Application of indole-3-propionic acid in preparation of medicine for preventing and treating pulmonary ischemia reperfusion injury

Abstract

The invention relates to application of indole-3-propionic acid in preparation of drugs for preventing and treating pulmonary ischemia reperfusion injury. Through in-vivo animals, the inventor finds that indole-3-propionic acid can reduce the lung injury score in a concentration-dependent manner, reduce the wet weight / dry weight ratio of lungs, relieve active oxygen accumulation after lung ischemia reperfusion, and reduce the levels of total protein, tumor necrosis factor alpha and interleukin 6 in alveolar lavage fluid; the cell level also shows that the indole-3-propionic acid can improve the vitality of alveolar epithelial cells after ischemia / reperfusion injury simulation in a concentration-dependent manner, reduce release of inflammatory factors such as tumor necrosis factors alpha, interleukin 6 and interleukin 1beta, and reduce accumulation of active oxygen in the cells. Results of integrating in-vitro cells and in-vivo animals show that the indole-3-propionic acid has the effect of preventing and treating the pulmonary ischemia reperfusion injury and has a good clinical application prospect.

Description

Technical Field

[0001] The present invention relates to a new application of indole-3-propionic acid, and in particular to an application of indole-3-propionic acid in preventing and treating pulmonary ischemia-reperfusion injury. Background Art

[0002] Indole-3-propionic acid is a metabolite produced by intestinal bacteria. Existing technology reports that it has the effects of reducing inflammation, anti-oxidation, scavenging free radicals, reducing oxidative stress, protecting neurons, and regulating intestinal microbiota and metabolism. Summary of the invention

[0003] The inventors found in vivo that indole-3-propionic acid can reduce lung injury scores, reduce lung wet weight / dry weight ratio, reduce reactive oxygen accumulation after lung ischemia-reperfusion, and reduce the levels of total protein, tumor necrosis factor α and interleukin 6 in bronchoalveolar lavage fluid; at the cellular level, it was also found that indole-3-propionic acid can increase the viability of alveolar epithelial cells after simulated ischemia / reperfusion injury in a concentration-dependent manner, reduce the release of inflammatory factors such as tumor necrosis factor α, interleukin 6 and interleukin 1β, and reduce the accumulation of intracellular reactive oxygen. The results of in vitro cells and in vivo animals show that indole-3-propionic acid has the effect of preventing and treating lung ischemia-reperfusion injury and has a good prospect for clinical application.

[0004] Based on this, the present invention provides the use of indole-3-propionic acid and pharmaceutically acceptable salts thereof for preparing a drug for preventing and treating pulmonary ischemia-reperfusion injury.

[0005] Optionally, the drug is an intravenous injection preparation, an oral preparation or an aerosol inhalation preparation. BRIEF DESCRIPTION OF THE DRAWINGS

[0006] Figure 1 The results of the invention are related to the effect of indole-3-propionic acid on reducing lung ischemia-reperfusion injury in mice; A: ratio of lung wet weight to dry weight; B: score of mouse lung injury; C: detection of reactive oxygen content in lung tissue; D: total protein content in lung lavage fluid; E: content of tumor necrosis factor α in lung lavage fluid; F: content of interleukin-6 in lung lavage fluid; W / D, wet weight / dry weight; BALF, bronchoalveolar lavage fluid; TNF-α, tumor necrosis factor α; IL-6, interleukin-6; DMSO, dimethyl sulfoxide; Sham, sham operation group; LI / R, lung ischemia / reperfusion. *P<0.05, *P<0.01.

[0007] Figure 2These are the results related to indole-3-propionic acid alleviating simulated ischemia-reperfusion injury of alveolar epithelial cells in the embodiments of the present invention; wherein: A cell viability detection; B flow cytometry detection of cell apoptosis level; C cell reactive oxygen staining; D tumor necrosis factor α content in cell culture supernatant; E interleukin-1β content in cell culture supernatant; F interleukin-6 content in cell culture supernatant; TNF-α, tumor necrosis factor α; IL-1β, interleukin-1β; interleukin-6, IL-6; DMSO, dimethyl sulfoxide; Con, control group; H / R, hypoxia-reoxygenation; *P<0.05, *P<0.01. DETAILED DESCRIPTION

[0008] Unless otherwise specified, the scientific and technical terms used herein are understood according to the knowledge of ordinary technicians in the relevant fields.

[0009] Pulmonary ischemia / reperfusion injury refers to a pathological phenomenon in which the degree of lung tissue damage rapidly increases after the lung tissue has been ischemic for a certain period of time and then blood perfusion (reperfusion) is restored. Clinically, lung transplantation, hemorrhagic shock, pulmonary embolism, CPB, etc. can all lead to pulmonary ischemia / reperfusion injury. The commonly used models of pulmonary ischemia and reperfusion injury in drug and clinical research are the lung ex vivo reperfusion model and the hilar clamping model. The following examples use the hilar clamping model, and the mice used are the inbred strain experimental mice C57BL / 6. The drug dosage in the following examples is ≥50 mg / kg body weight. The specific dosage is 50 mg / kg body weight to 100 mg / kg body weight.

[0010] The present invention is further described below by way of examples. The examples are provided to illustrate the present invention in detail, but the present invention is not limited thereto. The animals, cells, drugs and reagents used in the following examples are all commercially available products.

[0011] Example 1: Indole-3-propionic acid (IPA) reduces lung ischemia-reperfusion injury in mice

[0012] Group settings: sham operation group (Sham), ischemia reperfusion + vehicle group (control group, L1 / R+Vehicle), ischemia reperfusion + low-dose IPA group (L1 / R+Low) and ischemia reperfusion + high-dose IPA group (L1 / R+High).

[0013] Drug: IPA was dissolved in dimethyl sulfoxide (DMSO) to prepare IPA mother solution (IPA concentration in the mother solution was 189.21 mg / mL), and when used, it was diluted to the required concentration with PBS solution. The control group was added with the corresponding solvent (Vehicle,) which was PBS solution containing 10% DMSO.

[0014] Construction of ischemia-reperfusion model (L1 / R model): After anesthesia and endotracheal intubation, the left lung hilum (including pulmonary artery, pulmonary vein and left main bronchus) was ligated with surgical thread, and the ligature was loosened for reperfusion after 60 minutes to establish the mouse lung ischemia-reperfusion injury model. The mice in the sham group also underwent anesthesia, endotracheal intubation and surgical thread through the hilum, but no ligation and loosening were performed.

[0015] Administration: 3 days before the small animal surgery, IPA was supplemented to mice by oral gavage three times (24 hours apart each time) at a dose of 50 mg IPA / kg body weight / day (low-dose group) and 100 mg IPA / kg body weight / day (high-dose group); the control group was given an equal amount of solvent (DMSO+PBS solution).

[0016] After 2 hours of reperfusion, the mice were anesthetized and killed for relevant tests.

[0017] The results are as follows Figure 1 As shown in Figure 2, after IPA treatment, the ratio of lung wet weight to dry weight decreased with increasing IPA dose ( Figure 1 A); lung injury scores were also significantly reduced ( Figure 1 DHE staining showed that ROS accumulation in lung tissue increased after LI / R, while with the increase of IPA supplementation level, ROS accumulation in lung tissue decreased significantly ( Figure 1 C); In addition, the content of total protein and inflammatory factors (TNF- and IL-6) in lung lavage fluid also decreased with the increase of IPA supplementation level ( Figure 1 DF). It can be seen that after IPA treatment, lung injury in LI / R mice was alleviated in a dose-dependent manner.

[0018] Example 2: Indole-3-propionic acid reduces simulated ischemia-reperfusion injury in alveolar epithelial cells

[0019] Group settings: blank + solvent group (Con + DMSO), simulated ischemia reperfusion + solvent group (H / R + DMSO), simulated ischemia reperfusion + low-dose IPA group (H / R + 0.1mM) and simulated ischemia reperfusion + high-dose IPA group (H / R + 1mM).

[0020] Simulated ischemia-reperfusion model: MLE-12 mouse alveolar epithelial cells (purchased from ATCC Collection Center) were inoculated in 96-well plates / 6-well plates or confocal dishes; after culturing with serum-containing medium for 36 hours, the cells covered 80% of the culture bottom area, and then replaced with serum-free medium and continued to be cultured for 8 hours. The cells treated as above were placed in an oxygen-free box and nitrogen was passed to establish an oxygen-free environment. After 3 hours, the cells were transferred to an incubator and reoxygenated in a conventional environment for 8 hours.

[0021] Administration: IPA of corresponding concentration (low-dose group: IPA 0.1 mM, high-dose group: IPA 1 mM) was added to the culture medium until the end of the experiment; corresponding amount of DMSO was added to the control group.

[0022] Cell viability assay showed that the viability of MLE-12 cells was significantly increased after IPA treatment, and the viability of cells treated with high-dose IPA was higher than that of cells treated with low-dose IPA ( Figure 2 A); Flow cytometry showed that IPA significantly reduced the apoptosis level of MLE-12 cells ( Figure 2 B); The results of reactive oxygen species staining showed that IPA could reduce the accumulation of intracellular reactive oxygen species caused by simulated ischemia-reperfusion in a dose-dependent manner ( Figure 2 C); Further tests showed that the levels of inflammatory factors such as TNF-α, IL-1β and IL-6 in the culture supernatant also decreased with the increase of IPA dose ( Figure 2 DF).

Claims

1. Use of indole-3-propionic acid and its pharmaceutically acceptable salts for preparing drugs for preventing and treating pulmonary ischemia-reperfusion injury.

2. The use according to claim 1, characterized in that: The medicine is an intravenous injection preparation, an oral preparation or an aerosol inhalation preparation.

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