Intravascular stent for regulating neutrophil phenotype and preparation method thereof
By adding neutrophil chemotaxis active substances and phenotypic regulatory substances to the outer wall coating of the vascular stent, the transformation of neutrophils to the N2 phenotype was solved, and the problem that existing vascular stents were unable to effectively regulate the inflammatory microenvironment was achieved, and the regeneration and repair of the endometrial tissue and the improvement of the immune microenvironment were achieved.
Patent Information
- Application Number
- CN202510375814.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-03-27
- Publication Date
- 2025-06-27
AI Technical Summary
Existing vascular stents cannot specifically regulate the lesion's inflammatory microenvironment, resulting in a series of complications in clinical practice.
A vascular scaffold is designed, and its outer wall coating contains neutrophil chemotaxis active substances and neutrophil phenotype regulatory substances. A uniform drug-loaded polymer coating is prepared by spraying method, which induces neutrophils to infiltrate into the lesion and converts to the N2 phenotype, thereby regulating the immune microenvironment.
By regulating the neutrophil phenotype, inhibiting the inflammatory response of AS lesions, promoting the regeneration and repair of vascular endometrial tissue, improving the immune microenvironment, and achieving endothelialization on the surface of the scaffold.
Smart Images
Figure CN120204482A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of cardiovascular implant materials, and more particularly, to a vascular stent for regulating neutrophil phenotype and a preparation method thereof. Background Art
[0002] Atherosclerosis (AS), as the main pathological basis of highly disabling and lethal diseases such as ischemic heart disease and ischemic stroke, accounts for more than 70% of the total deaths from cardiovascular diseases. As an important means for treating critically ill AS patients, vascular stents have been clinically applied for more than 30 years. However, current vascular stents cannot specifically regulate the inflammatory microenvironment at the lesion site, resulting in a series of clinical complications.
[0003] In recent years, studies have found that neutrophils, as the most abundant immune cells in the blood circulation, play an important role in the occurrence and development of AS. For example, neutrophils promote the migration of myeloid cells such as monocytes to the lesion area by secreting cathelicidin and cathepsin G. In addition, NETs secreted by neutrophils can activate the NLRP3 and AIM2 inflammasomes of macrophages, inducing the secretion of pro-inflammatory factors IL-1β and IL-18, and exacerbating the inflammatory response at the lesion site. More importantly, the intimal mechanical injury caused by the stent implantation process will massively recruit neutrophils and further exacerbate the neutrophil-related inflammatory response.
[0004] However, neutrophils are heterogeneous. N1 phenotype neutrophils have a pro-inflammatory effect, while N2 phenotype neutrophils play an important role in promoting tissue regeneration and repair. In addition, N2 neutrophils can induce macrophages to transform from the pro-inflammatory M1 phenotype to the anti-inflammatory and pro-repair M2 phenotype and enhance their efferocytosis function. Therefore, regulating the phenotype of self-recruited neutrophils after stent implantation is of great significance for improving the local inflammatory microenvironment.
[0005] In view of this, the present invention is specifically proposed. Summary of the Invention
[0006] The purpose of the present invention is to provide a vascular stent for regulating neutrophil phenotype and a preparation method thereof, providing a vascular stent for regulating neutrophil phenotype to improve the immune microenvironment related to vascular intimal tissue regeneration and repair.
[0007] The present invention is implemented as follows:
[0008] In a first aspect, the present invention provides a vascular stent for regulating neutrophil phenotype, including a substrate and a polymer coating loaded on the substrate, and the polymer coating contains neutrophil chemotactic active substances and neutrophil phenotype regulating substances.
[0009] In an alternative embodiment, the neutrophil chemotactic substance is selected from at least one of chemokines, complement cleavage products, antimicrobial peptides, and bacteria-derived chemotactic peptides; wherein, the chemokines include CXCL8 and CXCL4, the complement cleavage products include C5a and C3a, the antimicrobial peptide includes LL-37, and the bacteria-derived chemotactic peptide includes fMLP;
[0010] Preferably, the neutrophil chemotactic substance is selected from at least one of CXCL8, LL37, and fMLP.
[0011] In an alternative embodiment, the neutrophil phenotype regulatory substance is selected from at least one of adrenocortical steroids, cyclooxygenase-2 inhibitors, agonists of chemokines or their receptors, Omega-3 fatty acids, interferon-β, cytokines and signal pathway regulators, transforming growth factor-β, and plant polyphenols; wherein, the adrenocortical steroids include dexamethasone and prednisone, the cyclooxygenase-2 inhibitor includes celecoxib, the agonists of chemokines or their receptors include agonists of CXCL1, CXCL2, and CXCR2, the Omega-3 fatty acids include eicosapentaenoic acid and docosahexaenoic acid, the cytokines and signal pathway regulators include IL-10, TGF-β, and VEGF, and the plant polyphenols include resveratrol, curcumin, and quercetin;
[0012] Preferably, the neutrophil phenotype regulatory substance is selected from at least one of the CXCL1 agonist, TGF-β, and resveratrol.
[0013] In an alternative embodiment, in the polymer coating, the mass ratio of the polymer, the neutrophil chemotactic substance, and the neutrophil phenotype regulatory substance is (0.5-25):(0.02-5):(0.1-15);
[0014] Preferably, the polymer in the polymer coating is selected from at least one of polyesters, polyethers, polyamino acids, polycarbonates, polyanhydrides, and polysaccharides; wherein, the polyesters include polylactic acid, polycaprolactone, and poly(butylene adipate-co-terephthalate); the polyethers include polyethylene glycol and poly(ethylene oxide); the polyamino acids include polyglutamic acid and polyaspartic acid; the polycarbonates include poly(butylene carbonate) and poly(butylene carbonate-co-tricyclodecane dimethanol carbonate); the polyanhydrides include poly(sebacic anhydride), poly(adipic anhydride), poly(fumaric anhydride), poly(citric anhydride); the polysaccharides include starch-based compounds and chitosan-based compounds;
[0015] Preferably, the vascular stent is a metal-based vascular stent.
[0016] Second aspect, the present invention provides a method for preparing a vascular stent according to any one of the foregoing embodiments, comprising: mixing a neutrophil chemotactic activity substance, a neutrophil phenotype regulation substance and a polymer to obtain a mixed solution;
[0017] Coating the mixed solution on the outer wall surface of the vascular stent, and then drying.
[0018] In an alternative embodiment, in the mixed solution, the mass fraction of the polymer is 0.5%-25%, the mass fraction of the neutrophil chemotactic activity substance is 0.02%-5%, and the mass fraction of the neutrophil phenotype regulation substance is 0.1%-15%.
[0019] In an alternative embodiment, when preparing the mixed solution, at least one of water and an organic solvent is used to dissolve the raw materials;
[0020] Preferably, the organic solvent is selected from at least one of hydrocarbons and their derivatives, esters, alcohols, ethers and ketones, wherein the hydrocarbons and their derivatives include n-hexane, chloroform, xylene and polydimethylsiloxane; the esters include dioctyl carbonate and tributyl phosphate; the alcohols include cetyl alcohol and stearyl alcohol; the ethers include diethyl ether and tetrahydrofuran; the ketones include acetophenone and methyl isobutyl ketone.
[0021] In an alternative embodiment, a rod-shaped tool is used to shield the inner wall of the stent, and then the mixed solution is sprayed onto the outer wall of the stent.
[0022] In an alternative embodiment, when spraying the mixed solution, the power of the ultrasonic atomizer is controlled to be 0.5W-3W, the liquid inlet speed is 0.01mL / min-0.3mL / min, and the distance between the nozzle and the substrate is 5mm-50mm;
[0023] In an alternative embodiment, after spraying the mixed solution, it is first placed at 15°C-35°C for 20h-30h, and then vacuum dried at a temperature of 15°C-35°C and a pressure of 0.01MPa-10MPa for 2h-48h.
[0024] The present invention has the following beneficial effects: The present invention forms a polymer coating loaded with neutrophil chemotactic activity substance and neutrophil phenotype regulation substance on the outer wall of the vascular stent. This coating can induce a large number of neutrophils recruited during the implantation of the vascular stent to infiltrate into the lesion and transform into the N2 phenotype. Subsequently, neutrophils play an upstream immune regulation role and induce The transformation from the pro-inflammatory M1 type to the pro-repair M2 type, and further secrete pro-repair related factors, improve the immune microenvironment related to the regeneration and repair of vascular intimal tissue, realize endothelialization of the stent surface, and finally realize the regeneration and repair of vascular intimal tissue. Description of the Drawings
[0025] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings required for the embodiments will be briefly introduced below. It should be understood that the following accompanying drawings only show certain embodiments of the present invention, and thus should not be regarded as limiting the scope. For those of ordinary skill in the art, without creative efforts, other related accompanying drawings can also be obtained based on these drawings.
[0026] Figure 1 Neutrophil-specific fluorescence staining images of the coating surfaces obtained in Comparative Example 1 and Example 1;
[0027] Figure 2 Neutrophil-specific fluorescence staining images of the coating surfaces obtained in Comparative Example 2 and Example 1;
[0028] Figure 3 Specific fluorescence staining images of macrophages on the coating surface after co-incubating neutrophils and macrophages on the coating surfaces obtained in Comparative Example 2 and Example 1. Detailed implementation manners
[0029] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. For those conditions not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. For reagents or instruments whose manufacturers are not specified, they are all conventional products that can be obtained through commercial purchase.
[0030] The present invention provides a vascular stent for regulating neutrophil phenotypes. By regulating the transformation of neutrophils into the N2 phenotype, the inflammatory response at the AS lesion site is effectively inhibited, and the immune microenvironment is regulated upstream to transform towards a pro-repair direction, ultimately achieving the regeneration and repair of the intimal tissue on the stent surface.
[0031] The embodiments of the present invention provide a preparation method for a vascular stent for regulating neutrophil phenotypes, and the steps are as follows:
[0032] S1. Prepare a mixed solution
[0033] A mixed solution is obtained by mixing and dissolving a neutrophil chemotactic active substance, a neutrophil phenotype regulating substance, and a polymer, and is reserved for use.
[0034] In some embodiments, the neutrophil chemotactic active substance is selected from at least one of chemokines, complement cleavage products, antimicrobial peptides, and bacterial-derived chemotactic peptides, and the neutrophil chemotactic active substance can be any one or several of the above. Among them, chemokines include CXCL8 and CXCL4, complement cleavage products include C5a and C3a, antimicrobial peptides include LL-37, and bacterial-derived chemotactic peptides include fMLP. CXCL8, CXCL4, C5a, C3a, LL-37, etc. are all commercially available raw materials.
[0035] In some embodiments, the neutrophil phenotype regulatory substance is selected from at least one of adrenocortical hormones, cyclooxygenase-2 (COX-2) inhibitors, agonists of chemokines or their receptors, Omega-3 fatty acids, interferon-β, cytokines and signal pathway regulators, transforming growth factor-β (TGF-β), and plant polyphenols, and the neutrophil phenotype regulatory substance can be any one or several of the above. Interferon-β can be low-dose (Interferon-β). Among them, adrenocortical hormones include dexamethasone and prednisone, cyclooxygenase-2 inhibitors include Celecoxib, agonists of chemokines or their receptors include agonists of CXCL1, CXCL2, and CXCR2 (belonging to agonists of receptors), Omega-3 fatty acids include eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), cytokines and signal pathway regulators include IL-10, TGF-β, and VEGF, and plant polyphenols include resveratrol, curcumin, and quercetin. Specifically, the above neutrophil phenotype regulatory substances are also commercially available raw materials.
[0036] In preferred embodiments, the neutrophil chemotactic active substance is selected from at least one of CXCL8, LL-37, and fMLP, and the neutrophil chemotactic active substance can be any one or several of the above; the neutrophil phenotype regulatory substance is selected from at least one of an agonist of CXCL1, TGF-β, and resveratrol, and the neutrophil phenotype regulatory substance can be any one or several of the above. By optimizing the types of the neutrophil chemotactic active substance and the neutrophil phenotype regulatory substance, the effect of regulating the transformation of neutrophils into the N2 phenotype can be enhanced, and the inflammatory response at the AS lesion site can be more effectively inhibited.
[0037] In some embodiments, the biodegradable polymer is selected from at least one of polyesters, polyethers, polyamino acids, polycarbonates, polyanhydrides, and polysaccharides, and can be any one or several of the above types. Among them, polyesters include polylactic acid (PLA), polycaprolactone (PCL), and poly(butylene adipate-co-terephthalate) (PBAT), but are not limited thereto; polyethers include polyethylene glycol (PEG) and polyethylene oxide (PEO), but are not limited thereto; polyamino acids include polyglutamic acid (PGA) and polyaspartic acid (PASP), but are not limited thereto; polycarbonates include poly(butylene carbonate) (PBC) and poly(butylene carbonate-co-tricyclodecane dimethanol carbonate) (PBTC), but are not limited thereto; polyanhydrides include poly(sebacic anhydride), poly(adipic anhydride), poly(fumaric anhydride), poly(citric anhydride), but are not limited thereto; polysaccharides include starch-based compounds and chitosan-based compounds, but are not limited thereto.
[0038] Furthermore, in the mixed solution, the mass fraction of the polymer is 0.5% - 25%, such as 0.5%, 1%, 5%, 10%, 25%, etc.; in the mixed solution, the mass fraction of the neutrophil chemotactic active substance is 0.02% - 5%, such as 0.02%, 0.5%, 1.0%, 2.0%, 3.0%, 4.0%, 5.0%, etc.; in the mixed solution, the mass fraction of the neutrophil phenotype regulatory substance is 0.1% - 15%, such as 0.1%, 0.5%, 1.0%, 5.0%, 10.0%, 15.0%, etc. By regulating the concentrations of the polymer, the neutrophil chemotactic active substance, and the neutrophil phenotype regulatory substance to control the content of each component in the prepared coating, it is possible to better regulate the transformation of neutrophils into the N2 phenotype and effectively inhibit the inflammatory reaction at the AS lesion site.
[0039] Furthermore, when preparing the mixed solution, the neutrophil chemotactic active substance, the neutrophil phenotype regulatory substance, and the polymer can be directly mixed and dissolved with the solvent, or the mixed solution can be obtained by the emulsion method. That is to say, a suitable solvent can be selected according to the solubility of the neutrophil chemotactic active substance, the neutrophil phenotype regulatory substance, and the polymer. When all the raw materials are water-soluble raw materials, water can be directly used as the solvent. If there are water-soluble and fat-soluble raw materials, a mixed solvent of water and an organic solvent is used. If water and an organic solvent are used simultaneously, a surfactant needs to be added additionally to form an emulsion.
[0040] Understandably, when preparing the mixed solution, at least one of water and organic solvents is used to dissolve the raw materials, and a suitable solvent can be selected according to the solubility of the raw materials. The organic solvent is a solvent that is poorly soluble or slightly soluble in water and is selected from at least one of hydrocarbons and their derivatives, esters, alcohols, ethers, and ketones. The solvent can be any one or several of the above. Among them, hydrocarbons and their derivatives include n-hexane, chloroform, xylene, and polydimethylsiloxane, but are not limited thereto; esters include dioctyl carbonate and tributyl phosphate, but are not limited thereto; alcohols include cetyl alcohol and stearyl alcohol, but are not limited thereto; ethers include diethyl ether and tetrahydrofuran, but are not limited thereto; ketones include acetophenone and methyl isobutyl ketone, but are not limited thereto.
[0041] S2. Coating
[0042] The mixed solution obtained in step S1 is coated on the surface of the vascular stent. A rod-shaped tool is used to shield the inner wall of the stent, and then the mixed solution is sprayed onto the outer wall of the stent by spraying.
[0043] In some embodiments, when spraying the mixed solution, the power of the ultrasonic nebulizer is controlled to be 0.5 W - 3 W (such as 0.5 W, 1.0 W, 2.0 W, 3.0 W, etc.), the liquid feeding speed is 0.01 mL / min - 0.3 mL / min (such as 0.01 mL / min, 0.02 mL / min, 0.03 mL / min, etc.), and the distance between the nozzle and the substrate is 5 mm - 50 mm (such as 5 mm, 10 mm, 20 mm, 30 mm, 40 mm, 50 mm, etc.) to form a uniform spraying coating.
[0044] Furthermore, the vascular stent can be a metal vascular stent, but is not limited thereto.
[0045] S3. Drying
[0046] After the spraying in step S2 is completed, drying is carried out to remove the solvent.
[0047] In some embodiments, after coating the mixed solution, it is first placed at 15°C - 35°C for 20 h - 30 h to play a role in preliminary drying, and then vacuum dried at a temperature of 15°C - 35°C and a pressure of 0.01 MPa - 10 MPa for 2 h - 48 h.
[0048] Specifically, the operating temperature during preliminary drying can be close to room temperature, such as 15°C, 20°C, 25°C, 30°C, 35°C, etc., and the placement time can be 20h, 23h, 25h, 28h, 30h, etc. When performing vacuum drying, the controlled drying temperature can be 15°C, 20°C, 25°C, 30°C, 35°C, etc., the pressure can be 0.01MPa, 0.10MPa, 1.00MPa, 5.00MPa, 10.00MPa, etc., and the vacuum drying time can be 2h, 5h, 10h, 20h, 30h, 40h, 48h, etc.
[0049] An embodiment of the present invention provides a vascular stent for regulating neutrophil phenotype, which includes a substrate and a polymer coating loaded on the substrate. The polymer coating is loaded with neutrophil chemotactic active substances and neutrophil phenotype regulating substances.
[0050] It should be noted that the vascular stent provided by the embodiment of the present invention can induce a large number of neutrophils recruited during the implantation of the vascular stent to infiltrate into the lesion and transform into the N2 phenotype. Subsequently, the neutrophils play an upstream immune regulation role and induce the transformation from the pro-inflammatory M1 type to the pro-repair M2 type, and further secrete pro-repair related factors, improve the immune microenvironment related to the regeneration and repair of the vascular intimal tissue, realize endothelialization of the stent surface, and finally realize the regeneration and repair of the vascular intimal tissue.
[0051] In some embodiments, in the polymer coating, the mass ratio of the polymer, the neutrophil chemotactic active substance, and the neutrophil phenotype regulating substance is (0.5 - 25):(0.02 - 5):(0.1 - 15), such as 0.5:0.02:0.1, 1.0:0.10:1.0, 5.0:1.00:5.0, 10.0:2.00:8.0, 20.0:4.00:10.0, 25.0:5.00:15.0, etc. It is appropriate that the mass ratio of the polymer, the neutrophil chemotactic active substance, and the neutrophil phenotype regulating substance is within the above range, which can better regulate the transformation of the upstream regulatory immune microenvironment towards the pro-repair direction and improve the regeneration and repair effect of the vascular intimal tissue.
[0052] The features and performance of the present invention will be further described in detail below in conjunction with embodiments.
[0053] Example 1
[0054] This example provides a preparation method for a vascular stent for regulating neutrophil phenotype, and the steps are as follows:
[0055] (1) Mix a polymer (polylactic acid, molecular weight of 50,000), a neutrophil chemotactic activity substance (fMLP, purchased from Merck, product number 59880 - 97 - 6), a neutrophil phenotype regulatory substance (resveratrol, purchased from Merck, product number 501 - 36 - 0), and a solvent (tetrahydrofuran) to obtain a mixed solution. In the mixed solution, the mass fraction of the polymer is 15%, the mass fraction of the neutrophil chemotactic activity substance is 0.2%, and the mass fraction of the neutrophil phenotype regulatory substance is 2%.
[0056] (2) Shield the inner wall of the rod-shaped tool. Coat the outer wall of the metal vascular stent (316L stainless steel, ) with the mixed solution obtained in step (2) by spraying. Control the power of the ultrasonic nebulizer to be 0.5 W, the liquid inlet speed to be 0.05 mL / min, and the distance between the nozzle and the substrate to be 10 mm.
[0057] (3) After the coating is completed, place the material at room temperature (about 25 °C) for 24 h, and then put it into a vacuum drying oven and vacuum dry it at 25 °C and a vacuum degree of 0.1 MPa for 24 h.
[0058] Example 2
[0059] The difference from Example 1 is only that: the neutrophil chemotactic activity substance is replaced with an equal amount of C5a, purchased from Merck, product number P01031; the solvent is replaced with a mixed emulsion of water and tetrahydrofuran.
[0060] Example 3
[0061] The difference from Example 1 is only that: the neutrophil phenotype regulatory substance is replaced with an equal amount of quercetin, purchased from Merck, product number 117 - 39 - 5.
[0062] Example 4
[0063] The difference from Example 1 is only that: the neutrophil phenotype regulatory substance is replaced with an equal amount of IL - 10, purchased from Merck, product number I9276; the solvent is replaced with a mixed emulsion of water and tetrahydrofuran.
[0064] Example 5
[0065] The difference from Example 1 is only that: in step (1), the mass fraction of the neutrophil chemotactic activity substance is 2%.
[0066] Example 6
[0067] The difference from Example 1 is only that: in step (1), the mass fraction of the polymer is 10%.
[0068] Example 7
[0069] The difference from Example 1 is only that: in step (1), the mass fraction of the neutrophil phenotype regulatory substance is 15%.
[0070] Example 8
[0071] The difference from Example 1 is only that: in step (1), the mass fraction of the neutrophil chemotactic activity substance is 0.02%.
[0072] Example 9
[0073] The difference from Example 1 is only that: in step (1), the mass fraction of the neutrophil phenotype regulatory substance is 0.2%.
[0074] Comparative Example 1
[0075] The difference between this comparative example and Example 1 is only that: the neutrophil chemotactic activity substance is not added, so that in the total mixed solution obtained in step (1), the mass fraction of the polymer is 15%, and the mass fraction of the neutrophil phenotype regulatory substance is 2%.
[0076] Comparative Example 2
[0077] The difference between this comparative example and Example 1 is only that: the neutrophil phenotype regulatory substance is not added, so that in the total mixed solution obtained in step (1), the mass fraction of the polymer is 15%, and the mass fraction of the neutrophil chemotactic activity substance is 0.2%.
[0078] Test Example 1
[0079] Compare the neutrophil recruitment and phenotype regulation of the coatings obtained in Example 1 and Comparative Examples 1 and 2.
[0080] Test method: Neutrophils were extracted from the tibial bone marrow of mice. The materials prepared in Example 1, Comparative Example 1 and Comparative Example 2 were co-incubated with neutrophils. Subsequently, a confocal microscope was used to observe the effect of fMLP on neutrophil recruitment and the effect of resveratrol on neutrophil phenotype changes.
[0081] Figure 1 The results showed that the number of neutrophils on the surface of Example 1 was significantly higher than that of Comparative Example 1, indicating that fMLP in the polymer coating could effectively promote the chemotactic behavior of neutrophils;
[0082] Figure 2 The results showed that the expression of neutrophil YM1 in Example 1 was significantly higher than that of Comparative Example 1, indicating that resveratrol in the polymer coating could effectively promote the polarization of neutrophils towards the N2 direction.
[0083] Test Example 2
[0084] Compare the regulation of macrophage phenotype by neutrophils after pretreatment of the coating obtained in Example 1 and Comparative Example 2.
[0085] Test method: The materials prepared in Example 1 and Comparative Example 2 were incubated with neutrophils for 2 h, and then macrophages were added and incubated for a further 24 h, and the polarization of the macrophages was observed using a confocal microscope.
[0086] Figure 3 The results showed that the surface macrophages of the material obtained in Example 1 mainly expressed CD206 (M2 type), while the surface macrophages of the material obtained in Comparative Example 2 mainly expressed CD86 (M1 type), indicating that the resveratrol in the polymer coating can induce neutrophils to transform into N2 phenotype, thereby inducing macrophages to polarize to M2.
[0087] Test Example 3
[0088] Compare the effects of the coatings obtained in Examples 1-7 on the recruitment and phenotype regulation of neutrophils.
[0089] Test method: The materials prepared in Examples 1-7 were co-incubated with neutrophils, and then confocal microscopy and Image J image analysis were used to evaluate the chemotactic effect of the materials on neutrophils, and qPCR was used to quantitatively evaluate the effect of the materials on the phenotypic changes of neutrophils.
[0090] Table 1 Performance test results of materials prepared in Examples 1-7
[0091] Group <![CDATA[Number of chemotactic cells (cells / mm 2 )]]> Relative expression level of YM1 Example 1 842.4±20.3 7.38±1.52 Example 2 602.8±10.4 6.94±1.37 Example 3 811.6±12.7 5.16±1.14 Example 4 824.4±20.9 4.83±1.62 Example 5 1032.3±40.4 3.11±0.67 Example 6 704.4±20.9 6.59±0.69 Example 7 516.5±28.4 1.83±1.41 Example 8 264.8±16.8 7.11±1.33 Example 9 815.4±35.7 2.18±0.72
[0092] The results in Table 1 show that compared with Examples 2 and 6, the material prepared in Example 1 has a more significant effect on the recruitment of neutrophils, indicating that the preferred neutrophil chemotactic factor fMLP in the present invention has a stronger chemotactic effect. At the same time, compared with Examples 3 and 4, the expression level of the neutrophil YM1 (N2) marker on the surface of the material in Example 1 was significantly improved, indicating that the preferred neutrophil phenotype regulating substance resveratrol in the present invention can effectively induce neutrophils to polarize to the N2 phenotype. It is worth noting that Example 8 (low concentration fMLP group) has a low efficiency in chemotactic neutrophils, while Example 5 (high concentration fMLP group) improves the neutrophil chemotactic efficiency but reduces its YM1 (N2) marker expression; Example 9 (low concentration resveratrol group) has a low efficiency in regulating neutrophils, while Example 7 (high concentration resveratrol group) has a decreased neutrophil chemotactic function and phenotype regulating function, suggesting that excessive concentrations may lead to decreased cell activity.
[0093] In summary, the present invention provides a vascular stent for regulating neutrophil phenotype and a preparation method thereof. By using a spraying method, a uniform drug-loaded polymer coating is prepared on the outer wall of the stent. The formed coating uses a neutrophil chemotactic activity drug to induce neutrophil infiltration into the lesion. The polymer carrier controls the release of the phenotype regulation drug, efficiently induces its transformation into the N2 phenotype, and realizes the directional regulation of the downstream immune microenvironment towards the pro-repair direction.
[0094] The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. For those skilled in the art, the present invention can have various modifications and changes. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.
Claims
1. A vascular stent for regulating neutrophil phenotype, characterized in that: The invention comprises a blood vessel stent and a polymer coating loaded on the outer wall of the blood vessel stent, wherein the polymer coating contains neutrophil chemotactic active substances and neutrophil phenotype regulating substances.
2. The vascular stent according to claim 1, characterized in that: The neutrophil chemotactic active substance is selected from at least one of chemokines, complement cleavage products, antimicrobial peptides and bacterial-derived chemotactic peptides; wherein the chemokines include CXCL8 and CXCL4, the complement cleavage products include C5a and C3a, the antimicrobial peptides include LL-37, and the bacterial-derived chemotactic peptides include fMLP; Preferably, the neutrophil chemotactic active substance is selected from at least one of CXCL8, LL-37 and fMLP.
3. The vascular stent according to claim 1, characterized in that: The neutrophil phenotype regulating substance is selected from at least one of adrenal glucocorticoids, cyclooxygenase-2 inhibitors, chemokines or agonists of their receptors, omega-3 fatty acids, interferon-β, cytokines and signaling pathway regulators, transforming growth factor-β and plant polyphenols; wherein the adrenal glucocorticoids include dexamethasone and prednisone, the cyclooxygenase-2 inhibitors include celecoxib, the chemokines or agonists of their receptors include agonists of CXCL1, CXCL2 and CXCR2, the omega-3 fatty acids include eicosapentaenoic acid and docosahexaenoic acid, the cytokines and signaling pathway regulators include IL-10, TGF-β and VEGF, and the plant polyphenols include resveratrol, curcumin and quercetin; Preferably, the neutrophil phenotype regulating substance is selected from at least one of CXCL1 agonist, TGF-β and resveratrol.
4. The vascular stent according to any one of claims 1 to 3, characterized in that: In the polymer coating, the mass ratio of the polymer, the neutrophil chemotactic active substance and the neutrophil phenotype regulating substance is (0.5-25): (0.02-5): (0.1-15); Preferably, the polymer in the polymer coating is selected from at least one of polyesters, polyethers, polyamino acids, polycarbonates, polyanhydrides and polysaccharides; wherein the polyesters include polylactic acid, polycaprolactone and polybutylene terephthalate-adipate; polyethers include polyethylene glycol and polyethylene oxide; polyamino acids include polyglutamic acid and polyaspartic acid; polycarbonates include polybutylene carbonate and poly(butylene carbonate-co-tricyclodecane dimethanol carbonate); polyanhydrides include poly(sebacic anhydride), poly(adipic anhydride), poly(fumaric anhydride) and poly(citric anhydride); polysaccharides include starch-based compounds and chitosan-based compounds; Preferably, the vascular stent is a metal-based vascular stent.
5. A method for preparing the vascular stent according to any one of claims 1 to 4, characterized in that: include: Mixing the neutrophil chemotactic active substance, the neutrophil phenotype regulating substance and a polymer to obtain a mixed solution; The mixed solution is applied to the outer wall of the vascular stent and then dried.
6. The preparation method according to claim 5, characterized in that: In the mixed solution, the mass fraction of the polymer is 0.5%-25%, the mass fraction of the neutrophil chemotactic active substance is 0.02%-5%, and the mass fraction of the neutrophil phenotype regulating substance is 0.1%-15%.
7. The preparation method according to claim 5, characterized in that: When preparing the mixed solution, at least one of water and an organic solvent is used to dissolve the raw materials; Preferably, the organic solvent is selected from at least one of hydrocarbons and their derivatives, esters, alcohols, ethers and ketones, wherein the hydrocarbons and their derivatives include n-hexane, chloroform, xylene and polydimethylsiloxane; the esters include dioctyl carbonate and tributyl phosphate; the alcohols include hexadecanol and octadecyl alcohol; the ethers include diethyl ether and tetrahydrofuran; the ketones include acetophenone and methyl isobutyl ketone.
8. The preparation method according to claim 5, characterized in that: A rod-shaped tool is used to mask the inner wall of the stent, and then the mixed solution is sprayed onto the outer wall of the stent.
9. The preparation method according to claim 8, characterized in that: When spraying the mixed solution, the power of the ultrasonic atomizer is controlled to be 0.5W-3W, the liquid feeding speed is controlled to be 0.01mL / min-0.3mL / min, and the distance between the nozzle and the substrate is controlled to be 5mm-50mm.
10. The preparation method according to claim 8, characterized in that: After spraying the mixed solution, it is placed at 15°C-35°C for 20h-30h, and then vacuum dried at a temperature of 15°C-35°C and a pressure of 0.01MPa-10MPa for 2h-48h.