Application of aspirin eugenol ester in preparation of medicine for relieving gastric mucosal lesion

Abstract

The invention discloses application of aspirin eugenol ester in preparation of a medicine for relieving gastric mucosal lesion. The invention finds that the aspirin eugenol ester can significantly relieve alcohol-induced rat gastric mucosa lesion and play a role in protecting gastric mucosa by reducing inflammatory response, enhancing antioxidant capacity, protecting mucosa barrier function, inhibiting apoptosis reaction and ferroptosis reaction and the like. In addition, after the aspirin eugenol ester is prepared into any oral administration dosage according to a conventional preparation method in the field, the alcohol-induced gastric mucosal lesion can be remarkably relieved.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical development technology, and in particular relates to the application of aspirin eugenol ester in the preparation of a drug to alleviate alcohol-induced gastric mucosal damage. Background Technology

[0002] Gastric mucosal injury is an important pathological basis for digestive system diseases, and its mechanism involves multiple factors such as oxidative stress, inflammatory response, and mucosal barrier dysfunction; it is closely related to Helicobacter pylori infection, the use of nonsteroidal anti-inflammatory drugs (NSAIDs), and alcohol consumption. Alcohol-induced gastric mucosal injury is one of the common causes of gastrointestinal diseases, and its pathogenesis is closely related to the direct cytotoxicity of alcohol, oxidative stress, and mucosal barrier damage. Globally, excessive alcohol consumption has led to a significant increase in the incidence of gastrointestinal diseases, especially among alcoholics, where the prevalence of gastric mucosal injury can reach over 30%, of which 10% to 20% may develop into peptic ulcers or chronic atrophic gastritis, and in severe cases, it can induce gastric bleeding or even gastric cancer.

[0003] Currently, clinical prevention and treatment mainly rely on proton pump inhibitors (PPIs), H2 receptor antagonists, and mucosal protectants. While these can provide short-term symptom relief, they cannot reverse deep pathological changes, and long-term use can lead to side effects such as drug resistance and osteoporosis. In recent years, the development of novel drugs for alcoholic gastric injury has focused on targeting and regulating inflammatory factors, enhancing antioxidant capacity, and promoting mucosal repair; however, most of these efforts are still in the preclinical research stage. Therefore, developing innovative drugs that combine highly effective repair, low side effects, and long-term protective properties has significant social and clinical value.

[0004] Anhydrous ethanol-induced gastric mucosal injury models have been widely used since the 1980s. Anhydrous ethanol is a necrotic medium that can directly corrode the gastric mucosa, rapidly disrupting the mucus-bicarbonate barrier, leading to hydrogen ion backdiffusion, reduced mucosal blood flow, and tissue ischemia and hypoxia. It also induces oxidative stress and gastric mucosal ulcers by generating highly cytotoxic free radicals. Compared to other gastric mucosal injury models, the anhydrous ethanol model is characterized by its simplicity, low cost, and high time efficiency, thus its widespread use.

[0005] Aspirin (ASP), a nonsteroidal anti-inflammatory drug (NSAID), not only possesses antipyretic, analgesic, and anti-inflammatory effects, but also antirheumatic and platelet-activating inhibitory effects. However, its associated gastrointestinal adverse reactions limit its clinical application. Eugenol (EUG), a natural product, exhibits significant anti-inflammatory, antibacterial, and antioxidant activities, but its chemical instability and irritating odor limit its application. To eliminate these side effects, a novel pharmaceutical compound, aspirin eugenol ester (AEE), was synthesized through esterification using the prodrug principle. Studies have shown that AEE, while maintaining the pharmacological activities of ASP and EUG, shields the carboxyl group of ASP, reducing gastrointestinal irritation and other side effects when ASP enters the body, and overcoming the irritating odor and instability of EUG. AEE, as a novel NSAID, has been proven to possess good anti-inflammatory, antithrombotic, anti-atherosclerotic, antioxidant, antipyretic, and analgesic activities and is currently in the clinical development stage. Summary of the Invention

[0006] To overcome the shortcomings and deficiencies of the prior art, the present invention aims to provide the application of aspirin eugenol ester in the preparation of a drug to alleviate alcohol-induced gastric mucosal damage.

[0007] This invention is achieved by using aspirin eugenol ester in the preparation of drugs for the prevention or treatment of alcohol-induced gastric mucosal damage.

[0008] Preferably, the dosage form of the gastric mucosal injury drug is a powder, tablet, granule, capsule, oral liquid, emulsion, or suspension.

[0009] Preferably, the gastric mucosal injury drug further comprises a carrier and / or excipients.

[0010] Preferably, in the application described, aspirin eugenol ester reduces the inflammatory response of human gastric mucosal epithelial cells, enhances antioxidant capacity, protects mucosal barrier function, and inhibits apoptosis and ferroptosis.

[0011] This invention overcomes the shortcomings of existing technologies and provides an application of aspirin eugenol ester in the preparation of drugs to alleviate alcohol-induced gastric mucosal damage. The compound structure of aspirin eugenol ester is shown below:

[0012]

[0013] In this invention, aspirin eugenol ester can significantly alleviate alcohol-induced gastric mucosal damage in rats. It mainly works by reducing inflammatory response, enhancing antioxidant capacity, protecting mucosal barrier function, inhibiting apoptosis and ferroptosis, etc., thus protecting the gastric mucosa.

[0014] Compared to the shortcomings and deficiencies of existing technologies, this invention has the following beneficial effects: This invention discovers that aspirin eugenol ester can significantly alleviate alcohol-induced gastric mucosal damage in rats by reducing inflammatory responses, enhancing antioxidant capacity, protecting mucosal barrier function, and inhibiting apoptosis and ferroptosis, thus playing a protective role for the gastric mucosa. Furthermore, when aspirin eugenol ester is prepared into any oral dosage form using conventional methods in the art, administration of the drug can significantly alleviate alcohol-induced gastric mucosal damage. Attached Figure Description

[0015] Figure 1 These are the cell transcriptomics analysis results in the embodiments of the present invention; wherein, Figure A shows differentially expressed genes, Figure B shows GO analysis, and Figure C shows KEGG analysis.

[0016] Figure 2 The figures show the surface changes and histopathological changes of rat gastric tissue induced by alcohol prevention using AEE in this embodiment of the invention; wherein, Figure A is the gastric tissue appearance, Figure B is HE staining, Figure C is AB-PAS staining, Figure D is histopathological score, and Figure E is AB-PAS score.

[0017] Figure 3 The figures show the results of AEE in preventing alcohol-induced oxidative stress, apoptosis, tight junctions, and inflammatory responses in rat gastric tissue in this embodiment of the invention; wherein, Figure A is MDA, Figure B is SOD, Figure C is p65, Figure D is PUMA, Figure E is ZO-1, Figure F is EGF, Figure G is IL-1β, and Figure H is TNF-α.

[0018] Figure 4 The figures show the histopathological changes in rat gastric tissue induced by alcohol-induced AEE treatment in this embodiment of the invention; Figure A shows the appearance of the gastric tissue, and Figure B shows the histopathological score.

[0019] Figure 5 The figures show the results of detection related to oxidative stress, apoptosis, tight junctions and inflammatory responses in rat gastric tissue induced by AEE treatment in this embodiment of the invention; wherein, Figure A is IL-1β, Figure B is MDA, Figure C is TP53, Figure D is ZO-1, and Figure E is VEGF. Detailed Implementation

[0020] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0021] Example 1: Cell transcriptomics analysis showed that AEE can alleviate alcohol-induced gastric mucosal damage.

[0022] Human gastric mucosal epithelial cells (GES-1) were divided into a model group and an AEE group, with three replicates in each group. Cells in the model group were treated with 5% absolute ethanol for 3 hours, while cells in the AEE group were first pretreated with AEE for 24 hours, then the culture medium was discarded, and the cells were treated with 5% absolute ethanol for 3 hours. Cell samples were collected, total RNA was extracted, and transcriptome analysis was performed.

[0023] Statistical analysis of differentially expressed genes was performed using a fold change absolute value ≥1.5 and a p-value <0.05 as criteria. Results are shown below. Figure 1 -A. Compared with the AEE group, the model group showed significantly upregulated expression of 309 genes and significantly downregulated expression of 414 genes.

[0024] GO clustering analysis was performed, and all GO terms with p < 0.05 were filtered out. The analysis results are shown in [link to analysis]. Figure 1 -B. Differentially expressed genes are mainly enriched in the positive regulation of cellular biological processes, regulation of apoptosis, regulation of programmed cell death, responses to stimuli, cellular responses to stimuli, positive regulation of metabolic processes, regulation of responses to stimuli, positive regulation of programmed cell death, positive regulation of apoptosis, responses to oxygen-containing compounds, signal transduction, and cellular responses to chemical stimuli.

[0025] KEGG signal pathway analysis was performed, and signal pathways with P < 0.05 were screened. The analysis results are shown below. Figure 1 -C. Differentially identified genes are mainly enriched in antigen processing and presentation, apoptosis, arachidonic acid metabolism, autophagy, cAMP signaling pathway, complement and coagulation cascade, ferroptosis, gastric acid secretion, glutathione metabolism, regulation of TRP channels by inflammatory mediators, mismatch repair, NF-κB signaling pathway, p53 signaling pathway, PI3K-Akt signaling pathway, PPAR signaling pathway, tight junctions, TNF signaling pathway, and VEGF signaling pathway.

[0026] Example 2: AEE can prevent alcohol-induced gastric mucosal damage in rats.

[0027] Forty-eight male SPF-grade SD rats (8 weeks old, weighing 220–240 g) were provided by the Lanzhou Veterinary Research Institute of the Chinese Academy of Agricultural Sciences. Animal grouping and administration are shown in Table 1.

[0028] Table 1. Grouping and administration of experimental animals

[0029]

[0030] All test drugs were suspended in 0.5% CMC-Na. Animals in each group were administered the drug once daily for 14 consecutive days. On day 15 of the experiment, after fasting for 12 hours, all rats were administered 5 mL / kg of EtOH (anhydrous ethanol) by gavage, and sacrificed 1 hour later. Blood samples were collected via the abdominal aorta. Tissue samples from organs such as the stomach were then carefully obtained, flash-frozen in liquid nitrogen and stored at -80°C, or fixed in tissue fixative for further analysis.

[0031] The results are as follows Figure 2 As shown, compared with the Control group, the EtOH group rats exhibited severe gastric ulceration, surface hemorrhage, and edema. Compared with the EtOH group, the pathological changes in the gastric tissue of rats in each drug group were alleviated, with the AEE L group showing the best effect. Therefore, in this embodiment of the invention, the low-dose AEE group was selected for subsequent experiments.

[0032] Paraffin sections were prepared from rat gastric tissue, and the effects of the drug on the pathological changes of the gastric tissue were further evaluated by HE staining. In the control group, the gastric tissue of rats showed relatively intact structures of the mucosa, submucosa, muscularis propria, and serosa. The mucosa was covered with a single layer of columnar epithelium, and tubular gastric glands were visible in the lamina propria with neatly arranged cells. Blood vessels and nerves were visible in the loose connective tissue of the submucosa, and no obvious fibrous tissue hyperplasia or inflammatory cell infiltration was observed. In the EtOH group, the gastric tissue of rats showed more severe lesions. The single layer of columnar epithelium in the mucosa was sloughed off, and many tubular gastric glands in the lamina propria showed degeneration and necrosis, with glandular atrophy, cell shedding and disintegration, accompanied by hemorrhage. Red blood cells were seen filling the extravascular interstitium, and edema fluid exudation was visible in the submucosa, accompanied by a small amount of inflammatory cell infiltration. The AEE L group and the Omeprazole group showed milder gastric tissue lesions, with a small amount of columnar epithelial shedding in the mucosa and fewer tubular gastric glands in the lamina propria showing degeneration and necrosis. Glandular atrophy, cell shedding and disintegration were observed, accompanied by a small amount of hemorrhage. Red blood cells were seen filling the extravascular interstitium, and edema fluid exudation was observed in some submucosa, accompanied by a small amount of inflammatory cell infiltration. However, the AEE L group showed better efficacy than the Omeprazole group.

[0033] In summary, compared with the Control group, the EtOH group showed more obvious pathological changes in the gastric tissue of rats, indicating successful model establishment; compared with the EtOH group, the Omeprazole group showed a reduction in the degree of lesions, and the AEE L group showed a significant reduction in the degree of lesions in the gastric tissue of rats.

[0034] AB-PAS staining was used to detect histopathological changes in the gastric epithelial mucosa, further evaluating the effect of drugs on gastric histopathological changes. In the Control group, the gastric mucosal layer of rats showed intact structure, with visible mucus secretion glycoproteins, and a superficial purplish-red color. In the EtOH group, the gastric mucosal layer of rats showed necrosis, gastric gland cell shedding, and loss of mucus cells, with significantly reduced superficial mucus secretion, obvious hemorrhage, and a large amount of erythrocyte leakage. In the AEE L and Omeprazole groups, localized mucosal necrosis, gastric gland cell shedding, and decreased mucus secretion were observed, with lighter local staining. However, the AEE L group showed better efficacy than the Omeprazole group.

[0035] In summary, compared with the Control group, the EtOH group showed more obvious pathological changes in the gastric tissue of rats, with a significant reduction in mucus secretion; compared with the EtOH group, the Omeprazole group showed a less severe lesion and a deeper mucus staining, while the AEEL group showed the mildest lesion, with more mucus secretion and a deeper staining.

[0036] The results are as follows Figure 3 As shown, compared with the Control group, the EtOH group showed significantly increased levels of MDA, p65, PUMA, IL-1β, and TNF-α in the gastric tissue of rats (P<0.05), and significantly decreased levels of SOD, ZO-1, and EGF (P<0.05). Compared with the EtOH group, the AEE group showed significantly decreased levels of MDA, p65, PUMA, IL-1β, and TNF-α in the gastric tissue of rats (P<0.05), and significantly increased levels of SOD, ZO-1, and EGF (P<0.05). The experimental results indicate that EtOH can induce oxidative stress, inflammation, apoptosis, tight junction disruption, and weakened recovery function in rat gastric tissue; while AEE can effectively protect the gastric mucosa, promote tissue repair, and alleviate the damage caused by EtOH to rat gastric tissue.

[0037] Example 3: AEE can treat alcohol-induced gastric mucosal damage in rats.

[0038] Twenty-four male SPF-grade SD rats (8 weeks old, weighing 250–270 g) were provided by the Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences. The experimental animals were randomly divided into three groups: a Control group, an EtOH group (60% EtOH 5 mL / kg), and an AEE group (60% EtOH + AEE 54 mg / kg). Rats in the EtOH and AEE groups were administered 60% EtOH (5 mL / kg) by gavage, while the Control group was administered the same volume of ultrapure water, once daily for 5 consecutive days. After model establishment, starting on day 6, rats in the AEE group were administered AEE (54 mg / kg) by gavage, while rats in the Control and EtOH groups were administered the same volume of 0.5% CMC-Na, once daily for 3 consecutive days. At the end of the experiment, blood samples were collected from the animals under anesthesia via the abdominal aorta. Then, tissue samples from organs such as the stomach were carefully obtained, flash-frozen in liquid nitrogen and stored at -80°C or fixed in tissue fixative for further analysis.

[0039] Experimental results are as follows Figure 4 As shown, in the Control group, the gastric tissue of rats exhibited relatively intact structures of the mucosa, submucosa, muscularis propria, and serosa. The mucosa was covered with a single layer of columnar epithelium, and tubular gastric glands were visible within the lamina propria, with cells arranged neatly. Blood vessels and nerves were visible within the loose connective tissue of the submucosa, without significant fibrous tissue hyperplasia or inflammatory cell infiltration. In the EtOH group, localized atrophy and necrosis of the gastric mucosa cells were observed, with thinning of the mucosa, disappearance of gastric glands in necrotic areas, and inflammatory cell infiltration. In the AEE group, submucosal edema, widened interstitial spaces, sparse fiber arrangement, and a small amount of inflammatory cell infiltration were observed in the submucosal layer. In conclusion, compared with the Control group, the EtOH group showed significant pathological changes; compared with the EtOH group, the severity of lesions in the AEE group was significantly reduced.

[0040] Experimental results are as follows Figure 5 As shown, compared with the Control group, the EtOH group showed significantly increased levels of IL-1β, MDA, and TP53 in the gastric tissue of rats (P<0.05), and significantly decreased levels of ZO-1 and VEGF (P<0.05). Compared with the EtOH group, the AEE group showed significantly decreased levels of IL-1β, MDA, and TP53 in the gastric tissue of rats (P<0.05), and significantly increased levels of ZO-1 and VEGF (P<0.05). The experimental results indicate that EtOH can induce oxidative stress, inflammation, apoptosis, disruption of tight junctions, and weakened recovery function in rat gastric tissue; while AEE can effectively protect the gastric mucosa, promote tissue repair, and alleviate the damage caused by EtOH to rat gastric tissue.

[0041] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. Application of aspirin eugenol ester in the preparation of drugs to alleviate alcohol-induced gastric mucosal damage.

2. The application as described in claim 1, characterized in that, The dosage form of the gastric mucosal injury drug is powder, tablet, granule, capsule, oral liquid, emulsion or suspension.

3. The application as described in claim 1, characterized in that, The gastric mucosal injury drug also includes a carrier and / or excipients.

4. The application as described in claim 1, characterized in that, In the aforementioned application, aspirin eugenol ester reduces the inflammatory response of human gastric mucosal epithelial cells, enhances antioxidant capacity, protects mucosal barrier function, and inhibits apoptosis and ferroptosis.

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