Composite nano-particles for targeting mitochondria to prevent and treat liver injury as well as preparation method and application of composite nano-particles

Abstract

The invention relates to a composite nanoparticle VZTM-Gal and a preparation method thereof.According to the VZTM-Gal, efficient loading of a VDAC1 inhibitor VBIT-4 is achieved on the basis of a ZIF-8 nano system, then tannic acid is introduced to be coordinated with exogenous Mg < 2 + > and Zn < 2 + > on the surface of ZIF-8, a metal-polyphenol layer on the surface of ZIF-8 is constructed, and the composite nanoparticle is rapidly disintegrated and released in a lysosome environment; the surface-coupled GalNAc ligand can significantly enhance the specific recognition and uptake of the drug to parenchymal hepatic cells; the preparation process is mild in conditions and simple and convenient in steps, and the obtained nanoparticles are uniform in particle size; the invention further relates to application of the VZTM-Gal in treatment of cadmium-induced acute liver injury and non-alcoholic steatohepatitis induced by combined choline deficiency L-amino acid limited high fat diet exposure, experiments prove that VBIT-4 released by the VZTM-Gal can inhibit VDAC1 oligomerization and inhibit mitochondrial excessive autophagy, tannic acid can remove active oxygen, and the VZTM-Gal can be used for treating cadmium-induced acute liver injury and non-alcoholic steatohepatitis induced by choline deficiency L-amino acid limited high fat diet exposure. Mg < 2 + > synergistically reduces calcium overload caused by cadmium, recovers mitochondrial morphology and functions, and improves liver functions.

Description

Technical Field

[0001] This invention belongs to the field of nanobiomedicine technology, and in particular relates to a composite nanoparticle that targets mitochondria to prevent liver damage, its preparation method, and its application. Background Technology

[0002] Cadmium (Cd) is a heavy metal pollutant widely present in the environment. It can accumulate in the body through the food chain, mainly in organs such as the liver and kidneys, causing pathological changes such as oxidative stress, calcium homeostasis imbalance, and mitochondrial dysfunction, leading to acute or chronic liver damage. Recent studies have found that cadmium exposure not only directly induces hepatocyte necrosis but also promotes the occurrence and progression of non-alcoholic steatohepatitis (NASH), posing a serious threat to public health.

[0003] Current treatments for cadmium-induced liver injury primarily focus on chelating agents for detoxification and antioxidant interventions, such as dimercaprol (BAL) and ethylenediaminetetraacetic acid (EDTA). However, traditional chelating agents suffer from drawbacks including poor selectivity, weak targeting, significant side effects, and difficulty in penetrating cells and mitochondria, making it difficult to effectively block the core cellular damage pathways caused by cadmium toxicity. Furthermore, most existing treatment strategies target acute toxic reactions, lacking effective interventions for metabolic disorders induced by cadmium exposure (such as NASH).

[0004] At the pathological mechanism level, recent studies have shown that mitochondrial voltage-dependent anion channel protein 1 (VDAC1) plays a crucial role in cadmium toxicity. Oligomerization and ubiquitination of VDAC1 can promote excessive mitophagy, leading to energy metabolism disorders and apoptosis. However, effective drugs that can specifically regulate VDAC1 function and restore mitochondrial homeostasis are currently lacking. Furthermore, cadmium exposure can also lead to intracellular and mitochondrial calcium overload, triggering oxidative stress and the accumulation of mitochondrial reactive oxygen species (mtROS), further exacerbating liver damage and inflammatory responses.

[0005] In recent years, metal-organic frameworks (MOFs), especially ZIF-8 nanocarriers, have been widely studied for drug delivery due to their tunable pore structure, biodegradability, and pH-responsive properties. However, traditional ZIF-8 systems generally suffer from insufficient targeting and limited biological functions of degradation products, and cannot yet achieve synergistic regulation of multi-mechanism liver injury.

[0006] Therefore, there is an urgent need for a nanomedicine system that is liver-targeting, controllable in release, and capable of multi-mechanism synergistic intervention, to block key pathways of cadmium toxicity at the source, restore mitochondrial function and ion homeostasis, and thus achieve dual prevention and treatment of acute liver injury and NASH. Summary of the Invention

[0007] To address the shortcomings of existing technologies, this invention proposes a composite nanoparticle for targeting mitochondria to prevent liver damage, along with its preparation method and applications.

[0008] The technical solution of this invention is as follows: A method for preparing composite nanoparticles includes the following steps: S1. After ultrasonically mixing ZIF-8 dispersion and VBIT-4 dispersion, the mixture was stirred in an ice bath and then washed by centrifugation to obtain ZIF-8 nanoparticles VZ loaded with VBIT-4. S2. The VZ nanoparticles obtained in step S1 are mixed with tannic acid and MgCl2, reacted, and then centrifuged and washed to obtain VZTM; S3. After activating GalNAC with EDC / NHS, it is mixed with VZTM prepared in step S2, reacted in the dark, washed by centrifugation, and then freeze-dried to obtain liver-targeting composite nanoparticles VZTM-Gal.

[0009] Furthermore, the drug loading rate of VBIT-4 loaded in VZ in step S1 is 4%-6%.

[0010] Furthermore, the technical parameters for ultrasonic mixing in step S1 are: ultrasonication at 60W for 10-20 minutes; the technical parameters for ice bath stirring in step S1 are: stirring under ice bath conditions for 10-20 hours.

[0011] Further, in step S2, the mass ratio of VZ nanoparticles: tannic acid: MgCl2 is (8-12):(1-2):(0.2-0.3); the technical parameters for the reaction of VZ nanoparticles with tannic acid and MgCl2 in step S2 are: reaction at 25±1℃ for 1-3 hours.

[0012] Furthermore, the technical parameters for EDC / NHS activation of GalNAC in step S3 are as follows: reaction in MES buffer at pH 6.0 for 30-60 min.

[0013] Further, the mass ratio of GalNAC:VZTM in step S3 is (0.8-1.2):(8-12); Furthermore, the reaction medium for GalNAC and VZTM in step S3 is a PBS buffer at pH 7.4.

[0014] The composite nanoparticles were prepared according to the preparation method described above.

[0015] Application of the composite nanoparticles in the preparation of drugs for treating liver damage caused by cadmium poisoning.

[0016] Furthermore, the liver injury is cadmium-induced acute liver injury or non-alcoholic steatohepatitis induced by combined choline deficiency and L-amino acid-restricted high-fat diet exposure.

[0017] Compared with the prior art, the present invention has at least the following advantages: 1. This invention relates to the composite nanoparticle VZTM-Gal and its preparation method. The VZTM-Gal is based on the ZIF-8 nanosystem to achieve efficient loading of the VDAC1 inhibitor VBIT-4, and then tannic acid and exogenous Mg are introduced. 2+ and Zn on ZIF-8 surface 2+ Coordination was used to construct a metal-polyphenol layer on the surface of ZIF-8, enabling the composite nanoparticles to rapidly disintegrate and release drugs in an acidic lysosomal environment (pH 4.5-5.5). In addition, the surface-coupled GalNAc ligand significantly enhanced the specific recognition and uptake of drugs by hepatocytes. The preparation process described in this invention is mild and simple, and the resulting nanoparticles have uniform particle size and good drug development potential.

[0018] 2. This invention also relates to the application of the composite nanoparticles VZTM-Gal in the preparation of drugs for treating cadmium-induced acute liver injury or non-alcoholic steatohepatitis induced by L-amino acid-limited high-fat diet exposure combined with choline deficiency, which differs from traditional cadmium poisoning treatments that are limited to metal chelation. Experiments have demonstrated that the VBIT-4 released by the composite nanoparticles VZTM-Gal prepared in this invention inhibits VDAC1 oligomerization, suppresses excessive mitochondrial autophagy, and the tannic acid layer can scavenge reactive oxygen species. Furthermore, the Mg released by the composite nanoparticles... 2+ It can reduce calcium overload caused by cadmium and restore mitochondrial morphology and function. In terms of efficacy, in vivo experiments have confirmed that the drug can significantly reduce transaminase (ALT / AST) levels and improve liver function. In particular, for NASH induced by cadmium combined with choline deficiency and L-amino acid-restricted high-fat diet exposure, it can significantly reduce intrahepatic lipid deposition, downregulate the expression of inflammatory factors (TNF-α, IL-1β) and fibrotic proteins, and reverse metabolic lesions. Attached Figure Description

[0019] To more clearly illustrate the specific embodiments of the present invention, the accompanying drawings used in the description of the specific embodiments or the prior art will be briefly introduced below.

[0020] Figure 1 This is a schematic diagram of the preparation method of VZTM-Gal nanoparticles of the present invention; Figure 2 This is a TEM image of VZTM-Gal nanoparticles from Example 2; Figure 3 The hydrated particle size of VZTM-Gal nanoparticles in Example 2; Figure 4 The zeta potential of VZTM-Gal nanoparticles in Example 2; Figure 5 These are TEM images of VZTM-Gal nanoparticles under different pH conditions in Example 3; Figure 6 This is a liver-targeted in vivo imaging image of VZTM-Gal nanoparticles in Example 4; Figure 7 Example 5: Western blot analysis of the expression and quantification of autophagy-related proteins p62 and LC3; Figure 8 Example 5: Quantitative fluorescence analysis of Rhod-2AM; Figure 9 Example 5: MitoSOX staining results and quantitative analysis; Figure 10 Example 5: TEM detection and quantitative analysis of mitochondrial morphology in liver tissue; Figure 11 Example 5: Western blot detection and quantitative analysis of VDAC1 oligomerization; Figure 12 The levels of AST and ALT in the serum of a mouse model of acute cadmium exposure in Example 6; Figure 13 This is an HE staining image of liver tissue from Example 6; Figure 14 The levels of AST, ALT, and HDL-C in serum in Example 7; Figure 15 Example 7: HE staining and quantitative analysis of liver tissue; Figure 16 Example 7: Oil Red O staining and quantitative analysis of liver tissue. Detailed Implementation

[0021] The present invention will now be described in further detail. It should be noted that the following specific embodiments are only used to further illustrate the present invention and should not be construed as limiting the scope of protection of the present invention. All other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0022] This invention provides a general and / or specific description of the materials and experimental methods used in the experiments. Unless otherwise specified, all experimental or testing methods are conventional methods; all reagents and instruments used, unless otherwise specified, are commercially available conventional products prepared or used using conventional methods, and unless otherwise specified, the raw materials used in parallel experiments are from the same batch.

[0023] Source of materials for this invention Zinc nitrate hexahydrate (Zn(NO3)2·6H2O), 2-methylimidazole (2-mIM), tannic acid (TA), N-acetylgalactosamine (GalNAC), and VBIT-4 (VDAC1-specific oligomerization inhibitor) were all purchased from Sigma-Aldrich.

[0024] Anhydrous methanol, DMSO, 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), and N-hydroxysuccinimide (NHS) were all of analytical grade.

[0025] Experimental animals: C57BL / 6 male mice, SPF grade, 6-8 weeks old.

[0026] Hepatocellular carcinoma cell line: HepG2 was purchased from the Chinese Academy of Sciences Cell Bank.

[0027] Example 1: Preparation of the nanomedicine VZTM-Gal This embodiment provides a method for preparing ZIF-8 nanomedicine loaded with a VDAC1 inhibitor and modified with a liver-targeting ligand. A schematic diagram of the preparation scheme is shown below. Figure 1 As shown.

[0028] Synthesis of S1. ZIF-8 nanoframework: 0.295 g Zn(NO3)2·6H2O was accurately weighed and dissolved in 20 mL of anhydrous methanol to prepare solution A; 0.656 g 2-mIM was dissolved in 20 mL of anhydrous methanol to prepare solution B; solutions A and B were rapidly mixed under magnetic stirring at room temperature and reacted for 2 h. The mixture became turbid and a white precipitate was formed. After the reaction was completed, the suspension was centrifuged at 10,000 rpm for 10 min, the precipitate was collected, and washed three times with anhydrous methanol to remove unreacted raw materials. After vacuum drying at 60 °C, ZIF-8 nanoparticles were obtained. S2. VBIT-4 loading (VZ): 10 mg of ZIF-8 nanoparticles prepared in step S1 were weighed and dispersed in 10 mL of methanol to obtain a ZIF-8 dispersion; 1 mg of VBIT-4 was dissolved in 1 mL of DMSO and added dropwise to the ZIF-8 dispersion. The mixture was sonicated at 60 W for 10 min and then placed in an ice bath and stirred for 12 h to allow the drug to fully enter the ZIF-8 pores; the precipitate was collected by centrifugation (10,000 rpm, 10 min) and washed with methanol to remove the free drug adsorbed on the surface, obtaining ZIF-8 nanoparticles loaded with VBIT-4, named VBIT-4@ZIF-8, abbreviated as VZ. The drug loading rate of VBIT-4 loaded in this example was found to be 5%. S3. Modification with Tannic Acid and Functional Ions (VZTM): 10 mg of VZ nanoparticles were redispersed in 10 mL of deionized water. 100 μL of tannic acid (TA) solution (10 mg / mL) was added to the dispersion, and the mixture was stirred for 5 min. Then, 20 μL of MgCl2 solution (10 mg / mL) was added, and the mixture was reacted at room temperature for 1 h. The phenolic hydroxyl groups of TA react with Zn... 2+ and exogenous Mg 2+ Through coordination, a stable metal-TA network layer is formed on the surface of ZIF-8; finally, centrifugation and washing are performed to obtain TA-coated nanoparticles, which are named VZTM. S4. Conjugation of liver-targeting ligand GalNAC (VZTM-Gal): Using the EDC / NHS activation method, 1 mg GalNAC containing amino groups, 11.5 mg EDC, and 6.9 mg NHS were dissolved in 10 ml of MES buffer (pH 6.0) and activated for 30 min. Then, the activated GalNAC solution was added to 10 mg VZTM in PBS dispersion (pH 7.4), and the reaction was carried out at room temperature in the dark with stirring for 4 h. After centrifugation, the product was washed three times with deionized water and freeze-dried to obtain the final product VZTM-Gal.

[0029] Example 2 Material Characterization Morphology and particle size: Observations by transmission electron microscopy (TEM) are as follows Figure 2 As shown, VZTM-Gal exhibits a uniform, regular polyhedral structure; dynamic light scattering (DLS) results are as follows. Figure 3 As shown, its average hydrated particle size was measured to be 321.4 nm, indicating good dispersibility.

[0030] Potential analysis: such as Figure 4 As shown, the Zeta potential of VZTM-Gal is -27.5 mV.

[0031] Example 3: pH-responsive degradation and drug release performance test To verify the responsive release capability of the nanomedicine VZTM-Gal in the tumor microenvironment or lysosomal environment, release experiments were conducted under simulated pH conditions.

[0032] Experimental method: 2 mg of VZTM-Gal was placed in 10 mL of PBS buffer at pH 7.4 (simulating physiological blood environment), pH 6.8 (simulating tumor tissue extracellular environment), and pH 4.5 (simulating lysosomal acidic environment) and incubated. A simulated release experiment was carried out in a 37℃ constant temperature water bath shaker, and the drug concentration was measured after 48 h.

[0033] Experimental results: In the pH 7.4 group, drug release was slow within 48 hours, with a cumulative release rate of only 43.9%, indicating that the system is relatively stable at physiological pH and can effectively prevent premature drug leakage in the bloodstream; in the pH 4.5 group, drug release was significantly accelerated, with a cumulative release rate of 95.9% within 12 hours; and Figure 5 TEM results show that the VZTM-Gal particle structure undergoes significant collapse and disintegration under pH 4.5 conditions, realizing the synthesis of VBIT-4 drug and Zn. 2+ / Mg 2+ Rapid release of ions.

[0034] Example 4: Verification of Liver-Targeting Characteristics This embodiment uses mice to verify the targeted uptake of nanomedicines.

[0035] Liver targeting evaluation: VZTM (non-targeted group) and VZTM-Gal (targeted group) nanomaterials were labeled with Cy5 fluorescent dye to prepare Cy5-VZTM and Cy5-VZTM-Gal. These were injected into mice via the tail vein at a dose of 10 mg / kg. In vivo imaging results of the small animals are shown below. Figure 6 As shown in the figure, the Cy5-VZTM-Gal fluorescence signal rapidly increased within 2 hours and could still be detected within 24 hours, indicating that the targeted particles prepared in this invention have the characteristic of prolonged retention in liver tissue.

[0036] Example 5: In vitro cell anti-cadmium toxicity and mechanism verification This embodiment uses the human hepatocellular carcinoma cell line HepG2 to verify the molecular regulatory mechanism of nanomedicine.

[0037] Excessive autophagy inhibition: Cellular experiments were conducted. A cadmium damage model was established in HepG2 cells by inducing cadmium damage with CdCl2 (20 μM), followed by intervention with VZTM-Gal. Western blot results are shown below. Figure 7 As shown in the figure, the LC3Ⅱ / LC3Ⅰ ratio of the Cd-treated group was significantly increased (9.689±1.582), and p62 decreased (0.309±0.052). After VZTM-Gal treatment, the LC3Ⅱ / LC3Ⅰ ratio decreased to 1.532±0.405, and p62 increased to 1.023±0.085, indicating that excessive autophagy was significantly inhibited.

[0038] Calcium homeostasis recovery: [Ca] was determined using Rhod-2AM (DoJinDo, R002). 2+To determine the concentration, prepare 2 μM Rhod-2AM working solution using HBSS. Remove the culture medium from the 24-well plate, wash three times with HBSS (Beyotime, C0218), and incubate at 37°C for 30 min. After incubation, wash three times with HBSS, then incubate for another 10 min with HBSS. Measure the fluorescence value at 557 nm / 581 nm (Ex / Em) using a microplate reader. Figure 8 As shown, compared with the Cd-treated group, the fluorescence intensity of mitochondrial calcium Fluo-2AM was also significantly reduced after VZTM-Gal treatment (0.719±0.028 vs. 0.956±0.018), indicating that VZTM-Gal inhibited calcium overload caused by Cd exposure and restored calcium homeostasis.

[0039] Antioxidant assay: After removing the cell culture medium, wash once with PBS, add 100 nm MitoTracker Red CM-H2Xros (Thermo Fisher Scientific, M7513) working solution, and incubate at 37°C for 30 min. After washing, add fresh cell culture medium and observe the red fluorescence using a fluorescence microscope. The production of mitochondrial ROS is positively correlated with the intensity of the red fluorescence. Immunofluorescence results are as follows. Figure 9 As shown in the figure, VZTM-Gal treatment reduced the increase in mitochondrial ROS caused by Cd exposure, indicating that VZTM-Gal has antioxidant capacity.

[0040] Mitochondrial morphology restoration: Collected cells or liver tissue were fixed in 2.5% glutaraldehyde at 4°C, followed by post-fixation, infiltration embedding, polymerization, sectioning (ultrathin sections of 60-80 nm), staining, and then observation and image acquisition under a transmission electron microscope for further analysis. The transmission electron microscopy results are shown below. Figure 10 As shown, Cd exposure resulted in extensive mitochondrial damage in liver tissue, including cristae disruption, swelling, rounding, and mitophagosome aggregation. Quantitative assessment revealed a significantly increased proportion of damaged mitochondria and a relative decrease in mitochondrial area. Conversely, VZTM-Gal treatment significantly reduced mitochondrial damage and increased mitochondrial area, indicating a potent protection of mitochondrial morphology.

[0041] Inhibition of VDAC1 oligomerization: HA-VDAC1-overexpressing cells were washed with pre-chilled PBS, then collected into EP tubes with pre-chilled PBS and centrifuged to collect the pellet. Cells were resuspended in PBS containing 300 μM [ethylene glycolbis(succinimidylsuccinate)] (EGS, Thermo Fisher Scientific, 21565). The cells were incubated at 30°C for 20 min at 100 rpm on a shaker to allow for VDAC1 cross-linking. Cells were lysed with lysis buffer, and protein concentration was measured after centrifugation. The cells were then heated at 56°C for 10 min with denaturing non-reducing loading buffer. Subsequently, VDAC1 oligomerization was immunoblotted using HA antibody. Western blotting results are shown below. Figure 11 As shown, Cd exposure can induce VDAC1 oligomerization, while VZTM-Gal can alleviate VDAC1 oligomerization induced by Cd exposure. The expression level of VDAC1 dimer / monomers decreased to about 46% of that in the Cd group, indicating that VZTM-Gal can inhibit VDAC1 oligomerization.

[0042] Example 6: In vivo pharmacodynamic evaluation of the therapeutic effect of VZTM-Gal on cadmium-induced acute liver injury. VZTM-Gal alleviated cadmium-induced acute liver injury: Seven-week-old male C57BL / 6 mice were exposed to saline or 2.5 mg / kg CdCl2 via intraperitoneal injection once daily for seven consecutive days. Treatment drugs (VBIT-4, liver-targeting nanomaterials ZTM-Gal, VZTM-Gal) were administered intraperitoneally every other day. Seven days after modeling, mice were euthanized with sodium pentobarbital, and whole blood and liver tissue were collected. Histological and biochemical analyses of serum and liver tissue were performed to assess liver function and inflammatory cell infiltration. Serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) are two important liver enzymes; serum levels are used to assess liver function. Figure 12 As shown, compared with the Cd group, the VZTM-Gal nanomaterial group showed significantly lower AST (94.36 U / L ± 5.833 U / L vs. 160.9 U / L ± 10.23 U / L) and ALT (27.38 U / L ± 2.413 U / L vs. 57.06 U / L ± 3.637 U / L), indicating recovery of liver function. Similarly, the HE staining results are as follows... Figure 13 The results also showed that VZTM-Gal intervention significantly reduced inflammatory infiltration caused by Cd exposure. These results indicate that VZTM-Gal can alleviate Cd-induced acute liver injury.

[0043] Example 7: In vivo pharmacodynamic evaluation of the therapeutic effect of VZTM-Gal on cadmium- and high-fat diet-induced NASH. VZTM-Gal alleviated NASH induced by cadmium combined with a high-fat diet: (1) Model establishment and drug administration regimen: The choline-deficient, L-amino acid-defined, high-fat diet (CDA-HFD) induces strong histological and biochemical indicators of fibrotic NASH in a relatively short period of time, and is currently recognized as a mouse model that can simulate the pathological progression of NASLD. Seven-week-old male C57BL / 6 mice were fed the CDA-HFD diet for 16 weeks to induce NASH. At the same time, they were fed normal drinking water or drinking water with 10 mg / L CdCl2 to establish a mouse model of combined exposure to Cd and NASH. They were treated with intraperitoneal injection of VBIT-4, liver-targeting nanomaterials ZTM-Gal, and VZTM-Gal three times a week. After 16 weeks, the mice were euthanized with sodium pentobarbital, and blood was collected from the orbital cavity. ALT and AST were measured in the serum to assess liver function. Liver tissue was collected, and HE staining and Oil Red O staining were performed on the liver to evaluate the infiltration of inflammatory cells, steatosis, and degree of fibrosis. The therapeutic effects of different drugs were observed, and the results are as follows.

[0044] (2) Serological and biochemical index analysis: Serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and high-density lipoprotein cholesterol (HDL-C) expression levels were detected. The results are as follows: Figure 14 As shown, compared with the combined exposure group, VZTM-Gal treatment significantly reduced AST and ALT, indicating its restorative effect on liver function. VZTM-Gal treatment significantly increased HDL-C, confirming its potential to treat NASH promoted by Cd combined exposure.

[0045] (3) Pathological examination: HE staining results are shown in the figure. Figure 15 HE staining results showed that VZTM-Gal treatment significantly reduced the degree of hepatic steatosis, ballooning degeneration, and inflammatory infiltration, and analysis indicated a reduction in the severity of NASH. To assess hepatic steatosis, we performed Oil Red O staining on liver sections, as shown... Figure 16 As shown, the Cd+CDA-HFD group exhibited significant lipid deposition in the liver, but VZTM-Gal treatment significantly reduced lipid accumulation in fatty liver tissue. These results suggest that VZTM-Gal has the potential to mitigate NASH induced by combined cadmium exposure.

[0046] The above results indicate that VZTM-Gal can effectively inhibit cadmium-promoted excessive mitophagy and inflammatory-fibrotic response, and improve liver metabolic homeostasis.

[0047] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention, and they should all be covered within the scope of the claims and specification of the present invention.

Claims

1. A method for preparing a composite nanoparticle, characterized by, Comprising the following steps: S1. After ultrasonic mixing of ZIF-8 dispersion liquid and VBIT-4 dispersion liquid, ice bath stirring, and centrifugal washing, VZ of VBIT-4 loaded ZIF-8 nanoparticles is obtained; S2. After mixing reaction, centrifugal washing of VZ nanoparticles prepared in step S1 with tannic acid and MgCl2, VZTM is obtained; S3. After mixing of EDC / NHS activated GalNAC with VZTM prepared in step S2, light-avoiding reaction, centrifugal washing, and freeze-drying, VZTM-Gal of liver targeting composite nanoparticles is obtained.

2. The production method according to claim 1, characterized by, The drug loading rate of VZ of VBIT-4 loaded in step S1 is 4%-6%.

3. The production method according to claim 1, characterized by, The technical parameters of ultrasonic mixing in step S1 are: ultrasonic mixing for 10-20 min under 60 W; and the technical parameters of ice bath stirring in step S1 are: stirring for 10-20 h under ice bath condition.

4. The method of claim 1, wherein, The mass ratio of VZ nanoparticles: tannic acid: MgCl2 in step S2 is (8-12):(1-2):(0.2-0.3); and the technical parameters of reaction of VZ nanoparticles with tannic acid and MgCl2 in step S2 are: reaction for 1-3 h under 25±1℃.

5. The preparation method according to claim 1, characterized in that, The technical parameters of EDC / NHS activated GalNAC in step S3 are: reaction for 30-60 min in MES buffer solution with pH=6.

0.

6. The method of claim 1, wherein, The mass ratio of GalNAC: VZTM in step S3 is (0.8-1.2):(8-12).

7. The preparation method according to claim 1, characterized in that, The reaction medium of GalNAC and VZTM in step S3 is PBS buffer solution with pH 7.

4.

8. The composite nanoparticles prepared by the preparation method according to any one of claims 1-7.

9. The use of the composite nanoparticles according to claim 8 in the preparation of a drug for treating liver damage induced by cadmium poisoning.

10. Use according to claim 9, characterized in that, The liver damage is acute liver damage induced by cadmium or non-alcoholic steatohepatitis induced by combined choline deficiency L-amino acid defined high-fat diet exposure.

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