Application of compound indole acrylic acid in preparation of medicine for regulating intestinal epithelium necrotic apoptosis and preventing and treating neonatal necrotic enterocolitis

Abstract

The invention discloses an application of a compound indoleacrylic acid in preparation of a medicine for regulating intestinal epithelial necrotic apoptosis and preventing and treating neonatal necrotic enterocolitis, and belongs to the technical field of medicines. The compound indoleacrylic acid is used for relieving related pathological characteristics of NEC mice, including increasing the survival rate of the NEC mice, increasing the body weight of the NEC mice, reducing small intestine shortening, reducing disease activity index scores and relieving histological injury, and in addition, the indoleacrylic acid can inhibit intestinal epithelium necrotic apoptosis of the NEC mice to relieve and relieve disease symptoms; the compound indole acrylic acid is used for preparing a medicine for regulating intestinal epithelial necrotic apoptosis and preventing intestinal inflammation and intestinal necrosis.

Description

Technical Field

[0001] The invention belongs to the field of medical technology, and specifically relates to the use of a compound, indole acrylic acid, in the preparation of a drug for regulating intestinal epithelial necroptosis to prevent and treat neonatal necrotizing enterocolitis. Background Art

[0002] Necrotizing enterocolitis (NEC) is a disease characterized by intestinal inflammation and necrosis. It usually occurs in the neonatal population, especially in premature infants. It is one of the most common and serious intestinal inflammatory diseases in neonates. In extremely low-birth-weight premature infants, the mortality rate of NEC can be as high as 30-50%. This makes the prognosis of the disease extremely severe. Clinically, it mainly manifests as abdominal distension, inflammation in the body, blood in the stool, and intestinal necrosis. NEC develops rapidly and progresses rapidly, usually quickly affecting the health of newborns. The intestinal epithelium is a key barrier against pathogens and toxins in the intestinal cavity. Epithelial damage is a hallmark of NEC. The pathogenesis of the disease involves local necrosis of intestinal tissue caused by various reasons. When the intestinal flora is disturbed, the endotoxins released by bacteria can induce the production of inflammatory factors, further mediate the death of intestinal epithelial cells, and ultimately lead to intestinal inflammation and necrosis. Necroptosis is a key mechanism that drives intestinal epithelial cell death and damage, involving receptor-interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL). In addition, short bowel syndrome, intestinal stenosis, and neurodevelopmental disorders are common sequelae in surviving children after recovery, which seriously affect their quality of life and physical and mental development. Therefore, regulating intestinal epithelial necroptosis will provide a new idea for preventing and treating the occurrence and development of neonatal necrotizing enterocolitis.

[0003] In recent years, NEC-related research and clinical data have shown that traditional drug treatment accounts for less than 15% of the treatment methods for NEC. As a serious gastrointestinal emergency in the neonatal period, the incidence and mortality of NEC are high in the premature infant population, but the progress of drug development is very slow. Most of the drugs currently used for NEC in clinical practice have significant side effects, such as intestinal flora disorders, immune function suppression, affecting the normal growth and development of newborns, causing electrolyte disorders and other adverse reactions, which greatly limit their wide application in clinical treatment. For children with severe NEC, surgical treatment is mainly relied on, such as intestinal resection and anastomosis, intestinal fistula, etc. These operations are not only physically traumatic to the children, but also have a high risk of complications after surgery, such as anastomotic leakage, short bowel syndrome, etc. At the same time, the cost of treatment is high, which brings a heavy economic burden to the family. Therefore, the exploration of new drugs and innovative treatment strategies that can effectively inhibit intestinal inflammatory overreactions, promote the repair of damaged intestinal mucosa, and are highly safe is still a major neonatal health problem that needs to be solved urgently. Summary of the invention

[0004] In order to overcome the deficiencies of the prior art, the present invention provides a pharmaceutical application of a compound indole acrylic acid for preventing and treating neonatal necrotizing enterocolitis by inhibiting intestinal epithelial necroptosis. The compound indole acrylic acid is an indole compound produced by intestinal microorganisms metabolizing tryptophan, and its therapeutic effect in necrotizing enterocolitis has not been reported.

[0005] The present invention provides application of a compound of indoleacrylic acid or a pharmaceutically acceptable salt thereof in preparing a medicine for preventing and treating neonatal necrotizing enterocolitis.

[0006] The present invention provides the use of the compound indole acrylic acid or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and treating intestinal-related diseases mediated by intestinal epithelial cell necrosis.

[0007] The present invention also provides the use of the compound indoleacrylic acid or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating and relieving the relevant clinical symptoms after necrotizing enterocolitis.

[0008] In one embodiment of the present invention, the related clinical symptoms include short bowel syndrome, intestinal stenosis, and intestinal epithelial necrosis.

[0009] In one embodiment of the present invention, during the application, the compound indole acrylic acid exhibits inhibition of intestinal epithelial necroptosis, proving that the compound plays a key role in the development of necrotizing enterocolitis and intestinal epithelial cell necrosis. Therefore, the compound indole acrylic acid of the present invention can be used to prepare drugs for treating and relieving neonatal necrotizing enterocolitis, and can be used to prepare drugs for relieving intestinal diseases related to intestinal epithelial necrosis.

[0010] The present invention provides the use of a compound of indoleacrylic acid or a pharmaceutically acceptable salt thereof in the preparation of a drug for regulating intestinal epithelial necroptosis and preventing and treating neonatal necrotizing enterocolitis.

[0011] In one embodiment of the present invention, the structural formula of the compound indole acrylic acid is as follows:

[0012]

[0013] In one embodiment of the present invention, the pharmaceutically acceptable salt is an inorganic salt or an organic salt; wherein the inorganic salt is selected from hydrochloride, hydrobromide, hydroiodide, perchlorate, sulfate, bisulfate, nitrate, phosphate, acid phosphate, sodium salt, calcium salt, potassium salt, magnesium salt, silver salt, and lithium salt; and the organic salt is selected from formate, acetate, trifluoroacetate, propionate, pyruvate, glycolate, oxalate, malonate, succinate, glutarate, fumarate, maleate, lactate, malate, citrate, tartrate, methanesulfonate, ethanesulfonate, benzenesulfonate, salicylate, p-toluenesulfonate, ascorbate, meglumine salt, tromethamine salt, diethylamine salt, lysine salt, choline salt, arginine salt, terbutylamine salt, and N,N-dibenzylethylenediamine salt.

[0014] In one embodiment of the present invention, the compound indoleacrylic acid is used to prepare a drug for inhibiting necroptosis.

[0015] In one embodiment of the present invention, the compound indoleacrylic acid is used to prepare a drug for inhibiting intestinal epithelial necroptosis.

[0016] In the scheme of the present invention, the indoleacrylic acid, a compound derived from intestinal microorganisms, has the purpose of inhibiting intestinal epithelial necroptosis and is used in the preparation of a drug for regulating intestinal epithelial necroptosis to prevent and treat neonatal necrotizing enterocolitis.

[0017] The present invention found that indole acrylic acid dose-dependently reduced NEC-related pathological features at the animal level, including improving the survival rate of NEC mice, increasing the weight of NEC mice, reducing small intestinal shortening and disease activity index score reduction, and reducing histological damage. Indole acrylic acid can significantly inhibit the expression of necrosis-related proteins in the ileum tissue of NEC mice. Western blot results showed that indole acrylic acid can significantly inhibit the phosphorylation levels of RIPK3 and MLKL in the ileum tissue of NEC mice; immunofluorescence experiments showed that indole acrylic acid can reduce the number of p-RIPK3 positive cells in intestinal epithelial cells. Therefore, the compound indole acrylic acid of the present invention has a good application prospect and can be used to prepare a drug for regulating intestinal epithelial necroptosis to treat necrotizing enterocolitis.

[0018] In one embodiment of the present invention, the dosage form of the drug includes decoction, pill, powder, ointment, pill, wine, syrup, extract, lozenge, stick, suppository, quji, and roasted medicine.

[0019] In one embodiment of the present invention, the dosage form of the drug includes tablets, granules, bags, oral liquids, capsules, pills, mixtures, tinctures, aerosols, films, injections, and injections.

[0020] In one embodiment of the present invention, the drug further contains medically acceptable excipients, and the medically acceptable excipients include adhesives, fillers, disintegrants, lubricants, antioxidants, flavoring agents, solubilizers, emulsifiers, solubilizers, osmotic pressure regulators, and colorants.

[0021] In one embodiment of the present invention, the drug further contains a pharmaceutically acceptable carrier.

[0022] In one embodiment of the present invention, the pharmaceutically acceptable carrier includes microcapsules, microspheres, nanoparticles and liposomes.

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

[0024] (1) The compound indoleacrylic acid has a strong effect in alleviating NEC-related pathological features;

[0025] (2) The compound indoleacrylic acid can inhibit intestinal epithelial necroptosis and can be used to prepare drugs for the prevention and treatment of necrotizing enterocolitis;

[0026] (3) The compound indoleacrylic acid showed the effect of alleviating NEC-related pathological characteristics, proving that it plays a key role in the occurrence and development of intestinal inflammation and short bowel syndrome. Therefore, the compound indoleacrylic acid of the present invention can be used to prepare drugs for alleviating intestinal inflammation, and can be used to prepare drugs for treating and alleviating short bowel syndrome in the prognosis of NEC. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 The results of the effect of the compound indole acrylic acid on the survival rate, weight change and disease index of NEC mice. (A) The survival rate of NEC mice treated with indole acrylic acid was recorded every 6 hours; (B) The weight change of NEC mice treated with indole acrylic acid was recorded every 6 hours; (C) The disease index score of NEC mice treated with indole acrylic acid. The results showed that indole acrylic acid had a significant effect on improving the survival rate of NEC mice, improving weight loss and disease index scores, that is, indole acrylic acid can alleviate the disease symptoms of NEC mice. *P<0.05, **P<0.01, ***P<0.001.

[0028] Figure 2 The results of the effect of the compound indole acrylic acid on the intestinal pathological damage of NEC mice are shown in Figure 1. The results of the blank group, NEC model group, and indole acrylic acid treatment group in improving the length of the small intestine of NEC mice (A); and the results of improving the shortening of villi and crypt loss in the ileum of NEC mice, that is, alleviating the intestinal pathological damage of NEC mice (B). *P<0.05, **P<0.01, ***P<0.001.

[0029] Figure 3 This is a graph showing the effect of the compound indoleacrylic acid on necroptosis-related proteins in the ileum of NEC mice.

[0030] Western Blot was used to detect the phosphorylation levels of necroptosis markers MLKL and RIPK3. IA treatment significantly reduced

[0031] Phosphorylation levels of RIPK3 and MLKL in the ileum of NEC mice. The results showed that indoleacrylic acid could inhibit necroptosis in the ileum of NEC mice. *P<0.05, **P<0.01, ***P<0.001.

[0032] Figure 4 The results of the effect of the compound indole acrylic acid on proteins related to necroptosis of intestinal epithelial cells in NEC mice. Immunofluorescence staining of p-RIPK3 and FABP2 (intestinal epithelial cell marker protein) showed that p-RIPK3 was mainly present in the intestinal epithelial cells of NEC mice, and the number of positive cells decreased after indole acrylic acid treatment. The results show that indole acrylic acid inhibits intestinal epithelial cell necroptosis to alleviate NEC. *P<0.05, **P<0.01, ***P<0.001. DETAILED DESCRIPTION

[0033] The present invention is further described below in conjunction with examples. Due to space constraints, the description of the experimental process cannot be very detailed. All parts not described in detail in the experiment are routine operations well known to those skilled in the art, but the examples are by no means any limitation to the present invention.

[0034] The methods used in the following examples are all conventional methods unless otherwise specified, and the reagents required in the following implementations are all purchased from the market unless otherwise specified.

[0035] The data analysis in the examples were all performed using Graphpad Prism 10.0 statistical analysis software for One-Way ANOVA multi-group comparisons.

[0036] Example 1: Effects of compounds on survival rate, body weight change and disease index of NEC mice

[0037] Newborn mice were randomly divided into five groups, namely control group, NEC group, and IA group (25, 50, and 75 mg / kg). From day 7 to day 10, mice were continuously given low, medium, and high doses of IA, and TNBS (50 mg / kg) was given on day 10 to induce the NEC model. After modeling, the survival, weight changes, and DAI scores of the five groups of mice were observed and recorded every 3 hours.

[0038] The results are as follows Figure 1 The results showed that indoleacrylic acid had a significant effect on improving the survival rate of NEC mice (A), improving weight loss (B) and disease index score (C), that is, indoleacrylic acid can alleviate the disease symptoms of NEC mice. *P<0.05, **P<0.01, ***P<0.001.

[0039] Example 2: Effect of the compound indoleacrylic acid on intestinal pathological damage in NEC mice

[0040] After 12 hours of TNBS-induced NEC model, the changes in the length of the small intestine of the five groups of mice were detected. It was found that the length of the small intestine of NEC mice was significantly shorter than that of the control group. Indoleacrylic acid treatment significantly improved the shortening of the small intestine of NEC mice. The terminal ileum tissue was taken for paraffin section and H&E staining.

[0041] The results are as follows Figure 2 As shown, (A) shows that indole acrylic acid treatment significantly improved the shortening of the small intestine of NEC mice; (B) indole acrylic acid can significantly improve the shortening of ileal villi and crypt loss in NEC mice, that is, indole acrylic acid alleviates the intestinal pathological damage of NEC mice. *P<0.05, **P<0.01, ***P<0.001. The results showed that the length of the small intestine of NEC mice was significantly shortened compared with that of the control group mice, and indole acrylic acid treatment significantly improved the shortening of the small intestine of NEC mice; H&E staining results showed that the villi of the ileum of NEC group mice were disordered and significantly shortened or even fell off, the muscular layer was broken, and the crypt structure was missing and significantly damaged; the villi of the ileum of mice in the IA (25mg / kg) + NEC group were more disordered and the villi were shorter, and the crypt structure was less damaged; the villi of the ileum of mice in the IA (50 and 75mg / kg) + NEC group were neatly arranged and the villi were longer, the muscular layer was intact with slight damage, and the crypt structure was relatively intact. The pathological damage scores of the ileum tissue of each group of mice were scored by H&E staining. The results showed that the scores of the NEC group were significantly higher than those of the control group, while the scores after IA treatment were significantly lower than those of the NEC group. The morphological changes in the ileum tissue of mice showed that IA could alleviate the pathological damage of the ileum caused by NEC.

[0042] Example 3: Effects of the compound indoleacrylic acid on necroptosis-related proteins in the ileum of NEC mice

[0043] 12 hours after TNBS induced the NEC model, the terminal ileum tissue of each group of mice was taken out, and the phosphorylated protein expression of receptor interacting serine / threonine kinase (RIPK3) and mixed lineage kinase domain-like protein (MLKL) was detected by Western blot. The Western blot method was briefly described as follows: the total protein of ileum tissue was extracted and quantified, 30 μg of the total protein was separated by SDS-PAGE and transferred to a nitrocellulose membrane, blocked with 5% skim milk, incubated with the above antibodies at 4°C overnight, and detected with chemiluminescence reagent after incubation with secondary antibodies for 2 hours.

[0044] The results are as follows Figure 3 As shown, indoleacrylic acid treatment significantly reduced the phosphorylation levels of RIPK3 and MLKL in the ileum tissue of NEC mice. The results showed that indoleacrylic acid could inhibit necroptosis in the ileum tissue of NEC mice. *P<0.05, **P<0.01, ***P<0.001.

[0045] Example 4: Effect of the compound indoleacrylic acid on necroptosis-related proteins in intestinal epithelial cells of NEC mice

[0046] 12 hours after TNBS induced the NEC model, the terminal ileum tissue of each group of mice was removed and fixed and embedded for paraffin sectioning to detect the effect of the compound indoleacrylic acid on necroptosis-related proteins of intestinal epithelial cells in NEC mice.

[0047] The results are as follows Figure 4 As shown, immunofluorescence staining of p-RIPK3 and FABP2 (intestinal epithelial cell marker protein) showed that p-RIPK3 was mainly present in the intestinal epithelial cells of NEC mice, and the number of positive cells decreased after indoleacrylic acid treatment. The results showed that indoleacrylic acid inhibited intestinal epithelial cell necroptosis to alleviate NEC. *P<0.05, **P<0.01, ***P<0.001.

Claims

1. Use of the compound indoleacrylic acid or a pharmaceutically acceptable salt thereof in the preparation of a medicament for preventing and treating neonatal necrotizing enterocolitis.

2. Use of the compound indoleacrylic acid or a pharmaceutically acceptable salt thereof in the preparation of a drug for preventing and treating intestinal-related diseases mediated by intestinal epithelial cell necrosis.

3. Use of the compound indoleacrylic acid or a pharmaceutically acceptable salt thereof in the preparation of a drug for treating and relieving clinical symptoms associated with neonatal necrotizing enterocolitis.

4. The use according to claim 3, characterized in that: The related clinical symptoms include short bowel syndrome, intestinal stricture, and intestinal epithelial necrosis.

5. The use according to any one of claims 1 to 3, characterized in that: The pharmaceutically acceptable salt is an inorganic salt or an organic salt; wherein the inorganic salt is selected from hydrochloride, hydrobromide, hydroiodide, perchlorate, sulfate, bisulfate, nitrate, phosphate, acid phosphate, sodium salt, calcium salt, potassium salt, magnesium salt, silver salt, and lithium salt; and the organic salt is selected from formate, acetate, trifluoroacetate, propionate, pyruvate, glycolate, oxalate, malonate, succinate, glutarate, fumarate, maleate, lactate, malate, citrate, tartrate, methanesulfonate, ethanesulfonate, benzenesulfonate, salicylate, p-toluenesulfonate, ascorbate, meglumine salt, tromethamine salt, diethylamine salt, lysine salt, choline salt, arginine salt, terbutylamine salt, and N,N-dibenzylethylenediamine salt.

6. The use according to any one of claims 1 to 3, characterized in that: The dosage forms of the medicine include decoction, pill, powder, ointment, pill, wine, syrup, extract, lozenge, stick, suppository, quji and roasted medicine.

7. The use according to any one of claims 1 to 3, characterized in that: The dosage forms of the medicine include tablets, granules, bags, oral liquids, capsules, pills, mixtures, tinctures, aerosols, films, injections and injections.

8. The use according to any one of claims 1 to 3, characterized in that: The medicine also contains medically acceptable excipients, which include adhesives, fillers, disintegrants, lubricants, antioxidants, flavoring agents, solubilizers, emulsifiers, solubilizers, osmotic pressure regulators, and colorants.

9. The use according to any one of claims 1 to 3, characterized in that: The drug also contains a pharmaceutically acceptable carrier.

10. The use according to claim 9, characterized in that: The pharmaceutical carriers include microcapsules, microspheres, nanoparticles and liposomes.

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