CN108853147B - Polypeptide nanofiber hydrogel for slowly releasing exosomes and preparation method and application thereof - Google Patents
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Abstract
The invention belongs to the technical field of medicines, and discloses a self-assembly polypeptide, a polypeptide nanofiber hydrogel prepared from the self-assembly polypeptide NapFF and an exosome, and a preparation method and application of the polypeptide nanofiber hydrogel. The amino acid sequence of the self-assembly polypeptide is shown as SEQ ID No. 1. The self-assembly polypeptide has a heart protection function. The polypeptide nanofiber hydrogel is prepared from the self-assembly polypeptide, NapFF and exosome, wherein the self-assembly polypeptide and NapFF form hydrogel, and the exosome is encapsulated in the hydrogel. The polypeptide nanofiber hydrogel disclosed by the invention can improve the residence time of exosomes with a heart protection function in tissues, improve the curative effect of the exosomes on improving the heart function, directly provide the heart protection function, further improve the heart function of a myocardial infarction region, and can be used for treating cardiovascular diseases such as myocardial infarction.
Description
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to a polypeptide nanofiber hydrogel as well as a preparation method and application thereof, in particular to a polypeptide nanofiber hydrogel for slowly releasing exosomes as well as a preparation method and application thereof.
Background
Myocardial infarction is defined pathologically as the death of cardiomyocytes due to prolonged ischemia. Ischemic heart diseases, which are generally myocardial cell injury and death due to persistent coronary ischemia, oxygen deficiency and myocardial ischemia trigger inflammatory reactions, myocardial cell necrosis and apoptosis, leading to myocardial remodeling and cardiac function reduction which is difficult to reverse, even cardiac insufficiency, sudden death and the like, which seriously threaten human health. Myocardial damage due to ischemia and hypoxia is also known as acute coronary syndrome. According to statistics, about 70 ten thousand patients die of the acute coronary syndrome every year in China, and the incidence of the acute coronary syndrome is on the rise due to unreasonable nutrition collocation, changes of life and working rules and the like in recent years.
At present, the main methods for clinically treating myocardial infarction comprise drug therapy, coronary intervention therapy, coronary artery bypass transplantation and the like, but the methods can relieve initial myocardial damage in the acute stage, and because adult myocardial cells hardly proliferate, infarcted myocardium cannot be regenerated; in addition, after an acute myocardial infarction period, subsequent treatment is still affected by heart failure, cardiogenic shock and other problems caused by cardiac remodeling. Therefore, new therapeutic approaches to address these problems are urgently needed. At present, a plurality of researches hope that stem cells are proliferated and differentiated into new myocardial cells to make up for lost myocardium and improve the cardiac function after myocardial infarction, and certain effect is achieved. Recent studies have shown that stem cells have a low survival and differentiation rate in transplanted hearts and that stem cell transplantation also has a potential risk of neoplasia. It is appreciated that current studies indicate that stem cell transplantation improves heart function by improving endogenous heart cells primarily through paracrine action, and that exosomes secreted by stem cells are thought to be factors that may play a major role. Several studies have also shown that exosomes derived from embryonic stem cells, mesenchymal stem cells, cardiac progenitor cells and cardiac spherocytes increase cardiomyocyte proliferation, reduce myocardial infarct size, reduce inflammatory response and improve cardiac function. The exosome myocardial injection can improve the microenvironment of myocardial infarction, improve the cardiac function, promote the proliferation of myocardial cells and reduce the myocardial infarction area. Therefore, a new idea is developed for the treatment of myocardial infarction based on the protective effect of exosome on the cardiovascular system. However, both exosome and stem cell therapy have the disadvantage of short in vivo half-life, and studies report that the half-life of intravenous exosome in blood circulation is only 2min, while the signal in vivo reaches the strongest 3 days after the fluorescent-labeled exosome is injected at the myocardial infarction, and the signal is basically not detected after 7 days. Since one of the biggest problems faced with this exosome therapy is its short residence time in the body, multiple injections are generally required to improve the therapeutic effect of exosomes. Because the half-life of intravenous injection is too short and the heart homing rate is relatively low, the heart tissue cannot be targeted 100%; for many internal organs such as the heart, multiple in situ injections mean multiple thoracotomy, etc., increasing the risk of infection. Therefore, if the residence time of the exosome in the in-situ tissue can be prolonged, the clinical transformation of the exosome for treating myocardial infarction is promoted by a huge step. Therefore, exosome myocardial injections of the "once-for-all" type are required.
The polypeptide hydrogel has potential application value in the aspect of slow release of medicines and bioactive substances. The self-assembly polypeptide hydrogel is an artificially synthesized gel with high water content and a structure similar to a natural extracellular matrix (ECM), can be spontaneously assembled into reticular nanofibers in an aqueous solution, quickly forms hydrogel when meeting multivalent salt ions, and has an internal water environment suitable for polar molecules to diffuse in a large range and suitable for being used as a carrier of medicines, particularly bioactive substances. In addition, the self-assembled polypeptide hydrogel also has the advantages of stable structure, good biocompatibility, controllable degradation, no immunogenicity, sequence designability, small formed pore size and the like. However, since self-assembling polypeptides generally do not have biological activity, they are generally engineered on the basis of self-assembling polypeptides for clinical transformation. Professor Yong-sup Yoon reports that nanofiber hydrogel formed by functional self-assembly polypeptides (C16-GTAGLIGQS and C16-GTAGLIGQRGDS, respectively abbreviated as PA-S and PA-RGDS) can obviously prolong the retention time of endothelial cells in injured hind limbs in transferring the endothelial cells, does not influence the functions of the endothelial cells, and can further improve angiogenesis and hind limb blood flow recovery. However, the self-assembled polypeptide hydrogel does not provide cardioprotective function.
Disclosure of Invention
In view of the above, the present invention provides a self-assembled polypeptide nanofiber hydrogel with a heart protection function, and a preparation method and an application thereof, aiming at the defects of the prior art.
In order to realize the purpose of the invention, the invention adopts the following technical scheme:
a self-assembly polypeptide has an amino acid sequence of Palmitoyl-Gly-Thr-Ala-Gly-Leu-Ile-Gly-Gln-Gly-Gly-His-D-2-methyl-Trp-Ala-Trp-D-Phe-Lys-Ser, which is shown in SEQ ID No. 1. The self-assembly polypeptide comprises a growth hormone releasing peptide GHRP-6 (Hexarelin with the sequence of His-D-2-methyl-Trp-Ala-Trp-D-Phe-Lys) with a heart protection function, and is connected to the C terminal of a basic self-assembly amphiphilic polypeptide PA through two glycines so as to ensure that the structure and the function of the polypeptide are not influenced. Wherein the amphiphilic polypeptide PA comprises two parts: palmitoyl is a hydrophobic fatty chain, providing a hydrophobic chain; Gly-Thr-Ala-Gly-Leu-Ile-Gly-Gln is a matrix metalloproteinase recognition sequence, provides a hydrophilic chain, two different hydrophilic parts jointly promote the self-assembly of molecules, and simultaneously adds a serine at the C terminal to increase the water solubility of the polypeptide, wherein the polypeptide is named as PA-GG-Hexarelin-S. The self-assembly polypeptide PA-GG-Hexarelin-S has a heart protection function.
The invention also provides a polypeptide nanofiber hydrogel which is prepared from the self-assembly polypeptide, NapFF and exosome, wherein the self-assembly polypeptide, NapFF and exosome jointly form hydrogel, and the exosome is encapsulated in the hydrogel.
The NapFF in the polypeptide nanofiber hydrogel disclosed by the invention is a strong gelling agent, so that the polypeptide nanofiber hydrogel not only has strong gelling capability, but also can promote other polypeptides with weak gelling capability to gel. Therefore, the self-assembly polypeptide and NapFF mixed solution can form hydrogel after being mixed according to a certain proportion. The hydrogel has high water content, contains a matrix metalloproteinase recognition sequence GTAGLIGQ, and can highly simulate an extracellular matrix; the self-assembly polypeptide in the hydrogel contains a polypeptide sequence Hexarelin with heart protection, is released in the process of slow degradation of the hydrogel, provides a heart protection function, and improves the heart function of a myocardial infarction region; in addition, the hydrogel has small pores and can encapsulate exosomes, so that the exosomes are slowly released, and the residence time of the exosomes in the heart is prolonged.
Preferably, in the polypeptide nanofiber hydrogel, the mass ratio of the NapFF, the self-assembly polypeptide and the exosome is 4 (2-0.002): 1, and all the components can be gelatinized.
Furthermore, in the polypeptide nanofiber hydrogel, the mass ratio of the NapFF, the self-assembly polypeptide and the exosome is preferably 4 (0.02-0.002): 1, so that the effect of protecting myocardial cells is improved.
Exosomes are vesicles with a diameter of about 30-100nm which are released to the outside of cells after fusion of intracellular vesicles and cell membranes, are widely present in various body fluids, contain various substances including cytokines, proteins, lipids, messenger RNA, micro RNA (mirna), ribosomal RNA and the like, and play an important role in intercellular signal communication. Preferably, in the polypeptide nanofiber hydrogel, the sources of the exosomes are embryonic stem cells, mesenchymal stem cells, cardiac progenitor cells and cardiac spherocytes.
Further, preferably, the mesenchymal stem cell is an umbilical cord mesenchymal stem cell.
In some embodiments, the mesenchymal stem cell is an umbilical cord mesenchymal stem cell. Because the exosome of the mesenchymal stem cell can improve the microenvironment after the myocardial infarction and improve the cardiac function, the polypeptide nanofiber hydrogel can further improve the cardiac function after the myocardial infarction.
The invention also provides a preparation method of the polypeptide nanofiber hydrogel, and specifically relates to a polypeptide nanofiber hydrogel capable of encapsulating exosomes by self-assembly, which is prepared by mixing the self-assembly polypeptide with NapFF to obtain a polypeptide mixed solution, heating the polypeptide mixed solution, cooling the polypeptide mixed solution to room temperature, and uniformly mixing the polypeptide mixed solution with exosomes. The polypeptide mixed liquor is heated, so that the solubility of the polypeptide can be improved, and energy can be provided for forming the colloid of the NapFF polypeptide. The polypeptide mixed liquor is heated, rapidly cooled to room temperature, and can be gelatinized within 2-5 min.
In some embodiments, the heating is a boiling water bath for 30 seconds.
Preferably, the volume ratio of the self-assembly polypeptide to NapFF and exosome is 8:1: 1.
Therefore, the invention also provides the application of the polypeptide nanofiber hydrogel in preparing a medicament for treating cardiovascular diseases.
Wherein the cardiovascular disease is myocardial infarction.
According to the technical scheme, the invention provides a self-assembly polypeptide, a polypeptide nanofiber hydrogel prepared from the self-assembly polypeptide NapFF and an exosome, and a preparation method and application of the polypeptide nanofiber hydrogel. The amino acid sequence of the self-assembly polypeptide is shown as SEQ ID No. 1. The self-assembled polypeptide has a heart protection function and high water solubility. The polypeptide nanofiber hydrogel is prepared from the self-assembly polypeptide, NapFF and exosome, wherein the self-assembly polypeptide, NapFF and exosome jointly form hydrogel, and the exosome is encapsulated in the hydrogel. The polypeptide nanofiber hydrogel disclosed by the invention can improve the residence time of exosomes with a heart protection function in tissues, improve the curative effect of the exosomes on improving the heart function, directly provide the heart protection function and further improve the heart function of a myocardial infarction region. Therefore, the polypeptide nanofiber hydrogel can be used for treating cardiovascular diseases such as myocardial infarction.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below.
FIG. 1 shows a technical roadmap;
FIG. 2 shows MS (A) and HPLC (B) identification of PA-GG-Hexarelin-S polypeptides;
FIG. 3 shows MS (A) and HPLC (B) identification of NapFF polypeptides;
FIG. 4 shows the effect of different concentrations of PA-GG-Hexarelin-S on cardiomyocyte viability;
FIG. 5 shows a morphological observation of a polypeptide hydrogel;
FIG. 6 shows the effect of varying amounts of PA-GG-Hexarelin-S of examples 3-7 on cardiomyocyte viability of the resulting hydrogels;
FIG. 7 shows the sustained release effect of polypeptide hydrogels on exosomes;
figure 8 shows the effect of a polypeptide hydrogel encapsulating exosomes on cardiac function.
Detailed Description
The invention discloses a self-assembled polypeptide nanofiber hydrogel and a preparation method and application thereof. Those skilled in the art can modify the process parameters appropriately to achieve the desired results with reference to the disclosure herein. It is expressly intended that all such similar substitutes and modifications which would be obvious to one skilled in the art are deemed to be included in the invention. While the methods and products of the present invention have been described in terms of preferred embodiments, it will be apparent to those of ordinary skill in the art that variations and modifications of the methods described herein, as well as other suitable variations and combinations, may be made to implement and use the techniques of the present invention without departing from the spirit and scope of the invention.
In order to further understand the present invention, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the embodiments of the present invention, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Unless otherwise specified, the reagents involved in the examples of the present invention are all commercially available products, and all of them are commercially available.
The specific technical route in the embodiment of the invention is shown in fig. 1.
Example 1 isolation of exosomes derived from umbilical cord mesenchymal Stem cells
(1) Obtaining adherently growing human umbilical cord mesenchymal stem cells UMSC (UMSC) by resuscitation. When the degree of polymerization of cells in 10cm cell culture dishes reached 90% or more, cell passaging was performed, digestion was performed using 0.125% trypsin, and then digestion was terminated using complete medium, and the obtained cells were resuspended after centrifugation and were cultured by dividing into 3 10cm cell culture dishes on average.
(2) Normal fetal calf serum is ultracentrifuged for 8h at 4 ℃ at 110000g to remove exosomes, and the centrifuged serum is filtered by using a disposable sterile filter membrane of 0.22 mu m to obtain the exosome-removed serum.
(3) Exosomes were collected from 6-8 passages of cells, digested and centrifuged cells were resuspended in 10% α -MEM medium with exosome-depleted fetal bovine serum, passaged at normal ratio and cultured for 48 h, and the supernatant was collected.
(4) The supernatant was then centrifuged at 2000 g at 4 ℃ for 30 min, the pellet discarded, the supernatant gently transferred to a new centrifuge tube, and 1/2 volumes of total exosome isolation reagent (Invitrogen) were added.
(5) Standing at 4 ℃ for 12 h after uniform mixing, then centrifuging at 10000g at 4 ℃ for 60 min, obtaining UMSC exosomes as white precipitates on the tube wall, completely discarding supernatant, then resuspending with PBS, obtaining the UMSC exosomes, and adjusting the exosome concentration to 10 mg/mL.
Example 2 Synthesis and characterization of self-assembling Polypeptides
1. The synthetic polypeptides PA-GG-Hexarelin-S and NapFF.
The polypeptide PA-GG-Hexarelin-S has the sequence as follows: Palmitoyl-Gly-Thr-Ala-Gly-Leu-Ile-Gly-Gln-Gly-Gly-His-D-2-methyl-Trp-Ala-Trp-D-Phe-Lys-Ser. The polypeptide is synthesized in Nanjing Kingsrei Biotechnology GmbH, and the specific synthesis steps are as follows: seventeen amino acid polypeptides containing MMP-2 sensitive sequence (GTAGLIQQE), myocardial protection polypeptide (Hexarelin) and the like are synthesized by a standard FMOC chemical synthesis method, and then the N end carries out alkylation reaction on the peptides, and the method comprises the following specific steps: the polypeptide can be alkylated at the N-terminus by two 12-hour reactions with palmitic acid in the presence of a mixture of O-benzotriazole-N, N' -tetramethylureidohexafluorophosphate (HBTU) and Diisopropylethylamine (DIPEA) dissolved in Dimethylformamide (DMF). The polypeptide was then cleaved from the resin using a cleavage solution consisting of trifluoroacetic acid (TFA), deionized water, Triisopropylsilane (TIPS) and anisole at 40:1:1:1 for 2h, and the filtrate was collected by filtration. The collected samples were subjected to rotary evaporation to remove excess TFA and precipitated in ether and lyophilized.
The sequence of the polypeptide NapFF is Naphtalene acetyl-Phe-Phe. Polypeptide Synthesis by Haiyao Biotechnology Ltd, detailed Synthesis procedure the polypeptide detailed Synthesis procedure is described in the literature (Y. Zhang, Y. Kuang, Y. Gao, B. Xu, Versatile small-molecular motion for selected-assembly in water and the formation of biological sub-assembly carbohydrates, Langmuir, 27 (2011) 529. 537.)
2. Polypeptide mass fraction and purity identification
(1) Method for identifying self-assembled polypeptide by mass spectrum
a, sample dissolving treatment of MS: the sample was dissolved with formic acid and diluted to about 1mg/mL with purified water.
b, ionization method: electrospray ionization is adopted, and sample flow passes through a capillary tube, and is atomized under the action of atomizing gas N2 at the moment of flowing out of the capillary tube. Under the action of an applied high voltage (<5KV), original liquid drops generated by polar samples contain ions with two polarities under the action of an applied electric field. The charged liquid drop moves rapidly under the action of the electric field, the volume of the liquid drop is reduced along with the rapid evaporation of the solvent under the atmospheric pressure, the ions move to the surface of the liquid drop, and the electric field intensity of the liquid drop per se is increased. When the critical electric field is reached, the repulsive force of the electric field on the surface of the liquid drop is larger than the surface tension of the maintaining liquid drop, coulomb explosion is generated, the process is repeated, and finally, sample ions are resolved out and are introduced into a mass spectrum detector under the action of the electric field.
Successful synthesis of the polypeptide was confirmed by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry.
(2) HPLC identification of purity of self-assembled polypeptides
a, HPLC sample dissolution treatment: the sample was dissolved with formic acid and diluted to about 1mg/mL with purified water.
b, HPLC: wherein Pump A is 0.065% trifluoroacetic acid in water, Pump B is 0.05% trifluoroacetic acid in acetonitrile, and the flow rate is 1mL
And/min, the detection wavelength is 220 nm.
The molecular weight and purity of the functional polypeptide PA-GG-Hexarelin-S are identified in figure 2 by mass spectrometry and HPLC; the mass spectrum of the NapFF self-assembly polypeptide and the identification of molecular weight and purity by HPLC are shown in fig. 3. The result shows that the molecular weight of the synthesized two polypeptide sequences is in accordance with expectation, the purity is more than 95 percent, and subsequent experiments can be carried out.
Example 3 protective Effect of the self-assembling polypeptide PA-GG-Hexarelin-S on cardiac muscle cells
Different concentrations of PA-GG-Hexarelin-S and cardiomyocytes were preincubated for 4 hours, followed by 100. mu. M H2O2After 4h of treatment, CCK-8 experiment is carried out to detect the activity of the cells, and the result shows that 10 percent of the activity is detected-6-10-8The PA-GG-Hexarelin-S with mol/L concentration can improve the activity of the myocardial cells (figure 4), and the self-assembly polypeptide PA-GG-Hexarelin-S has the protective function of the myocardial cells.
Example 4 preparation and morphological Observation of functional self-assembled polypeptide nanofiber hydrogels encapsulating exosomes
Two synthesized short peptides were prepared as an aqueous solution of 0.02% PA-GG-Hexarelin-S (g/mL, w/v) and 0.5% NapFF (g/mL, w/v) polypeptides, and a polypeptide mixture was obtained by mixing the two polypeptides PA-GG-Hexarelin-S and NapFF in a ratio of 1:8 (v: v).
Heating 90 mu L of polypeptide mixed liquor in a boiling water bath for 30 seconds, cooling to room temperature, uniformly mixing with 10 mu L of 1% (in g/mL, w/v) exosome, and standing at room temperature to self-assemble the polypeptide nanofiber hydrogel for encapsulating exosomes. The hydrogel was observed for morphology, as shown in FIG. 5.
Examples 5-8 preparation of exosome-encapsulating functional self-assembling polypeptide nanofiber hydrogels and comparison of the effects on cardiomyocytes
Polypeptide nanofiber hydrogels encapsulating exosomes were prepared according to the method of example 4, in the formulation of table 1 below, comparing gelling effects.
TABLE 1 dosage of the components of the polypeptide nanofiber hydrogel for forming encapsulated exosomes
The results show that examples 4 to 8 all gel.
10 μ L of each of the hydrogels and cardiomyocytes of examples 4-8 were then preincubated for 4 hours, followed by 100 μ M H2O2After 4h of treatment, the CCK-8 experiment detects the cell activity, and the results show that the example 4 and the example 7-8 can obviously improve the activity of the myocardial cells, namely NaPFF: PA-GG-Hexarelin-S: the weight ratio of the exosome to the exosome is 4 (0.02-0.002) to 1, and the result is shown in figure 6.
Example 9 sustained Release of functional self-assembled polypeptide nanofiber hydrogels on exosomes
(1) By exchanging the exosomes for PKH 26-labeled exosomes at the same concentration and volume according to example 3, a hydrogel encapsulating PKH 26-labeled exosomes could be formed.
(2) mu.L of each of the hydrogels encapsulating PKH 26-labeled exosomes was placed in a 96-well cell culture plate, 100. mu.L of PBS was added thereto, PBS was collected every 2 days, and new PBS was added. A total of 18 collections were made. Used for detecting the release of the hydrogel to exosomes.
(3) And (3) uniformly mixing the latex microspheres, adding 10 mu L of microsphere (excessive) suspension into the repeatedly collected incubation PBS (phosphate buffer solution) of the polypeptide hydrogel encapsulating the PKH 26-labeled exosomes, uniformly blowing and beating the mixture by using a pipette, and incubating the mixture for 15 min at room temperature.
(4) PBS was added to the EP tube to 1mL and incubation continued at room temperature for 2h with intermittent mixing.
(5) Add 110. mu.L of 1M glycine solution to the EP tube to a final concentration of 0.1M, mix well and incubate at room temperature for 30 min.
(6) Centrifugation was carried out at 4000 rpm for 3 min, the supernatant was discarded, and after resuspension using 200. mu.L of PBS containing 0.5% BSA, the number of beads bound with red fluorescent-labeled exosomes was detected by flow assay.
As a result, as shown in FIG. 7, red-labeled exosomes were detected from day 4 to day 36, and the cumulative release rate was positively correlated with time (R)2= 0.979), indicating that the polypeptide hydrogel can provide stable sustained release of exosomes.
Example 10 functional self-assembling polypeptide nanofiber hydrogels improve cardiac function after myocardial infarction
(1) After constructing the rat myocardial infarction model, when the blood supply area becomes white, the polypeptide nanofiber hydrogel encapsulating exosomes prepared in example 3 (50 μ L each site) was implanted at 2 positions in the peripheral area of the white area, and the injection position was kept away from the vascular position. Then closing the chest, drawing out the tracheal catheter, placing on an electric hot plate at 37 ℃, and placing in a mouse cage for normal culture after the rat revives.
(2) When the rats after myocardial infarction grow to 1d and 28d, the rats are anesthetized by intraperitoneal injection with 10% chloral hydrate according to the proportion of 0.3mL/100g of body weight, hairs in the chest front area are removed by depilatory cream, then the rats are fixed on a heart super detection platform, and the coupling agent is coated on the joint of the limbs and the metal sheet on the plate in advance.
(3) M-mode was selected by applying a couplant over the heart site of the rat and dropping the probe to combine with the couplant at the heart site.
(4) Positioning a probe: the probe is vertically placed, and the incisure faces the head of the animal; the rat heart was rotated counterclockwise by about 35 degrees to obtain an image of the long axis (PLAX) section of the rat heart. The parasternal short axis section is then obtained by rotating the probe 90 degrees clockwise at a PLAX section. Further adjustment of the y-axis displacement is often required to obtain the correct slice.
(5) The index indicating the heart function of the rat can be obtained through data analysis. Thus, comparisons of cardiac function between different groups were made.
The results are shown in FIG. 8. With time, the cardiac function (ejection fraction and left ventricular short axis shortening rate) of the exosome group, the polypeptide nanofiber hydrogel containing PA-GG-Hexarelin-S, and the polypeptide nanofiber hydrogel encapsulating exosomes group was improved, and the cardiac function of the polypeptide nanofiber hydrogel encapsulating exosomes was more significantly improved at 28d compared with the other groups. The polypeptide nanofiber hydrogel group encapsulating the exosomes is shown to have a more significant effect on improving the cardiac function.
Sequence listing
<110> Suzhou university
<120> polypeptide nanofiber hydrogel for slow release of exosome, and preparation method and application thereof
<130> MP1810542
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 17
<212> PRT
<213> Artificial Sequence (Artificial Sequence)
<220>
<221> UNSURE
<222> (1)..(17)
<223> Xaa(1)=Palmitoyl-Gly;Xaa(12)=D-2-methyl-Trp;Xaa(16)=D-Phe;
<400> 1
Xaa Thr Ala Gly Leu Ile Gly Gln Gly Gly His Xaa Ala Trp Xaa Lys
1 5 10 15
Ser
Claims (6)
1. A polypeptide nanofiber hydrogel, which is prepared from a self-assembled polypeptide shown in SEQ ID No.1, NapFF and exosomes, wherein the self-assembled polypeptide and NapFF form hydrogel, and the exosomes are encapsulated in the hydrogel;
the source of the exosome is umbilical cord mesenchymal stem cells.
2. The polypeptide nanofiber hydrogel according to claim 1, wherein the mass ratio of NapFF, the self-assembled polypeptide and the exosome is 4 (0.0002-2): 1.
3. The polypeptide nanofiber hydrogel according to claim 1, wherein the mass ratio of NapFF, self-assembled polypeptide and exosome is 4 (0.0002-0.02): 1.
4. The method for preparing the polypeptide nanofiber hydrogel as claimed in claim 2 or 3, wherein the self-assembly polypeptide as claimed in claim 1 is mixed with NapFF to obtain a polypeptide mixed solution, and the polypeptide mixed solution is heated, cooled to room temperature and then uniformly mixed with exosomes to form the polypeptide nanofiber hydrogel for self-assembly encapsulation of exosomes.
5. The preparation method according to claim 4, wherein the volume ratio of the self-assembly polypeptide to NapFF and exosome is 8:1: 1.
6. Use of the polypeptide nanofiber hydrogel of claim 3 for the preparation of a medicament for the treatment of cardiovascular disease; the cardiovascular disease is myocardial infarction.
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| CN113321709A (en) * | 2021-05-18 | 2021-08-31 | 南方医科大学 | Self-assembled polypeptide, slow-release exosome polypeptide hydrogel and preparation method and application thereof |
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