Application of micromolecular antioxidant to alleviation of type 2 diabetes mellitus vascular injury

Abstract

The invention belongs to the technical field of medicines, and particularly relates to application of a small-molecule antioxidant to alleviation of type 2 diabetes vascular injury. In order to develop an effective scheme for dealing with vascular injury of a diabetic patient, research finds that Lithospermoside can specifically inhibit XO in a cell membrane anchoring state and effectively reduce generation of ROS in thoracic aorta, so that BUN, E-selectin, MDA and UACR are remarkably reduced, meanwhile, the fluorescence intensity of nitrotyrosine staining is inhibited, and the blood vessel injury of the diabetic patient is inhibited. Therefore, vascular endothelial injury under the state of type 2 diabetes accompanied by continuous hyperglycemia is relieved, the antioxidation effect is exerted to slow down vascular injury of diabetes and protect blood vessels, and an effective material basis and a new thought are provided for prevention and treatment of vascular injury of type 2 diabetes.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of medicine, and particularly relates to application of a small-molecule antioxidant in slowing down blood vessel injury of type 2 diabetes. BACKGROUND

[0002] Patients with type 2 diabetes have an increased risk of cardiovascular disease (CVD) and are more likely to die from CVD. Oxidative stress is considered to be one of the causes of blood vessel injury in patients with diabetes. Factors that trigger oxidative stress mainly include increased generation of glycosylation end products, activation of the polyol pathway, enhanced activity of protein kinase C and xanthine oxidase (XO), and decoupling of endothelial nitric oxide synthase. Therefore, inhibition of oxidative stress is crucial for preventing the progression of vascular disease. However, there is currently a lack of effective strategies to deal with CVD.

[0003] Xanthine oxidoreductase (XOR) catalyzes two key reaction steps: one is the synthesis of xanthine from hypoxanthine, and the other is the further synthesis of uric acid from xanthine. Under physiological conditions, XOR mainly exists in the form of xanthine dehydrogenase (XDH), which uses nicotinamide adenine dinucleotide as an electron acceptor. However, under certain conditions, XDH can be converted into XO through limited proteolysis. The latter uses molecular oxygen as an electron acceptor and generates superoxide anions during the reaction. XO is also believed to be associated with endothelial dysfunction, hypertension and heart failure, which is mainly due to the active oxygen (ROS) produced by it, both intracellularly and extracellularly. Among them, the extracellular active oxygen has two forms: one is free, which can exist in circulating blood; the other is anchored, which is combined with the proteoglycans on the surface of endothelial cells.

[0004] Studies have shown that the injection of allopurinol into cholesterol-loaded rabbits improves the vascular endothelial cell-dependent relaxation response, and this effect is not related to the serum uric acid concentration. It has also been reported that the XOR activity in the diseased coronary arteries of patients with coronary artery disease is significantly higher than that in the healthy control group. These research results suggest that the overactivation of XOR may be involved in the process of functional abnormalities of large artery endothelial cells.

[0005] Lithospermoside is extracted from the roots of Lithospermum purpurocaeruleum and L. officinale of the Boraginaceae family, and Thalictrum rugosum and T. dasycarpum of the Ranunculaceae family, and has antioxidant and antitumor effects.

[0006] Although Lithospermoside (a small molecule antioxidant) has been used to treat hyperuricemia and gout, studies have found that the administration of febuxostat or Lithospermoside to db / db mice can inhibit albuminuria, but only Lithospermoside shows a dose-dependent effect, and this effect is related to the activity of XOR in plasma. Plasma XOR can be anchored on the proteoglycans on the surface of endothelial cells, resulting in the local production of superoxide anions, which react with nitrite (NO) in blood vessels to generate peroxynitrite with cytotoxicity.

[0007] It is speculated that Lithospermoside can improve vascular endothelial function by inhibiting circulating XOR, anchored XOR, and XOR in vascular endothelial cells; however, the degree of its association with these three types of XOR is currently unknown. In addition, there is currently no research indicating that Lithospermoside is associated with type 2 diabetes. SUMMARY

[0008] In order to overcome the shortcomings of the prior art, the present application explores the effect of the small molecule antioxidant Lithospermoside on vascular ROS in type 2 diabetic rats. In a streptozotocin-induced type 2 diabetic rat model, XO increases or is activated, and the amount of ROS generated in large blood vessels (thoracic aorta) increases; Lithospermoside can effectively reduce this reaction, thereby protecting blood vessels in a diabetic state from damage.

[0009] In order to achieve the above purpose, the technical scheme adopted by the present application is:

[0010] The present application provides the use of Lithospermoside in the preparation of a drug for slowing down the vascular damage of type 2 diabetes.

[0011] Preferably, the Lithospermoside exerts an antioxidant effect by inhibiting XO in an anchored state, thereby slowing down the vascular damage of type 2 diabetes and protecting blood vessels.

[0012] The present application has been researched and confirmed that Lithospermoside can improve the vascular endothelial damage of the aorta in a type 2 diabetic animal model by inhibiting XO in an anchored state. The research results suggest that Lithospermoside may have a protective effect on patients with microvascular diseases caused by type 2 diabetes. Next, prospective clinical trials need to be carried out to evaluate the protective effect of lithospermoside on large vessel diseases caused by type 2 diabetes.

[0013] Preferably, the effective dose of the Lithospermoside is 1-5 mg / kg.

[0014] Preferably, the medicine further comprises a pharmaceutically acceptable excipient.

[0015] More preferably, the excipient comprises at least one of an excipient, a propellant, a solubilizer, a co-solvent, an emulsifier, a colorant, a binder, a disintegrant, a filler, a lubricant, a wetting agent, an osmotic pressure regulator, a stabilizer, a glidant, a flavoring agent, a preservative, a suspending agent, a coating material, an aromatic agent, an anti-adhesion agent, an integrating agent, a penetration enhancer, a pH regulator, a buffer, a plasticizer, a surfactant, a foaming agent, an antifoaming agent, a thickening agent, an inclusion agent, a humectant, an absorbent, a diluent, a flocculating agent and a deflocculating agent, an antioxidant, an adsorbent, a filter aid, a release retardant.

[0016] Preferably, the dosage form of the medicine comprises a tablet, a capsule, an aerosol, a pill, a powder, a solution, a suspension, an emulsion, a granule, a liposome, a transdermal agent or a suppository.

[0017] Compared with the prior art, the present application has the following beneficial effects:

[0018] The present application finds that the small molecule antioxidant Lithospermoside can specifically inhibit the XO in the anchored state of the cell membrane, effectively reduce the generation of ROS in the thoracic aorta, and thus significantly reduce blood urea nitrogen (BUN), E-selectin, urinary malondialdehyde (MDA) and urinary albumin / creatinine ratio (UACR), while inhibiting the fluorescence intensity of nitrotyrosine staining, thereby reducing the vascular endothelial injury in type 2 diabetes with persistent hyperglycemia, playing an antioxidant role, and slowing down the diabetic vascular injury and protecting the blood vessels.

[0019] This breakthrough discovery of the present application can improve diabetes-related vascular lesions in a targeted manner, provides a highly targeted solution for improving diabetes-related vascular lesions, and provides an effective material basis and a new idea for the prevention and treatment of type 2 diabetes vascular injury. At the same time, the small molecule antioxidant Lithospermoside is expected to be developed as a new drug or intervention preparation, which can be applied in clinical treatment through oral administration, injection and other convenient methods, and provides more accurate and efficient treatment options for type 2 diabetes patients. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1 The purine body concentration in the plasma; A: the column chart of the detection results of the concentration of uric acid; B: the column chart of the detection results of hypoxanthine; C: the column chart of the detection results of xanthine; D: the column chart of the detection results of xanthine oxidoreductase (XOR) activity.

[0021] Figure 2For the effect of Lithospermoside on anchoring XOR, the effect of Lithospermoside on anchoring XOR was detected by chemiluminescence method; the data in the figure was based on the standard (5 μU / mL xanthine oxidase), and after dry weight correction, the difference in luminescence intensity between the sample group and the blank control group was shown; black column chart: no heparin added; gray column chart: 1000 U / mL heparin added; the figure shows the detection results column chart of superoxide production of different groups, the vertical coordinate is the superoxide production (measured by the standard ratio of chemiluminescence method), and the horizontal coordinate is different groups, including Control group, Streptozotocin (STZ) group, and different doses (0.3 mg / kg, 1 mg / kg, 3 mg / kg) of Lithospermoside group.

[0022] Figure 3 For the effect of Lithospermoside on intracellular XOR, the effect of Lithospermoside on intracellular superoxide dismutase (XOR) was detected by dihydroethidium oxidation fluorescence staining method; A: observation of dihydroethidium fluorescence signal in aortic tissue section under confocal microscope, B: quantitative analysis of dihydroethidium fluorescence intensity in the intima region, C: quantitative analysis of dihydroethidium fluorescence intensity in the media region.

[0023] Figure 4 For the effect of Lithospermoside on cytotoxicity, the effect of Lithospermoside on cytotoxicity was detected by fluorescence immunostaining method (aortic frozen sections were immunostained with anti-3-NT antibody and CD31 antibody, and co-stained with DAPI); A: observation of fluorescence signals of 3-NT (green), CD31 (red) and DAPI (blue) in aortic tissue sections under confocal microscope, B: quantitative analysis of fluorescence intensity in the overlapping region of CD31 and 3-NT, scale bar 50 μm. DETAILED DESCRIPTION

[0024] The specific embodiments of the present application will be further described below. It should be noted that the description of these embodiments is used to help understand the present application, but does not constitute a limitation on the present application. In addition, the technical features involved in each embodiment of the present application described below can be combined with each other as long as they do not conflict with each other.

[0025] The experimental methods in the following examples are all conventional methods unless otherwise specified. The test materials used in the following examples are all commercially available unless otherwise specified.

[0026] In the following examples, the composition of the Krebs solution was as follows: 137.4 mM NaCl, 5.9 mM KCl, 1.2 mM MgS04, 2.5 mM CaCl2, 15.5 mM NaHC03, 1.2 mM KH2P04, and 11.5 mM glucose. The solution was gassed with 95% 02and 5% C02, and the pH was adjusted to 7.3-7.4. Modified Krebs-N-(2-hydroxyethyl)piperazine N'-2-ethanesulfonic acid (HEPES) buffer contained 99 mM NaCl, 4.7 mM KCl, 1.9 mM CaCl2, 1.2 mM MgS04, 20 mM HEPES, 1.03 mM KH2P04, 25 mM NaHC03, and 11.1 mM glucose (pH 7.4). Krebs-Henseleit buffer contained 118.3 mM NaCl, 4.7 mM KCl, 2.5 mM CaCl2, 1.2 mM MgS04, 1.2 mM KH2P04, 25 mM NaHC03, and 11 mM glucose (pH 7.4).

[0027] XOR exists both in the intracellular and extracellular, inducing vascular injury by generating ROS. Based on this, the present application uses an animal model of type 2 diabetes with persistent hyperglycemia to study the effect of a small molecule antioxidant Lithospermoside on ROS and its mechanism of action. 6-week-old male Sprague-Dawley rats were induced to diabetes by 50mg / kg Streptozotocin; at 8 weeks of age, the animals were given Lithospermoside (0.3mg / kg, 1mg / kg or 3mg / kg) by mixed feeding for 2 weeks, and then the aortic samples were taken. Compared with the Streptozotocin group, Lithospermoside 3mg / kg significantly reduced BUN, E-selectin, urinary MDA and UACR. Compared with the Streptozotocin group, Lithospermoside at a dose of 1mg / kg and 3mg / kg significantly reduced the superoxide anion produced by cell membrane-anchored XO. 3mg / kg Lithospermoside significantly inhibited the fluorescence intensity of nitrotyrosine staining. The above results show that Lithospermoside can reduce the production of ROS in the thoracic aorta by inhibiting the anchored XO, thereby reducing the vascular endothelial injury in the diabetic state. It can be seen that Lithospermoside plays an antioxidant role by inhibiting the anchored XO, thereby slowing down the vascular injury in type 2 diabetes and protecting the blood vessels.

[0028] To present the technical solutions of the present application and its significant advantages comprehensively and clearly, the present application will be described in detail below with specific embodiments.

[0029] 1. Experimental methods

[0030] 1.1. Experimental animals and their treatment

[0031] The present study was conducted in strict accordance with the Animal Experimental Operation Guide of the Second People's Hospital of Shenzhen and was approved by the Animal Experimental Ethics Committee of the hospital. Thirty-five Sprague-Dawley rats were purchased from the Guangdong Provincial Animal Center and were fed with standard laboratory feed and housed individually in a room with controlled temperature (23±2℃) and light (12 hours light / dark cycle) and free access to water. At 6 weeks of age, diabetes was induced by a single intravenous (i.v.) injection of 50 mg / kg Streptozotocin in normal saline, and the control group was injected with the same volume of normal saline. The rats were monitored for hyperglycemia for 7 days, and blood samples were taken from the tail vein at 7 weeks of age to determine the fasting blood glucose. According to the body weight and fasting blood glucose level, the diabetic rats were divided into four groups: control group, Streptozotocin group, and three Lithospermoside treatment groups (0.3 mg / kg / d, 1 mg / kg / d, and 3 mg / kg / d). The rats received intervention from 8 weeks of age for 2 weeks. Before tissue collection, the rats were fasted overnight and sacrificed by bloodletting under sevoflurane anesthesia. After cardiac blood collection, the plasma was separated by centrifugation at 4℃ and 3000 x g for 15 minutes and stored at -80℃ for testing. The aortas were immediately removed and placed in Krebs solution, and the connective tissue was removed and stored in 4℃ Krebs solution until use.

[0032] 1 mM Lithospermoside stock solution preparation: 1 mg of Lithospermoside was dissolved in 3.0367 mL of double-distilled water, and the stock solution was aliquoted and stored in a -80℃ refrigerator to avoid repeated freezing and thawing. When used, it was diluted to the working solution concentration.

[0033] 1.2. Biochemical analysis

[0034] Whole blood was used for glucose concentration determination. Fasting blood glucose was measured using the glucose oxidase method with a glucose meter (Sanwa Kagaku Kenkyusho Co., Ltd., Nagoya, Japan); plasma blood urea nitrogen (BUN) and urinary creatinine concentrations were measured using the L-type Wako UN kit (Catalog Nos. 416-55192, 412-55292, 419-41691, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan) and the L-type Wako Cre kit (Catalog Nos. 469-07594, 465-07694, 413-41591, FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan), respectively; urinary albumin concentration was measured using an enzyme-linked immunosorbent assay kit (Catalog No. E111-125, Bethyl Laboratory, AL, USA); and urinary malondialdehyde (MDA) concentration was measured using an MDA assay kit (Catalog No. E111-125, Bethyl Laboratory, AL, USA). Urinary MDA and albumin concentrations were measured using an ELISA kit (Catalog No. NWK-MDA01, Northwest Life Science Specialties LLC, OR, USA). Urinary MDA and albumin concentrations were expressed as the urinary creatinine ratio (UACR). Plasma E-selectin concentrations were measured using an ELISA kit (Catalog No. ELR-Eselectin, RayBiotech Life, GA, USA).

[0035] 1.3 Purine bodies and drug concentration

[0036] When determining purine bases, plasma was taken and added to Tris buffer (pH 8.5) containing sodium chloride, along with […]. 15 N2]-xanthine and [ 15 [N2]-uric acid was used as an internal standard, and the mixture was then heated at 95°C for 5 minutes. The resulting suspension was centrifuged at 4°C and 15000×g for 10 minutes. The supernatant was filtered through an ultrafiltration membrane and analyzed by liquid chromatography-mass spectrometry (LC / MS).

[0037] When determining the concentration of shikonin, plasma was taken and acetonitrile containing F10460 as an internal standard was added. After filtration through a membrane filter, the filtrate was evaporated, redissolved in 10% methanol, and used for liquid chromatography-tandem mass spectrometry (LC / MS / MS) analysis.

[0038] 1.4 XOR plasma activity assay

[0039] XOR activity determination: Plasma was added to a 20 mmol / L Tris buffer (pH 8.5), which contains [ 15 [N2]-xanthine (0.8 mmol / L), nicotinamide adenine dinucleotide (1 mmol / L), and oxaloacetic acid (0.013 mmol / L) were incubated at 37°C for 30 minutes. Then [ 13 C2, 15 [N2]-uric acid was used as an internal standard. The mixture was heated at 95°C for 5 minutes, and then centrifuged at 4°C and 15000×g for 10 minutes. The supernatant was filtered through an ultrafiltration membrane and the determination was performed by LC / MS. 15 N2]-uric acid concentration, activity as [ 15 [N2]-uric acid nmol / min / mg protein.

[0040] 1.5 Anchoring XOR activity

[0041] The effect of shikonin on anchored XOR activity was investigated by detecting the aortic segment (3 mm long) using the superoxide-sensitive chemiluminescent dye 8-amino-5-chloro-7-phenylpyrrole[3,4-d]pyrazine-1,4-(2H,3H)dione sodium salt and a chemiluminescence analyzer. The specific steps were as follows: The aortic segment was equilibrated at 37°C in a Krebs solution with 95% O2 / 5% CO2 for 30 minutes; a scintillation bottle containing modified Krebs–HEPES buffer (containing 100 μM L-012 and 50 μM xanthine) was placed in the chemiluminescence analyzer to determine the background signal; subsequently, the aortic segment was placed in the scintillation bottle, and the chemiluminescence signal was continuously tracked at 37°C for 30 minutes; after removal, the aortic segment was dried at 90°C for 24 hours and weighed.

[0042] During the detection process, the blank signal was subtracted from the chemiluminescence signal of the sample, and the difference was used to calculate the luminescence rate per minute. Chemiluminescence was expressed as a standard ratio (5 μU / mL xanthine oxidase) and corrected for dry weight. Furthermore, the effects of heparin (1000 U / mL) and superoxide dismutase (SOD) were evaluated to confirm whether these drugs could inhibit the effects of reactive oxygen species (ROS).

[0043] 1.6 Intracellular XOR activity

[0044] The effect of shikonin on intracellular XOR activity was investigated using ethidium dihydroacetate oxidation fluorescence staining. The experiment used aortic segments stored in the compound at 25°C (OCT). The specific procedures were as follows: Embedded frozen aortic segments were cut into 8 μm thick sections using a cryostat and fixed onto MAS-coated slides. The arterial sections were incubated in a CO2 incubator at 37°C for 20 minutes in Krebs-Henseleit buffer (pH 7.4) with or without 50 μM xanthine. Subsequently, they were incubated in Krebs-Henseleit buffer (pH 7.4) containing 1 μM ethidium dihydroacetate at 37°C for 30 minutes.

[0045] Images were acquired using a confocal laser scanning microscope system. Fluorescence intensity was measured in eight randomly selected regions from each slice, and the average value was calculated using digital image analysis software.

[0046] 1.7 Cytotoxicity test

[0047] The tissue section preparation procedure is as follows: The aorta was cut into 3 mm thick sections, fixed with 4% paraformaldehyde, and then incubated sequentially in 10% and 15% sucrose solutions for 4 hours each, followed by overnight incubation in 20% sucrose solution. After embedding with an optimal cutting temperature (OCT) compound, the specimen was rapidly frozen in liquid nitrogen. The frozen aorta was then cut into 6 μm thick sections using a cryostat and attached to MAS-coated glass slides (Matsunami Glass, Osaka, Japan).

[0048] 1.8. Fluorescent Immunostaining

[0049] Antigen activation was performed using HistoVT One solution. After rinsing with phosphate-buffered saline (PBS), sections were blocked with HistoOne blocking solution and then incubated overnight at 4°C with two primary antibodies: a mouse monoclonal antibody against 3-nitrotyrosine (3-NT) and a rabbit monoclonal antibody against the endothelial marker CD31.

[0050] The following day, the slides were washed with PBS and incubated at room temperature for 1 hour. The secondary antibody, goat anti-mouse immunoglobulin G (IgG) H&L, was used to label 3-NT (appearing green), and goat anti-rabbit IgG H&L was used to label CD31 (appearing red). A control experiment was also set up, in which PBS was used to treat the slides instead of the primary antibody as a secondary antibody control.

[0051] Finally, the sections were mounted and stained with a Vector TrueVIEW autofluorescence quenching kit containing 4,6-diamidinyl-2-phenylindole (DAPI). Images were acquired using a confocal laser scanning microscope system, and fluorescence intensity was measured in four randomly selected overlapping regions of CD31 and 3-NT fluorescence from each section. The average value was calculated using ImageJ software.

[0052] 1.9 Statistical Analysis

[0053] All results are reported as mean ± standard deviation, where n represents the number of rats used (each rat was used to provide only one segment for a specific experiment). Multiple group comparisons were performed using one-way ANOVA followed by Tukey's post-hoc test. Statistical significance was defined as p < 0.05.

[0054] 2. Experimental Results

[0055] 2.1 Biochemical characteristics and plasma purine body concentration

[0056] At 10 weeks of age, the body weight of rats in the control group was significantly higher than that in the Streptozotocin-treated group. Fasting blood glucose levels were significantly higher in the Streptozotocin-treated group compared to the control group. Fecal urea nitrogen (BUN) concentrations were significantly higher in the Streptozotocin group, while fecal BUN concentrations were significantly lower in the Lithospermoside 3 mg / kg group compared to the Streptozotocin group. Urinary MDA concentrations, corrected for creatinine, were significantly higher in the Streptozotocin group compared to the control group, but significantly lower in the Lithospermoside 1 mg / kg and 3 mg / kg groups. The urinary albumin-to-creatinine ratio (UACR) was significantly higher in the Streptozotocin group compared to the control group. Lithospermoside showed a dose-dependent significant decrease in UACR. Compared with the control group, the plasma E-selectin concentration in the Streptozotocin group was significantly increased; while compared with the Streptozotocin group, the plasma E-selectin concentration in the 1 mg / kg and 3 mg / kg Lithospermoside groups was significantly decreased (Table 1).

[0057] Meanwhile, no difference in uric acid concentration was observed between the Control group and the Streptozotocin group, but the concentration in the Lithospermoside 3 mg / kg group was significantly lower than that in the Streptozotocin group. Hypoxanthine and xanthine were detected only in the Lithospermoside 1 mg / kg and 3 mg / kg groups, as shown in the figure. Figure 1 As shown in AC, XOR activity in plasma was significantly higher in the Streptozotocin group than in the Control group, but significantly lower in the Lithospermoside 1 mg / kg and 3 mg / kg groups than in the Streptozotocin group. Figure 1 D).

[0058] Table 1 Biochemical characteristics of rats treated with Lithospermoside

[0059]

[0060]

[0061] Note: STZ stands for Streptozotocin group, and Lto stands for Lithospermoside group.

[0062] 2.2 Anchoring XOR activity

[0063] Compared with the control group, superoxide production was significantly increased in the streptozotocin group; while compared with the streptozotocin group, superoxide production was significantly decreased in the lithospermoside 1 mg / kg and 3 mg / kg groups. Furthermore, superoxide production was significantly inhibited in the following groups: control group, streptozotocin group, lithospermoside 0.3 mg / kg group, and lithospermoside 1 mg / kg group. Figure 2 This indicates that Lithospermoside significantly inhibits the activity of xanthine oxidase (XOR) in the circulatory system.

[0064] 2.3 Effects of intracellular XOR

[0065] Compared to the control group, the Streptozotocin group showed a significant increase in intracellular XOR-active superoxide production, while the Lithospermoside group showed no change. In contrast, Lithospermoside did not inhibit intracellular reactive oxygen species (ROS) produced by xanthine oxidase. Figure 3 AC).

[0066] Besides xanthine oxidase, various other enzyme systems in the blood vessel wall can produce reactive oxygen species (ROS), including nicotinamide adenine dinucleotide phosphate oxidase, mitochondrial respiratory chain-related enzymes, and endothelial nitric oxide synthase uncoupling enzymes. Lithospermoside alone may not be sufficient to inhibit intracellular ROS generation induced by oxidative stress from these complex factors. Figure 2 Chemiluminescence results and Figure 3 Dihydroethidium oxidation fluorescence staining results showed that the level of ROS generated by intracellular XOR was not affected. These results suggest that Lithospermoside can inhibit the increase of ROS generated by anchored XOR, and Lithospermoside may improve vascular injury by inhibiting anchored XOR.

[0067] 2.4 Cytotoxic Effects

[0068] Cytotoxicity assay results showed that, compared with the control group, the fluorescence intensity of nitrotyrosine in the Streptozotocin group was significantly increased; compared with the Streptozotocin group, the fluorescence intensity of nitrotyrosine in the Lithospermoside 3 mg / kg group was significantly decreased. Figure 4 AB).

[0069] Oxidative stress is a major contributing factor to vascular damage in diabetes. XOR activation promotes the uptake of low-density lipoprotein by macrophages, indicating that XOR is directly involved in the progression of atherosclerosis. Figure 4 Nitrotyrosine staining results suggest that Lithospermoside can improve aortic endothelial cell damage. These findings indicate that Lithospermoside may reduce aortic vascular damage by inhibiting reactive oxygen species (ROS), a change that has potential benefits for managing cardiovascular disease in diabetic patients.

[0070] In summary, Lithospermoside significantly reduces blood urea nitrogen (BUN), E-selectin, urinary malondialdehyde (MDA), and the urinary albumin / creatinine ratio (UACR) by inhibiting anchored XO enzymes and reducing the resulting reactive oxygen species (ROS). It also inhibits the fluorescence intensity of nitrotyrosine staining, thereby effectively improving endothelial damage in the aorta of diabetic animal models. The results of this study suggest that this drug may have a vascular protective effect in patients with diabetic macrovascular disease. Further prospective clinical trials targeting diabetic macrovascular disease should be conducted to further validate the vascular protective effect of Lithospermoside.

[0071] The embodiments of the present invention have been described in detail above, but the present invention is not limited to the described embodiments. For those skilled in the art, various changes, modifications, substitutions, and variations can be made to these embodiments without departing from the principles and spirit of the present invention, and these variations still fall within the protection scope of the present invention.

Claims

1. Application of shikonin in the preparation of drugs that alleviate vascular damage in type 2 diabetes.

2. The application according to claim 1, characterized in that, The shikonin exerts its antioxidant effect by inhibiting anchored xanthine oxidase, thereby slowing down vascular damage in type 2 diabetes and protecting blood vessels.

3. The application according to claim 1, characterized in that, The effective dose of the shikonin is 1-5 mg / kg.

4. The application according to claim 1, characterized in that, The drug also includes pharmaceutically acceptable excipients.

5. The application according to claim 4, characterized in that, The excipients include at least one of the following: excipients, propellants, solubilizers, cosolvents, emulsifiers, colorants, binders, disintegrants, fillers, lubricants, wetting agents, osmotic pressure regulators, stabilizers, flow aids, flavoring agents, preservatives, suspending agents, coating materials, fragrances, anti-adhesion agents, binding agents, penetration promoters, pH adjusters, buffers, plasticizers, surfactants, foaming agents, defoamers, thickeners, encapsulating agents, humectants, absorbents, diluents, flocculants and anti-flocculation agents, antioxidants, adsorbents, filter aids, and release inhibitors.

6. The application according to claim 4, characterized in that, The dosage forms of the drug include tablets, capsules, aerosols, pills, powders, solutions, suspensions, emulsions, granules, liposomes, transdermal preparations, or suppositories.

Images