CN114705871A - Application of 8-isoprostaglandin F2alpha in diagnosis of severe asthma - Google Patents

Abstract

The invention discloses the application of 8-isoprostaglandin F2α in the diagnosis of severe asthma. The invention provides the application of 8-isoprostaglandin F2α as a biomarker in the preparation of products for diagnosing or assisting in the diagnosis of severe asthma. The present invention detects the concentration of 8-isoprostaglandin F2α in the plasma and induced sputum supernatant of asthma patients, and finds that 8 The concentration of isoprostaglandin F2α was significantly different in the test samples of patients with severe asthma (SA) and patients with non-severe asthma (NSA). The pulmonary function index is in a negative correlation trend; the biomarker 8-iso-PGF2α provided by the present invention can be conveniently detected by ELISA, and the present invention can diagnose severe asthma in a simpler manner.

Description

Application of 8 isoprostaglandin F2α in the diagnosis of severe asthma

technical field

The invention relates to the application of 8-isoprostaglandin F2α in the diagnosis of severe asthma, and belongs to the technical field of biomedical detection.

Background technique

The diagnosis of severe asthma is carried out according to the expert consensus of GINA and the asthma group of the Respiratory Branch of the Chinese Medical Association. Generally, the disease control can only be achieved by the application of grade 4 or 5 anti-asthma treatment or the disease is difficult to control once the treatment grade is lowered. Therefore, the judgment of severe asthma is generally delayed, and a period of follow-up observation of specific patients is required to accurately diagnose.

8-iso-prostaglandin F2α (8-iso-prostaglandin F2alpha, 8-iso-PGF2α) is derived from free radical-catalyzed arachidonic acid peroxidation, and its concentration can reflect the level of oxidative stress damage in the body, and is the most reliable oxidative stress. one of the biomarkers. Biofilms exist widely in the body, and lipids are easily attacked by free radicals, resulting in the production of peroxidative damage products. The peroxidation of membrane lipids can lead to the scission of fatty acid chains, aggravate airway inflammation and cause tissue damage and morphological changes.

Lipid peroxidation is also considered to be closely related to asthma, and elevated concentrations of 8-iso-PGF2α and its congeners have been found in exhaled condensate, induced sputum, peripheral blood and even urine of asthmatic patients. Its concentration in the blood of asthmatic patients was significantly increased in the acute exacerbation period of asthma, and although it was reduced in the remission period, it was still significantly higher than that in the healthy population. However, there is no report on the use of 8-isoprostaglandin F2α as a diagnostic marker for severe asthma.

SUMMARY OF THE INVENTION

The present invention aims to provide an oxidative stress biomarker capable of assessing the severity of asthma, so as to conveniently and accurately diagnose severe asthma.

In order to achieve the above object, the present invention provides the application of 8-isoprostaglandin F2α as a biomarker in preparing a product for diagnosing or assisting in the diagnosis of severe asthma. The application of the biomarker is: by detecting the concentration of 8-isoprostaglandin F2α in the biological sample to determine whether the patient is a severe asthma patient, the method for detecting the concentration of 8-isoprostaglandin F2α can adopt GC-MS, LC-MS and ELISA method. etc. Among them, ELISA method is the most convenient and fast.

Preferably, the product includes a reagent capable of detecting the concentration of 8-isoprostaglandin F2α in a sample.

Preferably, the sample comprises patient plasma and/or induced sputum supernatant.

Preferably, the product includes a reagent for detecting the concentration of 8-isoprostaglandin F2α in a sample by ELISA.

Preferably, the reagent for detecting the concentration of 8-isoprostaglandin F2α in the sample by ELISA is an ELISA kit

Compared with the prior art, the beneficial effects of the present invention are:

The present invention proposes for the first time that 8-isoprostaglandin F2α is used as a diagnostic marker for severe asthma. The present invention finds that the concentration of 8-isoprostaglandin F2α is significantly higher in severe asthma by detecting the concentration of 8-isoprostaglandin F2α in the plasma and induced sputum supernatant of asthma patients. There were significant differences in the test samples of patients with asthma (SA) and non-severe asthma patients (NSA). At the same time, by detecting the pulmonary function indexes of asthma patients, it was found that the concentration of 8-iso-PGF2α in the test samples was negatively correlated with the pulmonary function indexes. The biomarker 8-iso-PGF2α provided by the invention can be conveniently detected by ELISA, and the invention can diagnose severe asthma in a simpler manner.

Description of drawings

Figure 1 shows the relationship between the concentration of 8-iso-PGF2α in induced sputum and pulmonary function parameters in asthma patients.

Detailed ways

In order to make the present invention more obvious and comprehensible, preferred embodiments are described in detail below with reference to the accompanying drawings.

In the embodiments of the present invention, asthma patients are recruited by the following methods: the inclusion criteria are in line with my country's "Guidelines for the Prevention and Treatment of Bronchial Asthma" (2016 edition), and the condition is stable within the past 4 weeks, that is, there are no respiratory tract infections, acute asthma attacks, etc. Adult asthma patients. For confirmed asthma patients who meet the inclusion criteria, according to their treatment status and control level, they are divided into severe asthma patients and mild-to-moderate asthma patients with reference to my country's "Guidelines for the Prevention and Treatment of Bronchial Asthma" (2016 edition). There were 25 cases in asthma (SA) group and 42 cases in non-severe asthma (NSA) group.

In the embodiment of the present invention, the plasma and induced sputum supernatant of asthma patients were collected by the following methods: 2 mL of peripheral venous blood of asthma subjects was extracted and stored in a 3.6 mg EDTA anticoagulant tube at room temperature, and the upper plasma was drawn by centrifugation. Referring to the "Guidelines for the Diagnosis and Treatment of Cough" (2015 edition) of the Asthma Group of the Respiratory Disease Branch of the Chinese Medical Association, the method for detecting inflammatory cells in the sputum induced by hypertonic saline aerosolization was performed to collect the induced sputum and collect the supernatant.

In the embodiment of the present invention, the concentration of 8-iso-PGF2α is detected by ELISA: operate according to the instructions of the ELISA kit: add 100uL standard substance or sample to be tested to each well and mix well, measure 3 replicate wells for each sample, and set the reaction The plate was incubated at a constant temperature of 37 °C for 40 min, and then the reaction plate was fully washed 6 times with washing solution and dried with filter paper. Add 50uL of double-distilled water and primary antibody working solution to each well and mix well (except blank control wells), incubate the reaction plate at 37°C for 20 minutes, wash the plate, then add 100uL of enzyme-labeled antibody working solution, mix well, set the reaction plate Wash the plate after incubating at 37°C for 10 min. Add 100uL of substrate working solution to each well, react at 37°C in the dark for 15min, add 100uL of stop solution and mix well. Measure the absorbance value at 450nm with a microplate reader within 30min, and calculate the corresponding concentration according to the OD value of the sample.

In the embodiment of the present invention, the lung function of asthma patients is detected by the following method: within the first 24 hours after the patient completes the lung function test, long-acting β2 receptor agonists or anticholinergic drugs shall not be used, and short-term β2 receptor agonists or anticholinergic drugs shall not be used within the first 12 hours. potent β2-receptor agonists. Operated by a professional technician in the Pulmonary Function Room of the Department of Respiratory Medicine, Zhongshan Hospital, using the Master Screen-PFT pulmonary function instrument (Jaeger, Germany), the test is performed at about 25 °C from 8:00 am to 11:00 am every day, and the instrument is calibrated before the measurement. And calibration, in strict accordance with the operating standards, and the measurement results meet the quality control requirements at least three times, take the best results in three consecutive tests, record FEV1%pred, FEV1/FVC, FVC%pred, DLCO%pred and other indicators.

Example

This example provides the application of 8-isoprostaglandin F2α in the diagnosis of severe asthma:

The plasma and induced sputum supernatants were collected from the recruited severe and non-severe asthma patients, and 8-iso-PGF2α was detected by ELISA. The results showed that the concentrations of 8-iso-PGF2α in peripheral blood (162.41±307.32vs.61.39±58.78, P=0.057) and induced sputum (239.92±234.13vs.106.39±97.78, P=0.022) of patients with severe asthma were higher than those in non- For patients with severe asthma, the specific data are shown in Table 1. In order to further evaluate the effect of 8-iso-PGF2α concentration on pulmonary function in asthma patients, we detected the pulmonary function indexes of asthma patients, and performed linear regression on the 8-iso-PGF2α concentration in peripheral blood and induced sputum of asthma patients and pulmonary function parameters. It showed that the concentration of 8-iso-PGF2α in induced sputum was negatively correlated with FEV1/FVC, DLCO%pred (β=-0.030, P=0.003) (β=-0.046, P=0.013), and was significantly correlated with FVC%pred, FEV1%pred The negative correlation trend is shown in Figure 1. That is, the higher the sputum 8-iso-PGF2α concentration of asthmatic patients, the more serious the obstructive ventilation function and diffusion dysfunction. Therefore, plasma/induced sputum supernatant 8-iso-PGF2α has a good application prospect for the diagnosis of severe asthma. The present invention can diagnose severe asthma in a simpler manner, and based on the characteristics of the biomarkers, it is expected to prepare a more reliable and sensitive kit for diagnosing severe asthma.

Table 1 Comparison of 8-iso-PGF2α concentrations in SA and NSA patients

The above-mentioned embodiments are only the preferred embodiments of the present invention, and are not intended to limit the present invention in any form and substance. It should be pointed out that those of ordinary skill in the art can make Several improvements and supplements should also be regarded as the protection scope of the present invention.

Claims (5)

1.8 The application of isoprostaglandin F2α as a biomarker in the preparation of a product for diagnosing or assisting in the diagnosis of severe asthma. 2. The use of claim 1, wherein the product comprises a reagent capable of detecting the concentration of 8-isoprostaglandin F2α in a sample. 3. The use of claim 2, wherein the sample comprises patient plasma and/or induced sputum supernatant. The application according to any one of claims 1 to 3, wherein the product comprises a reagent for detecting the concentration of 8-isoprostaglandin F2α in the sample by ELISA. 5 . The application according to claim 4 , wherein the reagent for detecting the concentration of 8-isoprostaglandin F2α in the sample by ELISA is an ELISA kit. 6 .

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