A polyoxovanadomolybdate nanocluster, a preparation method and application thereof

Abstract

The present application relates to a kind of polyoxovanadomolybdate nanoclusters and its preparation method and application.The chemical general formula of the polyoxovanadomolybdate nanocluster is Na6[V 10‑x Mo x O 28 ]·nH2O, wherein 1≤x≤4, n is the number of crystallization water, the value range is 8~12.The polyoxovanadomolybdate nanocluster provided by the present application is by Mo substitution part V atom in dodecanovanadate lattice, into lattice in substitutional doping form, on the basis of retaining parent body POV insulin-like activity, give treatment agent excellent antioxidant function, and through the combined action of antioxidant and insulin-like activity, realize the multiple regulation of diabetic wound microenvironment, promote wound healing.

Description

Technical Field

[0001] This invention belongs to the field of biomaterials science and relates to a polyoxovanadium molybdate nanocluster, its preparation method, and its application. Background Technology

[0002] Chronic diabetic wounds are a serious complication of diabetes, with a high incidence and difficult treatment, placing a heavy burden on patients and the healthcare system. Statistics show that approximately 15%-25% of diabetic patients develop foot ulcers, with about 20% ultimately requiring amputation. The pathological mechanisms of delayed wound healing in diabetes are extremely complex, involving the synergistic effects of multiple factors, including persistent hyperglycemia, oxidative stress, and dysregulation of the inflammatory response. First, persistent hyperglycemia leads to disordered energy metabolism in wound-resident cells (such as fibroblasts and endothelial cells), impaired glucose uptake, and reduced cellular ATP production, which cannot support normal proliferation and migration. Second, the high-glucose environment induces excessive production of reactive oxygen species (ROS) through pathways such as the polyol pathway and mitochondrial dysfunction. Excessive ROS not only directly oxidizes and damages cellular components but also activates inflammatory pathways such as NF-κB, triggering a cascade release of pro-inflammatory factors (such as IL-6 and TNF-α), forming a vicious cycle of oxidative stress and inflammation. Furthermore, diabetic wounds are often accompanied by bacterial colonization and biofilm formation, further exacerbating the inflammatory response and tissue damage. The above factors together disrupt the inflammatory, proliferative, and remodeling phases of wound healing, ultimately leading to refractory wounds.

[0003] Current clinical management strategies for chronic diabetic wounds mainly include local debridement, negative pressure wound therapy, infection control, and growth factor therapy. However, most of these interventions alleviate local symptoms or single pathological processes, failing to address the underlying pathological mechanism leading to impaired healing—namely, cellular dysfunction caused by glucose metabolism disorders. For example, debridement and negative pressure therapy can only improve the physical environment of the wound and cannot reverse cellular energy metabolism disorders; while topical application of growth factors can temporarily promote cell proliferation, the efficacy is often limited due to insulin resistance in the cells themselves. Therefore, existing therapies only provide palliative effects, and there is an urgent need to develop novel treatment strategies that can simultaneously target hyperglycemia, oxidative stress, and inflammation.

[0004] Insulin, as the gold standard drug for lowering blood sugar, has also attracted attention for its application in wound treatment. Local insulin can promote glucose uptake by cells and improve energy metabolism by activating insulin receptors and their downstream signaling pathways. However, the direct application of insulin to wound treatment has several limitations: First, insulin has poor chemical stability and is easily degraded and inactivated at room temperature; second, peripheral tissues in diabetic states generally exhibit insulin resistance, and cellular responsiveness to insulin is significantly reduced; third, insulin itself lacks antioxidant capacity and cannot clear excessive ROS accumulated in the wound microenvironment. These limitations make insulin difficult to use as an ideal drug for treating diabetic wounds.

[0005] Vanadium (V)-based compounds have attracted widespread attention in recent years as insulin mimics. Vanadium compounds can mimic insulin signaling by inhibiting the activity of protein tyrosine phosphatase (PTP1B), thus prolonging the phosphorylation state of the insulin receptor and its substrates, and promoting GLUT4 translocation and glucose uptake. Among them, polyoxovanadates are a class of cluster compounds composed of vanadium-oxygen polyhedra, possessing advantages such as well-defined structure, tunable composition, and high stability. Studies have shown that decavanadates (V... 10 O 28 6- Typical polyoxovanadates, such as α-phosphoric acid vanadates, exhibit good anti-hyperglycemic effects. However, the functions of traditional polyoxovanadates are relatively limited, mainly focusing on insulin-like activity, and they lack effective regulatory capabilities for the oxidative stress and inflammation simultaneously present in diabetic wounds. Therefore, developing a multifunctional therapeutic agent that can simultaneously target hyperglycemia, oxidative stress, and inflammation in diabetic wounds has significant clinical implications. Summary of the Invention

[0006] To address the shortcomings of existing diabetic chronic wound treatments, which often lack specific functions and struggle to simultaneously address multiple pathological factors such as high glucose levels, oxidative stress, and inflammation, this invention provides a polyoxovanadium molybdate nanocluster, its preparation method, and its applications. This nanocluster incorporates molybdate by substituting some V atoms in the decavanadate lattice with Mo, thus entering the lattice as substitutional doping. While retaining the insulin-like activity of the parent POV, it endows the treatment with excellent antioxidant properties. Through the combined effect of antioxidant and insulin-like activity, it achieves multi-level regulation of the diabetic wound microenvironment, promoting wound healing.

[0007] In a first aspect, the present invention provides a polyoxovanadium molybdate nanocluster, wherein the general chemical formula of the polyoxovanadium molybdate nanocluster is Na6[V]. 10-x Mo x O 28The formula is ·nH₂O, where 1 ≤ x ≤ 4, and n is the number of water molecules of crystallization, ranging from 8 to 12. Here, x is the actual doping ratio determined by calculation based on the mass concentrations of V and Mo in the sample detected by inductively coupled plasma optical emission spectroscopy (ICP-OES). It should be noted that although the materials are fed according to a specific molar ratio during synthesis, due to the incomplete conversion characteristics of the dopant, the actual amount of Mo entering the crystal lattice (i.e., x in the general formula) differs from the proportion of Mo in the feed. Therefore, the actual doping ratio x in the polyoxovanadium molybdate nanoclusters described in this invention is based on the value determined by ICP-OES. Typically, under conventional drying conditions, the value of n can fluctuate within a certain range depending on factors such as the degree of drying during the preparation process and the humidity of the storage environment.

[0008] In this invention, the polyoxovanadium molybdate nanoclusters possess a decavanadate framework structure, wherein some V atoms are replaced by Mo atoms, which enter the crystal lattice in the form of substitutional doping, occupying the original V atom lattice positions. By introducing Mo, the polyoxovanadium molybdate nanoclusters maintain the stable framework of decavanadate while acquiring new functional properties.

[0009] Preferably, 2 ≤ x ≤ 3.6, and more preferably, x = 3.

[0010] In this invention, the antioxidant properties of nanoclusters can be optimized by adjusting the substitution amount of Mo. When x < 1, the Mo content is too low, and the improvement in antioxidant properties is not significant; when x > 4, excessive Mo may affect the stability of the decavanadate framework. When x = 3, it is the optimal solution, at which point the nanoclusters maintain good structural stability while exhibiting the best overall performance.

[0011] Preferably, the particle size of the polyoxovanadium molybdate nanoclusters is 1–6 nm, more preferably 3–5 nm. The polyoxovanadium molybdate nanoclusters are uniformly dispersed nanoparticles and exhibit good dispersibility in water.

[0012] The vanadium molybdate nanoclusters described in this invention possess broad-spectrum antioxidant activity, capable of scavenging substances selected from hydrogen peroxide (H2O2), hydroxyl radicals (•OH), and superoxide anions (O2•). -The nanoclusters contain at least three reactive oxygen species or free radicals, including DPPH free radicals and ABTS free radicals; the nanoclusters possess insulin-like activity, promoting glucose uptake in insulin-resistant cells (e.g., adipocytes, endothelial cells) and upregulating the expression of glucose transporter GLUT4; the nanoclusters possess anti-inflammatory activity, inhibiting the expression of pro-inflammatory factor IL-6 and promoting the expression of anti-inflammatory factor IL-10; the antioxidant activity and insulin-like activity of the nanoclusters have a combined effect, protecting the insulin signaling pathway by scavenging reactive oxygen species, thereby enhancing its glucose uptake effect; the nanoclusters possess pro-angiogenic activity, directly promoting endothelial cell migration, proliferation, and tube formation, and indirectly promoting angiogenesis by acting on adipocytes to improve the local high-glucose microenvironment.

[0013] Secondly, the present invention provides a method for preparing polyoxovanadium molybdate nanoclusters, comprising: dissolving a V source and a Mo source in a solvent at a molar ratio, adjusting the pH of the solution to acidic, and performing an acidification precipitation reaction; after the reaction is completed, collecting the precipitate by centrifugation, and washing and drying it to obtain the polyoxovanadium molybdate nanoclusters.

[0014] Preferably, the V source is at least one of sodium metavanadate (NaVO3) and ammonium metavanadate (NH4VO3); the Mo source is sodium molybdate (Na2MoO4) or ammonium molybdate ((NH4)6Mo7O3). 24 At least one of the following: V source and Mo source. The purity of V source and Mo source is preferably analytical grade or higher to ensure the purity and performance of the product.

[0015] Preferably, the molar ratio of the Mo source to the V source is 1:2 to 2:1.

[0016] Preferably, the reagent used to adjust the pH of the solution to acidity is at least one of hydrochloric acid, nitric acid, and sulfuric acid, with hydrochloric acid being the most preferred. Preferably, the pH of the solution is adjusted to 3.0–6.0, more preferably 4.0–5.0.

[0017] Preferably, the acidification precipitation reaction is carried out at a temperature of 30–80°C, more preferably 50–70°C, and for a reaction time of 6–48 hours, more preferably 12–24 hours.

[0018] Preferably, the acidification precipitation reaction is carried out under stirring conditions at a stirring speed of 200–1000 rpm to ensure the uniformity of the reaction system.

[0019] Preferably, the washing involves washing with deionized water and ethanol 2 to 5 times in sequence to remove unreacted raw materials and byproducts and to help precipitate out.

[0020] Preferably, the drying is vacuum freeze-drying or ordinary vacuum drying; the drying time for vacuum freeze-drying is 12 to 48 hours, and the drying temperature is -50 to -80°C; the drying time for ordinary vacuum drying is 40 to 60°C, and the drying time is 12 to 24 hours; preferably, the drying is vacuum freeze-drying, which can better maintain the dispersion of nanoclusters.

[0021] Thirdly, the present invention provides the application of the above-mentioned polyoxovanadium molybdate nanoclusters in the preparation of therapeutic compositions for diabetic chronic wounds.

[0022] Fourthly, the present invention provides a therapeutic composition for diabetic chronic wounds, comprising the above-mentioned polyoxovanadium molybdate nanoclusters and an acceptable carrier.

[0023] Fifthly, the present invention provides a liquid therapeutic agent comprising the above-mentioned polyoxovanadium molybdate nanoclusters and an acceptable solvent. The liquid therapeutic agent is a therapeutic composition in a specific dosage form.

[0024] Preferably, the concentration of polyoxovanadium molybdate nanoclusters in the liquid therapeutic agent is 10–100 μg / mL, more preferably 40 μg / mL.

[0025] Preferably, the acceptable solvent is physiological saline, phosphate buffer, or water for injection.

[0026] Beneficial effects: This invention introduces Mo into the decavanadate lattice through a simple transition metal substitution method, obtaining a multifunctional polyoxovanadium molybdate nanocluster (hereinafter referred to as POMoV). Experimental results show that this nanocluster has at least the following beneficial effects: (1) Insulin-like activity: POMoV significantly promoted glucose uptake in insulin-resistant adipocytes and endothelial cells and upregulated the expression of glucose transporter GLUT4. In vitro experiments showed that after treating insulin-resistant adipocytes at a concentration of 40 μg / mL for 24 h, the fluorescence intensity of glucose uptake measured by 2-NBDG fluorescence method was more than 30% higher than that of the insulin-resistant control group and 14% higher than that of the POV-treated group; immunofluorescence staining showed that the fluorescence intensity of GLUT4 in the cell membrane was 60% higher than that of the insulin-resistant control group and 11% higher than that of the POV-treated group. When treating insulin-resistant endothelial cells, 40 μg / mL POMoV increased the fluorescence intensity of glucose uptake by 11% compared with the POV-treated group, and upregulated GLUT4 expression by 16% compared with the POV-treated group. The above results indicate that the insulin-like activity of POMoV is superior to that of the unsubstituted parent POV. (2) Antioxidant function: POMoV has excellent scavenging ability against various free radicals and can effectively reduce intracellular reactive oxygen species levels. In vitro chemical scavenging experiments showed that POMoV at a concentration of 40 μg / mL achieved a scavenging rate of over 34% for DPPH free radicals, over 35% for ABTS free radicals, over 35% for hydroxyl radicals (•OH), and over 35% for superoxide anions (O2• - The clearance rate of POMoV reached over 40%, and the clearance rate of hydrogen peroxide (H2O2) also reached over 40%. Cellular level experiments showed that in a diabetic microenvironment simulated by insulin resistance and H2O2, treatment of adipocytes with 40 μg / mL POMoV for 24 h resulted in a reduction of intracellular ROS levels (detected by the DCFH-DA probe) of over 60% compared to the model group; treatment of endothelial cells also resulted in a reduction of ROS levels of over 60%. However, the same concentration of POMoV had no significant effect on intracellular ROS levels. (3) Anti-inflammatory function: POMoV can inhibit the expression of pro-inflammatory factor IL-6 and promote the secretion of anti-inflammatory factor IL-10, effectively reversing the inflammatory response induced by the diabetic microenvironment. In vitro experiments showed that in adipocytes cultured in a diabetic simulated microenvironment, after treatment with 40 μg / mL POMoV for 24 h, ELISA detection showed that the IL-6 content in the cell culture supernatant was reduced by more than 50% compared with the model group, and the IL-10 content was increased by more than 10 times. In vivo experiments further confirmed that immunohistochemical staining of wound tissue in diabetic mice showed that on day 7, the IL-6 positive staining area in the POMoV treatment group was reduced by more than 50% compared with the model group, and the IL-10 positive staining area was increased by more than 2 times. The above results indicate that POMoV has a good ability to regulate inflammation; (4) Combined Effect: The antioxidant activity and insulin-like activity of POMoV have a combined effect, which can protect the insulin signaling pathway by scavenging reactive oxygen species, thereby enhancing its glucose uptake effect. In vitro experiments showed that in a diabetic simulated microenvironment, treatment of adipocytes with 40 μg / mL POMoV increased the glucose uptake fluorescence intensity by more than 1.4 times, while the same concentration of POV only increased it by 1.2 times. This indicates that the antioxidant function conferred by Mo substitution has an auxiliary effect on the inherent insulin-like activity: on the one hand, Mo scavenges ROS, reducing the damaging modification of key molecules in the insulin signaling pathway by oxidative stress; on the other hand, the retained insulin-like activity directly activates the pathway, and the combined effect of the two achieves a better glucose uptake effect. (5) Angiogenesis-promoting activity: POMoV can directly promote endothelial cell migration, proliferation, and tube formation, and can also indirectly promote angiogenesis by acting on adipocytes to improve the local high-glucose microenvironment. In terms of direct effects, treatment of HUVECs at a concentration of 40 μg / mL for 24 h showed that the cell migration rate was more than 2.5 times higher than that of the model group in the scratch assay; the cell proliferation activity was more than 1.3 times higher after 5 days as detected by CCK-8 assay; and the number of branching points was more than 2 times higher in the Matrigel tube formation assay. In terms of indirect effects, in the adipocyte-endothelial cell co-culture system, adipocytes pretreated with 40 μg / mL POMoV increased the migration rate of lower ventricular endothelial cells by more than 2.5 times, indicating that POMoV can indirectly create a microenvironment conducive to angiogenesis by regulating the metabolic function of adipocytes. (6) In vivo wound healing effect: In a full-thickness skin defect model experiment in diabetic mice, the wound healing rate of the POMoV treatment group reached 79.6±7.2% on day 6, significantly higher than that of the blank control group (56.3±8.4%) and the insulin group (62.3±8.5%). On day 14, the healing rate of the POMoV group reached 98.1±1.0%, and the wound was basically closed, while about 12% of the wounds in the blank control group remained unhealed. Histological analysis showed that the newly formed epidermis in the POMoV group was intact, the epithelialization was complete, the granulation tissue was mature, and the collagen deposition was dense. CD31 immunofluorescence staining showed that CD31 expression in the wound tissue of the POMoV group was significantly enhanced, about 1.3 times that of the blank control group. GLUT4 immunofluorescence staining showed that GLUT4 expression in the wound tissue of the POMoV group was significantly upregulated to 2 times that of the blank control group, indicating that local cellular glucose metabolism was improved. The above results fully demonstrate that POMoV can significantly promote the healing of diabetic chronic wounds through the combined effects of insulin-like activity, antioxidant, anti-inflammatory and angiogenesis-promoting effects; (7) Good biocompatibility: In vitro cytotoxicity experiments showed that, within the concentration range of 10–50 μg / mL, the cell viability of endothelial cells and adipocytes treated with POMoV for 24 h remained above 90% as detected by the CCK-8 assay. In vivo biocompatibility evaluation showed that, 14 days after intraperitoneal injection of POMoV dispersion (40 μg / mL) into diabetic mice, there were no significant differences in blood biochemical indicators compared with the normal control group, and no obvious pathological damage was observed in the H&E staining of major organs, indicating that POMoV has good in vitro and in vivo biocompatibility; In summary, the therapeutic agent of the present invention has a simple preparation method and good biocompatibility. Through the combined effects of insulin-like activity, antioxidant activity, anti-inflammatory activity and angiogenesis-promoting activity, it can synergistically improve the microenvironment of diabetic wounds from multiple levels, significantly promote wound healing, provide a new strategy for the treatment of chronic diabetic wounds, and has good prospects for clinical translation. Attached Figure Description

[0027] Figure 1 This is a schematic diagram of the crystal structure of the polyoxovanadium molybdate nanoclusters of the present invention; Figure 2 The following figures show the screening results of POMoV with different Mo / V molar ratios; where A is the XRD pattern; B is the scavenging curve of DPPH free radical by different proportions of samples; C is the scavenging curve of ABTS free radical by different proportions of samples; and D is the effect of different samples on the cell viability of HUVECs. Figure 3 The diagram shows the structural characterization of POMoV; where A is the FTIR spectrum; B is the XPS full spectrum; C is the TEM image of POMoV; D is the EDS elemental distribution map of POMoV; and E is the particle size distribution statistical map of POMoV. Figure 4 This is a graph evaluating the antioxidant activity of POMoV; where A is the DPPH radical scavenging rate as a function of concentration; B is the ABTS radical scavenging rate as a function of concentration; C is the hydroxyl radical (•OH) scavenging rate as a function of concentration; and D is the superoxide anion (O2• - E is the scavenging rate versus concentration curve; F is the scavenging rate versus concentration curve of hydrogen peroxide (H2O2); F is the representative fluorescence image of ROS staining in adipocytes and the quantitative statistics of fluorescence intensity. Figure 5 Figure 1 shows the results of POMoV promoting glucose uptake and GLUT4 expression in adipocytes; where A represents the quantitative results of 2-NBDG fluorescence after treatment with different concentrations of POMoV; B represents the representative images of 2-NBDG fluorescence; C represents the representative images of GLUT4 immunofluorescence staining; and D represents the quantitative statistics of GLUT4 fluorescence intensity. Figure 6 The image shows the combined antioxidant and insulin-like effects of POMoV; where A represents the quantitative results of 2-NBDG fluorescence under different treatments in a diabetes-simulated microenvironment; B represents representative images of 2-NBDG fluorescence; C represents representative images of GLUT4 immunofluorescence staining; and D represents the quantitative statistics of GLUT4 fluorescence intensity. Figure 7 Representative images showing the results of POMoV in improving endothelial cell dysfunction; where A is a representative image of the scratch assay; B is a representative image of the tube formation assay; and C is a representative image of the endothelial cell scratch assay in the adipocyte-endothelial cell co-culture system. Figure 8 A statistical graph showing the results of POMoV in improving endothelial cell dysfunction; where A represents... Figure 7 A corresponds to the migration rate statistics; B is... Figure 7 B corresponds to quantitative statistics of branch points; C is the result of cell proliferation detection using the CCK-8 assay; D is... Figure 7 The migration rate statistics corresponding to C; Figure 9 The results of POMoV promoting wound healing in diabetic mice are shown in the figure below. A shows photos of wound healing at different time points; B shows the quantitative statistical results of wound healing rate; C shows H&E staining of wound tissue on day 14; D shows Masson staining of wound tissue on day 14; E shows CD31 immunofluorescence staining; F shows GLUT4 immunofluorescence staining; and G shows IL-6 and IL-10 immunohistochemical staining of wound tissue on day 7. Figure 10 Figure 1 shows the biocompatibility evaluation of POMoV. In this figure, A represents the survival rate of endothelial cells after treatment with different concentrations of POMoV for 24 h; B represents the survival rate of adipocytes after treatment with different concentrations of POMoV for 24 h; C represents the survival rate of fibroblasts after treatment with different concentrations of POMoV for 24 h; D represents the blood biochemical indicators of mice in each group during the in vivo safety evaluation; and E represents the H&E staining images of major organs. Figure 11 The graph shows a comparison of the scavenging rates of DPPH and ABTS free radicals by POV and POMoV0.8, respectively. Detailed Implementation

[0028] To further illustrate the invention's content, features, and practical effects, the invention will be described in detail below with reference to embodiments. It should be noted that the modification methods of the invention are not limited to these specific implementation methods. Equivalent substitutions and modifications made by those skilled in the art based on their reading of the invention's content, without departing from the spirit and essence of the invention, are also within the scope of protection claimed by this invention.

[0029] First, this invention provides a polyoxovanadium molybdate nanocluster, which has a decavanadate framework structure, wherein some V atoms are replaced by Mo atoms, which enter the crystal lattice in the form of substitutional doping, occupying the original V atom lattice positions. By introducing Mo, the polyoxovanadium molybdate nanocluster maintains the stable framework of the decavanadate while acquiring new functional properties. The general chemical formula of the polyoxovanadium molybdate nanocluster is Na6[V] 10-x Mo x O 28The molybdenum content is 1 ≤ x ≤ 4, preferably 2 ≤ x ≤ 3.6, and more preferably x = 3. n is the number of water molecules of crystallization, ranging from 8 to 12. Typically, under normal drying conditions, the value of n can fluctuate within a certain range depending on factors such as the degree of drying during preparation and the humidity of the storage environment. The antioxidant properties of the nanoclusters can be optimized by adjusting the amount of Mo substitution. When x < 1, the Mo content is too low, and the improvement in antioxidant properties is not significant; when x > 4, excessive Mo may affect the stability of the decavanadate framework. When x = 3, it is the optimal solution, where the nanoclusters maintain good structural stability while exhibiting the best overall performance.

[0030] In an optional embodiment, the particle size of the polyoxovanadium molybdate nanoclusters is 1–6 nm, preferably 3–5 nm.

[0031] In this invention, the mechanism by which Mo substitutes some V atoms in the decavanadate lattice to enhance the performance of nanoclusters is summarized as follows: Mo and V have similar atomic radii (Mo is 139 pm, V is 134 pm), electronegativity, and coordination preference, which allows Mo to exist stably in the decavanadate lattice as a substitutional dopant without disrupting its framework structure. At the electronic level, Mo exists in multiple reversible oxidation states and possesses excellent redox regulation capabilities; its introduction can significantly modulate the electronic structure of the nanoclusters. On one hand, Mo mimics antioxidant enzyme activity, achieving broad-spectrum free radical scavenging through an electron transfer mechanism, thus endowing the material with excellent antioxidant function. On the other hand, Mo substitution alters the spatial configuration or charge distribution binding to the protein tyrosine phosphatase (PTP1B) active site by regulating the electronic structure, enhancing affinity and thus more effectively inhibiting PTP1B activity, prolonging insulin signal transduction, and exerting insulin-like function. At the functional level, these two effects work together, not only giving the nanoclusters insulin-like and antioxidant capabilities but also enabling effective anti-inflammatory effects, ultimately achieving multiple regulation of diabetic wounds and accelerating healing.

[0032] The polyoxovanadium molybdate nanoclusters provided by this invention have the following excellent properties: (1) The polyoxovanadium molybdate nanoclusters have broad-spectrum antioxidant activity and can scavenge free radicals selected from hydrogen peroxide (H2O2), hydroxyl radicals (•OH), and superoxide anions (O2•OH). - At least three reactive oxygen species or free radicals, including DPPH free radicals and ABTS free radicals, are present. Broad-spectrum antioxidant activity is evaluated using the following methods: free radical scavenging rate is determined by the DPPH method, and the scavenging rate is calculated from the change in absorbance at 520 nm; •OH is generated using the Fenton system, and the scavenging rate is determined using tetramethylbenzidine; O2• is measured using WST-8. -Scavenging rate; the H2O2 scavenging rate was determined using the titanium salt colorimetric method. In this invention, the polyoxovanadium molybdate nanoclusters exhibited a scavenging rate of over 34% for DPPH free radicals, over 35% for ABTS free radicals, and over 35% for •OH at a concentration of 40 μg / mL, and also showed a scavenging rate of over 35% for O2• - The scavenging rate reached over 40%, and the scavenging rate of H2O2 also reached over 40%. Furthermore, the nanoclusters could reduce the level of reactive oxygen species (ROS) in cells (such as adipocytes and endothelial cells) under a simulated diabetic microenvironment (insulin resistance + H2O2). Detection using the DCFH-DA probe method showed that treatment with 40 μg / mL POMoV for 24 h reduced intracellular ROS levels by over 60%. This antioxidant activity stems from the alteration of the electronic structure of the nanoclusters after the introduction of Mo, enabling them to quench various free radicals through an electron transfer mechanism. (2) The polyoxovanadium molybdate nanoclusters possess insulin-like activity, which can promote glucose uptake in insulin-resistant cells (such as adipocytes and endothelial cells) and upregulate the expression of the glucose transporter GLUT4. The insulin-like activity was evaluated using the following methods: cellular glucose uptake capacity was measured using the 2-NBDG fluorescent probe method and observed under a fluorescence microscope; GLUT4 expression was detected using immunofluorescence staining. In this invention, when the polyoxovanadium molybdate nanoclusters were treated with insulin-resistant adipocytes at a concentration of 40 μg / mL for 24 h, the 2-NBDG fluorescence intensity increased by more than 1.3 times, and the cell membrane GLUT4 fluorescence intensity increased by more than 1.5 times; when treating insulin-resistant endothelial cells, the 2-NBDG fluorescence intensity increased by more than 1.4 times, and the cell membrane GLUT4 fluorescence intensity increased by more than 1.4 times. This insulin-like activity originates from the enhanced PTP1B inhibitory activity of the nanoclusters after Mo substitution, which activates the downstream PI3K / Akt signaling pathway by prolonging the phosphorylation state of the insulin receptor and its substrate. (3) The polyoxovanadium molybdate nanoclusters possess anti-inflammatory activity, inhibiting the expression of pro-inflammatory factor IL-6 and promoting the expression of anti-inflammatory factor IL-10. The anti-inflammatory activity was evaluated by the following method: the content of IL-6 and IL-10 in the cell culture supernatant was detected using an ELISA kit. In this invention, in adipocytes cultured in a diabetic simulated microenvironment, treatment with 40 μg / mL POMoV for 24 h reduced IL-6 secretion by more than 50% and increased IL-10 secretion by more than 10 times; in the wound tissue of diabetic mice, the IL-6 positive staining area in the POMoV treatment group decreased by more than 50% on day 7, and the IL-10 positive staining area increased by more than 2 times. This anti-inflammatory activity stems from the inhibitory effect of the nanoclusters on oxidative stress after the introduction of Mo, blocking the activation of inflammatory pathways such as NF-κB by clearing ROS, and may also promote anti-inflammatory expression by directly regulating cellular immune function; (4) The antioxidant activity and insulin-like activity of the polyoxovanadium molybdate nanoclusters have a combined effect, which can protect the insulin signaling pathway by scavenging reactive oxygen species, thereby enhancing its glucose uptake effect. The combined effect was evaluated by the following method: in a diabetic simulated microenvironment, the protective effect of the nanoclusters on glucose uptake and GLUT4 expression in insulin-resistant cells was detected and compared with the effect of POV without antioxidant activity. In adipocytes, treatment with 40 μg / mL POMoV increased glucose uptake by more than 1.4 times, which was significantly better than POV (1.2 times). This indicates that the antioxidant function conferred by Mo substitution has an auxiliary effect on the inherent insulin-like activity: on the one hand, Mo scavenges ROS and reduces the damaging modification of key molecules in the insulin signaling pathway by oxidative stress; on the other hand, it retains the insulin-like activity to directly activate the pathway. (5) The polyoxovanadium molybdate nanoclusters possess angiogenic activity, directly promoting endothelial cell migration, proliferation, and tube formation. Angiogenic activity was evaluated using the following methods: a scratch assay was used to detect endothelial cell migration ability and calculate the migration rate; the CCK-8 assay was used to detect cell proliferation activity; and the Matrigel tube formation assay was used to detect endothelial cell tube formation ability and count the number of branch points. In this invention, treating HUVECs with the polyoxovanadium molybdate nanoclusters at a concentration of 40 μg / mL for 24 h increased the scratch healing rate by more than 2.5 times; after 5 days, cell proliferation activity increased by more than 1.3 times, and the number of tube formation branch points increased by more than 2 times. Furthermore, the nanoclusters can indirectly promote angiogenesis by acting on adipocytes to improve the local high-glucose microenvironment: in an adipocyte-endothelial cell co-culture system, adipocytes pretreated with POMoV significantly enhanced endothelial cell migration and tube formation ability, indicating that POMoV can indirectly create a microenvironment conducive to angiogenesis by regulating adipocyte metabolic function. (6) The polyoxovanadium molybdate nanoclusters exhibit good biocompatibility, maintaining cell viability of over 90% in mammalian cells (such as endothelial cells and adipocytes) within a concentration range of 10–50 μg / mL. Biocompatibility was evaluated using the following methods: the CCK-8 assay was used to detect cell viability after 24 h of treatment with different concentrations of nanoclusters; and the live / dead cell staining method was used to observe cell status. In this invention, the polyoxovanadium molybdate nanoclusters, after treating endothelial cells at a concentration of 40 μg / mL for 24 h, still maintained a cell viability of over 96%. In vivo biocompatibility evaluation showed that, 14 days after intraperitoneal injection of POMoV dispersion into diabetic mice, there were no significant differences in blood biochemical indicators (ALT, AST, ALP, CREA) compared to the normal control group, and no obvious pathological damage was observed in H&E staining of major organs (heart, liver, spleen, lung, kidney), indicating that the nanoclusters have good in vitro and in vivo biocompatibility.

[0033] The following exemplifies the preparation method of polyoxovanadium molybdate nanoclusters provided by the present invention.

[0034] V source and Mo source are dissolved in solvent at a certain molar ratio, the pH of the solution is adjusted to acidic, and an acidification precipitation reaction is carried out. After the reaction is completed, the precipitate is collected by centrifugation, washed and dried to obtain the polyoxovanadium molybdate nanoclusters.

[0035] In alternative embodiments, the deposition method includes spin coating, blade coating, spray coating, dip coating, or chemical bath coating.

[0036] In an optional embodiment, the V source is at least one of sodium metavanadate (NaVO3) and ammonium metavanadate (NH4VO3); the Mo source is sodium molybdate (Na2MoO4) or ammonium molybdate ((NH4)6Mo7O3). 24 At least one of the following: V source and Mo source. The purity of the V source and Mo source is preferably analytical grade or higher to ensure the purity and performance of the product.

[0037] In an optional embodiment, the molar ratio of the Mo source to the V source is 1:2 to 2:1. This invention allows for the preparation of polyoxovanadium molybdate nanoclusters with different Mo contents by adjusting the molar ratio of the Mo source to the V source. These nanoclusters are labeled POMoV1, POMoV2, and POMoV3, respectively. After screening for antioxidant activity, POMoV2 exhibits the best antioxidant performance.

[0038] In an optional embodiment, the reagent used to adjust the pH of the solution to acidity is at least one of hydrochloric acid, nitric acid, and sulfuric acid, preferably hydrochloric acid; more preferably, the pH of the solution is adjusted to 3.0 to 6.0, and more preferably 4.0 to 5.0. A pH that is too low (<3.0) may result in the inability to dope, while a pH that is too high (>6.0) may prevent the formation of a precipitate or reduce the purity of the product.

[0039] In an optional embodiment, the acidification precipitation reaction is carried out at a temperature of 30–80°C, preferably 50–70°C, and for a reaction time of 6–48 hours, preferably 12–24 hours. Too low a reaction temperature (<30°C) may lead to incomplete reaction; too high a reaction temperature (>80°C) may lead to product agglomeration. Too short a reaction time may lead to incomplete reaction and poor product crystallinity; too long a reaction time has no significant improvement on product performance.

[0040] In an optional embodiment, the acidification precipitation reaction is carried out under stirring conditions at a speed of 200–1000 rpm to ensure a uniform reaction system. Too slow a stirring speed may lead to uneven local concentrations, while too fast a stirring speed may generate bubbles that affect the precipitation quality.

[0041] In an optional embodiment, the centrifugation speed is 8000-15000 rpm, and the centrifugation time is 5-15 minutes.

[0042] In an optional embodiment, the washing involves sequentially washing with deionized water and ethanol 2 to 5 times to remove unreacted raw materials and byproducts and to aid in precipitation. Too few washes may result in insufficient product purity, while too many washes may lead to product loss.

[0043] In an optional embodiment, the drying is vacuum freeze-drying or ordinary vacuum drying; the drying time for vacuum freeze-drying is 12 to 48 hours, and the drying temperature is -50 to -80°C; the drying time for ordinary vacuum drying is 40 to 60°C, and the drying time is 12 to 24 hours; preferably, the drying is vacuum freeze-drying, which can better maintain the dispersion of nanoclusters.

[0044] As a preferred preparation method, the preparation method includes: dissolving sodium metavanadate and sodium molybdate in deionized water at a Mo:V molar ratio of 1:1 and stirring until completely dissolved; adjusting the pH to 4.5 with hydrochloric acid and stirring the reaction at 60°C for 24 hours; after the reaction is completed, collecting the precipitate by centrifugation at 12000 rpm for 10 minutes, washing it three times each with deionized water and ethanol; and freeze-drying the precipitate under vacuum for 24 hours to obtain polyoxovanadium molybdate nanoclusters POMoV2.

[0045] In this invention, during the preparation of polyoxovanadium molybdate nanoclusters by replacing V with Mo, the following measures are taken: First, to address the risk of structural damage due to the difference in atomic radii between Mo and V, the pH is precisely controlled at 4.0–5.0 and the reaction temperature at 50–70°C to ensure that Mo successfully enters the crystal lattice without damaging the original structure. Second, to address the deviation between the feed ratio and the actual doping amount, this invention precisely determines the actual doping amount by controlling the feed molar ratio and combining it with ICP-OES. It was found that within the range of 1 ≤ x ≤ 4, x=3 was determined to be the optimal ratio for antioxidant activity, biocompatibility, and insulin-like activity. To avoid particle agglomeration, the reaction time is controlled at 12–24 hours and the stirring speed at 200–1000 rpm to obtain uniformly dispersed nanoclusters with a particle size of 1–6 nm.

[0046] Furthermore, the present invention also provides the application of the above-mentioned polyoxovanadium molybdate nanoclusters in the preparation of diabetic chronic wound treatment compositions.

[0047] The following examples further illustrate the present invention in detail. It should also be understood that the following examples are only for further explanation of the present invention and should not be construed as limiting the scope of protection of the present invention. Any non-essential improvements and adjustments made by those skilled in the art based on the above description of the present invention are within the scope of protection of the present invention. The specific process parameters, etc., in the following examples are merely examples within a suitable range; that is, those skilled in the art can make appropriate selections within the appropriate range based on the description herein, and are not intended to be limited to the specific values ​​in the examples below. Example 1

[0048] In Example 1, polyoxovanadium molybdate nanoclusters (chemical formula Na6[V8Mo2O]) were prepared with a Mo / V molar ratio of 1:2. 28 The preparation process of ·n H2O (x=2, labeled POMoV1) includes the following steps: Weigh 3 g of NaVO3 and 2.98 g of sodium molybdate Na2MoO4•2H2O according to the molar ratio Mo:V = 1:2, place them in a 250 mL round-bottom flask, add 200 mL of deionized water, and stir at room temperature until completely dissolved; adjust the pH to 4.5 with hydrochloric acid, place in a 60℃ oil bath, and stir at 800 rpm for 24 hours; after the reaction is completed, centrifuge at 12000 rpm for 10 minutes to collect the precipitate, wash it three times each with deionized water and anhydrous ethanol, and finally freeze-dry the precipitate under vacuum at -60℃ for 24 hours to obtain POMoV1 yellow-green powder. Example 2

[0049] In Example 2, polyoxovanadium molybdate nanoclusters (chemical formula Na6[V7Mo3O4]) were prepared with a Mo / V molar ratio of 1:1. 28 The preparation process of ·n H2O (x=3, labeled as POMoV2) includes the following steps: Weigh 3 g of NaVO3 and 5.95 g of sodium molybdate Na2MoO4•2H2O according to the molar ratio Mo:V=1:1, place them in a 250 mL round-bottom flask, add 200 mL of deionized water, and stir at room temperature until completely dissolved; adjust the pH to 4.5 with hydrochloric acid, place in a 60℃ oil bath, and stir at 800 rpm for 24 hours; after the reaction is completed, centrifuge at 12000 rpm for 10 minutes to collect the precipitate, wash it three times each with deionized water and anhydrous ethanol, and finally freeze-dry the precipitate under vacuum at -60℃ for 24 hours to obtain a yellow-green powder of POMoV2. Example 3

[0050] In Example 3, polyoxovanadium molybdate nanoclusters (chemical formula Na6[V]) were prepared with a Mo / V molar ratio of 2:1. 6.4 Mo 3.6O 28 The preparation process of ·n H2O (x=3.6, labeled as POMoV3) includes the following steps: Weigh 3 g of NaVO3 and 11.9 g of sodium molybdate Na2MoO4•2H2O according to the molar ratio Mo:V=2:1, place them in a 250 mL round-bottom flask, add 200 mL of deionized water, and stir at room temperature until completely dissolved; adjust the pH to 4.5 with hydrochloric acid, place in a 60℃ oil bath, and stir at 800 rpm for 24 hours; after the reaction is completed, centrifuge at 12000 rpm for 10 minutes to collect the precipitate, wash it three times each with deionized water and anhydrous ethanol, and finally freeze-dry the precipitate under vacuum at -60℃ for 24 hours to obtain a yellow-green powder of POMoV3. Comparative Example 1

[0051] In Comparative Example 1, polyoxovanadium molybdate nanoclusters (chemical formula Na6[V] vanadium molybdate) were prepared with a Mo / V molar ratio of 1:3. 9.2 Mo 0.8 O 28 The preparation process of ·n H2O (x≈0.8, labeled as POMoV0.8) includes the following steps: Weigh 3 g of NaVO3 and 1.98 g of sodium molybdate Na2MoO4•2H2O according to the molar ratio Mo:V = 1:3, place them in a 250 mL round-bottom flask, add 200 mL of deionized water, and stir at room temperature until completely dissolved; adjust the pH to 4.5 with hydrochloric acid, place in a 60℃ oil bath, and stir at 800 rpm for 24 hours; after the reaction is completed, centrifuge at 12000 rpm for 10 minutes to collect the precipitate, wash it three times each with deionized water and anhydrous ethanol, and finally freeze-dry the precipitate under vacuum at -60℃ for 24 hours to obtain a yellow-green powder of POMoV0.8.

[0052] The free radical scavenging rate was determined using the DPPH method and ABTS (see Example 5 for details of the method), and the results are as follows: Figure 11 As shown, compared with POV, POMoV0.8 did not significantly improve the scavenging rate of DPPH and ABTS free radicals (< 10%), indicating that the low Mo content cannot effectively endow the nanoclusters with antioxidant activity. Comparative Example 2

[0053] In Comparative Example 2, polyoxovanadium molybdate nanoclusters (chemical formula Na6[V] vanadium molybdate) were prepared with a Mo / V molar ratio of 3:1. 5.2 Mo 4.8 O 28 The preparation process of ·n H2O (x≈4.8, labeled as POMoV4.8) includes the following steps: Weigh 3 g of NaVO3 and 17.85 g of sodium molybdate Na2MoO4•2H2O according to the molar ratio Mo:V = 3:1, place them in a 250 mL round-bottom flask, add 200 mL of deionized water, and stir at room temperature until completely dissolved; adjust the pH to 4.5 with hydrochloric acid, place in a 60 °C oil bath, and stir at 800 rpm for 24 hours; after the reaction, centrifuge at 12000 rpm for 10 minutes to collect the precipitate, wash it three times each with deionized water and anhydrous ethanol, and finally freeze-dry the precipitate under vacuum at -60 °C for 24 hours to obtain POMoV4 blue-green powder. Its appearance is significantly different from the color of the POMoV series samples. It is preliminarily judged that the excessive Mo doping caused the framework structure to collapse, so no further performance tests and characterization were carried out.

[0054] The mass concentrations of V and Mo in Examples 1-3 and Comparative Examples 1-2 were detected by inductively coupled plasma optical emission spectroscopy (ICP-OES). The actual doping ratio was determined by calculation. The specific calculation method for x in the chemical formula is as follows: Based on the mass concentration of the elements in the same solution and the relative atomic masses of V and Mo (V=50.94, Mo=95.94), their atomic molar ratio was calculated. Since the number of metal atoms in the basic building block of decavanadate is 10, the calculated atomic molar ratio was normalized to 10, and the value of x could be calculated. Taking POMoV2 as an example: n(V)=10.12 / 50.94≈0.1987, n(Mo)=8.17 / 95.94≈0.08516, n(V) / n(Mo)=0.1987 / 0.08516=2.33 / 1, x=10×(1 / (2.33+1))=3.00. Table 1 lists the mass concentrations of V and Mo obtained from the corresponding detections of the polyoxovanadium molybdate nanoclusters prepared in Examples 1-3 and Comparative Examples 1-2.

[0055] Table 1: .

[0056] X-ray diffraction analysis was performed on the three samples prepared in Examples 1-3, and the results are as follows: Figure 2 As shown in Figure A; the free radical scavenging rates of the three samples were determined using the DPPH and ABTS methods (details of the methods are in Example 5), and the results are as follows. Figure 2 As shown in BC; the cell viability of HUVECs in three samples was detected using the CCK-8 assay, and the results are as follows. Figure 2 As shown in D, based on the results of structural confirmation, antioxidant performance screening, and cell viability screening, POMoV2 was selected as the optimal ratio for use in all subsequent embodiments (i.e., POMoV as described below).

[0057] Figure 1This is a schematic diagram of the crystal structure of the polyoxovanadium molybdate nanoclusters of the present invention, showing that Mo atoms occupy some of the positions of V atoms in the decavanadate lattice in the form of substitutional doping.

[0058] Figure 2 The figure shows the screening results of POMoV with different Mo / V feed ratios; where A is the XRD pattern; B is the scavenging curve of DPPH free radicals by different sample ratios; C is the scavenging curve of ABTS free radicals by different sample ratios; and D is the effect of different samples on the cell viability of HUVECs. As shown in the figure, the antioxidant performance of the nanoclusters increases with increasing Mo content, and POMoV3 has a slightly higher scavenging rate of DPPH and ABTS than POMoV2. However, from POMoV2 to POMoV3, the Mo feed amount doubled, but the improvement in antioxidant performance was far less than the increase in feed amount, showing a clear diminishing marginal effect. Meanwhile, cell viability testing results showed that the endothelial cell viability of the POMoV3 treatment group was lower than that of the POMoV2 group, indicating that excessively high Mo content may have adverse effects on specific cells. Considering antioxidant performance, biocompatibility, and cost-effectiveness, POMoV2 maintains excellent antioxidant activity while exhibiting the best cell safety, and is therefore the optimal choice. Example 4: Structural Characterization of POMoV

[0059] The POMoV2 sample prepared in Example 1 was subjected to structural characterization. The FTIR spectrum of the sample was measured using a Fourier transform infrared spectroscopy (FTIR) instrument, and the results are as follows: Figure 3 As shown in Figure A, in addition to the characteristic absorption peaks of decavanadate (752, 877, 917, 956 cm⁻¹), new absorption peaks appear at 985 cm⁻¹ and 1405 cm⁻¹, which are attributed to the Mo=O vibration, confirming the successful incorporation of Mo into the decavanadate framework. The XPS full spectrum of the sample was measured using X-ray photoelectron spectroscopy, and the results are as follows. Figure 3 As shown in Figure B, characteristic peaks for Na, V, O, and Mo are visible in the full spectrum. The sample morphology was observed using transmission electron microscopy (TEM), and the TEM images (…) are shown below. Figure 3 C) shows that POMoV is in the form of uniformly dispersed spherical particles with no obvious agglomeration, and the particle size distribution ranges from 1 to 6 nm. Figure 3 E). Elemental surface scanning analysis was performed using energy-dispersive X-ray spectroscopy, and the elemental distribution map was obtained using EDS (Energy Dispersive X-ray Spectroscopy). Figure 3 D) The results show that V, Mo, and O elements are uniformly distributed within the nanoclusters, with no elemental segregation observed. These results indicate the successful preparation of Mo-doped, uniformly sized, and structurally stable polyoxovanadium molybdate nanoclusters. Example 5: In vitro free radical scavenging ability and concentration screening of POMoV

[0060] The free radical scavenging ability of POMoV was determined using the DPPH method. POMoV was prepared at different concentrations (0, 10, 20, 40, 50 μg / mL), mixed with DPPH solution, and reacted in the dark for 30 min. The absorbance at 520 nm was then measured, and the scavenging rate was calculated. The results are as follows: Figure 4 As shown in Figure A, the scavenging of DPPH by POMoV is dose-dependent, with the scavenging rate increasing with increasing concentration. The scavenging rate reaches 34.9% at 40 μg / mL and 46.6% at 50 μg / mL. The ABTS radical scavenging rate was determined using the ABTS method. The above-mentioned concentrations of POMoV were mixed with the ABTS working solution, and the absorbance at 730 nm was measured. The results are as follows. Figure 4 As shown in Figure B, the scavenging rate reached 36.1% at 40 μg / mL and 50.9% at 50 μg / mL. Hydroxyl radicals (•OH) were generated using the Fenton system, and the scavenging rate was determined by the tetramethylbenzidine colorimetric method. The results are shown in Figure B. Figure 4 As shown in Figure C, the scavenging rate reached 37.4% at 40 μg / mL and 47.1% at 50 μg / mL. Superoxide anion (O2•) was determined using the WST-8 method. - The clearance rate and results are as follows: Figure 4 As shown in Figure D, the removal rate reached 40.7% at 40 μg / mL and 48.5% at 50 μg / mL. The removal rate of hydrogen peroxide (H2O2) was determined using the titanium salt colorimetric method, and the results are as follows: Figure 4 As shown in Figure E, the scavenging rate reached over 42.8% at 40 μg / mL and 50.9% at 50 μg / mL. These results indicate that POMoV possesses broad-spectrum free radical scavenging ability, and the scavenging effect is concentration-dependent, with a higher scavenging rate at 50 μg / mL than at 40 μg / mL. However, considering cell safety, cytotoxicity experiments (…) Figure 10 C) showed that fibroblast viability was significantly reduced at a concentration of 50 μg / mL compared to 40 μg / mL, indicating that excessively high concentrations may have adverse effects on cells. Therefore, to ensure excellent antioxidant activity while also considering cell safety, 40 μg / mL was selected as the preferred working concentration for POMoV in subsequent experiments. Example 6: POMoV reduces intracellular ROS levels

[0061] 3T3-L1 preadipocytes were seeded into culture plates and cultured for 8 days in a differentiation-inducing medium containing IBMX, dexamethasone, and insulin to induce differentiation into mature adipocytes. Subsequently, an insulin resistance model was established by treatment with high insulin (0.1 μM) for 48 h. H2O2 (100 μM) was added to simulate the oxidative stress microenvironment of diabetes. The cells were incubated with the DCFH-DA probe at 37℃ in the dark for 30 min, washed with PBS, and observed and photographed under a fluorescence microscope. The results are as follows: Figure 4 As shown in Figure F, the intensity of green fluorescence in the cells of the POMoV-treated group was significantly weaker than that of the model control group. Quantitative statistics showed that the ROS level was reduced by more than 60%, indicating that POMoV can effectively remove excess ROS in cells. Example 7: POMoV promotes glucose uptake by adipocytes

[0062] An insulin-resistant adipocyte model was established according to the method in Example 6. The model cells were co-cultured with different concentrations of POMoV (0, 10, 20, 30, 40, 50 μg / mL) for 24 h, incubated with 50 μM 2-NBDG for 30 min, washed with PBS, and observed and photographed under a fluorescence microscope. The results are as follows: Figure 5 As shown in AB, POMoV enhances glucose uptake in a dose-dependent manner, with fluorescence intensity at 40 μg / mL increasing by more than 30% compared to the model control group, indicating that POMoV has insulin-like activity. Example 8: POMoV upregulates GLUT4 expression

[0063] GLUT4 expression was detected using immunofluorescence staining. Insulin-resistant adipocytes treated with POMoV for 24 h in Example 7 were fixed with 4% paraformaldehyde for 15 min, blocked with 5% BSA for 1 h, and then incubated overnight at 4°C with GLUT4 primary antibody (1:1000 dilution). After washing with PBS, fluorescently labeled secondary antibody (1:500 dilution) was added and incubated at room temperature for 1 h. The nuclei were counterstained with DAPI. After mounting, the cells were observed and photographed under a fluorescence microscope, and the fluorescence intensity of the cell membrane region was counted using ImageJ software. The results are as follows: Figure 5 As shown in CD, the fluorescence intensity of GLUT4 in the cell membrane of the POMoV-treated group was more than 40% higher than that of the model control group, indicating that POMoV can upregulate GLUT4 expression. Example 9: Combined effect of antioxidant and insulin-like substances

[0064] In an insulin-resistant adipocyte model, H2O2 (100 μM) was added to simulate the oxidative stress microenvironment. The experiment was divided into four groups: a model control group, an insulin group, a POV group (25 μg / mL, consistent with the V concentration of POMoV), and a POMoV group (40 μg / mL). After 24 h of treatment, 2-NBDG glucose uptake and GLUT4 expression were detected according to the methods in Examples 7 and 8. The results are as follows: Figure 6As shown in the AD diagram, under oxidative stress, the fluorescence intensity of glucose uptake in the POV group only increased to 1.2 times that of the model group, while that in the POMoV group increased to more than 1.4 times. GLUT4 fluorescence intensity quantification showed that the POMoV group increased by 1.6 times compared to the model group, significantly better than the 1.4 times increase in the POV group. These results indicate that the antioxidant function of POMoV can protect the insulin signaling pathway and enhance its insulin-like activity. Example 10: POMoV directly promotes endothelial cell function

[0065] HUVECs were seeded in culture plates and cultured in ECM medium containing 10% FBS. Scratch assay: After cell confluence, scratches were performed using a 200 μL pipette tip. Exfoliated cells were washed with PBS, and serum-free medium containing different concentrations of POMoV (0, 10, 20, 30, 40, 50 μg / mL) was added. Images were taken under a microscope at 0 h and 24 h, and migration rate was calculated using ImageJ. Results are shown below. Figure 7 As shown in A and 8A, the migration-promoting effect increased with increasing concentration, with the migration rate of 40 μg / mL POMoV treatment for 24 h increasing by 2.8 times compared to the control group. Tube formation experiment: Matrigel was spread into 24-well plates and allowed to solidify. HUVECs were then inoculated and POMoV was added. After 6 h of culture, tube formation was observed under a microscope, and the number of branch points was counted. The results are as follows... Figure 7 As shown in B and 8B, the tube formation promotion effect increased with increasing concentration, with the number of branch points increasing 2.6 times after treatment with 40 μg / mL POMoV compared to the control group. Proliferation experiment: HUVECs were seeded in 96-well plates, cultured with POMoV for 1, 3, and 5 days, and CCK-8 reagent was added to each well for 2 h of incubation before measuring absorbance at 450 nm. The results are as follows. Figure 8 As shown in C, the proliferation-promoting effect increased with increasing concentration. The cell proliferation activity of the 40 μg / mL POMoV treatment group was significantly higher than that of the control group, increasing by 1.3 times at 5 days. Example 11: POMoV indirectly promotes angiogenesis (co-culture system)

[0066] Adipocytes were seeded in the upper chamber of a Transwell culture medium and treated with different concentrations of POMoV for 24 h, then replaced with fresh medium. HUVECs were seeded in the lower chamber and co-cultured with the upper chamber for 24 h. The HUVECs from the lower chamber were then removed for migration measurement (scratch assay, method as in Example 10). The results are as follows: Figure 7 As shown in C and 8D, the migration-promoting effect increased with increasing concentration. Among them, after co-culturing with adipocytes pretreated with 40 μg / mL POMoV, the migration rate of HUVECs increased by 2.6 times compared with the co-culture group of untreated adipocytes, indicating that POMoV can indirectly promote endothelial cell migration by regulating adipocyte function. Example 12: Establishment and Treatment of Full-Thickness Skin Defect Model in Diabetic Mice

[0067] Type 2 diabetes was established in 8-week-old male C57 mice by intraperitoneal injection of STZ combined with a high-fat diet. After anesthesia, the back was shaved and disinfected, and a full-thickness skin defect with a diameter of 10 mm was created using a skin punch. Mice were randomly divided into four groups: non-diabetic group, blank control group, insulin group, and POMoV group. The insulin group received an intraperitoneal injection of 1 U / kg / day, while the POMoV group received 60 μL of the corresponding drug solution applied topically to the wound every other day. Wound healing was recorded by photographing the wound on days 0, 4, 6, 10, and 14 using a digital camera. The wound area was calculated using ImageJ software, and the healing rate was calculated as (initial area - current area) / initial area × 100%. Results are as follows: Figure 9 As shown in AB, the healing rate of the POMoV group reached 79.6±7.2% on day 6, which was significantly higher than that of the blank control group (56.3±8.4%) and the insulin group (62.3±8.5%); the healing rate of the POMoV group reached 98.1±1.0% on day 14, and the wound was basically closed. Example 13: Histological Analysis

[0068] Mice in each group were sacrificed on days 7 and 14, respectively. Wound tissue and surrounding skin were collected, fixed in 4% paraformaldehyde, embedded in paraffin, and sectioned. Sections from day 14 were stained with H&E and Masson staining. After dewaxing and antigen retrieval, CD31 primary antibody (1:500) and GLUT4 primary antibody (1:500) were added and incubated overnight at 4°C. After incubation with fluorescent secondary antibody, DAPI was used for counterstaining. The sections were then mounted and observed under a fluorescence microscope. Sections from day 7 were subjected to immunohistochemical staining: IL-6 primary antibody (1:500) and IL-10 primary antibody (1:500) were added and incubated overnight at 4°C. After DAB development and hematoxylin counterstaining, the sections were dehydrated, mounted, and observed and photographed under a microscope. Results are as follows: Figure 9 As shown in CG, the POMoV group showed intact new epidermis, dense collagen deposition, CD31 positive expression increased by 1.3 times compared with the blank control group, GLUT4 fluorescence intensity increased by 2 times, IL-6 positive area decreased by more than 50%, and IL-10 positive area increased by more than 2 times. Example 14: In vitro cell compatibility

[0069] HUVECs, 3T3-L1 adipocytes, and L929 fibroblasts were seeded into 96-well plates and cultured for 24 h. Different concentrations of POMoV (0, 10, 20, 30, 40, 50 μg / mL) were then added to each well for an additional 24 h of culture. After incubation with CCK-8 reagent for 2 h, the absorbance at 450 nm was measured, and cell viability was calculated. Results are as follows: Figure 10As shown in AC, within the concentration range of 10–50 μg / mL, the survival rates of both HUVEC and 3T3-L1 cells remained above 90%, while the survival rate of L929 fibroblasts remained above 80%. Example 15 In vivo biocompatibility

[0070] Healthy ICR mice were randomly divided into two groups (n=5): a control group (saline) and a POMoV group (40 μg / mL). Each group received an intraperitoneal injection of 0.2 mL of the corresponding drug solution. Blood samples were collected from the orbital sinus 14 days later to measure ALT, AST, ALP, CREA, and other indicators. Results are as follows: Figure 10 As shown in Figure D, there were no significant differences in any indicators between the POMoV group and the control group. Mice were sacrificed after blood collection, and the heart, liver, spleen, lungs, and kidneys were harvested, fixed in 4% paraformaldehyde, embedded in paraffin, sectioned, stained with H&E, and observed under a microscope for histopathological changes. The results are as follows: Figure 10 As shown in Figure E, the structures of all organs and tissues in the POMoV group were normal, and no obvious pathological damage was observed, indicating that POMoV has good biocompatibility in vivo.

[0071] The above description represents only some preferred embodiments of the present invention and is not intended to limit the invention in any other way. Any person skilled in the art may make changes or modifications to the disclosed technical content to create equivalent embodiments. However, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention, without departing from the content and spirit of the present invention, shall still fall within the protection scope of the present invention.

Claims

1. A polyoxovanadium molybdate nanocluster, characterized in that, The general chemical formula of the polyoxovanadium molybdate nanoclusters is Na6[V]. 10-x Mo x O 28 ]·nH2O, where 1≤x≤4, and n is the number of water molecules of crystallization, ranging from 8 to 12.

2. The polyoxovanadium molybdate nanoclusters according to claim 1, characterized in that, 2≤x≤3.6, preferably, x=3.

3. The polyoxovanadium molybdate nanoclusters according to claim 1 or 2, characterized in that, The particle size of the polyoxovanadium molybdate nanoclusters is 1–6 nm, preferably 3–5 nm.

4. A method for preparing polyoxovanadium molybdate nanoclusters according to any one of claims 1-3, characterized in that, include: The V source and Mo source are dissolved in a solvent according to the molar ratio of the feed, the pH of the solution is adjusted to acidic, and an acidification precipitation reaction is carried out. After the reaction was completed, the precipitate was collected by centrifugation, washed, and dried to obtain the polyoxovanadium molybdate nanoclusters.

5. The preparation method according to claim 4, characterized in that, The V source is at least one of sodium metavanadate (NaVO3) and ammonium metavanadate (NH4VO3); the Mo source is sodium molybdate (Na2MoO4) or ammonium molybdate ((NH4)6Mo7O3). 24 At least one of the following; Preferably, the molar ratio of the Mo source to the V source is 1:2 to 2:

1.

6. The preparation method according to claim 4 or 5, characterized in that, The reagent used to adjust the pH of the solution to acidity is at least one of hydrochloric acid, nitric acid, and sulfuric acid, preferably hydrochloric acid; Preferably, the pH of the solution is adjusted to 3.0–6.0, more preferably 4.0–5.

0.

7. The preparation method according to any one of claims 4-6, characterized in that, The reaction temperature of the acidification precipitation reaction is 30-80℃, preferably 50-70℃, and the reaction time is 6-48 hours, preferably 12-24 hours. Preferably, the acidification precipitation reaction is carried out under stirring conditions, with a stirring speed of 200–1000 rpm.

8. The preparation method according to any one of claims 4-7, characterized in that, The drying process is either vacuum freeze-drying or conventional vacuum drying, preferably vacuum freeze-drying; The vacuum freeze-drying time is 12 to 48 hours, and the drying temperature is -50 to -80°C. The drying time for the ordinary vacuum drying is 40-60℃ and 12-24 hours.

9. The use of polyoxovanadium molybdate nanoclusters according to any one of claims 1-3 in the preparation of therapeutic compositions for diabetic chronic wounds.

10. A therapeutic composition for chronic diabetic wounds, comprising any one of claims 1-3 vanadium molybdate nanoclusters and an acceptable carrier.

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