Differential marker heptadecanoic acid based on metabolomics technique to detect plaque rupture and plaque erosion

Abstract

The application discloses a biomarker for identifying plaque rupture (PR) and plaque erosion (PE) in acute coronary syndrome (ACS) based on positive ion mode metabolomics technology and application thereof. Serum samples of patients are analyzed in a positive ion mode (ESI+) by liquid chromatography-mass spectrometry (LC-MS), and it is found that heptadecanoic acid has significant expression difference (VIP>1, P<0.05, Fold Change>2) between PR and PE patients. Based on this, the application provides a kit and method for distinguishing PR and PE by detecting the heptadecanoic acid level in the positive ion mode. The method has high sensitivity and simple operation, and can provide a basis for accurate typing and individualized treatment of ACS.

Description

Technical Field

[0001] This invention relates to cardiovascular disease diagnostic techniques, specifically a method for detecting heptadecanoic acid based on positive ion mode metabolomics and its application in identifying ACS plaque types. The method utilizes metabolomics to detect differential markers of plaque rupture and erosion. Background Technology

[0002] (1) Plaque erosion and plaque rupture are important pathogenic mechanisms of ACS. Acute coronary syndrome (ACS) is a leading cause of cardiovascular death and hospitalization worldwide, primarily affecting individuals aged 50 to 70. With socioeconomic development and improved living standards, unhealthy lifestyles have led to an increase in high-risk factors, resulting in a year-on-year rise in ACS incidence, placing a significant burden on healthcare and society. ACS refers to a syndrome of acute ischemic heart disease, encompassing three subtypes: Unstable Angina (UA), Non-ST-segment Elevation Myocardial Infarction (NSTEMI), and ST-segment Elevation Myocardial Infarction (STEMI).

[0003] The main pathogenesis of ACS is plaque rupture (PR, accounting for about 2 / 3) and plaque erosion (PE, accounting for about 1 / 3).

[0004] Plaque rupture refers to the breaking of the fibrous cap within a plaque, exposing its components to the bloodstream, which in turn triggers platelet aggregation and thrombus formation. In this situation, the thrombus often forms within the coronary artery, obstructing blood flow and leading to acute myocardial ischemia and / or necrosis. Plaque rupture is typically caused by factors such as inflammation and apoptosis within the plaque. These processes can lead to the rupture of the fibrous cap within the plaque and promote platelet aggregation and thrombus formation.

[0005] Plaque erosion refers to the destruction of endothelial cells on the surface of atherosclerotic plaques, exposing the glassy material beneath the vascular intima to the bloodstream. These substances can stimulate platelet aggregation and vasoconstriction, further destabilizing the plaque and leading to myocardial ischemia. Plaque erosion is most common in the proximal and mid-segments of the coronary arteries, especially when the inflammatory response within the plaque is active.

[0006] (2) Accurate identification of PR and PE is of great significance for "intervention without implantation". Plaque rupture and plaque erosion have different pathological bases and therefore require different clinical treatment strategies. Previously, stenting was the first-line treatment for ACS patients, regardless of whether the cause was partial rupture (PR) or partial erosion (PE). However, among patients who received stents, those with plaque erosion had a 3.38 times higher rate of delayed intimal healing after stent implantation compared to those with plaque rupture. In contrast to plaque rupture, plaque erosion is often accompanied by negative remodeling that leads to further stenosis of the vascular lumen, and the inflammatory response and severity of luminal stenosis are lower in plaque rupture patients.

[0007] Therefore, patients with plaque erosion should carefully consider stent implantation, as thrombolytic therapy may be more effective. A prospective study by Professor Yu Bo's team in the Department of Cardiology at the Second Affiliated Hospital of Harbin Medical University found that ACS patients with plaque erosion can effectively reduce thrombus volume and increase blood flow area through antithrombotic therapy alone. Follow-up results showed that antithrombotic therapy alone is safe and effective for plaque erosion. This study is the first in the world to confirm the safety and feasibility of conservative antithrombotic therapy for ACS caused by plaque erosion, and proposes the concept of "intervention without stents and intervention without residues." This will change the current "one-size-fits-all" interventional treatment for ACS. Professor Peter Libby of Harvard Medical School published an article in the journal Eur Heart, pointing out that this study is of great significance for effectively controlling thrombosis caused by plaque erosion.

[0008] Furthermore, in-stent thrombosis has a mortality rate as high as 50%, making it a significant adverse factor threatening patient prognosis. In 2010, Professor Yu Bo's team from the Department of Cardiology at the Second Affiliated Hospital of Harbin Medical University pioneered the discovery internationally that in-stent lipid plaque rupture is a major factor contributing to late-stage in-stent thrombosis, with an incidence rate of approximately 16%. For patients with plaque erosion, the "intervention without implantation" strategy can completely avoid the risks of in-stent restenosis and in-stent thrombosis. Therefore, accurate identification of plaque rupture and plaque erosion is of great guiding significance for the selection of clinical treatment strategies.

[0009] (3) OCT is the best method to distinguish between PR and PE, but it has certain limitations. Optical coherence tomography (OCT) has become a crucial technique in the diagnosis and treatment of coronary heart disease. It provides high-resolution images and can measure various parameters of the blood vessel walls, enabling doctors to develop more accurate diagnostic and treatment plans.

[0010] OCT technology can display the characteristic structures inside plaques through high-resolution images, including plaque thickness, internal composition, and surface conditions. OCT can observe structures such as fibrous discs, calcifications, and fibrous fractures within plaques on high-resolution images, while also revealing details such as minute protrusions and surface coverings on the plaque surface. OCT-based plaque fracturing identification mainly includes the following aspects: 1) Surface characteristics of the plaque: The surface of the fractured plaque often has cracks or unevenness. 2) Internal characteristics of plaques: The internal structure of ruptured plaques often exhibits characteristics such as fibrous disc rupture, calcified plaque rupture, and internal hemorrhage.

[0011] 3) Thin-shell region inside the plaque: The thin-shell region before plaque rupture is an important precursor to plaque rupture. OCT can observe the thin-shell region inside the plaque and predict whether the plaque is about to rupture. 4) Lipid plaques inside the plaque: Cavities within lipid plaques are a danger signal of plaque rupture. OCT can observe lipid plaques and cavities inside lipid plaques.

[0012] Although OCT technology has shown high sensitivity and specificity in identifying plaque rupture, it still has some limitations and problems: 1) OCT cannot identify plaque composition: OCT can only show the internal structural features of plaques, but cannot determine the specific components inside the plaque, such as whether it contains a certain proportion of lipids and calcifications. [9-12] 2) OCT cannot determine plaque stability: OCT can only observe the internal structure and surface features of plaques, and it is difficult to determine the stability of plaques and the risk of rupture.

[0013] 3) OCT has difficulty identifying large plaques: Due to the limitations of OCT probe size, it is difficult to identify large plaques, and multiple angles and multiple scans are required to obtain sufficient information.

[0014] 4) OCT artifacts: OCT images contain artifacts and interference that may affect plaque identification and analysis.

[0015] (4) Through positive ion mode metabolomics analysis, the team of this invention screened 47 differential components from 1132 metabolites and found for the first time that heptadecanoic acid was significantly highly expressed in PR patients and could be used as a specific biomarker. Summary of the Invention

[0016] Key findings: In positive ion mode, the concentration of heptadecanoic acid (m / z 271.3) in the serum of the PR group was significantly higher than that in the PE group (VIP>1, P<0.05).

[0017] Technical solution Sample processing: Serum samples were subjected to methanol precipitation of proteins, centrifuged, and the supernatant was collected. Mass spectrometry conditions: Ionization mode: electrospray ionization positive ion (ESI+); Detection ion: m / z 271.3 [M+H]+; Column: C18 reversed-phase column, mobile phase: 0.1% formic acid-acetonitrile gradient elution. Data analysis: OPLS-DA analysis was performed using MetaboAnalyst 6.0 software, with thresholds of VIP>1, P<0.05, and FC>2 (PPT 2.5). Attached Figure Description

[0018] Figure 1: OPLS-DA score plots of the PR and PE groups under positive ion mode: The OPLS-DA score plots show a clear separation trend between the two groups, indicating differences in metabolites between the serum samples of the two groups. The displacement test result R2Y=0.785 indicates strong interpretability of the model, and Q2=0.568 indicates acceptable predictability. The p-values ​​of Q° and R2Y <0.01 indicate that the results are reliable; Figure 2: Mass spectrum of heptadecanoic acid in positive ion mode (m / z 271.3). Detailed Implementation

[0019] 1. Serum samples were collected from 135 patients with acute coronary syndrome (ACS); 2. Heptadecanoic acid (m / z 271.3) was detected by positive ion mode LC-MS. The average concentration in the PR group was 3.2 ± 0.8 μM, and in the PE group it was 1.1 ± 0.3 μM (P < 0.01). 3. ROC curve analysis showed that the AUC of heptadecanoic acid for distinguishing PR from PE was 0.89 (95% CI: 0.82-0.96).

Claims

1. A biomarker for differentiating between plaque rupture (PR) and plaque erosion (PE), characterized in that, The biomarker is heptadecanoic acid (chemical formula C17H34O2), which is detected by positive ion mode mass spectrometry (ESI+) with a mass-to-charge ratio (m / z) of 271.3 [M+H]+.

2. The method for detecting biomarkers according to claim 1, comprising the following steps: collecting serum samples from ACS patients; Metabolomics analysis was performed using positive ion mode liquid chromatography-mass spectrometry (LC-ESI+-MS); differentially expressed metabolites with VIP>1, P<0.05 and Fold Change>2 were screened using orthogonal partial least squares discriminant analysis (OPLS-DA). Quantitative detection of heptadecanoic acid (m / z 271.3) levels showed that elevated levels indicated PR and decreased levels indicated PE.

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