Search

CN-121971713-A - Preparation method and application of intelligent response hydrogel for programmed delivery of hypoxia-energized stem cell vesicles

CN121971713ACN 121971713 ACN121971713 ACN 121971713ACN-121971713-A

Abstract

The invention discloses a preparation method of intelligent response hydrogel for programmed delivery of hypoxia-energized stem cell vesicles and application of the intelligent response hydrogel in promotion of postoperative non-adhesion healing of peritoneum. The hydrogel is formed by crosslinking gallic acid modified chitosan of a first component loaded hypoxia energized stem cell vesicle and oxidized hyaluronic acid grafted by a second component amino phenylboronic acid, and a dual-network structure is constructed by phenylboronic acid ester bonds and dynamic covalent imine bonds. Hypoxia-energized stem cell vesicles cooperate with the hydrogel matrix to promote polarization of macrophages to an anti-inflammatory repair phenotype and to promote repair of the peritoneum. The phenylboronic acid ester bonds in the hydrogel respond to the high active oxygen level programming delivery hypoxia energized stem cell vesicles of the postoperative injury area, and the response of the forming material is coordinated with the biological intervention, so that the problem that the stem cell vesicles are difficult to accurately regulate and control the key links of adhesion formation is solved, and a comprehensive solution with physical barrier and tissue repair functions is provided for postoperative peritoneal non-adhesion healing.

Inventors

  • ZHAO CHUNSHUN
  • XIAO DANNI
  • HUANG YANJUAN

Assignees

  • 中山大学

Dates

Publication Date
20260505
Application Date
20260210

Claims (8)

  1. 1. The intelligent response hydrogel for programmed delivery of the hypoxia-enhanced stem cell vesicles is characterized by being prepared by mixing a first component solution and a second component solution to perform a crosslinking reaction, wherein the first component is a gallic acid modified chitosan solution mixed with the hypoxia-enhanced stem cell vesicles, the second component is an amino phenylboronic acid grafted oxidized hyaluronic acid solution, and the first component solution and the second component solution form a double crosslinking network with dynamic covalent imine bonds through a phenylboronic acid ester bond network.
  2. 2. A smart responsive hydrogel for programmed delivery of hypoxia-enhanced stem cell vesicles according to claim 1 wherein the hypoxia-enhanced stem cell vesicles have the ability to promote macrophage M2 polarization and to promote peritoneal mesothelial cell proliferation and migration and wherein the first component gallic acid modified chitosan solution imparts active oxygen scavenging ability to the hypoxia-enhanced stem cell vesicles.
  3. 3. A method of preparing a smart responsive hydrogel for the programmed delivery of hypoxia-energized stem cell vesicles according to any one of claims 1 to 2 comprising the steps of: step 1, preparing hypoxia-energized stem cell vesicles; step 2, synthesizing gallic acid modified chitosan; step 3, synthesizing oxidized hyaluronic acid of aminophenylboronic acid branches; And 4, dissolving the materials in an aqueous medium to prepare a gel precursor solution, mixing the hypoxia-energized stem cell vesicles with a gallic acid modified chitosan solution to prepare a first component, mixing an amino phenylboronic acid grafted oxidized hyaluronic acid solution of a second component with the first component, and forming a phenylboronic acid ester bond network and a dynamic covalent imine bond to prepare the double-crosslinked hydrogel.
  4. 4. A method of preparing a smart responsive hydrogel for programmed delivery of hypoxia-energized stem cell vesicles according to claim 3 wherein the preparation of the hypoxia-energized stem cell vesicles comprises the steps of: Step 1, extracting primary mesenchymal stem cells from fat sources, selecting cells of 3 rd to 8 th generation, and placing the cells in an anoxic incubator with 5 percent of oxygen volume fraction and 10 percent of carbon dioxide volume fraction for pretreatment for 24 hours before cell digestion and collection; step 2, homogenizing for 5 minutes under the pressure of 200 bar by adopting a high-pressure homogenizing method to crush cells; step 3, cell debris was removed by centrifugation at 10,000 g followed by ultracentrifugation at 130000 g for 2 hours to pellet vesicles.
  5. 5. The method for preparing intelligent response hydrogel for programmed delivery of hypoxia-energized stem cell vesicles according to claim 3 wherein the mass concentration of the gallic acid-modified chitosan solution is 2% -5% (wt%) and the grafting rate is 5% -10%, the mass concentration of the oxidized hyaluronic acid of the aminophenylboronic acid is 1% -5% (wt%) and the grafting rate is 20% -70% and the oxidation degree is 20% -50%, and the loading amount of the hypoxia-energized stem cell vesicles is 0.5-2 mg protein per 1 mL hydrogel.
  6. 6. A smart responsive hydrogel for programmed delivery of hypoxia-energized stem cell vesicles according to claim 3 wherein the volume ratio of the first component solution to the second component solution is from 4:1 to 1:2, preferably the ratio is 2:1.
  7. 7. The method of claim 3, wherein the hyaluronic acid has a molecular weight of 10-200 kDa and the chitosan has a degree of deacetylation of not less than 85%.
  8. 8. A smart responsive hydrogel for the programmed delivery of hypoxia-energized stem cell vesicles according to any one of claims 1 to 7 for use in the prevention of post-operative adhesions including any one or more of post-operative abdominal adhesions, post-operative uterine adhesions.

Description

Preparation method and application of intelligent response hydrogel for programmed delivery of hypoxia-energized stem cell vesicles Technical Field The invention belongs to the technical field of biomedical materials, and relates to intelligent response hydrogel for programmed delivery of hypoxia-energized stem cell vesicles, a preparation method thereof and application thereof in promoting postoperative peritoneal non-adhesion healing. Background Postoperative abdominal adhesion is one of the most common and unavoidable complications after abdominal surgery, and the clinical incidence rate is as high as 63% -97%. The complications can not only cause severe consequences such as intestinal obstruction, chronic abdominal pain, female infertility and the like, but also obviously increase the readmission rate of patients, the risk of secondary operation and the overall medical burden. Thus, prevention and treatment of abdominal adhesions after surgery is extremely important. Postoperative adhesions occur from the peritoneum that is subjected to mechanical injury, ischemia, or desiccation irritation during the surgical procedure, resulting in the destruction of the integrity of the peritoneal mesothelial cell layer covering the surface of the abdominal cavity. Under normal conditions, mesothelial cells maintain lubrication and barrier functions between organs in the abdominal cavity by secreting glycosaminoglycans and surface active substances, and once damaged, lose their defensive and repairing abilities, and then initiate a series of pathological cascade reactions. First, vascular permeability increases and fibrin-rich plasma components exude, activating the coagulation system. At the same time, a large number of immune cells (especially macrophages) are recruited to the wound surface, polarized into pro-inflammatory M1 type under local high-active oxygen environment, and release a large number of inflammatory factors such as interleukin-1 beta, interleukin-6 and tumor necrosis factor-alpha, forming positive feedback circulation of 'oxidative stress-inflammatory reaction'. This process further exacerbates tissue damage and inhibits fibrinolytic system activity, rendering fibrin unable to be cleared in time and deposited in large amounts on the wound surface. The fibroblast cells simultaneously acquire myofibroblast phenotype, secrete a large amount of extracellular matrix, and finally construct compact and irreversible fibrous adhesion tissues together with deposited fibrin. Current strategies for clinically preventing postoperative abdominal adhesions mainly include optimizing surgical procedures, using anti-inflammatory drugs, and implanting physical barrier materials. Among them, physical barriers such as hyaluronic acid gel, chitosan washing solution, collagen film, etc. are widely used because of high safety and easy operation. However, existing products generally have the limitations of short residence time, lack of biological activity, inability to actively intervene in pathological microenvironments, and the like. For example, although the hyaluronic acid gel has good biocompatibility, the hyaluronic acid gel is easily and rapidly degraded by hyaluronidase in abdominal cavity, the critical window period (3 to 14 days after operation) for adhesion formation is difficult to cover, the collagen material can provide a certain mechanical isolation, but has no function of regulating inflammation or promoting repair, and part of biological source materials have immunogenicity or pathogen transmission risks. Most products only play a physical barrier role, and do not actively intervene in the core links (such as peritoneal cortex injury, oxidative stress and over-immune activation) in the pathogenesis of postoperative abdominal adhesions. The mesenchymal stem cell exosome therapy is a cell-free treatment strategy, and the extracellular vesicles secreted by the mesenchymal stem cells are extracted, and the bioactive molecules such as protein, lipid, mRNA, microRNA and the like carried by the extracellular vesicles are utilized to realize multiple treatment effects such as immunoregulation, antioxidation stress, tissue repair promotion, angiogenesis and the like. Compared with the traditional cell transplantation, the exosome therapy effectively avoids the risks of immune rejection, tumor formation, low cell survival rate and the like, has the advantages of higher biostability, lower immunogenicity and easier standardized production and storage, and has wide clinical application prospect in the fields of tissue engineering, inflammatory diseases, postoperative repair and the like. To further enhance the efficacy of cell-free therapies and to increase production efficiency, researchers have developed two optimization strategies, one is pretreatment of stem cells, where hypoxia pretreatment is of particular concern. Because the natural habitat of the mesenchymal stem cells in vivo is mostly a low-oxygen environme