the multivesicular body. At present, exosomes are widely used as pharmacological active substances for treating various diseases due to the advantages of stable properties, easy reaction, no immunogenicity, easy acquisition, convenient modification and the like.
Polylactic acid-glycolic acid copolymer (PLGA) and polylactic acid (PLA) have good mechanical property and biodegradability, and can be used as a novel pharmaceutical preparation, such as a carrier material of drug-loaded microspheres. However, when the two materials are used as carrier materials, the physiological balance of the microenvironment of the organism is easily adversely affected, and when the carrier materials are used for body surface wounds, the healing of the wounds is not facilitated.
Disclosure of Invention
Based on the above, there is a need for providing an exosome-based drug-loaded sustained release microsphere. The drug-loaded sustained-release microsphere has good biocompatibility, can not cause adverse effect on the physiological balance of the microenvironment of an organism in the degradation process, does not influence the healing of wounds when being applied to body surface wounds, can effectively load drugs and has good drug sustained-release effect.
The specific technical scheme is as follows:
a medicine-carrying sustained-release microsphere based on exosome comprises an inner core and a carrier material coated on the surface of the inner core;
the inner core is an exosome loaded with a medicament;
the carrier material is polylactic acid-glycolic acid copolymer or polylactic acid.
In one embodiment, the mass ratio of the exosome loaded with the drug to the carrier material is (0.01-5): 1000.
in one embodiment, the weight ratio of the drug to the exosomes is (1-10): 1.
in one embodiment, the exosomes are neural stem cell or mesenchymal stem cell-derived exosomes.
In one embodiment, the preparation method of the exosome comprises the following steps:
collecting supernatant of neural stem cell and/or mesenchymal stem cell culture solution, removing impurities and concentrating, extracting the obtained concentrated solution by using an exosome extraction column, and concentrating the obtained extracting solution.
In one embodiment, the contact angle of the drug-loaded sustained-release microsphere is more than 110 degrees.
The invention also provides a preparation method of the exosome-based drug-loaded sustained-release microsphere, which comprises the following steps:
mixing a solvent, the exosome loaded with the drug, and a polylactic acid-glycolic acid copolymer and/or polylactic acid to prepare a mixture;
and (3) carrying out electrostatic spray treatment on the mixture, and drying.
In one embodiment, the solvent is at least one of N, N-dimethylformamide, dichloromethane, acetone, and tetrahydrofuran.
In one embodiment, the concentration of the polylactic acid-glycolic acid copolymer and/or the polylactic acid in the solvent is 0.01-1 g/mL.
In one embodiment, the process conditions of the electrostatic spraying include: the positive voltage is 15-25 kV, the negative voltage is-0.5-1.5 kV, and the receiving distance between the spray head and the spinning is 10-15 cm.
Compared with the prior art, the invention has the following beneficial effects:
according to the medicine-carrying slow-release microsphere based on the exosome, the exosome containing the medicine is wrapped by polylactic acid-glycolic acid copolymer (PLGA) or polylactic acid (PLA), on one hand, the existence of the exosome can well relieve and neutralize an acidic environment formed by degrading the PLGA or the PLA, so that the adverse effect of the exosome on the healing of an in-vivo microenvironment or a body surface wound is avoided, and the biocompatibility of the medicine-carrying microsphere is improved; on the other hand, the inclusion of the drug in the exosome forms a drug-loaded exosome, and the drug-loaded slow-release system can be matched with PLGA or PLA to form a dual drug-loaded slow-release system, so that the exosome can play an excellent drug-loaded function and a drug slow-release function, can wrap the same drug and different drugs, and has good application value. In addition, PLGA or PLA can play a temporary storage role in exosomes, and the in-vitro activity and in-vivo drug delivery function of the exosomes are maintained; on the other hand, the medicine-carrying sustained-release microspheres containing exosomes improve the contact angle of the medicine-carrying sustained-release microspheres, and the prepared microspheres are stacked to form a layer of interface, so that the microspheres are easier to compound on the surface of a medical instrument and play a role in modification; with the increase of the contact angle, the hydrophobicity is improved, and the degradation performance of PLA is regulated and controlled to a certain degree; and the contact angle is increased, so that an antibacterial effect can be achieved, the antibacterial agent wrapped by the microspheres can be further cooperated, and the drug effect of the antibacterial agent is obviously improved.
Drawings
FIG. 1 shows the basic features of exosomes prepared in one example:
wherein A is the basic form of exosome under TEM microscope; b is the diameter distribution of the exosomes obtained by separation; c is Coomassie brilliant blue staining observation exosome protein distribution; d is exosome-specific marker expression (high expression of CD63, TSG101, CD81 and CD9, no expression of GAPDH);
FIG. 2 is an SEM interface morphology of PLGA drug-loaded sustained-release microspheres prepared in example 1;
FIG. 3 is an SEM interface morphology of PLA drug-loaded sustained-release microspheres prepared in example 2;
fig. 4 is a drug release profile of PLGA drug-loaded sustained release microspheres prepared in example 1 in PBS buffer (pH 7.4).
Detailed Description
The exosome-based drug-loaded sustained-release microsphere and the preparation method thereof are further described in detail in the following with reference to specific examples. The present invention may be embodied in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.
The invention provides a medicine-carrying sustained-release microsphere based on exosome, which comprises an inner core and a carrier material coated on the surface of the inner core; the inner core is an exosome loaded with drugs; the carrier material is polylactic-co-glycolic acid (PLGA) and/or polylactic acid (PLA).
In a specific example, the exosome-based drug-loaded sustained-release microsphere is prepared from an exosome loaded with a drug and a carrier material by an electrostatic spraying method.
In practice, the drugs in the inner cores may be the same or different. When different, the medicine can be loaded on exosomes respectively and then is prepared by electrostatic spraying together with a carrier material.
In one embodiment, the drug-loaded exosomes may be introduced into the exosomes by mixing the exosomes with the drug, using a method such as one of room temperature incubation, freeze-thaw cycling and sonication.
In one particular example, the drug may be one or more of an antibacterial, analgesic, antiviral, etc. drug. Specifically, the antibacterial agent may be antibiotic penicillin, roxithromycin, amoxicillin, norfloxacin, cefradycin, aminoglycoside, gentamycin, amikacin, macrolide, azithromycin, etc., semisynthetic antibiotic ampicillin, amikacin, doxycycline, rifampin, etc., artificially synthesized antibacterial agent flomoxifloxacin, ciprofloxacin lactate, norfloxacin, sulfamethoxazole, sulfisoxazole, sulfadiazine, etc.; the analgesic agent can be aspirin, ibuprofen, celecoxib, ereoxib, parecoxib, phenylbutazone, tramadol, etc.; the antiviral drug can be such as abacavir, bocavir, asunaprevir, vorciclovir, vipatavir, ledippivoxil, dalatavir hydrochloride, and bocavirenz.
In a specific example, the mass ratio of the exosome loaded with the drug to the carrier material is (0.01-5): 1000. the mass ratio of the inner core to the carrier material is reasonably controlled, so that the inner core is effectively coated by the carrier material, and the stability of the microsphere structure is improved. Specifically, the mass ratio of the drug-loaded exosomes to the carrier material includes, but is not limited to, the following ratios: 0.01: 1000. 0.02: 1000. 0.03: 1000. 0.04: 1000. 0.05: 1000. 0.1: 1000. 0.5: 1000. 1: 1000. 2: 1000. 3: 1000. 5: 1000.
in a specific example, the weight ratio of the drug to the exosomes is 1-10: 1. Further, the weight ratio of the medicine to the exosome is 1-3: 1. Specifically, the weight ratio of drug to exosome includes, but is not limited to, the following ratios: 1:1, 1.2:1, 1.5:1, 2:1, 3: 1.
In a specific example, the exosomes have a diameter of 80-150 nm.
In a specific example, the exosomes are neural stem cell or mesenchymal stem cell derived exosomes. The exosome using the source has the advantages that the proliferation capacity of the neural stem cells and/or the mesenchymal stem cells is strong, the exosome has strong immunogenicity, and a plurality of trophic factors can be secreted, so that exosomes are relatively easy to separate and obtain from the neural stem cells and/or the mesenchymal stem cells, and the exosomes can also carry certain trophic factors and have certain immunogenicity. Further, the exosomes are exosomes specifically expressing CD63, TSG101, CD81 and CD9 and not expressing GAPDH.
In one specific example, the preparation method of the exosome comprises the following steps:
collecting supernatant of neural stem cell and/or mesenchymal stem cell culture solution, removing impurities and concentrating, extracting the obtained concentrated solution by using an exosome extraction column, and concentrating the obtained extracting solution.
Further, concentration (including a concentration step after the impurity removal step and/or concentration of the extract) means concentration with an ultrafiltration membrane having a molecular weight cut-off of 50 to 150 kD. The exosome obtained by adopting the ultrafiltration membrane concentration can effectively remove impurity components, and the exosome obtained by enrichment is beneficial to effective loading of a medicament and has higher medicament loading rate. Specifically, the cut-off molecular weight of the ultrafiltration membrane includes, but is not limited to, the following molecular weights: 50kD, 70kD, 80kD, 85kD, 90kD, 95kD, 100kD, 105kD, 110kD, 115kD, 120kD, 130kD, 150 kD.
Further, the exosome extraction column is an izon Super EVs exosome extraction column.
Further, the impurities refer to cells and/or cell debris derived from neural stem cells and/or mesenchymal stem cells. Specifically, the step of removing impurities is to centrifuge the mixture for 5-15 min at 300-400 g and centrifuge the mixture for 25-35 min at 1500-2500 g.
In a specific example, the contact angle of the drug-loaded sustained-release microsphere is more than 110 degrees.
Further, the method for obtaining the exosome loaded with the drug is a method of combining low-temperature incubation with freeze-thaw cycle or an ultrasonic method, so that the drug enters the exosome.
Specifically, the low-temperature incubation refers to incubation for 16-20 hours at 1-5 ℃. The freeze-thaw cycle is to incubate at room temperature for 10-40 min and then to stand at-70 to-90 ℃ for 5-10min, so as to realize the cycle.
The ultrasonic treatment is carried out for 3-4 s under the condition of ice bath at 2-5 kHz, and then is stopped for 1-3 s, so as to circulate.
The invention also provides a preparation method of the drug-loaded sustained-release microsphere based on exosome, which comprises the following steps:
mixing a solvent, an exosome loaded with a drug, and a polylactic acid-glycolic acid copolymer and/or polylactic acid to prepare a mixture;
and (4) carrying out electrostatic spray treatment on the mixture, and drying.
In a specific example, the solvent is at least one of N, N-dimethylformamide, dichloromethane, acetone, and tetrahydrofuran. Further, the solvent comprises a solvent A, wherein the solvent A is N, N-dimethylformamide, and also comprises a solvent B, and the solvent B is selected from at least one of dichloromethane, acetone and tetrahydrofuran. Further, the ratio of solvent a to solvent B is 1: (0.5 to 5), preferably 1: (0.8 to 1.2).
In a specific example, the concentration of the polylactic acid-glycolic acid copolymer and/or the polylactic acid in the solvent is 0.01-0.05 g/mL.
In one specific example, the process conditions for electrostatic spraying include: the positive voltage is 15-25 kV, the negative voltage is-0.5-1.5 kV, and the receiving distance between the spray head and the spinning is 10-15 cm.
In a specific example, drying refers to natural drying at room temperature.
Hereinafter, specific examples are described, and the raw materials used in the examples are all commercially available products unless otherwise specified.
The exosomes loaded with drugs adopted in the examples are all exosomes loaded with norfloxacin, and the preparation method is as follows:
(1) preparing an exosome:
periodically collecting the supernatant of NSCs culture solution, centrifuging for 10min at 350g and 30min at 2000g, and removing precipitate (including cells and cell debris); filtering the cell supernatant by adopting a 50mL ultrafiltration tube of 100kD, and extracting exosomes from the obtained cell supernatant concentrated solution by using an izon Super EVs exosome extraction column; the obtained exosomes were then further concentrated using a 100kD 1.5mL ultrafiltration tube to obtain high concentration exosomes. Placing the mixture in an incubator at minus 80 ℃ for standby. The basic characteristics of exosomes are shown in fig. 1, and as can be seen from fig. 1, the diameter of exosomes is 80-150 nm.
(2) Loading a drug:
(2-1) Low-temperature incubation: the NSCs exosomes obtained from multiple separations (1526.4ng/mL) were further concentrated to obtain high-concentration NSCs exosomes (PBS dilution, final concentration of 0.01mg/mL), then 250uL was taken, and norfloxacin was added at a weight ratio of 1:1, was added to the exosome solution, placed at 4 ℃ on a shaker and incubated for 18 h.
(2-2) freeze-thaw cycle: on the basis of low-temperature incubation, the cells are incubated at room temperature for 30min, then quickly placed in a refrigerator at minus 80 ℃ for 7min, and repeated for 3 times.
Example 1
The embodiment provides a medicine-carrying sustained-release microsphere based on exosome, and the preparation method comprises the following steps:
(1) taking the PLGA out of the refrigerator, standing for about 10min, weighing 0.3g of PLGA after the water on the surface of the bag is evaporated, then taking a measuring cylinder of 10mL, weighing 10mL of N, N-dimethylformamide, pouring the weighed PLGA and 200uL of the exosome containing the fleroxacin (the concentration of the obtained exosome containing the fleroxacin is 50ug/mL) into the N, N-dimethylformamide organic solvent, stirring for 1.5h to obtain electrostatic spray liquid, and placing the electrostatic spray liquid at room temperature for later use;
(2) carrying out electrostatic spraying on the electrostatic spraying liquid: starting an electrostatic sprayer, preheating for 30min, adjusting a positive voltage of 20kV and a negative voltage of-1 kV, wherein the spray head and spray receiving distance is 12 cm; then sucking the electrostatic spraying liquid into a 20mL syringe, and carrying out electrostatic spraying by using a No. 21 needle; and then, placing the prepared PLGA electrospray microsphere interface layer at room temperature for natural drying.
An interface topography of the PLGA drug-loaded sustained-release microsphere prepared in this example is shown in fig. 2. As can be seen from FIG. 2, the interface mainly comprises PLGA microspheres and microfilaments, and the PLGA microspheres and the microfilaments are combined with each other in a cross manner to form a special silk-ball interface with high porosity, so that the silk-ball interface has high hydrophobicity and a contact angle of more than 130 degrees.
Example 2
The preparation method of the drug-loaded sustained-release microsphere based on exosome is the same as that of the microsphere in example 1, and the main difference is that: PLA was used instead of PLGA.
An interface topography of the PLA drug-loaded sustained-release microspheres prepared in this example is shown in fig. 3. As can be seen from fig. 3, the interface mainly comprises PLA microspheres and microfilaments, and the cross-bonding between the two forms a special silk-sphere interface with higher porosity, and compared with the PLGA microsphere interface, the interface has weaker hydrophobicity (contact angle is above 110 °).
Example 3
The preparation method of the drug-loaded sustained-release microsphere based on exosome is the same as that of example 1, and the main difference is that the adopted solvent is different. The method comprises the following specific steps:
(1) taking the PLGA material out of a refrigerator, standing for about 10min, weighing 0.3g of PLGA after the water on the surface of the bag is evaporated, then taking a measuring cylinder of 10mL, weighing 5mL of each of N, N-dimethylformamide/dichloromethane, pouring the weighed PLGA and 200uL of the exosomes containing the fleroxacin (the concentration of the obtained exosomes containing the fleroxacin is 50ug/mL) into the N, N-dimethylformamide/dichloromethane organic solvent, stirring for 1.5h, obtaining an electrostatic spray solution, and placing at room temperature for later use;
(2) carrying out electrostatic spraying on the electrostatic spraying liquid: starting an electrostatic sprayer, preheating for 30min, adjusting a positive voltage of 20kV and a negative voltage of-1 kV, wherein the spray head and spray receiving distance is 12 cm; then sucking the electrostatic spraying liquid into a 20mL syringe, and carrying out electrostatic spraying by using a No. 21 needle; and then, naturally airing the prepared PLGA electrostatic spraying microsphere interface layer at room temperature.
Example 4
The preparation method of the drug-loaded sustained-release microsphere based on exosome is the same as that of example 1, and the main difference is that the adopted solvent is different. The method comprises the following specific steps:
(1) taking the PLGA material out of a refrigerator, standing for about 10min, weighing 0.3g of PLGA after the water on the surface of the bag is evaporated, then taking a measuring cylinder with 10mL, weighing 5mL of each N, N-dimethylformamide/acetone, pouring the weighed PLGA and 200uL of the exosomes containing the fleroxacin (the concentration of the obtained exosomes containing the fleroxacin is 50ug/mL) into the N, N-dimethylformamide/acetone organic solvent, stirring for 1.5h, obtaining an electrostatic spray liquid, and placing at room temperature for later use;
(2) carrying out electrostatic spraying on the electrostatic spraying liquid: starting an electrostatic sprayer, preheating for 30min, adjusting a positive voltage of 20kV and a negative voltage of-1 kV, wherein the spray head and spray receiving distance is 12 cm; then sucking the electrostatic spraying liquid into a 20mL syringe, and carrying out electrostatic spraying by using a No. 21 needle; and then, naturally airing the prepared PLGA electrostatic spraying microsphere interface layer at room temperature.
Example 5
The preparation method of the drug-loaded sustained-release microsphere based on exosome is the same as that of example 1, and the main difference is that the adopted solvent is different. The method comprises the following specific steps:
(1) taking the PLGA out of a refrigerator, standing for about 10min, weighing 0.3g of PLGA after the water on the surface of the bag is evaporated, then taking a measuring cylinder of 10mL, weighing 5mL of each N, N-dimethylformamide/tetrahydrofuran, pouring the weighed PLGA and 200uL of exosomes containing the fleroxacin (the concentration of the obtained exosomes containing the fleroxacin is 50ug/mL) into the N, N-dimethylformamide/tetrahydrofuran organic solvent, stirring for 1.5h, obtaining electrostatic spray liquid, and placing at room temperature for later use;
(2) carrying out electrostatic spraying on the electrostatic spraying liquid: starting an electrostatic sprayer, preheating for 30min, adjusting a positive voltage of 20kV and a negative voltage of-1 kV, wherein the spray head and spray receiving distance is 12 cm; then sucking the electrostatic spraying liquid into a 20mL syringe, and carrying out electrostatic spraying by using a No. 21 needle; and then, naturally airing the prepared PLGA electrostatic spraying microsphere interface layer at room temperature.
The slow release effect test was performed on the PLGA electrostatic spray microspheres prepared in example 1.
The test method comprises the following steps: 0.1g of the prepared PLGA electrostatic spray microspheres is accurately weighed, placed in a 1.5mL EP tube at room temperature, added with 1mL of PBS buffer (pH 7.4) for soaking, and from the first day, 200uL of the soaked PBS buffer is quantitatively taken every day for concentration measurement, and meanwhile, the same amount of fresh PBS is supplemented in the EP tube.
Tests show that the drug release curve of the PLGA electrostatic spray microsphere prepared in example 1 in PBS buffer (pH 7.4) is shown in fig. 4, which shows that the release of the encapsulated drug is increased and the drug release can reach more than 14 days with the prolonged time, and the PLGA drug-loaded sustained-release microsphere has strong drug release capacity and long-lasting antibacterial effect.
The PLGA electrostatic spray microspheres prepared in example 1 were tested for antibacterial effect.
The test method comprises the following steps:
(1) preparing and sterilizing an agar culture medium;
(2) preparation of bacterial suspension: in a clean bench, Staphylococcus aureus and Escherichia coli colonies were scraped with a sterile cotton swab to a sterile NaCl-peptone solution at pH 7.0, 6X 108comparing cfu/mL turbidimetric tubes, performing gradient dilution after the concentration is reached, and obtaining the final bacterial liquid concentration of 6 x 105cfu/mL, followed by 6 x 10 dip with sterile cotton swab5Evenly smearing cfu/mL solution on a trypticase soy peptone agar culture medium culture dish;
(3) pasting and placing the bacteriostatic sample: and (3) shearing a sample sheet with the diameter area of 5mm from the prepared drug-loaded slow-release microsphere sheet layer, slightly pressing the sample sheet in a culture dish coated with bacteria, covering the culture dish, and placing the culture dish in an incubator for culturing for 17 hours.
And (3) judging the bacteriostatic action: if the antibacterial ring is larger than 7mm, the antibacterial effect is shown.
The results are shown in table 1:
TABLE 1
The technical features of the embodiments described above may be arbitrarily combined, and for the sake of brevity, all possible combinations of the technical features in the embodiments described above are not described, but should be considered as being within the scope of the present specification as long as there is no contradiction between the combinations of the technical features.
The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent should be subject to the appended claims.
Claims (10)
1. A medicine-carrying sustained-release microsphere based on exosome is characterized by comprising an inner core and a carrier material coated on the surface of the inner core;
the inner core is an exosome loaded with a medicament;
the carrier material is polylactic acid-glycolic acid copolymer or polylactic acid.
2. The exosome-based drug-loaded sustained-release microsphere according to claim 1, wherein the mass ratio of the drug-loaded exosome to the carrier material is (0.01-5): 1000.
3. the exosome-based drug-loaded sustained-release microsphere according to claim 1, wherein the weight ratio of the drug to the exosome is (1-10): 1.
4. the exosome-based drug-loaded sustained-release microsphere according to claim 1, characterized in that the exosome is a neural stem cell and/or mesenchymal stem cell derived exosome.
5. A drug-loaded sustained release microsphere based on exosome according to claim 4, characterized in that the preparation method of exosome comprises the following steps:
collecting supernatant of neural stem cell and/or mesenchymal stem cell culture solution, removing impurities and concentrating, extracting the obtained concentrated solution by using an exosome extraction column, and concentrating the obtained extracting solution.
6. A drug-carrying sustained release microsphere based on exosome according to any one of claims 1 to 5, characterized in that the contact angle of the drug-carrying sustained release microsphere is more than 110 °.
7. The preparation method of the exosome-based drug-loaded sustained-release microsphere according to any one of claims 1 to 6, which is characterized by comprising the following steps:
mixing a solvent, the exosome loaded with the drug, and a polylactic acid-glycolic acid copolymer and/or polylactic acid to prepare a mixture;
and (3) carrying out electrostatic spray treatment on the mixture, and drying.
8. The preparation method of exosome-based drug-loaded sustained-release microspheres according to claim 7, wherein the solvent is at least one of N, N-dimethylformamide, dichloromethane, acetone and tetrahydrofuran.
9. The preparation method of the exosome-based drug-loaded sustained-release microsphere according to claim 7, wherein the concentration of the polylactic acid-glycolic acid copolymer or the polylactic acid in the solvent is 0.01-1 g/mL.
10. A preparation method of exosome-based drug-loaded sustained-release microspheres according to any one of claims 7 to 9, wherein the technological conditions of electrostatic spraying comprise: the positive voltage is 15-25 kV, the negative voltage is-0.5-1.5 kV, and the receiving distance between the spray head and the spinning is 10-15 cm.
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