CN115569201A - Polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues and preparation method thereof - Google Patents

Polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues and preparation method thereof Download PDF

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CN115569201A
CN115569201A CN202211192244.5A CN202211192244A CN115569201A CN 115569201 A CN115569201 A CN 115569201A CN 202211192244 A CN202211192244 A CN 202211192244A CN 115569201 A CN115569201 A CN 115569201A
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张春乐
蒋琴
付平
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West China Hospital of Sichuan University
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Abstract

The invention discloses a polyphenol nanoparticle for navigating stem cell targeted kidney lesion tissues and a preparation method thereof, wherein the preparation method comprises the steps of preparing the polyphenol nanoparticle, preparing sulfhydrylated serine SH-S and sulfhydrylated polypeptide SH-E7, modifying SH-S and SH-E7 on the surface of the polyphenol nanoparticle and the like. The prepared nano-particles contain a large amount of phenolic hydroxyl groups and have the biological function of regulating tissue inflammation; meanwhile, the o-quinone group in the particles has strong protein adhesion performance and can be effectively combined with the cell membrane of the stem cell; the double bonds contained in the granules can be firmly modified on the surfaces of the granules by click chemistry reaction with L-serine modified by cysteine, and the navigation stem cells are targeted to the damaged kidney parts, so that the granules are used for treating acute/chronic kidney diseases.

Description

Polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues and preparation method thereof
Technical Field
The invention belongs to the technical field of biomedical functional materials, and particularly relates to polyphenol nanoparticles for navigating stem cells to target kidney lesion tissues and a preparation method thereof.
Background
Acute Kidney Injury (AKI) is one of the clinically common critical conditions, with mortality rates as high as 40% to 80%, with approximately 13% of patients progressing to chronic kidney disease within three years. Renal fibrosis is a common pathophysiological basis for Chronic Kidney Disease (CKD) of various causes, and is the ultimate result of all CKD. Currently, in clinical work, the treatment means for inhibiting or reversing acute and chronic kidney injury are very limited, and based on the limitations of the current treatment, new methods for treating kidney diseases need to be researched and developed.
The stem cells are progenitor cells capable of generating a wide and complex pluripotent cell lineage, have wide sources and low immunogenicity, have biological functions of resisting apoptosis, angiogenesis, inflammation, immunoregulation and the like, and can effectively repair damaged tissues. The stem cells are widely applied to treatment and research of various diseases, obtain better effects in treatment of various autoimmune diseases, metabolic diseases and nervous system diseases, and show good regeneration effects in organ injuries and tissue defects of different degrees through a tissue engineering technology. Stem cell therapy has proven to be a promising therapeutic strategy.
In addition to the need for high quality stem cells, it is necessary to focus on the safe and efficient homing rate of exogenous MSCs, i.e., the process by which MSCs are captured within the vasculature of the target tissue and subsequently migrate across the vascular endothelial cells to the target tissue. At present, stem cell therapy is mainly performed by circulatory system administration (including intravenous injection and arterial injection) and local injection, wherein the intravenous injection is most commonly used in treatment due to simple operation, small invasiveness and strong repeatability. However, intravenous injection has the problem that the number of cells reaching the damaged part for therapeutic action is reduced due to the cell retention in the lung. Therefore, how to increase the homing rate of the exogenous stem cells is an urgent problem to be solved.
Disclosure of Invention
Aiming at the prior art, the invention provides polyphenol nanoparticles for navigating stem cells to target kidney lesion tissues and a preparation method thereof. The nano-particle material of the invention firstly oxidizes, crosslinks and polymerizes polyamine and polyphenol compounds to form nano-particles with various active groups (amino, carboxyl, phenolic hydroxyl, double bonds and the like) in the presence of an oxidant; then, through click chemical reaction, the polypeptide E7 and L-serine subjected to sulfhydrylation treatment are simultaneously modified on the surfaces of the nanoparticles, the polypeptide E7 on the surfaces of the nanoparticles can be specifically combined with stem cells, polyphenol nanoparticles are modified on the surfaces of stem cell membranes, and the L-serine modified on the surfaces of the particles can guide the stem cells in blood to damaged kidney tissue parts, so that the corresponding biological functions of the particles can be exerted, and acute and chronic kidney diseases can be effectively treated.
In order to achieve the purpose, the invention adopts the technical scheme that: the preparation method of the polyphenol nanoparticles for navigating the stem cells to target kidney lesion tissues comprises the following steps:
s1: dissolving polyphenol compound, polyamino compound and oxidant in alkaline buffer solution, and stirring and reacting for 5-48 h at 15-35 ℃;
s2: centrifuging the solution after reaction, taking the precipitate, and then sequentially cleaning and freeze-drying to obtain polyphenol nano-particles;
s3: modifying serine and polypeptide E7 with cysteine to obtain sulfhydrylated serine SH-S and sulfhydrylated polypeptide SH-E7;
s4: dispersing SH-S, SH-E7 and polyphenol nanoparticles in water, irradiating for 0.5-60 min by ultraviolet, centrifuging, cleaning, and freeze-drying to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney focus tissues.
On the basis of the technical scheme, the invention can be further improved as follows.
Further, the final concentrations of the polyphenol compound, the polyamino compound and the oxidant in the alkaline buffer solution are 0.5-10 mg/mL, 0.5-10 mg/mL and 0.1-5 mg/mL respectively
Further, the polyphenol compound is at least one of catechol, ferulic acid, resorcinol, flavones, anthocyanins, flavonol and glycosides thereof, isoflavones, gallic acid, ellagic acid, tannic acid, procyanidins and pyrogallol.
Further, the polyamino compound is at least one of ethylenediamine, pentanediamine, 2, 4-trimethylhexamethylenediamine, 1, 8-diaminooctane, methylcyclohexanediamine, 1, 3-diaminomethylcyclohexane, 2,4, 6-triaminomethylcyclohexane, 1, 4-biscyclohexane and 2- (3, 4-dihydroxyphenyl) ethylamine.
Further, the oxidant is at least one of hydrogen peroxide, ammonium persulfate, concentrated nitric acid, sodium periodate, potassium permanganate and potassium dichromate.
Further, the alkaline buffer solution is a phosphate buffer solution with a pH value of 8-11, a tris buffer solution, a borate buffer solution, a glycine-NaOH buffer solution, a glycinamide buffer solution or a sodium bicarbonate-sodium carbonate buffer solution.
Further, the sulfhydrylated polypeptide SH-E7 is prepared by the following steps: dissolving the polypeptide E7 and cysteine in 20wt% piperidine according to the mass ratio of 1-3: 0.5-1, blowing nitrogen for 18-22 min, separating, washing with N, N-dimethylformamide for 5-7 times, and drying to obtain the polypeptide.
Further, the thiolated serine SH-S is prepared by the following steps: dissolving serine and cysteine in 20wt% piperidine according to the mass ratio of 2-4.
Further, the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in water are 0.05-1 mg/mL, 0.05-1 mg/mL and 0.5-10 mg/mL respectively; the ultraviolet wavelength of the ultraviolet radiation is 254 nm-365 nm, and the ultraviolet radiation time is 5-45 min.
The application also discloses the polyphenol nanoparticles of the navigated stem cell targeted kidney lesion tissues prepared by the method.
The invention has the beneficial effects that:
1. the invention leads polyphenol compounds and polyamine compounds to be oxidized, crosslinked and polymerized in the presence of an oxidant to form polyphenol nanoparticles with a plurality of active groups, and the active groups not only have synergistic effect, but also respectively exert corresponding biological functions. The carboxyl, the amino and the phenolic hydroxyl can cooperatively maintain the hydrophilicity of the nano particles, so that the hydrophilic nano particles have excellent biocompatibility; the phenolic hydroxyl can maintain the corresponding biological functions of polyphenol, such as regulating the inflammation of focal tissues; the double bond is used as a connecting group for further grafting a specific bioactive substance, can react with polypeptide E7 modified by cysteine and L-serine, and modifies two specific proteins on the surface of the nanoparticle through a click chemical reaction, so that the nanoparticle has the performance of a stem cell specifically captured, the stem cell is targeted to a kidney focus tissue, the homing rate of the stem cell is improved, and the corresponding biological function of the stem cell is further exerted.
2. The o-phenolic hydroxyl structure of the polyphenol compound and the amino group of the polyamino compound are subjected to various chemical reactions in the presence of an oxidant, such as Michael addition reaction, schiff base reaction, free radical polymerization reaction, hydrogen bonding reaction, pi-pi stacking and the like, so that the polyphenol nanoparticles with excellent biocompatibility are formed. Subsequently, under ultraviolet irradiation, double bonds on the surfaces of the nanoparticles can generate click chemical reaction with thiolated polypeptide E7 and L-serine, and the specific active protein is chemically grafted to the surfaces of the particles.
3. The preparation method provided by the invention constructs the polyphenol nanoparticles which have the specificity and can capture the stem cells and guide the stem cells to the damaged kidney tissue part for the first time, and is expected to improve the homing rate of exogenous high-quality stem cells; and the nano-particles have the performance of regulating inflammation of a diseased tissue part, so that stem cells can better play a biological function and repair diseased tissues. The whole nanoparticle preparation process is simple to operate, the reaction conditions are mild, the reaction medicine is healthy, green and natural, the prepared nanoparticles have excellent specificity for capturing stem cells and guiding the stem cells to acute and chronic kidney disease tissues, the homing rate of the stem cells is improved, meanwhile, the targeted treatment of the stem cells is realized, and an effective treatment strategy is provided for the treatment of the acute and chronic kidney diseases.
Drawings
Fig. 1 is a transmission electron microscope image of polyphenol nanoparticles navigating stem cells to target kidney lesion tissues;
FIG. 2 shows the results of fluorescent staining of kidney tissues.
Detailed Description
The following examples are provided to illustrate specific embodiments of the present invention.
Example 1
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: dissolving catechol, ethylenediamine and sodium periodate in a phosphate buffer solution with the pH value of 9 respectively, mixing the catechol solution, the ethylenediamine solution and the sodium periodate solution to obtain a mixed solution, wherein the final concentrations of the catechol, the ethylenediamine and the sodium periodate in the mixed solution are 1mg/mL, 1mg/mL and 0.5mg/mL respectively, and then continuously stirring and reacting for 30 hours at the constant temperature of 15 ℃;
s2: centrifuging the solution obtained after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual phosphate buffer and sodium periodate in the nanoparticles, and freezing the particles to obtain polyphenol nanoparticles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 2.8, dissolving the mixture into piperidine with the concentration of 20wt% according to the material-liquid ratio of 1.4g; mixing serine and cysteine according to the mass ratio of 3;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.05mg/mL, 0.05mg/mL and 2mg/mL respectively; then immediately irradiating for 5min by using ultraviolet rays with the wavelength of 254nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nano particles;
s5: centrifuging the mixed solution after ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and freeze-drying the residual nanoparticles at the bottom to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney focus tissue.
Example 2
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: respectively dissolving anthocyanin, pentanediamine and 30% hydrogen peroxide in a phosphate buffer solution with the pH value of 8.5, mixing the three solutions, respectively keeping the final concentrations of the anthocyanin, the pentanediamine and the hydrogen peroxide in the obtained mixed solution at 1mg/mL, 1mg/mL and 25%, and continuously stirring and reacting for 20 hours at the constant temperature of 20 ℃;
s2: centrifuging the solution after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual phosphate buffer and hydrogen peroxide in the nano particles, and freezing the particles to obtain polyphenol nano particles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 3, dissolving the mixture into piperidine with the concentration of 20wt% according to the mass ratio of 1.5g to 1mL, blowing nitrogen for 22min, separating, washing 7 times by using N, N-dimethylformamide, and drying to obtain the sulfhydrylated polypeptide SH-E7; mixing serine and cysteine according to the mass ratio of 2g to 1mL, dissolving the mixture into 20wt% piperidine according to the material-liquid ratio of 2g to 1mL, blowing nitrogen for 25min, separating, washing for 5 times by using N, N-dimethylformamide, and drying to obtain sulfhydrylated serine SH-S;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.1mg/mL, 0.25mg/mL and 2.5mg/mL respectively; then immediately irradiating for 15min by using ultraviolet rays with the wavelength of 254nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nanoparticles;
s5: centrifuging the mixed solution after ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and freeze-drying the residual nanoparticles at the bottom to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney focus tissue.
Example 3
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: respectively dissolving tannic acid, 2- (3, 4-dihydroxyphenyl) ethylamine and ammonium persulfate by using a trihydroxymethyl aminomethane buffer solution (Tris) with the pH value of 9, mixing the three solutions, respectively setting the final concentrations of the tannic acid, the 2- (3, 4-dihydroxyphenyl) ethylamine and the ammonium persulfate in the obtained mixed solution to be 2mg/mL, 1mg/mL and 2mg/mL, and continuously stirring and reacting for 15 hours at the constant temperature of 25 ℃;
s2: centrifuging the solution after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual Tris and ammonium persulfate in the nanoparticles, and freezing the particles to obtain polyphenol nanoparticles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 2, dissolving the mixture into 20wt% piperidine according to the material-liquid ratio of 2g (1 mL), blowing nitrogen for 18min, separating, washing with N, N-dimethylformamide for 5 times, and drying to obtain the sulfhydrylated polypeptide SH-E7; mixing serine and cysteine according to the mass ratio of 3;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.4mg/mL, 0.4mg/mL and 5mg/mL respectively; then immediately irradiating for 20min by using ultraviolet rays with the wavelength of 312nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nanoparticles;
s5: centrifuging the mixed solution after ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and freeze-drying the residual nanoparticles at the bottom to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney focus tissue.
Example 4
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: respectively dissolving ellagic acid, methylcyclohexanediamine and potassium permanganate in a borate buffer solution with the pH value of 10, mixing the three solutions, respectively adding the final concentrations of ellagic acid, methylcyclohexanediamine and potassium permanganate in the obtained mixed solution to 5mg/mL, 2.5mg/mL and 2.5mg/mL, and continuously stirring and reacting for 24 hours at the constant temperature of 30 ℃;
s2: centrifuging the solution after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual borate and potassium permanganate in the nano particles, and freezing the particles to obtain polyphenol nano particles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 2.8, dissolving the mixture into piperidine with the concentration of 20wt% according to the material-liquid ratio of 1.4g; mixing serine and cysteine according to the mass ratio of 3;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.5mg/mL, 0.5mg/mL and 8mg/mL respectively; then immediately irradiating for 25min by using ultraviolet rays with the wavelength of 312nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nanoparticles;
s5: and centrifuging the mixed solution subjected to ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and performing freeze drying treatment on the residual nanoparticles at the bottom to prepare the polyphenol nanoparticles of the navigated stem cell targeted kidney lesion tissues.
Example 5
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: dissolving isoflavone, 1, 3-diaminomethylcyclohexane and sodium periodate in Tris buffer solution with the pH value of 10 respectively, mixing the three solutions, and continuously stirring and reacting for 8 hours at the constant temperature of 32 ℃, wherein the final concentrations of the isoflavone, the 1, 3-diaminomethylcyclohexane and the sodium periodate in the obtained mixed solution are 8mg/mL, 4mg/mL and 4mg/mL respectively;
s2: centrifuging the solution after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual Tris and sodium periodate in the nanoparticles, and freezing the particles to obtain polyphenol nanoparticles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 3, dissolving the mixture in 20wt% piperidine according to the ratio of 1.5g to 1mL, blowing nitrogen for 22min, separating, washing with N, N-dimethylformamide for 7 times, and drying to obtain the sulfhydrylated polypeptide SH-E7; mixing serine and cysteine according to the mass ratio of 2:1, dissolving the mixture into 20wt% piperidine according to the material-liquid ratio of 2g;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.8mg/mL, 0.8mg/mL and 8mg/mL respectively; then immediately irradiating for 45min by using ultraviolet rays with the wavelength of 254nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nanoparticles;
s5: and centrifuging the mixed solution subjected to ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and performing freeze drying treatment on the residual nanoparticles at the bottom to prepare the polyphenol nanoparticles of the navigated stem cell targeted kidney lesion tissues.
Example 6
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: respectively dissolving procyanidine, 2,4, 6-triaminomethylcyclohexane and concentrated nitric acid in glycinamide buffer solution with the pH value of 10, mixing the three solutions, respectively keeping the final concentrations of procyanidine, 2,4, 6-triaminomethylcyclohexane and nitric acid in the mixed solution at 6mg/mL, 4mg/mL and 50%, and continuously stirring and reacting for 40 hours at the constant temperature of 20 ℃;
s2: centrifuging the solution after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual glycinamide and nitric acid in the nano particles, and freezing the particles to obtain polyphenol nano particles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 2, dissolving the mixture into 20wt% piperidine according to the material-liquid ratio of 2g (1 mL), blowing nitrogen for 18min, separating, washing with N, N-dimethylformamide for 5 times, and drying to obtain the sulfhydrylated polypeptide SH-E7; mixing serine and cysteine according to the mass ratio of 3;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.2mg/mL, 0.1mg/mL and 2.5mg/mL respectively; then immediately irradiating for 40min by using ultraviolet rays with the wavelength of 365nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nanoparticles;
s5: centrifuging the mixed solution after ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and freeze-drying the residual nanoparticles at the bottom to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney focus tissue.
Example 7
A polyphenol nanoparticle for navigating stem cells to target kidney lesion tissues is prepared by the following steps:
s1: respectively dissolving chlorogenic acid, 1, 4-bis (aminocyclohexane) and sodium periodate in Tris buffer solution with the pH value of 8.5, mixing the three solutions, respectively keeping the final concentrations of the chlorogenic acid, the 1, 4-bis (aminocyclohexane) and the sodium periodate in the mixed solution at 2mg/mL, 2mg/mL and 2mg/mL, and continuously stirring and reacting at the constant temperature of 30 ℃ for 24 hours;
s2: centrifuging the solution after the S1 reaction at 10000r/m for 5min, removing supernatant, dispersing and precipitating with UP water, centrifuging at 10000r/m for 5min again to remove residual Tris and sodium periodate in the nanoparticles, and freezing the particles to obtain polyphenol nanoparticles;
s3: mixing the polypeptide E7 (EPLQLKM) and cysteine according to the mass ratio of 3, dissolving the mixture into piperidine with the concentration of 20wt% according to the mass ratio of 1.5g to 1mL, blowing nitrogen for 22min, separating, washing 7 times by using N, N-dimethylformamide, and drying to obtain the sulfhydrylated polypeptide SH-E7; mixing serine and cysteine according to the mass ratio of 2g to 1mL, dissolving the mixture into 20wt% piperidine according to the material-liquid ratio of 2g to 1mL, blowing nitrogen for 25min, separating, washing for 5 times by using N, N-dimethylformamide, and drying to obtain sulfhydrylated serine SH-S;
s4: dissolving SH-S, SH-E7 and polyphenol nanoparticles by UP water respectively, and mixing the three solutions to obtain mixed solutions, wherein the final concentrations of SH-S, SH-E7 and polyphenol nanoparticles in the mixed solutions are 0.8mg/mL, 0.8mg/mL and 8mg/mL respectively; then immediately irradiating for 35min by using ultraviolet rays with the wavelength of 365nm to modify SH-S and SH-E7 on the surfaces of the polyphenol nanoparticles;
s5: centrifuging the mixed solution after ultraviolet irradiation at 10000r/m for 5min, removing supernatant, and freeze-drying the residual nanoparticles at the bottom to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney focus tissue.
Analysis of results
The properties of the polyphenol nanoparticles targeting kidney lesion tissues obtained in the above examples are similar, and the properties of the polyphenol nanoparticles prepared in example 7 are taken as an example to illustrate.
Fig. 1 is a transmission electron microscope image of a polyphenol nanoparticle for navigating stem cell targeting kidney lesion tissue, and it can be seen from the image that cysteine-modified polypeptide E7 (SH-E7) and cysteine-modified serine (SH-S) are successfully grafted to the surface of the polyphenol nanoparticle, that is, the surface of the obtained polyphenol nanoparticle for stem cell targeting kidney lesion tissue is loaded with polypeptide E7 and serine.
Modifying stem cells by polyphenol nanoparticles targeting kidney lesion tissues with navigated stem cells, injecting the modified stem cells into blood vessels from ear sources of rabbits, taking out the kidney tissues after 2 days, and performing specific fluorescent staining, wherein the results are shown in fig. 2, wherein fig. 2a is the kidney tissues, fig. 2b and fig. 2c are the immunofluorescent staining results of the kidney tissues under different scales, respectively, wherein the tissues in the dotted line of fig. 2b are glomeruli, and the off-white part indicated by the arrow of fig. 2c is a specific marker CD44 secreted by the stem cells in the glomerular tissues, which indicates that the polyphenol nanoparticles treated by SH-E7 and SH-S indeed have the biological function of navigating the stem cells to the kidney tissues.
While the present invention has been described in detail with reference to the embodiments, it should not be construed as limited to the scope of the patent. Various modifications and changes may be made by those skilled in the art without inventive work within the scope of the appended claims.

Claims (10)

1. A preparation method of polyphenol nanoparticles for navigating stem cells to target kidney lesion tissues is characterized by comprising the following steps:
s1: dissolving polyphenol compound, polyamino compound and oxidant in alkaline buffer solution, and stirring and reacting for 5-48 h at 15-35 ℃;
s2: centrifuging the solution after reaction, taking the precipitate, and then sequentially cleaning and freeze-drying to obtain polyphenol nano-particles;
s3: modifying serine and polypeptide E7 with cysteine to obtain sulfhydrylated serine SH-S and sulfhydrylated polypeptide SH-E7;
s4: dispersing SH-S, SH-E7 and polyphenol nanoparticles in water, irradiating for 0.5-60 min by ultraviolet, centrifuging, cleaning, and freeze-drying to obtain the polyphenol nanoparticles of the navigated stem cell targeted kidney lesion tissues.
2. The method of claim 1, wherein: the final concentrations of the polyphenol compound, the polyamino compound and the oxidant in the alkaline buffer solution are 0.5-10 mg/mL, 0.5-10 mg/mL and 0.1-5 mg/mL respectively.
3. The production method according to claim 1 or 2, characterized in that: the polyphenol compound is at least one of catechol, ferulic acid, resorcinol, flavonoid, anthocyanins, flavonol and its glycosides, isoflavone, gallic acid, ellagic acid, tannic acid, procyanidin and pyrogallol.
4. The production method according to claim 1 or 2, characterized in that: the polyamino compound is at least one of ethylenediamine, pentanediamine, 2, 4-trimethylhexamethylenediamine, 1, 8-diaminooctane, methylcyclohexanediamine, 1, 3-diaminomethylcyclohexane, 2,4, 6-triaminomethylcyclohexane, 1, 4-bis-diaminohexylcyclohexane and 2- (3, 4-dihydroxyphenyl) ethylamine.
5. The production method according to claim 1 or 2, characterized in that: the oxidant is at least one of hydrogen peroxide, ammonium persulfate, concentrated nitric acid, sodium periodate, potassium permanganate and potassium dichromate.
6. The production method according to claim 1 or 2, characterized in that: the alkaline buffer solution is phosphate buffer solution with the pH value of 8-11, tris buffer solution, borate buffer solution, glycine-NaOH buffer solution, glycinamide buffer solution or sodium bicarbonate-sodium carbonate buffer solution.
7. The method according to claim 1, wherein the thiolated polypeptide SH-E7 is produced by: the polypeptide E7 and cysteine are dissolved in piperidine with the concentration of 20wt% together according to the mass ratio of 1-3 to 0.5-1, nitrogen is blown for 18-22 min, and then the mixture is separated and washed by N, N-dimethylformamide for 5-7 times and dried to obtain the polypeptide.
8. The method according to claim 1, wherein the thiolated serine SH-S is prepared by the following steps: dissolving serine and cysteine in 20wt% piperidine according to the mass ratio of 2-4.
9. The method of claim 1, wherein: the final concentrations of the SH-S, SH-E7 and polyphenol nanoparticles in water are 0.05-1 mg/mL, 0.05-1 mg/mL and 0.5-10 mg/mL respectively; the ultraviolet wavelength of the ultraviolet radiation is 254 nm-365 nm, and the ultraviolet radiation time is 5-45 min.
10. The polyphenol nanoparticles of the navigated stem cell targeted kidney lesion tissue prepared by the preparation method of any one of claims 1 to 9.
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