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CN-121987666-A - Extracellular matrix-imitated hydrogel loaded with nano bait, preparation method and application

CN121987666ACN 121987666 ACN121987666 ACN 121987666ACN-121987666-A

Abstract

The application relates to the technical field of pharmacy, in particular to an extracellular matrix-imitated hydrogel loaded with nano bait, a preparation method and application thereof. The nanometer bait-loaded simulated extracellular matrix hydrogel comprises nanometer bait and extracellular matrix simulated hydrogel matrix. The nano bait (M1-NDs) is prepared from M1 type macrophage membrane stimulated by lipopolysaccharide and interferon-gamma. In an inflammatory environment, M1-NDs can selectively capture lipopolysaccharide and proinflammatory cytokines, thereby inhibiting NF- κB pathway activation and reducing inflammatory cascade. The ECM bionic hydrogel prepared by taking collagen, hyaluronic acid and sodium alginate as raw materials is taken as a carrier of M1-NDs, so that the biological activity of the nano bait can be effectively maintained, the slow release of the nano bait on the local part of a wound surface is realized, and meanwhile, the collagen deposition, the extracellular matrix remodeling and the angiogenesis are promoted, so that the healing of inflammatory wound surfaces is accelerated, and the synergistic effect of anti-inflammatory and tissue regeneration is realized.

Inventors

  • QIU YIHUI
  • WANG SHUNFU
  • TANG QI
  • HAN JINGANG
  • Xiao Tingfeng
  • ZHU KAIXIN
  • JIN WENZHANG
  • ZHANG WA

Assignees

  • 温州医科大学附属第一医院

Dates

Publication Date
20260508
Application Date
20260127

Claims (9)

  1. 1. The nanometer bait-loaded extracellular matrix-imitated hydrogel is characterized by comprising nanometer bait from an M1 type macrophage membrane and an extracellular matrix-imitated hydrogel matrix, wherein the M1 type macrophage membrane is M1 type phenotype macrophage induced by stimulating mouse bone marrow-derived macrophages with lipopolysaccharide and interferon-gamma, and the extracellular matrix-imitated hydrogel matrix is prepared from collagen, hyaluronic acid and sodium alginate.
  2. 2. The method for preparing the nano-bait loaded extracellular matrix-like hydrogel according to claim 1, comprising the following steps: (1) Co-culturing bone marrow-derived macrophages with lipopolysaccharide and interferon-gamma in a medium to induce differentiation into M1-type macrophages; (2) Separating and extracting the M1 type macrophages cultured in the step (1) to obtain an M1 type macrophage membrane, and extruding the M1 type macrophage membrane by using a micro extruder to obtain the nano bait; (3) Preparing a mixed system containing the methacryloyl hyaluronic acid, collagen, sodium alginate, the nano bait prepared in the step (2) and a cross-linking agent, and after a gelation reaction, cross-linking to prepare the extracellular matrix-imitated hydrogel carrying the nano bait.
  3. 3. The method of claim 2, wherein in step (1), the bone marrow-derived macrophages are derived from mice.
  4. 4. The method for preparing the nano-bait-loaded extracellular matrix-like hydrogel according to claim 3, wherein cells are isolated from the bone marrow of the mice and inoculated into a Du's modified eagle medium containing 10% fetal calf serum, 1% penicillin-streptomycin mixed solution and 20 ng/mL macrophage colony stimulating factor for culturing and differentiating, thereby obtaining the macrophages derived from the bone marrow of the mice.
  5. 5. The method of claim 2, wherein in step (1), 20 ng/mL lipopolysaccharide and 50 ng/mL interferon-gamma are added to a medium containing bone marrow-derived macrophages, and cells are stimulated for 24 hours to obtain M1 type macrophages.
  6. 6. The method for preparing the simulated extracellular matrix hydrogel loaded with the nano bait according to claim 2, wherein in the step (2), M1 type macrophages are washed by PBS buffer solution, then cell precipitates are collected by centrifugation of 700 Xg and 10min, the cell precipitates are resuspended in precooled extraction buffer solution A added with protease inhibitor mixture, then the cell suspensions are frozen in liquid nitrogen and thawed at room temperature, the freezing and thawing operation is repeated for a plurality of cycles, centrifugation of 700 Xg and 10min is carried out after freezing and thawing, supernatant is collected and further centrifugation of 14000 Xg and 30min is carried out to obtain membrane component precipitates, and a micro extruder is used for sequentially passing the membrane component precipitates through polycarbonate porous filter membranes of 400 nm, 100nm and 50 nm, and each pore-size filter membrane is extruded 10 times to obtain the nano bait.
  7. 7. The method for preparing the nano-bait-loaded extracellular matrix-like hydrogel according to claim 2, wherein in the step (3), an aqueous solution of methacryloylated hyaluronic acid is prepared, and then a PBS buffer, a sodium hydroxide solution, a collagen solution, a sodium alginate solution, a suspension containing the M1-type nano-bait and a cross-linking agent are added, and the mixture is gently blown to be uniformly mixed to obtain a mixed system.
  8. 8. The method for preparing a nano-bait-loaded extracellular matrix-like hydrogel according to claim 2, wherein in the step (3), the cross-linking agent is phenyl-2, 4, 6-trimethylbenzoyl lithium phosphinate, and the mixed system is incubated for 30min at 37 ℃ for gelation. Finally, the hydrogel is placed under ultraviolet light to crosslink for 40 seconds, and the simulated extracellular matrix hydrogel carrying the nano bait is prepared.
  9. 9. Use of the nano-bait loaded simulated extracellular matrix hydrogel according to claim 1 for preparing a medicament for treating inflammatory wounds.

Description

Extracellular matrix-imitated hydrogel loaded with nano bait, preparation method and application Technical Field The application relates to the technical field of pharmacy, in particular to an extracellular matrix-imitated hydrogel loaded with nano bait, a preparation method and application thereof. Background Inflammatory wounds refer to local tissue damage caused by factors such as infection, trauma or chronic disease, and are characterized by an impaired sustained inflammatory response and regeneration process. The pathological mechanisms of such wounds involve abnormal regulation of multiple signaling pathways, including excessive release of pro-inflammatory cytokines (e.g., tumor necrosis factor- α, interleukin-1 β, interleukin-6), exacerbation of neutrophil infiltration, and macrophage polarization disorders. These changes can lead to abnormal elevation of Reactive Oxygen Species (ROS) and Matrix Metalloproteinases (MMPs) within the tissue microenvironment. In recent years, camouflage of the nanoscopic baits by cell membranes has become a very potential therapeutic strategy. The cell membrane camouflage nano bait is a nano treatment platform based on cell bionics, and natural cell membranes are wrapped on the surface of a nano core to simulate the biological characteristics of target cells or tissues, so that harmful substances such as pathogens, toxins, autoantibodies or pro-inflammatory factors are competitively combined and removed, and the aim of treating diseases is achieved. The natural recognition function of the biological film and the adjustability of the nano material are ingeniously utilized, and the biological film becomes a research hot spot in the field of biomedicine in recent years. When a pathogen (e.g., virus, bacteria) or abnormal cell (e.g., cancer cell) attempts to bind to healthy cells, the nanobait will preferentially bind to it, thereby preventing its attack on the host normal cells. The action mechanism has great potential in two fields of anti-infection and immunoregulation. Has remarkable treatment effect in various disease models such as inflammatory diseases, pathogen infection, chemical substance poisoning and the like, and provides a promising treatment strategy. However, in theory, the cell membrane camouflage nano bait has a great application prospect in the application of inflammatory wounds, and the inflammatory wounds are characterized by continuous existence of inflammation and impaired healing function, and the pathogenesis of the inflammatory wounds is related to abnormal regulation of molecular signal channels and excessive release of proinflammatory cytokines. However, the application of the nano bait on the inflammatory wound surface is less studied at present, and an effective means is lacked, so that the nano bait can effectively and continuously act on the inflammatory wound surface. Disclosure of Invention Aiming at the defects of the prior art, the application provides an extracellular matrix-imitated hydrogel loaded with nano bait, a preparation method and application thereof. The technical scheme is that the nano-bait-loaded simulated extracellular matrix hydrogel comprises nano-bait derived from an M1 type macrophage membrane and an extracellular matrix simulated hydrogel matrix, wherein the M1 type macrophage membrane is M1 type phenotype macrophage induced by stimulating mouse bone marrow-derived macrophages with lipopolysaccharide and interferon-gamma, and the extracellular matrix simulated hydrogel matrix is prepared from collagen, hyaluronic acid and sodium alginate. The preparation method of the imitated extracellular matrix hydrogel loaded with the nano bait comprises the following steps: (1) Co-culturing bone marrow-derived macrophages with lipopolysaccharide and interferon-gamma in a medium to induce differentiation into M1-type macrophages; (2) Separating and extracting the M1 type macrophages cultured in the step (1) to obtain an M1 type macrophage membrane, and extruding the M1 type macrophage membrane by using a micro extruder to obtain the nano bait; (3) Preparing a mixed system containing the methacryloyl hyaluronic acid, collagen, sodium alginate, the nano bait prepared in the step (2) and a cross-linking agent, and after a gelation reaction, cross-linking to prepare the extracellular matrix-imitated hydrogel carrying the nano bait. Further, in step (1), the bone marrow-derived macrophages are derived from mice. Further, cells were isolated from the mouse bone marrow and inoculated in Du's modified eagle's medium containing 10% fetal calf serum, 1% penicillin-streptomycin mixture and 20 ng/mL macrophage colony stimulating factor for culture and differentiation to obtain mouse bone marrow-derived macrophages. Further, in the step (1), 20 ng/mL lipopolysaccharide and 50 ng/mL interferon-gamma are added to a medium containing bone marrow-derived macrophages, and the cells are stimulated for 24 hours to obtain M1 type macrophages. Further,