PM 2.5 Biomarkers of offspring cardiotoxicity and their applications

Abstract

The present invention discloses PM 2.5 Biomarkers of offspring developmental cardiotoxicity and their applications. 2.5 After the onset of leukemia, the expression of Angptl4 in the heart tissue of the offspring was significantly upregulated, while the expression of Sirt3 was significantly downregulated. 2.5 The impact of PM on offspring's cardiac development is 2.5 Provides methods for the diagnosis and evaluation of offspring developmental cardiotoxicity.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to PM 2.5 Biomarkers of offspring cardiac developmental toxicity and their applications. Background Art

[0002] Congenital heart disease (CHD) is the most common type of birth defect worldwide and an important factor in neonatal and child mortality. Therefore, understanding the environmental risk factors and mechanisms that cause the incidence and prevalence of CHD has important practical significance for global public health. The 2021 Global Burden of Disease analysis showed that PM 2.5 It is the primary cause of disease burden, accounting for 8.0% of the total disability-adjusted life years. A case-control study involving 1,434,998 newborns found that PM during pregnancy 2.5 For every 10 μg / m increase in exposure 3 , the risk of CHD will increase by 2%. Animal experimental studies have found that exposure to PM during pregnancy to weaning at 3 weeks of age 2.5 It can cause increased collagen deposition and histopathological changes in the myocardial tissue of offspring. 2.5 Exposure leads to abnormal expression of mitochondrial fusion / fission genes in offspring mice, which in turn leads to cardiac damage in offspring mice. 2.5 The precise underlying mechanisms and robust biomarkers of exposure leading to offspring developmental cardiotoxicity remain complex puzzles.

[0003] Angiopoietin-like protein 4 (Angptl4) interacts with CHD-related genes and known pathogenic genes, playing an important role in the regulation of CHD. Sirt3 is a deacetylase primarily located in the mitochondrial matrix. Existing studies have not reported the role of Angptl4 / Sirt3 as a biomarker in PM during pregnancy. 2.5 Application in developmental cardiac toxicity in offspring after exposure. Summary of the Invention

[0004] To clarify PM 2.5 Regarding the effects of cardiac developmental toxicity, and to better diagnose and evaluate cardiac developmental toxicity, the present invention provides the following technical solutions.

[0005] In the first aspect, the present invention provides a PM during pregnancy 2.5 A biomarker for offspring cardiac developmental toxicity, wherein the biomarker is Angptl4 or a combination of Angptl4 and Sirt3.

[0006] Furthermore, PM can be predicted and evaluated by detecting the expression of Angptl4 mRNA or Angptl4 protein. 2.5 Causes developmental cardiotoxicity in offspring.

[0007] More preferably, the expression level of Angptl4 mRNA or Angptl4 protein is upregulated.

[0008] Furthermore, while detecting the expression level of Angptl4 mRNA or Angptl4 protein, the expression level of Sirt3 mRNA or Sirt3 protein is auxiliary detected.

[0009] More preferably, the expression level of Sirt3 mRNA or Sirt3 protein is downregulated.

[0010] In a second aspect, the present invention provides Angptl4 as a biomarker for the preparation of diagnosis, monitoring severity assessment, efficacy assessment, prognosis assessment, prevention and / or treatment of PM during pregnancy. 2.5 Use of products that cause developmental cardiotoxicity in offspring.

[0011] Preferably, the diagnosis, monitoring severity assessment, efficacy assessment or prognostic assessment of PM during pregnancy 2.5 The offspring cardiac developmental toxicity includes detecting the expression level of Angptl4.

[0012] Preferably, the product comprises at least one of a reagent for detecting the expression level of Angptl4 mRNA, detecting the expression level of Angptl4 protein, down-regulating the expression level of Angptl4 mRNA, and down-regulating the expression level of Angptl4 protein.

[0013] Preferably, the agent for downregulating the expression of Angptl4 mRNA or Angptl4 protein is selected from at least one of the following:

[0014] 1) A viral vector, wherein Angptl4 is lowly expressed in the viral vector.

[0015] 2) Small molecule drugs that have the effect of reducing the expression of Angptl4 mRNA and / or Angptl4 protein in cardiac tissue.

[0016] 3) Small molecule drugs or antibodies that have the effect of reducing Angptl4 activity.

[0017] In a third aspect, the present invention provides Angptl4 and Sirt3 as combined biomarkers for the preparation of diagnosis, monitoring severity assessment, efficacy assessment, prognosis assessment, prevention and / or treatment of PM during pregnancy. 2.5 Use of products that cause developmental cardiotoxicity in offspring.

[0018] Preferably, the diagnosis, monitoring severity assessment, efficacy assessment or prognostic assessment of PM during pregnancy 2.5 The developmental cardiac toxicity in offspring included detecting the expression levels of Angptl4 and Sirt3.

[0019] Preferably, the product comprises at least one of reagents for detecting the expression levels of Angptl4 mRNA and Sirt3 mRNA, detecting the expression levels of Angptl4 protein and Sirt3 protein, downregulating the expression level of Angptl4 mRNA and upregulating the expression level of Sirt3 mRNA, and downregulating the expression level of Angptl4 protein and upregulating the expression level of Sirt3 protein.

[0020] Preferably, the agent that downregulates the expression of Angptl4 mRNA and upregulates the expression of Sirt3 mRNA, downregulates the expression of Angptl4 protein and upregulates the expression of Sirt3 protein is selected from at least one of the following:

[0021] 1) a viral vector that underexpresses Angptl4 and overexpresses Sirt3;

[0022] 2) small molecule drugs that reduce the expression of Angptl4 mRNA and / or Angptl4 protein in cardiac tissue and increase the expression of Sirt3 mRNA and / or Sirt3 protein in cardiac tissue;

[0023] 3) Small molecule drugs or antibodies that have the effects of reducing Angptl4 activity and increasing Sirt3 activity.

[0024] In a fourth aspect, the present invention provides a PM 2.5 A method for constructing an animal model of offspring cardiac developmental toxicity, comprising the following steps:

[0025] 1) Exposing the animals to PM 2.5 middle.

[0026] 2) Detecting the expression level of Angptl4 and / or the expression level of Angptl4 mRNA in the heart tissue of the offspring animals.

[0027] Preferably, in step 2), the expression level of Sirt3 and / or the expression level of Sirt3 mRNA in the heart tissue of the offspring animals is also detected.

[0028] Preferably, the animal is 2.5 The exposure time is 4-8 hours, more preferably 6 hours.

[0029] Preferably, the animal is a rodent.

[0030] More preferably, the rodent is a mouse, and even more preferably a C57BL / 6J mouse.

[0031] In a fifth aspect, the present invention provides a PM 2.5 The animal model of offspring cardiac developmental toxicity is constructed according to the method described in the fourth aspect.

[0032] Beneficial effects of the present invention:

[0033] The present invention uses Angptl4 or a combination of Angptl4 and Sirt3 as a marker to reveal PM 2.5 The impact of PM on offspring's cardiac development is 2.5 It provides a method for the diagnosis and evaluation of offspring cardiac developmental toxicity, which is of great significance for the early risk assessment of offspring cardiac developmental toxicity. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 PM shown 2.5 Effects on the expression levels of Angptl4 / Sirt3 protein and mRNA in the heart tissue of offspring mice. A: Western blot and statistical analysis of mouse heart tissue. B: qRT-PCR results.

[0035] Figure 2 PM shown 2.5 Effects on the interaction between Angptl4 and Sirt3 in cardiac tissue of offspring mice;

[0036] Figure 3 PM shown 2.5 Exposure-induced cardiac histopathological changes in mice.

[0037] Figure 4 PM shown 2.5 Effects of exposure on the expression of mitochondrial-related proteins and ATP content in mouse myocardial tissue. A: Western blotting and statistical analysis of mouse myocardial tissue. B: ATP content in mouse myocardial tissue.

[0038] Figure 5 The expression levels of Angptl4 / Sirt3 in umbilical cord serum and the corresponding ROC curves are shown. A: Angptl4 content in serum; B: Sirt3 content in serum; C: PM 2.5 Correlation analysis with Angptl4; D: PM 2.5 Correlation analysis with Sirt3; E: ROC curve of Angptl4; F: ROC curve of Sirt3;

[0039] In the figure, * indicates statistical difference, p < 0.05, ** indicates statistical difference, p < 0.01; # indicates the difference between Angptl4 knockout mice and wild-type PM 2.5 P < 0.05 compared with the treatment group, ## indicates that Angptl4 knockout mice were significantly different from wild-type PM 2.5 Compared with the treatment group, p<0.01. DETAILED DESCRIPTION

[0040] The present invention is further described below with reference to the accompanying drawings and specific examples. These examples are intended to illustrate the present invention only and are not intended to limit the scope of the invention. Experimental methods in the following examples, where specific conditions are not specified, are generally performed in accordance with conventional conditions in the art or the conditions recommended by the manufacturer. Unless otherwise specified, conventional methods are used. Unless otherwise defined, the technical and scientific terms used herein have the same meanings as those familiar to those skilled in the art.

[0041] As used herein, "treating" means slowing, interrupting, preventing, controlling, stopping, alleviating, or reversing the progression or severity of a sign, symptom, disorder, condition, or disease after the disease has begun to develop, but does not necessarily involve the complete elimination of all disease-associated signs, symptoms, conditions, or disorders.

[0042] The "prognostic assessment" of the present invention refers to predicting the possible course and outcome of a disease, including determining the specific consequences of the disease.

[0043] The "evaluation of therapeutic efficacy" mentioned in the present invention refers to the evaluation of a patient's response to treatment.

[0044] The "monitoring" mentioned in the present invention refers to observing the occurrence and development of a disease.

[0045] The term "prevention" as used in the present invention refers to an individual taking specific measures to prevent the occurrence of a disease before the disease is diagnosed or develops.

[0046] The term "diagnosis" in the present invention refers to finding out whether a patient has had a disease or condition in the past, at the time of diagnosis, or in the future, or to finding out the progression or possible future progression of a disease.

[0047] Some of the material sources and experimental methods involved in this application are as follows:

[0048] SPF wild-type male and female C57BL / 6J mice (8 weeks old) were obtained from Beijing Weitonglihua Laboratory Animal Technology Co., Ltd. Angptl4 systemic knockout mice were obtained from Saiye (Suzhou) Biotechnology Co., Ltd. Umbilical cord serum from healthy controls (n = 40) and children with congenital heart disease (n = 40) was obtained from the Beijing Obstetrics and Gynecology Hospital affiliated to Capital Medical University. Congenital heart disease was diagnosed using transthoracic color Doppler echocardiography.

[0049] Example 1 Detection of Angptl4 / Sirt3 protein and mRNA expression levels in cardiac tissue of offspring mice

[0050] 1.1 PM 2.5 Establishment of an animal model of pregnancy exposure

[0051] After SPF-grade wild-type male and female C57BL / 6J mice (8 weeks old) were acclimated for one week, males and females were mated at a ratio of 1:2 at night, and female mice were designated as embryonic day (E) 0 after vaginal plug was observed. Pregnant mice were divided into three groups: clean air group (Filtered air, FA, 1 μg / m 3 ), Environmental PM 2.5 Group (Ambient PM 2.5 , AP, 30.86μg / m 3 ), concentrated ambient PM 2.5 Group (Concentrated ambient PM 2.5 , CAP, 367.22 μg / m 3 Mice in the AP and CAP groups were exposed to PM2.5 daily from E6.5 to E18.5 using a small animal whole-body inhalation exposure system. 2.5 Pregnant mice in the FA group were housed in individually ventilated cages (IVCs) and allowed free movement and access to water during the exposure period. Mice were euthanized at day 1 after birth, and heart tissue was harvested for experiments.

[0052] 1.2 Detection of Angptl4 / Sirt3 protein expression levels in heart tissue of 1-day-old mice

[0053] The specific process is as follows:

[0054] (1) Add approximately 0.1 mL of pre-chilled lysis buffer to each tissue and homogenize in a pre-chilled homogenizer. Pipette the homogenate into a pre-chilled 1.5 mL centrifuge tube and centrifuge at 12,000 g for 5 min at 4°C. Transfer the supernatant to a fresh pre-chilled centrifuge tube.

[0055] (2) The protein concentration of the sample was determined using the BCA protein quantification kit (Dingguo Changsheng, BCA01).

[0056] (3) Proteins were separated by SDS-PAGE and then transferred to nitrocellulose membrane.

[0057] (4) The cells were blocked with skim milk for 1 h at room temperature and then incubated with the primary antibody against Angptl4 (Proteintech, 18374-1-AP) / Sirt3 (CST, D22A3) at 4°C overnight.

[0058] (5) Incubate with anti-rabbit IgG HRP-linked (CST, 7074S) secondary antibody at room temperature for 1 h.

[0059] (6) The protein bands were visualized using the ChemiDoc imaging system and their grayscale values ​​were analyzed using ImageJ software. Figure 1 .

[0060] 1.3 Detection of Angptl4 / Sirt3 mRNA expression levels in heart tissue of 1-day-old mice

[0061] The specific process is as follows:

[0062] (1) Place the magnetic beads and heart tissue in a 2.0 mL EP tube and place it in a standard homogenizer. Add 0.1 mL of Trizol and homogenize. After homogenization, place the EP tube on ice for 10 minutes to allow the sample to fully lyse.

[0063] (2) Add 0.2 volumes of chloroform to the Trizol lysis buffer, vortex mix for 15 seconds, and place on ice for 5 minutes. Centrifuge at 12,000g at 4°C for 10 minutes, and then aspirate the upper colorless aqueous phase containing total RNA into a new centrifuge tube.

[0064] (3) Add an equal volume of isopropanol, vortex to mix, and place on ice for 10 min to precipitate RNA. Centrifuge at 12,000 g at 4°C for 10 min until RNA precipitate is visible at the bottom of the tube. Discard the supernatant.

[0065] (4) Add 1 mL of 75% ethanol and vortex to mix. Centrifuge at 12,000 g for 5 minutes and discard the supernatant. Aspirate as much liquid as possible from the walls. Leave the tube open and air dry for 10 minutes. Once the RNA is slightly dry, add 20 μL of DEPC water to dissolve it.

[0066] (5) cDNA was generated from the extracted RNA using the PrimeScript RTMaster Mix (Takara) kit.

[0067] (6) cDNA was amplified using the SYBR Premix Ex Taq kit (Takara, Japan) and a real-time fluorescence quantitative PCR detection system. The results are shown in Figure 1 .

[0068] 1.4 Detection of the interaction between Angptl4 and Sirt3 in heart tissue of 1-day-old mice

[0069] The specific process is as follows:

[0070] (1) Add approximately 0.2 mL of pre-chilled lysis buffer to each tissue and homogenize in a pre-chilled homogenizer. Pipette the homogenate into a pre-chilled 1.5 mL centrifuge tube and centrifuge at 12,000 g for 5 min at 4°C. Transfer the supernatant to a fresh pre-chilled centrifuge tube.

[0071] (2) The interaction between Angptl4 and Sirt3 was detected using a Protein A / G immunoprecipitation kit (Beaver, 22202-20). 25 μL of magnetic bead suspension was placed in a 1.5 mL EP tube for magnetic bead pretreatment. Angptl4 (Proteintech, 18374-1-AP) antibody was prepared with binding buffer to a final concentration of 25 μg / mL in 200 μL of antibody working solution. The suspension was quickly resuspended and gently inverted in an EP tube on a rotary mixer at room temperature for 15 minutes before magnetic separation.

[0072] (3) Add 200 μL of the antigen sample prepared in step (1), gently pipette to evenly disperse the antigen and magnetic bead-antibody complex, and react at 4°C overnight.

[0073] (4) Remove the EP tube from the magnetic separator, add 25 μL of 1× SDS-PAGE Loading Buffer, mix well, and heat at 95°C for 5 min. Then perform magnetic separation and collect the supernatant for SDS-PAGE detection. The results are shown in the figure. Figure 2 .

[0074] like Figure 1 As shown in the results, compared with the FA group, the expression levels of Angptl4 protein and mRNA in the AP and CAP groups increased, and the expression levels of Angptl4 protein and mRNA increased in a dose-dependent manner; while the expression levels of Sirt3 protein and mRNA decreased, and the expression levels of Sirt3 protein and mRNA decreased in a dose-dependent manner. This indicates that Angptl4 or the combination of Angptl4 and Sirt3 can be used as PM 2.5 Biomarkers of developmental cardiotoxicity in offspring.

[0075] like Figure 2 As shown, there is an interaction between Angptl4 and Sirt3, and PM 2.5 Exposure significantly enhanced the binding of Angptl4 to Sirt3. Therefore, by downregulating Angptl4, reducing the binding of Angptl4 to Sirt3, and increasing the content of Sirt3 in mouse heart tissue, PM2.5 can be alleviated. 2.5 Developmental toxicity to the heart of offspring mice.

[0076] Example 2PM 2.5 Effects of Angptl4 knockout on cardiac development in offspring of pregnant mice

[0077] 2.1 Construction of animal model

[0078] Angptl4 is located on chromosome 17 of mice. Using CRISPR-AI gene editing technology and high-throughput electroporation of fertilized eggs, we obtained Angptl4 systemic gene knockout mice (C57BL / 6JCya-Angptl4 em1 / Cya gene knockout mice). At the same time, a wild-type (WT) mouse control group was set up. After the knockout mice and wild-type mice were raised to 8 weeks of age, males and females were mated at night in a ratio of 1:2. Female mice were designated as embryonic day (E) 0 after vaginal plug was observed. Pregnant mice were divided into two groups: clean air group (Filtered air, FA, 1 μg / m 3 ) and concentrated ambient PM 2.5 Group (Concentrated ambient PM 2.5 , CAP, 171.95 μg / m 3 Mice in the CAP group were exposed to PM2.5 daily from E6.5 to E18.5 using a small animal whole-body inhalation exposure system. 2.5 The mice were housed in individually ventilated cages (IVCs) for the remaining 6 hours. Pregnant mice in the FA group were housed in the IVCs and had free movement and access to water during the exposure period. Mice were euthanized at day 1 of age and their heart tissues were harvested for analysis.

[0079] 2.2 Hematoxylin-eosin (H&E) staining of heart tissue of 1-day-old mice

[0080] (1) The heart tissue of offspring mice was fixed with 4% paraformaldehyde and then immersed in 20% sucrose solution and placed at 4°C overnight.

[0081] (2) Prepare wax blocks and paraffin sections.

[0082] (3) Bake the slices at 60℃ for 30 minutes.

[0083] (4) Dewax the paraffin sections by immersing them in xylene, dehydrate them with gradient ethanol concentrations, and wash them with deionized water for 5 minutes.

[0084] (5) Treat with hematoxylin for 5 minutes and rinse with deionized water for 5 minutes.

[0085] (6) Differentiate with 1% hydrochloric acid and ethanol for 30 seconds and rinse with deionized water for 5 minutes.

[0086] (7) Stain with eosin solution for 2 minutes and rinse with deionized water for 5 minutes.

[0087] (8) After dehydrating the paraffin sections with gradient ethanol, they were transparentized with xylene.

[0088] (9) Seal the slides with neutral gum.

[0089] (10) GT450 fully automatic slice scanning system was used for scanning and analysis. The results are shown in Figure 3 .

[0090] 2.3 Detection of Sirt3, Drp1, and Mfn2 protein expression levels in heart tissue of 1-day-old mice

[0091] (1) Add approximately 0.1 mL of pre-chilled lysis buffer to each tissue and homogenize in a pre-chilled homogenizer. Pipette the homogenate into a pre-chilled 1.5 mL centrifuge tube and centrifuge at 12,000 g for 5 min at 4°C. Transfer the supernatant to a fresh pre-chilled centrifuge tube.

[0092] (2) The protein concentration of the sample was determined using the BCA protein quantification kit (Dingguo Changsheng, BCA01).

[0093] (3) Proteins were separated by SDS-PAGE and then transferred to nitrocellulose membrane.

[0094] (4) The sections were blocked with skim milk for 1 h at room temperature and then incubated with primary antibodies against Sirt3 (CST, D22A3) / Drp1 (CST, 8570S) / Mfn2 (CST, 9482S) at 4°C overnight.

[0095] (5) Incubate with anti-rabbit IgG HRP-linked (CST, 7074S) secondary antibody at room temperature for 1 h.

[0096] (6) The protein bands were visualized using the ChemiDoc imaging system and their grayscale values ​​were analyzed using ImageJ software. Figure 4 .

[0097] 2.4 Detection of ATP activity in heart tissue of 1-day-old mice

[0098] (1) Add approximately 0.5 mL of pre-chilled lysis buffer to each tissue and homogenize in a pre-chilled homogenizer. Pipette the homogenate into a pre-chilled 1.5 mL centrifuge tube and centrifuge at 12,000 g for 5 min at 4°C.

[0099] (2) ATP activity was detected using an ATP assay kit (Invitrogen, A22066). 10 μL of the supernatant from step 1 and 90 μL of the working solution were added to a 96-well plate.

[0100] (3) Incubate at 37°C in the dark for 15 min.

[0101] (4) Use a microplate reader to measure the fluorescence value at 560 nm and calculate the ATP content. Figure 4 .

[0102] like Figure 3 As shown, PM 2.5 Exposure significantly induced inflammatory cell infiltration in the heart tissue of wild-type mouse offspring. 2.5 Compared with the offspring of treated mice, Angptl4 - / - Inflammatory cell infiltration in the heart tissue of the mouse offspring was not obvious, indicating that Angptl4 knockout can alleviate PM 2.5 Caused cardiac inflammatory response in offspring mice.

[0103] like Figure 4 As shown, PM 2.5 Exposure can significantly cause the downregulation of mitochondrial deacetylase Sirt3 expression, upregulation of mitochondrial fission protein Drp1 expression, downregulation of mitochondrial fusion protein Mfn2 expression in the heart tissue of wild-type mouse offspring, and reduction of ATP activity in heart tissue. 2.5 The offspring mice treated with Angptl4 - / - In the heart tissue of mouse offspring, the expression of mitochondrial deacetylase Sirt3 was significantly upregulated, the expression of mitochondrial fission protein Drp1 was significantly downregulated, the expression of mitochondrial fusion protein Mfn2 was significantly upregulated, and the ATP activity of heart tissue was significantly increased. This indicates that Angptl4 knockout can alleviate PM. 2.5 Exposure caused mitochondrial dynamics disorder and ATP reduction in the heart tissue of offspring mice. In addition, the expression of Sirt3, which is located downstream of Angptl4, was significantly upregulated with Angptl4 knockout, indicating that Angptl4 and Sirt3 are closely related to PM. 2.5 Angptl4 or the combination of Angptl4 and Sirt3 can be used as a biomarker for PM 2.5 Diagnosis and treatment of developmental cardiotoxicity in offspring.

[0104] Example 3 Detection of Angptl4 and Sirt3 levels in serum of human samples and ROC curve

[0105] 3.1 Population studies

[0106] Healthy controls (n = 40) and patients with congenital heart disease (CHD) (n = 40) were recruited from Beijing Obstetrics and Gynecology Hospital, Capital Medical University. Transthoracic color Doppler echocardiography was used to diagnose CHD. This study has been reviewed by the Medical Ethics Committee of Beijing Obstetrics and Gynecology Hospital, Capital Medical University, with approval number 2018-KY-003-02. Written informed consent was obtained from all subjects included in the sampling process. Monthly average gridded concentration data of PM2.5 were obtained from the China Air Pollutant Tracking Dataset (http: / / tapdata.org.cn) and allocated according to residential and work addresses. PM2.5 concentrations during the exposure window period (12 months or 6 months before pregnancy and the first three months of pregnancy) were significantly higher than those during the first three months of pregnancy. 2.5 The average concentration is calculated as a weighted average of the monthly average concentrations, with weights based on the proportion of days covered in each month.

[0107] 3.2 Detection of Angptl4 / Sirt3 protein levels in umbilical cord serum of healthy controls and CHD patients

[0108] (1) Use the ELISA kit (RayBiotech, ELH-ANGPTL4-1 / ELH-SIRT3-1) and prepare all reagents, serum, and standards according to the instructions.

[0109] (2) Add 100 μL of standard or serum to each well and incubate at room temperature for 2.5 h.

[0110] (3) Add 100 μL of the prepared biotin antibody to each well and incubate at room temperature for 1 h.

[0111] (4) Add 100 μL of the prepared streptavidin solution to each well and incubate at room temperature for 45 min.

[0112] (5) Add 100 μL of TMB one-step substrate reagent to each well and incubate at room temperature for 30 min.

[0113] (6) Add 50 μL of stop solution to each well and read immediately at 450 nm.

[0114] like Figure 5 As shown in the figure, the serum Angptl4 level in CHD patients was significantly upregulated compared with that in healthy controls, while the Sirt3 level was significantly downregulated. 2.5 Correlation between exposure levels and biomarkers. The results showed that PM 2.5The concentration of Angptl4 was positively correlated with the content of Angptl4 (R = 0.35, p < 0.01), and negatively correlated with the content of Sirt3 (R = -0.34, p < 0.01). The effects of Angptl4 and Sirt3 on PM were analyzed using R software and pROC and ggplot2 software packages. 2.5 The ROC curve results showed that Angptl4 correctly distinguished 71% and 79% of patients in the training and test sets, respectively, and Sirt3 correctly distinguished 75% and 78% of patients in the training and test sets, respectively.

[0115] The above experimental results fully demonstrate that Angptl4 or the combination of Angptl4 and Sirt3 can be used as biomarkers to effectively detect PM 2.5 The related CHD was predicted.

[0116] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the scope of protection of the present invention.

Claims

1. The use of reagents for detecting Angptl4 in the preparation of diagnosis, monitoring severity assessment, efficacy assessment or prognosis assessment of PM during pregnancy 2.5 Use of products that cause developmental cardiotoxicity in offspring.

2. The use according to claim 1, characterized in that Diagnosis, monitoring, severity assessment, efficacy assessment, or prognostic assessment of PM during pregnancy 2.5 The offspring cardiac developmental toxicity includes detecting the expression level of Angptl4.

3. The use according to claim 1 or 2, characterized in that The product includes at least one of a reagent for detecting the expression level of Angptl4 mRNA or a reagent for detecting the expression level of Angptl4 protein.

4. Reagents for detecting Angptl4 and Sirt3 are used in the preparation of diagnostic, severity assessment, efficacy assessment, or prognostic assessment of PM during pregnancy. 2.5 Use of products that cause developmental cardiotoxicity in offspring.

5. The use according to claim 4, characterized in that Diagnosis, monitoring, severity assessment, efficacy assessment, or prognostic assessment of PM during pregnancy 2.5 The developmental cardiac toxicity in offspring included detecting the expression levels of Angptl4 and Sirt3.

6. The use according to claim 4 or 5, characterized in that The product includes at least one of reagents for detecting the expression levels of Angptl4 mRNA and Sirt3 mRNA or the expression levels of Angptl4 protein and Sirt3 protein.

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