A liposome nanoplatform with inflammation recognition ability, its preparation method and use

Abstract

The present invention provides a liposome nanoplatform with inflammation recognition ability, its preparation method and use. The liposome nanoplatform of the present invention takes liposome as the core and loads gold cluster enzymes. The in vitro and in vivo experimental results show that the biomimetic nanoplatform of the present invention not only utilizes the high drug loading capacity of liposomes, but also combines the targeting and long-circulation ability of cell membranes, etc., achieving a good anti-inflammatory effect, providing new ideas for individualized and precise treatment. Moreover, this biomimetic nanoplatform has good biocompatibility and the possibility of large-scale production, with high clinical practical significance, and it may be applicable to the precise treatment of a variety of inflammatory diseases.

Description

Technical Field

[0001] The present invention relates to the field of biomedical technologies, and in particular, to a liposome nanoplatform with inflammation recognition ability, a preparation method thereof, and uses thereof. Background Art

[0002] Ulcerative colitis (UC) is the most common chronic non-specific inflammatory bowel disease. The incidence and recurrence rate of this disease are relatively high, the disease course is long, and various complications are easily induced. If the long-term chronic ulcerative colitis inflammation cannot be effectively controlled, it will lead to damage to the large intestine structure and increase the risk of colon cancer. Coupled with the emergence of drug resistance and drug side effects of UC drugs, UC is listed as one of the modern intractable diseases by the World Health Organization. At present, the methods and drugs for treating UC have not reached an ideal state. Therefore, researchers are urgently in need of finding new safe and effective therapeutic drugs and methods.

[0003] Ulcerative colitis is a typical refractory inflammatory bowel disease accompanied by excessive production of reactive oxygen species (ROS). Reactive oxygen species are by-products of the oxidative reduction reaction of cell mitochondria. The normal balance of ROS can bidirectionally regulate cell apoptosis and proliferation and mediate a series of signal transduction pathways. Excessive ROS will damage the integrity of cells. Any redox imbalance with elevated ROS levels in cells will cause oxidative damage to living cells and lead to various long-term disorders. Research also shows that excessive ROS can lead to pro-inflammatory activation of macrophages, thereby inducing the continuous release of pro-inflammatory cytokines, accompanied by the accumulation and apoptosis resistance of T cells, further aggravating the inflammation. The dextran sulfate sodium (DSS)-induced murine ulcerative colitis model is a widely used chemically induced model in clinical practice, and its colonic epithelial cell damage is similar to that seen in human UC in terms of morphology and symptoms.

[0004] As is well known, intracellular anti-ROS utilizes intracellular enzymes, including superoxide dismutase (SOD) and catalase (CAT). To a certain extent, it will obtain balance by self-scavenging ROS. Some studies have attempted to use SOD / CAT to combat ROS-related diseases, but the ability of their own SOD / CAT to scavenge ROS in the diseased state is significantly insufficient, and the stability is low and the cost is high, prompting researchers to seek more economical and effective alternatives. Inspired by this, our team prepared artificial enzymes, namely gold cluster enzymes, based on Au 25 clusters by a single-atom substitution method. The experimental results show that such gold cluster enzymes have ultra-high antioxidant activity, which is 137-160 times higher than that of natural antioxidants. At the same time, the gold cluster enzymes exhibit preferential enzyme-mimicking catalytic activity, Au 24 Cu1 and Au 24Cd1 shows strong selectivity for the activities of catalase (CAT-like) and superoxide dismutase (SOD-like), respectively. We combined the two gold cluster enzymes and, relying on their antioxidant properties and reactive oxygen species scavenging ability, logically it is a good candidate for the treatment of UC. However, using gold clusters alone for inflammatory treatment may lead to random distribution of the material in the body, cause adverse side effects on normal tissues, reduce the drug dosage at the lesion site, and may also be captured by immune cells to a certain extent, severely weakening its therapeutic effect. Therefore, it is necessary to develop a drug delivery system with good biocompatibility that can bypass the clearance of the immune system and improve the targeting delivery ability. Effectively delivering drugs to the target tissue is a prerequisite for disease treatment.

[0005] Liposomes have been widely used in the delivery of genes, small molecule drugs, imaging agents, proteins, and nucleic acids due to their advantages such as strong drug loading capacity, slow drug release rate, and good biocompatibility. Liposomes are easy to functionalize and can introduce various properties, such as stimulus responsiveness, enhanced drug encapsulation, tissue targeting, reduced drug toxicity in non-target tissues, extended blood circulation, and diagnostic ability, thus facilitating the delivery process and proving to be an ideal delivery platform. Importantly, liposomes, as a drug carrier, have been widely used in clinical treatment in recent years and have certain potential for clinical translation.

[0006] Modifying and modifying the surface of liposomes using cells or cell membranes can provide the translocation of cell membranes and their peripheral proteins to the liposome surface, endowing the drug-loaded liposomes with targeting properties, enhancing their interaction with the target, and greatly reducing the degradation of drugs through degradation pathways such as liver and kidney metabolism and cell phagocytosis. At the same time, cell membranes are the targets of attack by various bacterial exotoxins. While neutralizing toxins, they can aggregate at the infection site, enabling the drugs they encapsulate to be released concentratedly at the infection site, achieving a targeting effect.

[0007] Neutrophils (NCs) are the first line of defense for the host against malignant tumors and invading pathogens. Due to the sensing of chemokines, neutrophils are recruited to the site of infection or inflammatory stimuli within minutes, with the response peaking within 24 - 48 hours. They sense the chemoattractant gradient in the blood and cross the vascular endothelium to reach the intestinal lamina propria. Given the innate inflammatory targeting ability of neutrophils, they are initially recognized as a reliable source for developing treatment regimens for inflammatory diseases. However, the direct use of neutrophils as drug delivery carriers is limited because the half-life of terminally differentiated neutrophils is only 7 hours and current in vitro culture techniques are not yet mature. Once isolated from the blood or bone marrow, neutrophils must be incubated, loaded with drugs, and reinjected within a few hours. This encapsulation process is both complex and time-consuming, and the entire process also requires strict preparation and storage conditions as well as disinfection methods. Additionally, the activity of neutrophils must be ensured during the isolation process because once the structure of isolated neutrophils is damaged, they will express "find me" or "eat me" signals, triggering phagocytosis by immune cells and resulting in severe off-target effects. Furthermore, it has been reported that in a model of ulcerative colitis, depletion of intestinal mononuclear phagocytes (macrophages and dendritic cells) increases neutrophil infiltration and the severity of injury. Different from using live cells, neutrophil membrane-coated nanoparticles provide a unique platform to achieve specific delivery for cancer and inflammation treatment. If neutrophil membranes are selected, they can selectively neutralize some pro-inflammatory factors such as TNF-α and IL-β, reduce the inflammatory storm caused by the cells themselves, and retain their own targeting ability.

[0008] Even so, mononuclear phagocytes may recognize the neutrophil membrane-coated nanocarriers, consuming the delivery effect and thus resulting in insufficient therapeutic effects. Red blood cells (RBCs) are relatively abundant cells in the human body, with approximately 5 billion RBCs per milliliter of human blood, which can provide abundant coating materials for functionalizing drug carriers. In addition, CD47 is overexpressed on the red blood cell membrane, which can bind to the N-terminus of signal regulatory protein α (SIRPα) on the surface of macrophages, triggering the "don't eat me" signal to inhibit the phagocytosis of red blood cells by immune cells, thereby enabling red blood cells to survive continuously during circulation. With this progress, the integration of red blood cell membranes is expected to be a direct method to mask nanocarriers to evade immune surveillance and improve the delivery process.

[0009] We combined two cell membranes, enabling this drug-loading system to extend blood circulation time, escape the immune system, enhance biocompatibility and biodegradability in vivo, improve drug-loading capacity, the stability of nanoparticles, extend the in vitro storage time of materials, inhibit aggregation, etc. Most importantly, it can effectively target the inflammatory site, making it possible for the therapeutic drug to actively accumulate at the infection site, increasing the drug concentration at the infection site, and thus achieving a better therapeutic effect. In addition, its size is controllable, the particle size can be controlled at the nanoscale, it has good biocompatibility, stable properties, and a simple preparation method.

[0010] Meanwhile, real-time imaging of the drug delivery process under non-invasive conditions can monitor the dynamic behavior of the nanocarrier in vivo, providing a more intuitive basis for understanding the therapeutic effect and being used to optimize the treatment plan. Based on deep tissue penetration, negligible autofluorescence, and low photon scattering, second near-infrared (NIR-II, 1000 - 1700 nm) fluorescence imaging has been widely adopted to achieve in vivo tracking of target substances with high spatio-temporal resolution. Experiments found that the Au cluster enzyme (regulated by MPA ligand) developed by the single-atom substitution method we proposed can achieve NIR-II imaging applications to assist in the treatment of UC. Summary of the Invention

[0011] The purpose of the present invention is to construct a preparation of a biomimetic nanoplatform with high inflammatory recognition ability and its application in the treatment of ulcerative colitis.

[0012] The technical solution of the present invention: Develop a cell membrane-mediated biomimetic nanoplatform, load the anti-inflammatory agent gold cluster enzyme onto liposomes, and deliver it to the inflammatory site after being "disguised" with natural and activated neutrophil cell membranes and red blood cell membranes for the prevention and treatment of ulcerative colitis.

[0013] The liposome nanoplatform with inflammatory recognition ability of the present invention takes liposomes as the core and loads gold cluster enzymes.

[0014] The gold cluster enzyme described is Au 24 Cd1 and Au 24 Cu1 mixture (named AU).

[0015] The liposome described is synthesized from 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane, and cholesterol in a chloroform solution.

[0016] The preparation method steps of the liposome nanoplatform with inflammatory recognition ability of the present invention are as follows:

[0017] Prepare gold cluster enzyme: Adopt a systematic single-atom substitution method to prepare artificial enzymes, namely cluster enzymes, based on Au 25 clusters, prepare Au 24 Cd1 and Au24 Cu1;

[0018] Dissolve 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane, and cholesterol in chloroform to prepare a chloroform solution;

[0019] Synthesis of drug-loaded liposomes: Rotate and evaporate the chloroform solution obtained in step (2) to form a thin film, add the aqueous solution of gold clusters prepared in step (1), ultrasonically hydrate the thin film, and then extrude to obtain drug-loaded liposomes (named AU-LIP).

[0020] In step (1), the specific preparation method of the gold cluster enzyme is as follows:

[0021] Add tetrachloroauric acid trihydrate and 3-mercaptopropionic acid aqueous solution to water, stir at room temperature, then add sodium hydroxide aqueous solution to the reaction solution, and then add sodium borohydride alkaline solution. The whole reaction is carried out in the dark, and after stirring at room temperature, Au 25 MPA 18 is collected, and finally the reaction solution is aged to obtain the gold cluster enzyme.

[0022] As one of the preferred technical solutions: Add tetrachloroauric acid trihydrate (HAuCl4·3H2O, 20 mM, 0.25 mL) and 3-mercaptopropionic acid (MPA, 5 mM, 2 mL) aqueous solution to water (2.35 mL), stir at room temperature for 5 min, then add 1 M, 0.3 mL of sodium hydroxide (NaOH) aqueous solution to the reaction solution, and then add 0.1 mL of sodium borohydride (NaBH4) solution (prepared by dissolving 43 mg of NaBH4 powder in 10 mL of 0.2 M NaOH solution). The whole reaction is carried out in the dark, and after stirring at room temperature for 3 h, Au 25 MPA 18 is collected. Finally, the reaction solution is aged at 4 °C for 12 h. The synthesis of various metal-substituted AuxM 25 -xSG 18 is also based on the same method. The only difference is that the Au atoms in HAuCl4 (20 mM, 0.25 mL) are replaced by different metal nitrate ions (Cu 2+ , Cd 2+ ) at a molar ratio of 4% (Au:M = 24:1). To further purify the gold cluster enzyme, we use 3 K and 10 K ultrafiltration tubes respectively, and ultrafiltrate and centrifuge at 3500 rpm / min to remove smaller organic ligands and larger clusters, and then lyophilize.

[0023] In step (2), the 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine is 16:0-18:1 PC, the 1,2-distearoyl-3-trimethylammonium-propane is the chloride salt, and is 18:0 TAP.

[0024] In step (2), the mass ratio of 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane, and cholesterol is 30-60:10-30:20-60; as one of the preferred technical solutions: the mass ratio of 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane, and cholesterol is 44:16:40.

[0025] In step (3), ultrasonic treatment is carried out at an ultrasonic frequency of 20-60 kHz for 1-5 min. After overnight, extrusion is repeatedly carried out using an Avanti mini-extruder, and the mixture is passed through 400 nm and 200 nm filter membranes in sequence; as one of the preferred technical solutions: ultrasonic treatment is carried out at an ultrasonic frequency of 40 kHz for 2 min. After overnight, extrusion is repeatedly carried out using an Avanti mini-extruder, and the mixture is passed through 400 nm and 200 nm filter membranes in sequence.

[0026] As one of the preferred technical solutions: after the liposome nanoplatform with inflammation recognition ability of the present invention is loaded with gold cluster enzyme, the outer layer is wrapped with neutrophil cell membrane and red blood cell membrane. The steps of the preparation method are as follows:

[0027] Preparation of gold cluster enzyme: A systematic single-atom substitution method is adopted to prepare artificial enzymes, namely cluster enzymes, based on Au 25 clusters, and prepare Au 24 Cd1 and Au 24 Cu1;

[0028] Dissolve 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane and cholesterol in chloroform to prepare a chloroform solution;

[0029] (3) Synthesis of drug-loaded liposomes: The chloroform solution obtained in step (2) is rotary evaporated to form a thin film, the gold cluster aqueous solution prepared in step (1) is added, the thin film is ultrasonically hydrated, and then extrusion is carried out to obtain drug-loaded liposomes;

[0030] (4) Ultrasonically mix the drug-loaded liposomes with neutrophil cell membrane and red blood cell membrane and then extrude repeatedly to prepare a biomimetic nanoplatform (named AU-LIP-CM).

[0031] In step (4), the preparation method of the neutrophil cell membrane and the red blood cell membrane is as follows: After inducing LPS in vivo in mice, neutrophils are isolated from the mouse bone marrow by the percoll density gradient centrifugation method, and red blood cells are obtained from the peripheral blood; the neutrophil cell membrane and the red blood cell membrane are obtained.

[0032] The liposome nano-platform with inflammation recognition ability of the present invention can be used to prepare drugs for treating ulcerative colitis.

[0033] The gold cluster enzyme in the present invention has strong antioxidant and reactive oxygen species scavenging abilities, and its toxicity can be almost ignored, so it is selected as an anti-inflammatory agent.

[0034] The liposome in the present invention has strong drug loading capacity and a sustained release effect, can increase the endocytosis ability of cells to the material, and enhance the scavenging ability of intracellular reactive oxygen species.

[0035] The neutrophils in the present invention are isolated from the mouse bone marrow by the percoll density gradient centrifugation method after being induced in vivo with the same species of mice, and the expression of related adhesion molecules on the cell membrane surface is up-regulated.

[0036] The species sources of the peripheral blood and bone marrow cells in the present invention are mice or rats.

[0037] The ability of the neutrophil cell membrane in the present invention to actively penetrate the lesion site and target inflammation, and the advantages of the red blood cell membrane such as prolonging blood circulation time, being extremely easy to obtain, and having a large content in the body.

[0038] After coating the neutrophil cell membrane in the present invention, the final particle size is about 500 nm. Compared with cells at the micron scale, the specific surface area is larger, and the role of the membrane structure can be better exerted.

[0039] The present invention uses a mouse ulcerative colitis model induced by DSS.

[0040] The experiments at the in vitro cell level of the present invention show that this drug-loading system has strong anti-inflammatory and reactive oxygen species scavenging abilities, and good biocompatibility.

[0041] The present invention studied the biocompatibility of the bionic nano-platform at the in vivo level. Combining all the toxicology results, at a dose of 50 mg / kg, the bionic nano-platform does not cause weight, inflammation, immune response or pathological sensitivity in mice, laying a foundation for the subsequent in vivo treatment process.

[0042] The gold cluster enzyme in the present invention can be used for near-infrared II region fluorescence imaging, and can non-invasively, dynamically, real-timely and visually monitor the physiological and pathological conditions and treatment effects of mice.

[0043] The present invention describes the preparation scheme of the bionic nanoplatform, and provides the results of its in vivo targeting, extended blood circulation time and the evaluation of pharmacodynamics.

[0044] In vivo near-infrared second window imaging experiments on a mouse living model of the present invention have demonstrated that the drug can specifically target the inflamed intestine and extend the blood circulation time. In vivo pharmacodynamic evaluation monitoring shows that compared with free drugs and other commonly used drugs, the bionic nanoplatform can significantly inhibit the occurrence and development of ulcerative colitis and has an obvious anti-inflammatory effect.

[0045] The results of in vivo experiments of the present invention show that after treatment with AU-LIP-CM, excessive ROS are attenuated, the intestinal barrier is repaired, and the number of goblet cells, the level of the mucus layer and tight junction-related proteins are significantly increased. The levels of malondialdehyde (MDA), myeloperoxidase (MPO) and pro-inflammatory factors (TNF-α, IL-6, IL-1β and IFN-γ) are decreased, while the levels of anti-inflammatory factors (IL-10, TGF-β) are increased. In addition, the diversity, richness and evenness of the intestinal flora of colitis mice can be restored to a certain extent. The relative abundance of beneficial flora increases, while the harmful flora synchronously decreases, successfully improving the ecological structure of the intestinal flora.

[0046] In the mechanism study of the present invention, the results of ROS immunofluorescence staining indicate that AU-LIP-CM has antioxidant properties and good ROS scavenging ability.

[0047] The results of RNA sequencing in the mechanism study of the present invention show that the PPAR signaling pathway, chemokine signaling pathway, thyroid hormone signaling pathway, FoxO signaling pathway and Fc epsilon RI signaling pathway are highly related to the treatment mechanism of AU-LIP-CM.

[0048] The novel targeted bionic nanoplatform prepared by the present invention and its administration method is intravenous injection, which can improve the drug utilization rate, reduce the damage to other organs and reduce the impact of side effects.

[0049] Compared with the prior art, the preparation of the bionic nanoplatform with high inflammation recognition ability and its application in the treatment of ulcerative colitis described in the present invention have the following advantages:

[0050] Gold cluster enzymes have super antioxidant properties and reactive oxygen species scavenging ability, and have good anti-inflammatory effects; their size is about 2 nm, which can penetrate the renal barrier and be excreted through the kidneys, avoiding long-term hepatotoxicity and multi-organ damage;

[0051] Liposomes have good biocompatibility, strong drug loading capacity and sustained release effect, can increase the endocytosis ability of cells to the material, and enhance the scavenging ability of intracellular reactive oxygen species;

[0052] Gold cluster enzymes can emit light in the second near-infrared region and can be used for bioimaging, enabling non-invasive, dynamic, real-time, and visual monitoring of the physiological and pathological conditions and treatment effects of mice;

[0053] Activated neutrophil membrane surface-related adhesion molecules can efficiently target inflammatory sites;

[0054] By using the camouflage of the erythrocyte membrane, it can avoid phagocytosis by immune cells and prolong the blood circulation time;

[0055] The combination of liposomes with the two cell membranes endows this biomimetic nanoplatform with multiple potentials such as neutralizing bacterial toxins, targeting inflammatory sites, prolonging blood circulation time, improving drug loading capacity, enhancing the stability of nanoparticles, drug slow release, prolonging the in vitro storage time of materials, inhibiting material aggregation, and being applicable to fluorescence imaging in the second near-infrared region;

[0056] This biomimetic nanoplatform significantly inhibited the occurrence and development of colitis and also improved the gut microbiota by scavenging excessive reactive oxygen species at the inflammatory site and regulating signaling pathways.

[0057] The novel biomimetic nanoplatform can achieve a fixed-point and controllable treatment process and has high safety and biocompatibility. Due to the extremely low immunogenicity of the cell membrane on the surface, the absence of cell organelles and genetic materials, etc., it has relatively high safety and prospects for medical translational applications. Description of the Drawings

[0058] Figure 1 It is the optimization of the amount of gold cluster enzyme loaded in liposomes by the hydrodynamic size method.

[0059] Figure 2 It is the optimization of the amount of membrane-coated protein of AU-LIP.

[0060] Figure 3 Daily body weight change graphs of different treatment groups, continuously monitored for 9 days.

[0061] Figure 4 It is a disease activity index scoring graph for daily monitoring of body weight and feces of mice in different treatment groups.

[0062] Figure 5 It is a statistical graph of the colon length of mice in different treatment groups.

[0063] Figure 6 It is a representative graph of the differences in colon graphs of mice in different treatment groups.

[0064] Figure 7 It is a graph showing the effect of treatment with different materials on the colon intestinal morphology (HE staining).

[0065] Figure 8It is a diagram showing the effects of treatments with different materials on goblet cells in the colon (AB-PAS staining).

[0066] Figure 9 It is a diagram showing the effects of treatments with different materials on myeloperoxidase (MPO) in the colon. Embodiment

[0067] The following examples provide further detailed descriptions of the present invention.

[0068] Preparation of the thin film: Using the thin film hydration method, dissolve the three reactants of 16:0-18:1 PC, 18:0 TAP, and cholesterol in chloroform to prepare a stock solution with a concentration of 10 mg / mL. Mix them together in proportion and react quickly for 1 minute, then evaporate all the organic solvents on a rotary evaporator. Seal it with a sealing film and puncture some small holes with a needle, and place it in a vacuum desiccator overnight to remove the residual organic solvents.

[0069] Hydration of the thin film:

[0070] ① Add 1.5 mL of 1× PBS solution containing AU to the dry lipid film, and use water bath sonication combined with vortex oscillation to help the lipids suspend in the solution.

[0071] ② Once all the lipid substances are suspended in the solution, let the suspension stand overnight at 4 °C to effectively hydrate the lipid substances.

[0072] ③ According to the required particle size, sequentially extrude through polycarbonate membranes with pore sizes of 400 nm and 200 nm. The average particle size of the extruded liposomes is about 200 nm.

[0073] ④ Protect the liposome sample with nitrogen (or argon) to reduce lipid oxidation and store it at 4 °C.

[0074] To maximize the loading amount of gold cluster enzyme beneficial to the treatment effect in the liposomes, we induced the assembly of a series of concentrations of gold cluster enzyme with a certain amount of dry lipid film by hydration. The particle size distribution of AU-LIP was measured by dynamic light scattering method. Through the change of the hydrated particle size of AU-LIP, the saturated loading amount of gold cluster enzyme in the subsequent experiments was determined, as Figure 1 shown.

[0075] Add the obtained NC and RBC to the hypotonic solution respectively for lysis, and obtain the fused cell membrane through a series of operations such as sonication, centrifugation, and extrusion with a microextruder. Subsequently, the membrane of AU-LIP was coated by sonication and continuous extrusion to obtain AU-LIP-CM. To ensure that the prepared biomimetic nanoplatform can well inherit the advantages of the cell membrane without causing waste of the membrane, we optimized the optimal coating amount of the membrane and analyzed the data according to the Zeta potential. As Figure 2As shown, as the content of membrane protein increases, the zeta potential gradually becomes negative, thus determining the finally selected amount of membrane protein.

[0076] The experimental animals in this invention are SPF-grade male C57BL6N mice (6 weeks old), purchased from Beijing Vital River Laboratory Animal Technology Co., Ltd. The breeding temperature is 23±2 °C and the humidity is 50±5%. The mice are allowed to adapt to the new environment for one week.

[0077] The modeling drug in this invention is dextran sulfate sodium, with a molecular weight of 36,000 - 50,000, purchased from MP Company.

[0078] Establishment of the mouse ulcerative colitis model: The experimental group of mice freely drank 3% DSS solution for 6 consecutive days to induce experimental ulcerative colitis in mice. The blank control group was given the same volume of mineral water.

[0079] On the second and fourth days after modeling, the mice in the drug administration group were respectively injected with PBS, 5-aminosalicylic acid (5-ASA), AU, AU-LIP, LIP-CM or AU-LIP-CM via the tail vein. The mice in the normal group were intravenously injected with 1*PBS as a control. The body weight, survival rate and disease activity index (DAI) of the mice were monitored every day. On the 9th day, after collecting feces, all the mice were sacrificed and the colon was taken out, the length was measured and photographed. Subsequently, a part of the colon at the same position near the anal end was intercepted and fixed in 4% paraformaldehyde solution, and the remaining colon was washed and collected and placed in a -80 °C refrigerator for later use. It was used for colon histopathological detection, histopathological scoring, goblet cell staining, myeloperoxidase (MPO) activity in colon tissue, and detection of related inflammatory factors, etc.

[0080] After modeling, the body weight of the mice decreased significantly ( Figure 3 ), the DAI score increased ( Figure 4 ), and the colon was significantly shortened and edematous ( Figure 5 , Figure 6 ). The colon sections of the mice were stained with H&E ( Figure 7 ) and AB-PAS ( Figure 8 ). The analysis results showed that obvious inflammatory reactions occurred in the colon of the modeling group mice, such as obvious damage marks, damage to the colon epithelium, severe damage to the colon mucosa, rupture of the crypt structure, and reduction in the abundance of goblet cells, etc. In addition, the levels of pro-inflammatory factors (TNF-α, IL-6, IL-1β, IFN-γ) in the colon tissue of the mice were significantly increased, and the levels of anti-inflammatory factors (IL-10, TGF-β) were significantly decreased.

[0081] After treatment with different materials, the results showed that the treatment group of AU-LIP had significantly improved treatment effects compared with other treatment groups (AU, LIP-CM, 5-ASA). For example, it inhibited the decrease in body weight, colonic edema and shortening, and reduced the DAI score. The results of HE staining, AB-PAS staining, and MPO staining also showed that AU-LIP inhibited UC inflammation to a certain extent. In addition, the treatment of AU-LIP-CM significantly inhibited the decrease in body weight, colonic edema and shortening, and the DAI score also decreased significantly. HE staining and AB-PAS staining showed that the inflammatory response of mice treated with AU-LIP-CM was significantly reduced, promoting the regular and compact arrangement of crypt structures, maintaining the abundance of goblet cells, and significantly alleviating the intestinal morphological damage of DSS-induced UC mice, restoring them to a normal state. This indicates that AU-LIP-CM plays a key role in the recovery of colonic epithelium, highlighting the importance of the fused NC / RBC membrane coating in improving the inflammatory targeting ability. After scoring the degree of colonic injury through histopathological analysis, it was clearly found that the treatment effect of AU-LIP-CM was the best. MPO catalyzes the production of ROS and causes colonic mucosal damage, which has been identified as an important biomarker for disease assessment in IBD patients. The MPO activity in the colon of mice in the model group increased significantly, and the exacerbation degree of MPO in the colon of mice treated with AU-LIP-CM decreased significantly ( Figure 9 ), indicating that the treatment of AU-LIP-CM inhibited neutrophil infiltration at the inflammatory site and successfully inhibited the further exacerbation of the UC inflammatory state. The secretion of inflammatory cytokines TNF-α, IL-1β, and IL-6 was also inhibited, while the anti-inflammatory factors IL-10 and TGF-β increased. This drug-loading system significantly improved the pathological symptoms of DSS-induced ulcerative colitis in mice, reduced the damage of mouse colonic tissue, and inhibited the infiltration of inflammatory cells and the secretion of inflammatory factors, significantly improving the symptoms of UC. Generally speaking, this biomimetic nanoplatform showed good treatment effects in the ulcerative colitis model.

[0082] Although the embodiments of the present invention have been shown and described, it will be understood by those of ordinary skill in the art that various changes, modifications, substitutions, and variations can be made to these embodiments without departing from the principles and spirit of the present invention. The scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A liposome nanoplatform with the ability to recognize inflammation, characterized in that: The described liposome nanoplatform has a liposome as the core. After loading the gold cluster enzyme, the outer layer is wrapped with neutrophil cell membranes and red blood cell membranes; the described gold cluster enzyme is Au 24 Cd1 and Au 24 Cu1 mixture; the liposome is synthesized from 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-stearoyl-3-trimethylammonium-propane, and cholesterol in a chloroform solution.

2. A method for preparing the liposome nanoplatform as described in claim 1, characterized in that: The steps of the preparation method are as follows: (1) Preparation of gold cluster enzymes: An artificial enzyme, i.e., a cluster enzyme, was prepared based on Au 25 clusters by using a systematic single-atom substitution method, and Au 24 Cd1 and Au 24 Cu1 were prepared; (2) Dissolve 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane, and cholesterol in chloroform to prepare a chloroform solution; (3) Synthesis of drug-loaded liposomes: Rotate and evaporate the chloroform solution obtained in step (2) to form a thin film, add the aqueous solution of gold cluster enzyme prepared in step (1), ultrasonically hydrate the thin film, and then extrude to obtain drug-loaded liposomes; (4) Ultrasonically mix the drug-loaded liposomes with neutrophil cell membranes and red blood cell membranes and extrude repeatedly to obtain a biomimetic nanoplatform.

3. The preparation method according to claim 2, characterized in that: In step (2), the mass ratio of 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine, 1,2-distearoyl-3-trimethylammonium-propane, and cholesterol is 30-60:10-30:20-60.

4. The preparation method according to claim 2, wherein: In step (3), ultrasonicate at an ultrasonic frequency of 20-60 kHz for 1-5 min, and after overnight, repeatedly extrude using an Avanti mini-extruder and pass through 400 nm and 200 nm filters in sequence.

5. Use of the liposome nanoplatform according to claim 1 for the preparation of a drug for treating ulcerative colitis.

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