Responsive prebiotics-based hydrogen-producing nanoparticles and their applications

Abstract

The present invention belongs to the field of carrier / medical technology for treating metabolic diseases, and specifically relates to responsive prebiotics-mimicking hydrogen production nanoparticles and their applications. The present invention constructs a plant exosome bionic oral nanoformulation HMS / A@GE, which fully combines the advantages of exosomes from edible prebiotics and mesoporous silica carriers, overcomes the limitations of existing microecological therapies, realizes the combination of microecological therapy and antioxidant therapy, and synergistically exerts the effects of lowering blood sugar and reducing fat, providing a safe, economical and controllable strategy for treating diabetes based on regulating intestinal microecology. The HMS / A@GE of the present invention improves the intestinal Lactobacillus spp. It can promote the release of GLP-1 from intestinal L cells and activate intestinal AhR receptors, promote the release of IL-22, improve insulin resistance and pancreatic β-cell function, and ultimately play a role in lowering blood sugar and reducing fat.

Description

Technical Field

[0001] The present invention belongs to the field of carrier / medicine technology for treating metabolic diseases, and specifically relates to responsive prebiotics-mimicking hydrogen production nanoparticles and applications thereof. Background Art

[0002] Type 2 diabetes mellitus (T2DM), as a multi-system, complex metabolic dysfunction disease, has become one of the most serious health problems in the world, accounting for more than 90% of all diabetes. T2DM is closely related to obesity, non-alcoholic fatty liver disease and cardiovascular disease. The prevalence of type 2 diabetes has become a heavy health burden on society. The pathophysiological characteristics of T2DM are highly related to reduced insulin sensitivity and subsequent pancreatic cell decompensation. It is generally believed that insulin injection and other oral drug treatments such as metformin, thiazolidinediones, sulfonylureas, sodium-dependent glucose transporter (SGLT2) inhibitors and dipeptidyl peptidase-4 (DPP-4) inhibitors are effective. However, these treatments have gradually become ineffective due to the development of insulin resistance and drug tolerance. In addition, there are side effects such as gastrointestinal reactions, hypoglycemia, renal impairment, and pancreatitis. Therefore, there is an urgent need to develop more effective and safer strategies that can control blood sugar, improve insulin resistance, and ultimately improve the diabetic disease state.

[0003] Gas therapy has attracted wide attention in disease diagnosis and treatment due to its strong diffusivity, high selectivity and biosafety. 2 ), nitric oxide (NO), carbon monoxide (CO) and hydrogen sulfide (H 2 S). As a gas with high safety, good bioreducibility and homeostasis regulation, hydrogen can easily pass through the intestinal mucosal barrier to act on the body, and has shown significant therapeutic effects in oxidative stress / inflammatory related diseases such as cancer, atherosclerosis, diabetes and pancreatitis, some of which have been confirmed in clinical trials. The main treatment modes are inhalation of hydrogen, oral hydrogen-rich water, injection of hydrogen-rich salt water, hydrogen bath, oral hydrogen-producing capsules and food, etc. The mechanism of action of hydrogen in reducing fat and blood sugar is mainly anti-oxidation, anti-apoptosis, regulation of liver FGF21 expression and regulation of intestinal flora composition. Summary of the invention

[0004] The present invention constructs a plant exosome bionic oral nanoformulation HMS / A@GE, which fully combines the advantages of exosomes from edible prebiotics and mesoporous silica carriers, overcomes the limitations of existing microecological therapy, realizes the combination of microecological therapy and antioxidant therapy, and synergistically exerts the effects of lowering blood sugar and reducing fat, providing a safe, economical and controllable strategy for treating diabetes based on regulating intestinal microecology.

[0005] The present invention adopts the following technical solutions:

[0006] The present invention first provides responsive prebiotic hydrogen-producing nanoparticles HMS / A@GE, which are prepared by the following method:

[0007] (1) Preparation of HMS using soft template method:

[0008] The micelle template was prepared by using hexadecyltrimethylammonium bromide (CTAB) surfactant and tetraethyl orthosilicate (TEOS) as silicon source. The HMS carrier was obtained by etching with anhydrous sodium carbonate and removing the template with concentrated hydrochloric acid.

[0009] (2) Preparation of HMS-NH 2 :

[0010] The HMS after template removal was amino-modified with 3-aminopropyltriethoxysilane (APTES) to obtain amino-modified hollow mesoporous silica nanoparticles. Specifically, the HMS nanoparticles were ultrasonically dispersed in anhydrous ethanol, 3-aminopropyltriethoxysilane was added and stirred at room temperature, and then heated to reflux at 70°C, centrifuged, rinsed, and dried to obtain HMS-NH 2 .

[0011] (3) Preparation of HMS / A:

[0012] Weigh A and add HMS-NH 2 The solution was stirred at 70°C for adsorption equilibrium (6 to 12 hours), and then a large amount of methanol solvent was removed, centrifuged, and the precipitate was collected to obtain HMS / A.

[0013] Among them, A and HMS-NH 2 The mass ratio is 1:4.

[0014] (4) GE exosome extraction:

[0015] Differential centrifugation was used to extract ginger exosomes (GE): fresh ginger was washed, juiced, and filtered with gauze to obtain a crude extract. The crude extract was centrifuged in a 4°C centrifuge, the supernatant was taken, and the second centrifugation was performed, and the supernatant was taken for the third centrifugation. The supernatant was taken as GE.

[0016] The gravitational acceleration of the second centrifugation is greater than the gravitational acceleration of the first centrifugation, and the gravitational acceleration of the third centrifugation is greater than the gravitational acceleration of the second centrifugation.

[0017] (5) Preparation of HMS / A@GE:

[0018] The HMS / A and GE were mixed, ultrasonicated in an ice bath, and then incubated in a 37°C incubator. The mixture was centrifuged, the precipitate was collected, and dried to obtain HMS / A@GE.

[0019] The present invention also found that HMS / A@GE can be used to improve glucose tolerance and insulin resistance in T2DM mice, thereby improving glucose homeostasis. The present invention also found that HMS / A@GE can be used to alleviate blood lipid and liver fatty degeneration in T2DM mice, reduce liver and subcutaneous fat deposition, and stimulate insulin secretion. Therefore, HMS / A@GE can be used to prepare blood sugar and fat-reducing drugs.

[0020] The present invention found that HMS / A@GE can also increase the level of anti-inflammatory cytokines and inhibit the secretion of pro-inflammatory cytokines, so HMS / A@GE can be used to prepare anti-inflammatory drugs.

[0021] Furthermore, the present invention found that HMS / A@GE also has antioxidant ability, therefore, HMS / A@GE can be used to prepare antioxidant drugs.

[0022] Specifically, the results of the percentage of weight gain of mice and the blood glucose value at the end of the experiment showed that HMS / A@GE had a good effect on hypoglycemia and weight loss; the ITT results showed that HMS / A@GE improved insulin sensitivity. The results of serum insulin and HOMA-IR showed that HMS / A@GE had the effect of improving insulin resistance. The results of ALT, AST, TG, and TC levels in the serum of each group of mice showed that HMS / A@GE could significantly improve lipid metabolism. HMS / A@GE could significantly improve the degree of fatty degeneration of the liver, reduce the area of ​​inguinal fat, and increase the expression of UCP-1. HMS / A@GE could increase the level of anti-inflammatory factors (IL-10) and reduce the level of pro-inflammatory cytokines (IL-6, IL-1β, TNF-α, LPS). The results of co-staining of Insulin and Glucagon showed that HMS / A@GE could improve the function of pancreatic β cells and reduce the dedifferentiation of pancreatic β cells. In addition, HMS / A@GE could also reduce the content of serum malondialdehyde (MDA), indicating that it has antioxidant capacity.

[0023] The beneficial effects of the present invention are:

[0024] The HMS / A@GE of the present invention improves the intestinal Lactobacillus spp. It can promote the release of GLP-1 from intestinal L cells and activate intestinal AhR receptors, promote the release of IL-22, improve insulin resistance and pancreatic β-cell function, and ultimately play a role in lowering blood sugar and reducing fat. Figure 8 shown. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 Characterization of prebiotic-responsive hydrogen-producing nanoparticles HMS / A@GE.

[0026] Figure 2 This is a graph showing the results of the fat-reducing and blood sugar-lowering effects of HMS / A@GE in a T2DM mouse model.

[0027] Figure 3 The results of HMS / A@GE improving liver steatosis, inguinal fat deposition and pancreatic islet function in the T2DM mouse model.

[0028] Figure 4 This is the result of HMS / A@GE regulating the intestinal flora of T2DM mouse model.

[0029] Figure 5 The results of HMS / A@GE improving the inflammation level in T2DM mouse model.

[0030] Figure 6 This is the result of HMS / A@GE improving the oxidative status of T2DM mouse model.

[0031] Figure 7 This is a diagram showing the mechanism by which HMS / A@GE regulates intestinal flora and tryptophan metabolites to reduce blood sugar and fat.

[0032] Figure 8 Diagram of the preparation steps and action mechanism of HMS / A@GE. DETAILED DESCRIPTION

[0033] The present invention is described in more detail below through specific implementation modes to facilitate understanding of the technical solution of the present invention, but is not intended to limit the protection scope of the present invention.

[0034] The A described in the embodiment is ammonia borane, and its CAS number is: 13774-81-7.

[0035] Example 1 Preparation of Responsive Prebiotic Hydrogen-producing Nanoparticles HMS / A@GE

[0036] The preparation steps are as follows ( Figure 8 ):

[0037] (1) Preparation of hollow mesoporous silica nanoparticles (HMS)

[0038] The HMS carrier was prepared by the soft template method: 74 mL of anhydrous ethanol and 10 mL of distilled water were measured and poured into a conical flask, stirred and mixed on a turntable, 3.15 mL of ammonia water was added, stirred for 15 min, 6.00 mL of tetraethyl orthosilicate (TEOS) was measured by a pipette, and quickly added to the above mixture. Stir for 1 h, centrifuge (11000 rpm, 10 min), collect the precipitate, wash it with anhydrous ethanol 3 times, and dry it in an oven to obtain solid silica spheres.

[0039] Weigh 300 mg of solid silica spheres, add them to a conical flask containing 60 mL of distilled water, and ultrasonicate. Weigh 450 mg of hexadecyltrimethylammonium bromide (CTAB) and dissolve it in a mixed solution consisting of 90 mL of distilled water and 90 mL of anhydrous ethanol. Use a pipette to measure 1.65 mL of ammonia water and pour it into the above solution. Stir at room temperature for 10 min, add the solid silica spheres that have been ultrasonically dispersed, stir at room temperature for 30 min, accurately add 0.75 mL of TEOS, stir at room temperature for 6 h, centrifuge (11000 rpm, 15 min), collect the precipitate, and rinse it with anhydrous ethanol and distilled water twice. Disperse the above precipitate with 60 mL of distilled water, pour it into a conical flask, add 1272 mg of anhydrous sodium carbonate, stir in a 50℃ water bath for 10 h, centrifuge (11000 rpm, 15 min), and wash the precipitate with distilled water 3 times. Disperse the precipitate with 300 mL of methanol, pour it into a round-bottom flask, add 10 mL of concentrated hydrochloric acid, reflux and stir in a water bath at 80°C for 24 h, centrifuge (11000 rpm, 15 min), wash the precipitate three times with anhydrous ethanol, and dry it in an oven to obtain HMS.

[0040] (2) Preparation of amino-modified hollow mesoporous silica nanoparticles (HMS-NH 2 )

[0041] The HMS after template removal was amino-modified with 3-aminopropyltriethoxysilane (APTES) to obtain amino-modified hollow mesoporous silica nanoparticles. The specific operation steps are as follows: 200 mg of HMS nanoparticles were ultrasonically dispersed in 500 mL of anhydrous ethanol, 800 μL of 3-aminopropyltriethoxysilane was added at room temperature and stirred for 10 min, then heated to reflux at 70 ° C for 14 h, centrifuged (11000 rpm, 15 min), and then rinsed with anhydrous ethanol for 3 times, and dried in an oven to obtain HMS-NH 2 .

[0042] (3) Preparation of HMS loaded with ammonia borane (HMS / A)

[0043] HMS / A was prepared by solvent evaporation method: 10 mg of A was weighed and 8 mL of 5 mg / mL HMS-NH 2 After the suspension is adsorbed at 70°C and 500 rpm for 6 to 12 hours, the solvent is evaporated to remove a large amount of methanol solvent, and the suspension is centrifuged to collect the precipitate to obtain HMS / A.

[0044] (4) Extraction of ginger exosomes (GE)

[0045] Extract GE by differential centrifugation: squeeze fresh ginger juice and filter with gauze to obtain crude extract. Centrifuge the crude extract at 3000g for 30min at 4℃, take the supernatant, continue centrifugation (13000g, 30min), take the supernatant and centrifuge (17000g, 60min), the supernatant is GE.

[0046] (5) Preparation of HMS / A@GE

[0047] The HMS / A prepared in step (3) was mixed with the GE prepared in step (4), and ultrasonicated in an ice bath at 20% power (ultrasonication for 5 seconds, pause for 2 seconds mode), each round of ultrasonication for 2 minutes, with an interval of 1 minute, for a total of 6 rounds of ultrasonication, and then incubated in a 37°C incubator for 30 minutes, centrifuged (11000 rpm, 5 minutes), and the precipitate was collected and dried to obtain HMS / A@GE.

[0048] Example 2 Characterization of GE, HMS, and Responsive Prebiotic Hydrogen Production Nanoparticles HMS / A@GE

[0049] The morphology of the prepared GE was observed using a transmission electron microscope (TEM). Figure 1 As shown in a, GE is a spherical bowl-shaped nanovesicle with an intact membrane and an average diameter of about 200 nm.

[0050] TEM examination showed that the HMS had a uniform morphology and a size of approximately 250 nm ( Figure 1 b). After GE modification, the pores of HMS are covered and the edges of HMS@GE become smooth ( Figure 1 c). To further verify the coating of GE, HMS and GE were labeled with FITC and TRITC, respectively. The co-localization of FITC-labeled HMS and TRITC-labeled GE was observed by fluorescence imaging ( Figure 1 d). At the same time, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to analyze the proteins on the surface of GE and HMS@GE. Figure 1 As shown in Figure e, the protein bands of GE and HMS@GE are similar. These results verify the feasibility of GE encapsulating HMS.

[0051] In addition, the present invention also evaluated the characteristics of HMS / A@GE. The zeta potential and particle size were measured using a Malvern instrument. Figure 1 f shows that the ζ-potential value of GE is about -14.5 mV. HMS and HMS-NH 2 The ζ potential values ​​of HMS / A@GE were -22.7 mV and 26.7 mV respectively. After GE modification, the ζ potential value of HMS@GE became -20 mV. After loading A, the ζ potential value became -24.0 mV. The reversal of ζ potential again indicated that HMS / A@GE was successfully prepared. The particle size results are shown in Figure 1As shown in Figure g, the particle size of GE is 265.4 nm and the polydispersity index (PDI) is 0.195. The particle size of HMS is 258 nm and the PDI is 0.417. After loading A, the particle size becomes 356.4 nm and the PDI is 0.193. The drug loading of A is 8.2% as determined by ICP-OES. The specific structural parameters of HMS and HMS / A@GE were tested using a specific surface area meter, as shown in Figure 2. Figure 1 h and Figure 1 As shown in Fig. 1, the specific surface area and pore volume of HMS decreased with the loading of A and GE. In addition, the acid response of HMS / A@GE was detected by hydrogen microelectrode. 2 Release behavior. Figure 1 As shown in j, after HMS / A@GE was incubated with simulated gastric fluid, H 2 The release rate and concentration were significantly enhanced.

[0052] In summary, these results further verified the successful preparation of acid-sensitive H 2 HMS / A@GE unleashes performance.

[0053] Example 3 Application of Responsive Prebiotic Hydrogen-producing Nanoparticles HMS / A@GE

[0054] 3.1 HMS / A@GE improves glucose homeostasis in T2DM mice

[0055] The animal model used in the experiment was the T2DM mouse model: 7-week-old male C57BL / 6J mice were adaptively fed for one week and then fed a high-fat diet (H10060 with a fat energy supply ratio of 60%) for 8 weeks. After the high-fat diet ended, the mice were fasted for 6 hours and then intraperitoneally injected with 35 mg / kg of STZ for 5 consecutive days. Random blood glucose was observed 72 hours after the injection. The model was successfully constructed when the random blood glucose was greater than 16.8 mmol / L. After the model was established, it was maintained with a high-fat diet (H10060 with a fat energy supply ratio of 60%). The mice were divided into 5 groups, namely: ①G1: blank control group, fed only with high-fat feed; ②G2: hollow mesoporous silica nanoparticles group (HMS group), gavage with 5 mg / kg HMS suspension; ③G3: ammonia borane group (also known as free A group): gavage with 0.4 mg / kg A solution; ④G4: ginger exosome-coated hollow mesoporous silica group (HMS@GE group), gavage with 5 mg / kg HMS@GE suspension; ⑤G5: HMS / A@GE group, gavage with 5 mg / kg HMS / A@GE suspension.

[0056] The therapeutic effect of HMS / A@GE was then evaluated in T2DM mice. Body weight was monitored during treatment, as shown in Figure 2As shown in a, the body weight of the blank control group increased, while A and HMS@GE moderately decreased the body weight. HMS / A@GE could significantly inhibit the body weight gain induced by high-fat diet.

[0057] This example also records fasting blood glucose, and finds that the blood glucose level of mice treated with HMS / A@GE is significantly decreased ( Figure 2 b).

[0058] One week before the end of the experiment, IPGTT was used to detect glucose homeostasis in mice. Figure 2 c It can be seen that the glucose tolerance of T2DM mice was significantly impaired, and HMS nanoparticles had no effect on glucose tolerance. In contrast, both HMS@GE and HMS / A@GE could improve glucose tolerance. Figure 2 d, The area under the IPGTT curve (AUC) of diabetic mice treated with HMS / A@GE was smaller than that of diabetic mice treated with the blank control group.

[0059] The ITT method was used to evaluate insulin resistance in T2DM mice. Figure 2 As shown in e and 2f, only HMS / A@GE can improve the insulin sensitivity of T2DM mice, while A or HMS@GE has almost no effect on the insulin resistance of T2DM mice. A, HMS@GE and HMS / A@GE can all reduce the fasting serum insulin level of T2DM mice, among which HMS / A@GE has the strongest reducing effect ( Figure 2 g). Therefore, the results of homeostasis model assessment (HOMA) to evaluate insulin sensitivity showed that HMS / A@GE treatment could improve insulin resistance in T2DM mice ( Figure 2 h).

[0060] The above results indicate that HMS / A@GE treatment can effectively improve glucose tolerance and insulin resistance in T2DM mice.

[0061] 3.2HMS / A@GE effectively alleviates blood lipid and liver fatty degeneration and reduces liver and subcutaneous fat deposition in T2DM mice

[0062] Compared with the blank control group, free A and HMS@GE nanoparticles could significantly reduce the liver fat coefficient ( Figure 2 i). Compared with other groups, the liver weight of the HMS / A@GE group decreased the most. Figure 2 As shown in j, HMS / A@GE treatment can reduce serum ALT and AST, two representative liver function indicators, in T2DM mice.

[0063] Further serum biochemical measurements showed that HMS / A@GE also reduced the levels of TG and TC ( Figure 2 k), suggesting that HMS / A@GE may affect lipid metabolism.

[0064] The results of H&E staining of liver pathological sections are shown in Figure 3 a, T2DM mice developed significant hepatic steatosis, but HMS / A@GE eliminated the accumulation of lipid droplets in the liver. Figure 3 As shown in Fig. 3b and 3f, Oil Red O staining of liver sections also showed that free A and HMS@GE significantly reduced the staining area, indicating that the cholesterol content in the liver was significantly reduced. Interestingly, the Oil Red O area of ​​the HMS / A@GE group was still smaller than that of the above two groups, confirming the excellent effect of HMS / A@GE in alleviating liver fatty degeneration.

[0065] This example evaluates the adipocyte size and UCP-1 expression in iWAT. Figure 3 As shown in Figures 3c and 3g, the adipocyte sizes of the blank control group and the HMS group were similar. The adipocyte sizes of the free A group and the HMS@GE group both tended to decrease, but only the adipocyte size of the free A group decreased significantly. In contrast, the average size of adipocytes in mice in the HMS / A@GE group was found to be significantly reduced.

[0066] UCP-1, a key protein for thermogenesis, was detected by detailed tissue analysis. Figure 3 d. Figure 3 h shows that free A, HMS@GE and HMS / A@GE all induced the expression of UCP-1 to varying degrees. The expression of UCP1 in the HMS / A@GE group was the highest.

[0067] The above results indicate that HMS / A@GE significantly improved hepatic steatosis and obesity, which was associated with decreased hepatic fat accumulation and increased UCP-1 expression.

[0068] In addition, histological analysis of pancreatic islets was performed by immunofluorescence staining. Figure 3 e and Figure 3 As shown in Fig. i, compared with other groups, HMS / A@GE treatment significantly increased the proportion of β-cells and decreased the proportion of α-cells in T2DM mice.

[0069] Taken together, these results suggest that HMS / A@GE has the ability to reduce liver and subcutaneous fat deposition and stimulate insulin secretion in T2DM mice.

[0070] 3.3 Effects of HMS / A@GE on intestinal flora

[0071] Shannon index, ASV number and Pielou index were used to evaluate the diversity, richness and uniformity of intestinal flora, respectively, to analyze the α-diversity of intestinal flora. The results showed that the diversity of intestinal flora among the five groups ( Figure 4a) Richness Figure 4 b) and uniformity ( Figure 4 c) There is no difference.

[0072] The principal component analysis based on Jaccard distance was used to evaluate the beta diversity of intestinal flora. The structures of the five groups of intestinal flora can be distinguished by Jaccard distance ( Figure 4 d).

[0073] In addition, among the five groups, the feces of mice in group G5 (i.e., HMS / A@GE) Lactobacillus The average relative abundance of bacterial genus was higher ( Figure 4 e). LEfSe was further used to select the differential ASVs between the two groups of mice, with LDA score>3 as the threshold. Among these five groups, mice in group G5 were significantly enriched for ASVs belonging to Lactobacillus murinus Four ASVs (ASV55, ASV40, ASV35, ASV31, ASV9) and Lactobacillus Two ASVs (ASV20, ASV34) and Akkermansia muciniphila ASV121, etc. ( Figure 4 f and Figure 4 g).

[0074] 3.4 Improvement of inflammatory response and oxidative stress by HMS / A@GE

[0075] The concentration of inflammatory markers in serum was further detected. The levels of IL-10, IL-1β, IL-6, and TNF-α in each group were detected by ELISA (kit purchased from Shanghai ELISA Biotechnology Co., Ltd.). The serum of mice was collected from the orbital vein and measured according to the instructions of the kit.

[0076] like Figure 5 As shown in a, compared with the blank control group and nano-HMS group, HMS / A@GE treatment significantly increased the level of anti-inflammatory cytokine IL-10. In addition, HMS / A@GE can significantly inhibit the secretion of pro-inflammatory cytokines such as IL-1β, IL-6, TNF-α, and IL-17A ( Figure 5 b, 5c, 5d, 5e). Among these cytokines, HMS / A@GE significantly reduced the production of TNF-α and IL-17A. The changes in these cytokines reflect that HMS / A@GE effectively improved the systemic inflammatory state of mice. In addition, HMS / A@GE can also reduce serum malondialdehyde (MDA) content, indicating that it has antioxidant capacity ( Figure 6 ).

[0077] 3.5 Study on the mechanism of HMS / A@GE in reducing blood sugar and fat

[0078] The correlation analysis between ASVs significantly enriched in HMS / A@GE and indicators such as body weight was performed ( Figure 7 a), the results showed that ASV9, ASV31, ASV35, ASV40, ASV87, and ASV126 were significantly negatively correlated with TC; ASV20, ASV35, ASV40, ASV87, and ASV126 were significantly negatively correlated with IL-17A. ASV34 and ASV55 were significantly negatively correlated with the area under the ITT curve. Further research on fecal targeted tryptophan metabolomics showed that HMS / A@GE could significantly increase the concentrations of indole (IND), indole-3-acetic acid (IAA), 5-hydroxyindoleacetic acid (5-HIAA), N-acetyl serotonin (NAS), and quinolinic acid (QA) in feces, while significantly reducing the concentration of kynurenine (KYN) ( Figure 7 b). Perform correlation analysis between these differential metabolites and indicators such as body weight ( Figure 7 c) The results showed that IAA, NAS and QA were significantly negatively correlated with body weight; IAA, 5-HIAA, NAS and TC were significantly negatively correlated. KYN was significantly positively correlated with HOMA-IR index, TG, TNF-α and IL-17A. Except for NAS, the other differential tryptophan metabolites were significantly negatively correlated with TNF-α.

[0079] The above results indicate that IND, IAA, 5-HIAA, NAS and QA are key metabolites of intestinal flora. In addition, the expression of intestinal tight junction protein ZO-1 and Occludin showed that HMS / A@GE can increase the expression level of Occludin ( Figure 7 d). The mRNA levels of colon AhR and IL-22 were further measured. The results showed that HMS / A@GE had no significant effect on the mRNA level of AhR, but significantly increased the mRNA level of IL-22 ( Figure 7 e). Further colon immunofluorescence results showed that HMS / A@GE could increase the expression level of colon AhR ( Figure 7 f).

[0080] The embodiments described above are only preferred embodiments of the present invention and are not intended to limit the scope of implementation of the present invention. Therefore, any equivalent changes or modifications made according to the structures, features and principles described in the patent scope of the present invention should be included in the scope of the patent application of the present invention.

Claims

1. Hydrogen-generating nanoparticles HMS / A@GE, characterized in that: The nanoparticles were prepared by the following method: (1) Preparation of HMS by soft template method: using hexadecyltrimethylammonium bromide surfactant to prepare micelle template, using tetraethyl orthosilicate as silicon source, and obtaining HMS carrier by etching and removing template, wherein the HMS is hollow mesoporous silica nanoparticles; (2) Preparation of HMS-NH2: The HMS after template removal was amino-modified with 3-aminopropyltriethoxysilane to obtain amino-modified hollow mesoporous silica nanoparticles HMS-NH2; (3) Preparation of HMS / A: Weigh A, add fully dispersed HMS-NH2 solution, stir at 70°C for adsorption equilibrium, remove the solvent, centrifuge, collect the precipitate, and obtain HMS / A; A is ammonia borane; the mass ratio of A to HMS-NH2 is 1:4; (4) GE exosome extraction: Differential centrifugation was used to extract ginger exosome GE: the ginger was washed, juiced, and filtered to obtain a crude extract; the crude extract was centrifuged in a 4°C centrifuge, the supernatant was taken, centrifuged, the supernatant was taken, and the supernatant was taken again as GE; the first centrifugation was performed at 3000 g, the second centrifugation was performed at 13000 g, and the third centrifugation was performed at 17000 g; (5) Preparation of HMS / A@GE: HMS / A and GE were mixed, sonicated on ice, incubated at 37°C, centrifuged, the precipitate was collected, and dried to obtain HMS / A@GE.

2. The hydrogen-generating nanoparticle HMS / A@GE according to claim 1, characterized in that: Step (2) is specifically as follows: ultrasonically dispersing HMS nanoparticles in anhydrous ethanol, adding 3-aminopropyltriethoxysilane and stirring at room temperature, then heating and refluxing at 70° C., centrifuging, eluting, and drying to obtain HMS-NH2.

3. Use of the hydrogen-producing nanoparticle HMS / A@GE according to any one of claims 1 to 2 in the preparation of hypoglycemic drugs.

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