Application of vitamin C in preparation of medicine for preventing and treating vascular injury caused by microbial infection

Abstract

The invention discloses application of vitamin C, or stereoisomers, geometric isomers, tautomers, nitrogen oxides, hydrates, solvates, metabolites, pharmaceutically acceptable salts or prodrugs thereof in preparation of drugs for preventing and treating vascular injury caused by microbial infection. The invention discloses a molecular mechanism that vitamin C regulates ATP11C expression and inhibits phosphatidylserine eversion so as to improve an endothelial barrier function, and provides a new action target for treatment of infection-related endothelial injury; cell experiments and animal experiments prove that the vitamin C has remarkable effects in the aspects of improving endothelial dysfunction caused by infection, reducing vascular leakage and increasing the survival rate of infected animals, and has good application prospects and clinical transformation values; the application of the vitamin C in vascular endothelial injury caused by staphylococcus aureus, hantavirus and dengue virus infection is clearly provided for the first time, and the new medical application of the vitamin C is expanded.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to the application of vitamin C in the preparation of drugs for preventing and treating vascular damage caused by microbial infections. Background Technology

[0002] The statements in this section are merely background information related to the present invention and do not necessarily constitute prior art.

[0003] Infectious diseases remain a major cause of high morbidity and mortality worldwide. Among them, viral infections such as Staphylococcus aureus, Hantavirus, and dengue virus can damage vascular endothelial cells directly or indirectly, leading to increased vascular permeability, endothelial barrier dysfunction, and multiple organ dysfunction syndrome (MODS), which have become an important pathological basis for infection-related pathological damage.

[0004] Vascular endothelial cells, as key cell types for maintaining vascular homeostasis and barrier function, are highly susceptible to inflammatory responses, pathogen invasion, and oxidative stress during infection. Studies have shown that Staphylococcus aureus infection can lead to endothelial cell damage and vascular leakage by releasing virulence factors and inducing inflammatory responses; Hantavirus and dengue virus can directly infect vascular endothelial cells, disrupting intercellular junction structures and causing severe vascular permeability abnormalities, clinically manifesting as bleeding tendency, shock, and even multiple organ failure. In the process of infection-induced vascular endothelial damage, the asymmetric disruption of endothelial cell membrane phospholipids is one of the important molecular events.

[0005] Phosphatidylserine (PS) is normally located on the inner side of the cell membrane, but it can evert under conditions of infection, inflammation, and oxidative stress, forming a procoagulant and pro-inflammatory endothelial phenotype. The occurrence of PS eversion is closely related to impaired endothelial barrier function, coagulation activation, and amplified inflammation. Among these, ATP11C, as a key enzyme in maintaining PS eversion, is considered to be one of the important molecular bases for infection-induced endothelial injury due to abnormal expression or function.

[0006] Currently, treatments for Staphylococcus aureus, Hantavirus, and dengue virus infections primarily focus on anti-infection, symptomatic support, and organ function maintenance. However, effective and specific drugs are lacking to address the vascular endothelial damage caused by these infections. Therefore, developing drugs that can directly protect vascular endothelial function and improve endothelial barrier integrity is of significant clinical importance for reducing infection-related complications and improving prognosis. Summary of the Invention

[0007] This invention aims to at least partially address the technical problem of the lack of effective and specific interventions for vascular endothelial damage caused by infection in the prior art. Therefore, the main objective of this invention is to provide the application of vitamin C in the preparation of drugs for preventing and treating vascular damage caused by Staphylococcus aureus, Hantavirus, and dengue virus infections.

[0008] The objective of this invention is achieved through the following technical solution: In their research, the inventors discovered that vascular endothelial cell function is significantly reduced after infection with Staphylococcus aureus, Hantavirus, and dengue virus, specifically manifested as decreased endothelial cell viability, disruption of endothelial barrier function, and a significant increase in vascular permeability. Further research indicates that during endothelial injury induced by these infections, the asymmetric nature of endothelial cell membrane phospholipids is disrupted, phosphatidylserine (PS) undergoes eversion, and the expression of its key regulatory enzyme ATP11C is significantly upregulated, thereby promoting a procoagulant, pro-inflammatory, and barrier dysfunction pathological phenotype in endothelial cells.

[0009] Based on transcriptomic analysis and small molecule drug screening of Staphylococcus aureus, Hantavirus, and dengue virus infections, this invention provides vitamin C as an effective drug for preventing and treating vascular endothelial damage caused by Staphylococcus aureus, Hantavirus, and dengue virus infections. The results show that vitamin C can significantly inhibit infection-induced upregulation of ATP11C expression, suppress endothelial cell apoptosis, improve endothelial cell viability, and restore endothelial barrier function, thereby alleviating infection-induced vascular endothelial damage.

[0010] Specifically, the present invention provides the following technical solutions: On the one hand, it provides the use of vitamin C (VC), or its stereoisomers, geometric isomers, tautomers, nitrogen oxides, hydrates, solvates, metabolites, pharmaceutically acceptable salts, or prodrugs, in the preparation of drugs for preventing and treating vascular damage caused by microbial infections, wherein the chemical structural formula of vitamin C is: .

[0011] In some specific embodiments, the vascular damage caused by the microbial infection includes one or more species of bacterial, fungal, or viral infection, as well as sepsis that develops therefrom.

[0012] Furthermore, the bacteria / fungus or virus includes, but is not limited to, one or more of Staphylococcus aureus, Hantavirus, and dengue virus.

[0013] In some specific embodiments, the vascular endothelial injury includes, but is not limited to, one or more of the following: impaired endothelial cell barrier function, increased permeability, or decreased endothelial cell viability.

[0014] In some specific embodiments, the microbial infection leads to upregulation of ATP11C expression and increased phosphatidylserine eversion in endothelial cells.

[0015] In some specific implementations, vitamin C improves vascular damage by inhibiting ATP11C enzyme, thereby inhibiting phosphatidylserine eversion and apoptosis.

[0016] In some specific embodiments, the drug further includes a pharmaceutically acceptable carrier, including but not limited to diluents, buffers, suspensions, emulsions, granules, encapsulation agents, excipients, fillers, binders, sprays, transdermal absorbents, humectants, disintegrants, absorption enhancers, surfactants, colorants, flavoring agents, or adsorbents.

[0017] In some specific embodiments, the drug may be prepared into dosage forms such as tablets, solutions, granules, patches, ointments, capsules, aerosols, or suppositories.

[0018] In some specific embodiments, the routes of administration of the drug include, but are not limited to, oral, intravenous, intramuscular, subcutaneous, sublingual, rectal, nasal spray, oral spray, topical or transdermal administration.

[0019] In some specific implementations, vitamin C may also be used in combination with other anti-infective drugs or drugs that improve endothelial function to further enhance the therapeutic effect.

[0020] Compared with the prior art, the present invention has at least the following beneficial effects: 1) This invention is the first to clearly propose the application of vitamin C in vascular endothelial damage caused by Staphylococcus aureus, Hantavirus and dengue virus infection, thus expanding the new medical applications of vitamin C; 2) This invention reveals the molecular mechanism by which vitamin C improves endothelial barrier function by regulating ATP11C expression and inhibiting phosphatidylserine eversion, providing a new target for the treatment of infection-related endothelial injury; 3) This invention has verified through cell experiments and animal experiments that vitamin C has significant effects in improving endothelial dysfunction caused by infection, reducing vascular leakage and improving the survival rate of infected animals, and has good application prospects and clinical translation value. Attached Figure Description

[0021] To more clearly illustrate the specific embodiments of the present invention, the accompanying drawings used in the description of the specific embodiments or the prior art will be briefly introduced below.

[0022] Figure 1In Example 1 of this invention, it was found that VC is a potential therapeutic molecule for vascular endothelial damage caused by Staphylococcus aureus, Hantavirus, and dengue virus infection. Figure 2 In Example 2 of this invention, VC improves lung tissue damage and apoptosis in mice infected with Staphylococcus aureus; Figure 3 In Example 3 of this invention, VC improves the survival rate of mice infected with Staphylococcus aureus; Figure 4 In Example 4 of this invention, VC inhibited the expression of ATP11C mRNA in lung microvascular endothelial cells and mouse lung tissue infected by Staphylococcus aureus, Hantavirus, and dengue virus. Figure 5 This is the result of VC improving vascular barrier damage in pulmonary microvascular endothelial cells of mice infected with Staphylococcus aureus, Hantavirus, and dengue virus in Example 5 of the present invention; Figure 6 This is the result of VC inhibiting lung tissue leakage in mice infected with Staphylococcus aureus, Hantavirus, and dengue virus in Example 6 of the present invention. Detailed Implementation

[0023] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be described in further detail below with reference to the accompanying drawings and embodiments, but this should not be construed as limiting the scope of protection of this invention. Any formal equivalent modifications made based on the concept of this invention should be considered within the scope of this invention.

[0024] This invention provides a general and / or specific description of the materials and experimental methods used in the experiments. Unless otherwise specified, all experimental or testing methods are conventional methods; all reagents or instruments used, unless otherwise specified, are commercially available conventional products prepared or used using conventional methods.

[0025] Example 1: Vitamin C is a potential therapeutic molecule for vascular endothelial damage caused by Staphylococcus aureus, Hantavirus, and dengue virus infections. Screening for genes co-regulated by Staphylococcus aureus, Hantavirus, and dengue virus infection.

[0026] This study included the following RNA-seq transcriptome datasets: GSE34628, GSE9378, and GSE139603 for dengue virus-infected endothelial cells; GSE133751 for Hantavirus-infected endothelial cells; and GSE82036 for Staphylococcus aureus-infected endothelial cells. Differentially expressed genes (DEGs) were screened using the online GEO database analysis tool GEO2R, ​​with a significance threshold (P-value < 0.05) and a fold change in expression |LogFC| ≥ 0.5. Veen analysis was performed using the Bioinformatics online platform (http: / / www.bioinformatics.com.cn) to identify common differentially expressed genes in endothelial cells infected by dengue virus, Hantavirus, and Staphylococcus aureus. Enrichment analysis of gene pathways co-regulated by Staphylococcus aureus, Hantavirus, and dengue virus infection. Using the common differentially expressed genes obtained in step (1), pathway enrichment analysis was performed using the Kyoto Encyclopedia of Genes and Genomes (KEGG). Screening for potential therapeutic molecules for vascular injury caused by Staphylococcus aureus, Hantavirus, and dengue virus infections. Based on differentially expressed gene features, the ConnectivityMap (CMap) platform was used for small molecule drug prediction analysis. Common differentially expressed genes from the three pathogens were used as input features. A pattern matching algorithm was used to calculate the similarity score between the compound and the perturbation gene expression profile of a reference drug. Finally, the candidate small molecule compounds with the highest scores were selected as potential small molecule drugs for treating vascular injury caused by microbial infections. Results and Discussion: The experimental results are as follows Figure 1 As shown, Veen analysis revealed 301 differentially expressed genes among endothelial cells infected by Hantavirus, dengue virus, and Staphylococcus aureus; pathway enrichment analysis showed that the most significantly regulated pathway was the apoptosis pathway; and analysis using the CMap platform revealed that VC had the best score.

[0027] Example 2: Effect of microorganism C on the survival rate of mice infected with Staphylococcus aureus A mouse model of Staphylococcus aureus infection was established: A solvent control group (intraperitoneal injection of sterile PBS, 200 μL / mouse, without Staphylococcus aureus infection, i.e., Vehicle or control group), a vitamin C intervention group (intraperitoneal injection of vitamin C solution, dose 180 mg / kg / day, 200 μL / mouse, solvent: sterile PBS, without Staphylococcus aureus infection, i.e., VC group), and an infection group (intraperitoneal injection of 2.5 × 10⁻⁶ ppm) were set up.9 CFU / single Staphylococcus aureus ATCC25926 PBS suspension, i.e., SA group), post-infection intervention group (intraperitoneal injection of 2.5×10 9 CFU / mouse of Staphylococcus aureus ATCC25926 PBS suspension was administered intraperitoneally immediately after infection with vitamin C solution at a dose of 180 mg / kg / day, in a volume of 200 μL / mouse, using sterile PBS as the solvent (VC+SA group). Fifteen 7-week-old mice weighing 22–25 g were randomly assigned to each group and allowed one week of acclimatization. Different concentrations (1×10⁻⁶ CFU / mouse) of vitamin C solution were administered intraperitoneally. 9 CFU / each, 2.5×10 9 CFU / each, 5 x 10 9 The optimal infection concentration of Staphylococcus aureus ATCC25926 (CFU / mouse) in PBS suspension was determined to construct a mouse model of Staphylococcus aureus infection. The animals used in the experiment were C57BL / 6 mice provided by the Experimental Animal Center of Daping Hospital, Army Medical University, and were housed in the SPF Animal Experiment Center of the Army Characteristic Medical Center. The indoor temperature was 23-25℃ and the relative humidity was 40%-60%. The mice had free access to water and food.

[0028] Mice were injected with Staphylococcus aureus and then immediately treated with intraperitoneal injection of vitamin C solution (180 mg / kg / day). Mice mortality was then observed within 48 hours.

[0029] The results are as follows Figure 2 As shown in the figure, the lethal dose of Staphylococcus aureus infection in mice is 2.5 × 10⁻⁶. 9 CFU and VC treatment significantly improved the survival rate of mice infected with Staphylococcus aureus. n = 20, a: P<0.05, comparison between SA group and Vehicle group; b: P<0.05, comparison between VC+SA group and SA group.

[0030] Example 3: Effects of Vitamin C on Lung Damage and Apoptosis Induced by Staphylococcus aureus, Hantavirus, and Dengue Virus Infections A mouse model of Staphylococcus aureus infection was established: A solvent control group, a vitamin C intervention group, an infection group, and a post-infection intervention group were set up (same as in Example 2). Six 7-week-old mice weighing 22–25 g were randomly selected for each group and allowed to acclimatize for one week. Mice were administered 2.5 × 10⁻⁶ mg / L via intraperitoneal injection. 9 A mouse model of Staphylococcus aureus infection was constructed using CFU / mouse ATCC25926. The animals used in the experiment were C57BL / 6 mice provided by the Experimental Animal Center of Daping Hospital, Army Medical University, and were housed in the SPF Animal Experiment Center of the Army Characteristic Medical Center. The indoor temperature was 23-25℃ and the relative humidity was 40%-60%. The mice had free access to water and food.

[0031] Pathological staining of lung tissue from infected mice: Mice were immediately injected intraperitoneally with vitamin C solution (180 mg / kg / d) for 24 hours after being injected with Staphylococcus aureus; they were then euthanized by an overdose of isoflurane, and lung tissue was subsequently removed from the infected mice. A portion of the liver tissue was also taken for histological section staining. Results are shown in […]. Figure 3 A. As shown in the figure, VC effectively reduces lung tissue leakage.

[0032] Apoptosis detection in lung tissue of infected mice: Mice were injected with Staphylococcus aureus via intraperitoneal injection of vitamin C solution (180 mg / kg / d) for 24 h immediately after injection. They were then euthanized under an overdose of isoflurane, and lung tissue was subsequently removed. Apoptosis of lung cells was detected using a Roche apoptosis ELISA kit (11544675001) according to the manufacturer's instructions. Results are shown below. Figure 3 B. As shown in the figure, VC effectively improves the apoptosis of lung cells in lung tissue.

[0033] Example 4: Vitamin C inhibits the upregulation of ATP11C expression induced by Staphylococcus aureus, Hantavirus, and dengue virus infections. Cell culture and intervention: Primary lung microvascular endothelial cells (HPMECs, CP-H001, Procell Biotechnology Co., Ltd.) were cultured at a concentration of 1×10⁻⁶ cells / mL. 7 Cells were seeded at a density in 10 cm cell culture dishes and cultured in DMEM medium containing 10% fetal bovine serum at 37°C and 5% CO2 until 80% confluence. HPMECs were infected with Staphylococcus aureus, Hantavirus, and dengue virus, respectively, and treated with 1 mM vitamin C. Untreated cells served as the control group.

[0034] Lung tissue acquisition from infected mice: The following groups were established: a solvent control group (intraperitoneal injection of sterile PBS, 200 μL / mouse, without Staphylococcus aureus infection, i.e., control group); a vitamin C intervention group (intraperitoneal injection of vitamin C solution, dose 180 mg / kg / day, dose 200 μL / mouse, solvent: sterile PBS, without Staphylococcus aureus infection, i.e., VC group); a Staphylococcus aureus infection group (same as in Example 2, i.e., SA group); a post-Staphylococcus aureus infection intervention group (immediately after Staphylococcus aureus infection, intraperitoneal injection of vitamin C solution, dose 180 mg / kg / day, dose 200 μL / mouse, solvent: sterile PBS, i.e., VC+SA group); and a hantavirus infection group (intraperitoneal injection of 1.0 × 10⁻⁶ ppm). 5The following groups were identified: HTNV group (CFU / animal 76-118 PBS suspension); Hantavirus infection intervention group (immediate intraperitoneal injection of vitamin C solution after Hantavirus infection, dose 180 mg / kg / day, volume 200 μL / animal, solvent: sterile PBS, i.e., VC+HTNV group); and dengue virus infection group (intraperitoneal injection of 1.0 × 10⁻⁶ CFU / animal). 8 The mice were divided into two groups: a CFU / mouse DENV-2 16681PBS suspension (DENV group) and a dengue virus infection intervention group (VC+DENV group, which consisted of 6 7-week-old mice weighing 22-25g each, administered intraperitoneally with 180mg / kg / day, 200μL / mouse, in sterile PBS immediately after dengue virus infection). Each group was randomly assigned to acclimatize for one week. Mice were treated with Staphylococcus aureus, Hantavirus, and dengue virus for 6 hours to establish infection models. They were then sacrificed with an overdose of anesthetic, and lung tissue was collected.

[0035] Reverse transcription PCR and real-time quantitative PCR: Total RNA was extracted from cells and lung tissue using TRIzol reagent according to the manufacturer's instructions; RNA purity (A260 / A280 ratio 1.8-2.0) and concentration were detected using a NanoDrop 2000 spectrophotometer; 2 μg of total RNA was used for reverse transcription using the PrimeScript™ RT kit (Takara, RR047A), with a reaction volume of 20 μL (4 μL of 5×PrimeScript Buffer, 1 μL of Random 6 mers, and 1 μL of PrimeScript RTase), and reaction conditions: 37℃ for 15 min, 85℃ for 5 sec; PCR amplification was performed using TB Green™ Premix Ex Taq™ (Takara, RR420A), with the following reaction program: 94℃ pre-denaturation for 2 min; 35 cycles (94℃ for 30 s, 58℃ for 30 s, 72℃ for 45 s); extension at 72℃ for 10 min; all primer sequences are as shown in SEQ ID No. 1 and SEQ ID No. 2. The results are shown in SEQ ID No. 2, SEQ ID No. 3, and SEQ ID No. 4, as detailed in Table 1. All were designed using NCBI Primer-BLAST. The relative expression levels of the target gene were calculated using the 2-ΔΔCt method, with GAPDH as an internal reference gene. The quantification results of cellular ATP11C mRNA are shown below. Figure 4 A, Quantitative results of ATP11C mRNA in lung tissue are shown in [Figure 1]. Figure 4B; As shown in the figure, VC reverses the upregulation of ATP11C caused by Staphylococcus aureus, Hantavirus and dengue virus infection in HPMECs and mice. Among them, a: P<0.05, compared with the control group; b: P<0.05, compared with the VC+ infection group and the simple infection group.

[0036] Example 5: Effects of Vitamin C on endothelial barrier damage induced by Staphylococcus aureus, Hantavirus, and dengue virus infections. (1) Cell viability and endothelial cell resistance measurement: CCK8 reagent (Bimake) and complete culture medium were added to a 96-well plate (Corning, USA) at a ratio of 1:9, and then incubated at 37°C for 1 hour before measuring absorbance at 450 nm.

[0037] HPMECs (1×10) 4 Cells (10 cells / well) were seeded in Transwell chambers (Corning, USA) with membranes having a 0.4 μm pore size and cultured until the cell monolayers reached confluence. Transmembrane resistance (TER) values ​​were measured using a Millicell-ERS voltmeter (Millipore, USA) to assess the effects of different pathogens and vitamin C on endothelial barrier function. All experiments were performed within a Class II biosafety cabinet in a BSL-2 laboratory, with six biological replicates per group.

[0038] The results are as follows Figure 5 As shown in Figure A, VC improves the barrier function impairment of HPMECs infected with Staphylococcus aureus, Hantavirus, and dengue virus. a: P<0.05, comparison between the infection group and the control group; b: P<0.05, comparison between the VC+ infection group and the simple infection group.

[0039] (2) Immunofluorescence staining to detect endothelial barrier integrity: Primary lung microvascular endothelial cells (HPMECs, CP-H001, Procell Biotechnology Co., Ltd.) were stained at a concentration of 1×10⁻⁶. 5Cells were seeded at a density in 12-well plates containing pre-coated cell spreaders and cultured in DMEM medium containing 10% fetal bovine serum at 37°C and 5% CO2 until 80% confluence. HPMECs were infected with Staphylococcus aureus, Hantavirus, and dengue virus, respectively, and treated with 1 mM vitamin C. Untreated cells served as the control group. Remove the culture medium, wash once with PBS buffer, fix with 4% paraformaldehyde (PFA) for 30 min, wash four times with PBS buffer, permeate with 0.25% Triton X-100 for 15 min, wash three times with PBS buffer, then transfer round coverslips containing cells to the well plates. Add 100 μL of ZO-1 antibody (Rabbit polyclonal Anti-ZO-1, 61-7300, 1:50) and VE-cadherin antibody (Goat polyclonal anti-VE-cadherin, 1:100 R&D, AF938) to each coverslip and incubate at room temperature in the dark for 3 hours. Wash five times with PBS buffer for 5 min each time. Subsequently, each coverslip was added with 100 μL of PBS diluted with Alexa Fluor 568 Donkey anti-Rabbit IgG (H+L) (A10042, 1:100) and Alexa Fluor 488-conjugated Donkey anti-Goat IgG (H+L) (A11055, 1:100) staining solution, and incubated at room temperature in the dark for 10 minutes. After staining, the coverslips were rinsed 5 times with PBS for 5 minutes each time. 100 μL of LDAPI staining solution (5 μg / mL) was added to each coverslip, and then the coverslips were fixed onto a slide using mounting medium (Mowio 14-88). After air-drying in the dark, the integrity of the endothelial cell barrier was observed under a Nikon ECLIPSETi microscope.

[0040] The results are as follows Figure 5 As shown in Figure B, VC improves the barrier function impairment of HPMECs infected by Staphylococcus aureus, Hantavirus, and dengue virus.

[0041] Example 6: Effects of Vitamin C on Pulmonary Vascular Leakage Caused by Staphylococcus aureus, Hantavirus, and Dengue Virus Infections (1) Establishment of a mouse infection model: The animals used in the experiment were C57BL / 6 mice, weighing 22-25g, provided by the Experimental Animal Center of Daping Hospital, Army Medical University. They were housed in the SPF Animal Experiment Center of the Army Special Medical Center, with an indoor temperature of 23-25℃ and a relative humidity of 40%-60%. The mice had free access to water and food. Staphylococcus aureus ATCC25926 was administered via intraperitoneal injection in PBS suspension (2.5×10⁻⁶).9 A mouse model of Staphylococcus aureus infection was constructed using CFU / mouse. All procedures were performed inside a biosafety cabinet.

[0042] (2) Evans Blue Staining. After establishing the Staphylococcus aureus infection model, mice were injected with Evans Blue Dye (50 mg / kg, 200 μL) via the tail vein for 2 hours. After anesthesia, the lungs of the mice were slowly flushed with 50 mL of heparinized saline through the right ventricle. Then, lung tissue from each group was placed in 1 mL of N,N-dimethylformamide and treated at 60°C for 24 hours. Finally, the lung tissue was homogenized, centrifuged at 12,000 r / min for 10 minutes, and the supernatant was collected. The absorbance was measured at 620 nm, and the Evans Blue content was calculated according to the standard curve.

[0043] See results Figure 6 A, 6B. From the figure, it can be seen that VC reduces vascular leakage in mice infected with Staphylococcus aureus. a: P<0.05, comparison between SA group and Vehicle group; b: P<0.05, comparison between VC+SA group and SA group.

[0044] (3) Detection of ZO-1 / VE-cadherin expression in lung tissue of infected mice: After establishing a Staphylococcus aureus infection model, mice were euthanized by an overdose of isoflurane. Lung tissue from infected mice was then removed, and approximately 50 mg was weighed and recorded. The tissue was washed 2-3 times with 1 mL of pre-cooled PBS, followed by 1.2 mL of RIPA tissue lysis buffer containing protease inhibitors and 2 mL of ceramic beads. The homogenate was homogenized in a Precellys24 tissue homogenizer (Bertin) at 5500 rpm for 10 seconds, repeated 3 times. After homogenization, the homogenate was placed on ice for 30 min, then centrifuged at 12000 rpm for 10 min at 4℃. 100 μL of the supernatant was transferred to a new 1.5 mL centrifuge tube. 4× Loading buffer was added proportionally and incubated at 100℃ for 10 min. The expression of ZO-1 / VE-cadherin in lung tissue was detected by standard Western blotting method. Primary antibodies were Rabbit polyclonal Anti-ZO-1, 61-7300, 1:1000, and Goat polyclonal anti-VE-cadherin, 1:1000 R&D, AF938. Secondary antibodies were Donkey Anti-Rabbit IgG (H+L) (711-035-152, 1:5000) and Donkey Anti-Goat IgG (H+L) (705-035-003, 1:5000). Finally, the expression of ZO-1 / VE-cadherin was detected by chemiluminescence immunoassay. Test results are shown Figure 6 CD, as shown in the figure, VC restores the expression of barrier-related proteins induced by Staphylococcus aureus infection in mice. a: P<0.05, comparison between SA group and Vehicle group; b: P<0.05, comparison between VC+SA group and SA group.

[0045] The above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit it. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention, and they should all be covered within the scope of the claims and specification of the present invention.

Claims

1. The use of the compound vitamin C shown in Formula I, or its stereoisomers, geometric isomers, tautomers, nitrogen oxides, hydrates, solvates, metabolites, pharmaceutically acceptable salts, or prodrugs, in the preparation of drugs for the prevention and treatment of vascular damage caused by microbial infections: Ⅰ。 2. The application according to claim 1, characterized in that, The vascular damage caused by the microbial infection includes one or more species of bacterial, fungal, or viral infection, as well as sepsis that develops therefrom.

3. The application according to claim 2, characterized in that, The vascular endothelial injury includes one or more of the following: impaired endothelial cell barrier function, increased permeability, or decreased endothelial cell viability.

4. The application according to claim 2, characterized in that, Vitamin C improves vascular damage by inhibiting ATP11C enzyme, thereby inhibiting phosphatidylserine eversion and apoptosis.

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