Relationship between Coronary VH-IVUS Plaque Characteristics and CTRP9, SAA, and Hcy in Patients with Coronary Heart Disease.

Coronary heart disease is a common disease threatening human health. In recent years, the incidence of coronary heart disease in China has only increased. It is the most common type of organ disease caused by coronary atherosclerosis, which is observed in the aorta, carotid artery, and femoral artery. The main clinical treatments for coronary heart disease include coronary artery bypass grafting and drug treatment. To investigate the relationship of serum adipocytokine C1q/tumor necrosis factor-related protein 9 (CTRP9), amyloid A (SAA), and plasma homocysteine (Hcy) with coronary artery plaque characteristics in patients with coronary heart disease. Overall, 143 patients with coronary heart disease admitted to our hospital are selected as research participants. The proportion of plaque necrosis core volume is higher in group A than in group B, and the differences are statistically significant (P < 0.05). In group A, necrotic core volume percentage is negatively correlated with CTRP9 levels and positively correlated with SAA and Hcy levels (P < 0.05). Logistic regression analysis revealed that increased systolic blood pressure, increased number of coronary artery lesions, decreased CTRP9 levels, and increased Hcy levels are independent risk factors for thin fibrous cap atherosclerosis in patients with coronary heart disease (P < 0.05). Decreased CTRP9 levels and increased Hcy levels are independent risk factors for coronary heart disease patients with thin fibrous cap atherosclerosis.

1. Introduction

Coronary heart disease includes stable angina pectoris and acute coronary syndrome (ACS). The occurrence of ACS is not associated with the severity of plaque stenosis but is closely related to the vulnerability of plaques, and inflammatory response is closely related to vulnerable plaque rupture [1]. Virtual-histology intravascular ultrasound (VH-IVUS) can evaluate plaque stability in vivo. According to the VH-IVUS classification, thin fibrous cap atherosclerosis is closely related to future cardiovascular events and is known as vulnerable plaques [2]. Changes in serum adipokine C1q/tumor necrosis factor-related protein 9 (CTRP9), amyloid A (SAA), and plasma homocysteine (Hcy) levels can be used as indicators to reflect the stability of coronary atherosclerotic plaque [3]. Hyperhomocysteinemia can cause coronary atherosclerotic plaque rupture via multiple mechanisms, which leads to cardiovascular risk events [4]. TRPP 9 is an accessory protein with the closest adiponectin structure in the CTRPP protein superfamily, and it plays an important role in anti-inflammatory activities, thereby improving endothelial function and regulating metabolism [5]. SAA belongs to the same gene cluster encoding polymorphic proteins and is a sensitive acute phase response protein that is involved in coronary heart disease development [6]. However, the relationship between coronary artery plaque and CTRP9, SAA, and Hcy in patients with coronary heart disease remains unclear.

Therefore, this study aims to examine the relationship between serum CTRP9, SAA, and Hcy levels and coronary artery plaque characteristics in patients with coronary heart disease to provide a basis for early detection and treatment of diseases and the selection of clinical diagnosis and treatment programs.

The remainder of this paper is organized as follows: Section 2 presents the proposed methods. Section 3 provides experimental data and analysis and shows the discussion in Section 4. Finally, the conclusions of this study and some future recommendations are given in Section 5.

2. Our Proposed Methods

2.1. Participants

This study included patients with coronary heart disease who were admitted to the Department of Cardiovascular Medicine at our hospital. Based on VH-IVUS findings, patients are divided into those having thin fibrous cap atherosclerosis (n = 67; group A) and those having thick fibrous cap plaque (n = 77; group B). The study participants were hospitalized from January 2017 to January 2019. Patients who met the diagnostic criteria for coronary heart disease according to the Guidelines for the Prevention and Treatment of Coronary Heart Disease [7]; those aged ≤79 years; those diagnosed based on ECG and coronary angiography findings and myocardial enzyme levels; and those who underwent VH-IVUS are included in the study [8-10]. Patients with malignant tumor; those with valvular heart disease; those with mental illness and cognitive dysfunction; those with serious liver and kidney function diseases; those blood diseases; those with thyroid function disease; or those with incomplete VH-IVUS data are excluded. This study complied with the relevant requirements of the Medical Ethics Committee and is confidential about information related to patient privacy [11, 12].

The patients included in the study are divided into groups A and B. Group A included patients aged 47–79 years (mean 65.8 ± 6.5 years) and comprised 37 male patients and 30 female patients [13]. Their BMI is 23.1 ± 2.3 kg/m2. Group B included patients aged 46–79 years (mean 64.5 ± 7.2 years) and comprised 46 male patients and 31 female patients. Their BMI is 22.9 ± 2.5 kg/m2. Age, gender, and BMI are compared between the two groups, and the differences are not statistically significant (P > 0.05) [14].

2.2. Methods

2.2.1. Coronary Angiography and VH-IVUS

After injecting 0.1 mg of nitroglycerin into the conventional coronary artery, an Eagle Eye IVUS catheter (Volcano, USA, probe 3.2 F, frequency 20 MHZ) is inserted toward the target vessel along a 0.014-inch guide wire until it reaches an area 20 mm distal to the lesion site. After collecting images, the IVUS catheter is then automatically retracted at 0.5 mm/s, and the images are recorded and saved for offline analysis. Elastic membrane volume, lumen volume, and plaque volume of the lesion sites are evaluated using the recorded images.

According to the IVUS analysis guidelines of the American College of Cardiology, external elastic membrane volume, lumen volume, and plaque volume are evaluated. Coronary atherosclerotic plaques are divided into four types: fibrous tissue (green), fibrous lipid (yellow-green), calcified tissue (white), and necrotic core (red). Based on the percentage of various components in the plaque, coronary plaques are divided into thin fibrous cap atherosclerosis, thick fibrous cap atherosclerosis, pathological intimal thickening, fibrous plaques, and fibrous calcification plaques.

Patients first underwent coronary angiography. Left coronary angiography can be performed in the right foot position, right anterior oblique foot position, right anterior oblique head position, and right head position, while right coronary angiography can be performed in the left anterior oblique head position to ensure that the contrast agent is filled in the entire vascular segment on imaging process. The vascular wall can be clearly visualized without laminar flow imaging.

2.2.2. Index Detection Method

On the morning of coronary angiography, 8 ml of fasting peripheral venous blood was collected from the study patients for laboratory testing and then centrifuged at 3000 r/min for 10 min. The serum is separated and stored in a low-temperature refrigerator at −80°C for later use.

The CTRP9, SAA, and Hcy levels are determined using a double antibody sandwich enzyme-linked immunosorbent assay. The experimental strips are removed from the sealed bag, and 100 μl of the sample or diluent is added to the blank microwell, and 100 μl of serum is added to the detection well. The experimental strip is sealed with adhesive paper and incubated at 36°C for 60–90 min. The strip/microplate is washed five times in phosphate buffer, followed by the addition of 100 μl of a biotin antibody diluent to the blank control well and 100 μl of a biotin antibody detection solution to the detection well. The experimental wells are sealed with adhesive paper and incubated at 36°C for 30–60 min. After incubation, 100 μl of an enzyme-binding diluent is added to the blank control well, and 100 μl of an enzyme-binding detection solution is added to the detection well. Adhesive papers are used to seal the test strip well. After washing with phosphate buffer five times, the TMB color base solution is added (YSRIBIO Huzhou Yingchuang Biotechnology Co., Ltd.), and the wells are incubated in the dark at 36°C for 15 min, after which a stop solution (100 μl) is added, and the plates are read at 450 OD within 3 min using a BioTek microplate reader (BioTek Instrument Co., Ltd.). TG, TC, HDL-C, and LDL-C levels are also measured (Hitachi automatic biochemical analyzer) according to the instructions.

2.3. Statistical Analyses

The SPSS 21.0 software is used for all statistical analyses. The diastolic blood pressure and TC, HDL-C, and LDL-C levels of groups A and B are presented as mean (SD), and the independent sample t-test is used for comparison between the two groups. The χ2 test is used to compare the enumeration data of combined diseases and gender composition. The Pearson method is used for correlation analysis, and logistic regression analysis is used for multivariate analysis. P values <0.05 indicated statistically significant differences.

3. Experimental Data and Analysis

3.1. Comparison of Basic Data of the Two Groups of Patients

Age, gender, BMI, smoking, diastolic blood pressure, TC, HDL-C, LDL-C, and target vessel distribution are compared between group A and group B, and the difference is not statistically significant (P > 0.05). In group A of patients with systolic blood pressure, fasting blood glucose, TG are higher than those in group B, and the difference is statistically significant (P > 0.05). There are 23.88% patients with triple vessel disease, 49.25% patients with double vessel disease, and 16.42% patients with single vessel disease in group A, and 6.49% patients with triple vessel disease, 63.64% patients with double vessel disease, and 29.87% patients with single vessel disease in group B. The difference is statistically significant (P < 0.05). Table 1 shows the comparison of basic data of groups A and B.

Table 1

Comparison of basic data of groups A and B.

Variables	Group A (n = 67)	Group B (n = 77)	t/χ2	P
Age (years)	65.8 ± 6.5	64.5 ± 7.2	1.130	0.260
Gender (%)			0.299	0.584
Male	37 (55.22)	46 (59.74)
Female	30 (44.78)	31 (40.26)
Smoking (%)			0.825	0.364
Yes	28 (41.79)	38 (49.35)
No	39 (58.21)	39 (50.65)
BMI (kg/m2)	23.1 ± 2.3	22.9 ± 2.5	0.497	0.620
Diastolic blood pressure (mmHg)	81.3 ± 7.0	80.6 ± 7.4	0.581	0.562
Systolic blood pressure (mmHg)	135.1 ± 8.9	132.0 ± 7.9	2.214	0.028
Fasting blood glucose (mmol/L)	5.72 ± 0.60	5.38 ± 0.68	3.160	0.002
TG (mmol/L)	2.31 ± 0.51	2.14 ± 0.40	2.239	0.027
TC (mmol/L)	5.28 ± 0.61	5.11 ± 0.48	1.869	0.064
HDL-C (mmol/L)	0.98 ± 0.20	1.04 ± 0.23	−1.658	0.099
LDL-C (mmol/L)	3.21 ± 0.48	3.14 ± 0.55	0.808	0.421
Target vessel (%)			0.689	0.709
Left coronary artery	22 (32.84)	32 (41.56)
Left anterior descending branch	26 (38.81)	28 (36.36)
Right coronary artery	12 (17.91)	17 (22.08)
Number of diseased branches (%)			6.863	0.032
Single-vessel disease	11 (16.42)	23 (29.87)
Double-vessel disease	33 (49.25)	49 (63.64)
Three-vessel disease	16 (23.88)	5 (6.49)

3.2. Comparison of the Characteristics of Fibrolipid Plaques in Diseased Segments between the Two Groups

The percentage of fibroliposomal volume, fibrolipid tissue, and calcified tissue volume is compared between group A and group B, and the difference is not statistically significant (P > 0.05). The percentage of plaque necrosis core volume in group A is higher than that in group B, and the difference is statistically significant (P < 0.05). Table 2 shows the comparison of the characteristics of fibrolipid plaques in the two groups of diseased segments.

Table 2

Comparison of characteristics of fibrolipid plaques in the two groups of diseased segments (x ± s).

Group	n	Fibrolipid volume (%)	Fibrolipid tissue (%)	Calcified tissue volume (%)	Necrotic core volume (%)
A	67	53.81 ± 7.20	8.73 ± 2.20	14.39 ± 3.01	23.81 ± 3.77
B	77	54.40 ± 7.88	9.18 ± 2.45	15.28 ± 3.54	19.56 ± 4.02
t		−0.466	−1.152	−1.612	6.513
P		0.642	0.251	0.109	≤0.001

3.3. Comparison of CTRP9, SAA, and Hcy Levels in the Two Groups

SAA and Hcy levels in group A are significantly higher than those in group B, and the difference is statistically significant (P < 0.05). CTRP9 levels in group A are significantly lower than those in group B, and the difference is statistically significant (P < 0.05). Table 3 shows the comparison of CTRP9, SAA, and Hcy levels between groups A and B.

Table 3

Comparison of CTRP9, SAA, and Hcy levels between groups A and B (x ± s).

Group	n	CTRP9 (10−2 mg/L)	SAA (mg/L)	Hcy (μmol/L)
A	67	3.32 ± 0.38	6.37 ± 1.33	14.29 ± 2.88
B	77	3.62 ± 0.34	5.58 ± 1.28	11.10 ± 2.61
t		−5.000	3.628	6.972
P		≤0.001	≤0.001	≤0.001

3.4. The Correlation between the Percentage of Necrotic Core Volume and CTRP9, SAA, and Hcy Levels

The simple linear correlation method is used for analysis. The percentage of necrotic core volume in the atherosclerotic plaque tissue of the thin fibrous cap is correlated with the levels of CTRP9, SAA, and Hcy, and is significantly negatively correlated with CTRP9, and is significantly positively correlated with SAA and Hcy (P < 0.05). Table 4 shows the linear correlation analysis results.

Table 4

Linear correlation analysis results.

Parameter	Correlation	CTRP9	SAA	Hcy
Plaque thickness	r	−0.396	0.442	0.486
	P	≤0.001	≤0.001	0.006

3.5. The Relationship with the Formation of Thin Fiber Cap Atherosclerosis in Patients with Coronary Heart Disease

With the formation of thin cap atherosclerosis in patients as the dependent variable and SAA, Hcy, CTRP9, systolic blood pressure, fasting blood glucose, TG, and the number of coronary artery lesions as independent variables, logistic regression analysis showed that increased systolic blood pressure, increased number of coronary artery lesions, decreased CTRP9, and increased Hcy level are independent risk factors for thin cap atherosclerosis in patients with coronary heart disease (P < 0.05). Table 5 shows the relationship of thin fibrous cap atherosclerosis in patients with coronary heart disease. Figure 1 depicts a VH-IVUS image showing the formation of thin cap atherosclerotic plaque in a 72-year-old man with left coronary artery disease. Figure 2 shows a VH-IVUS image showing pathological intimal thickening in a 66-year-old man after undergoing left anterior descending branch surgery.

Table 5

Relationship of thin fibrous cap atherosclerosis in patients with coronary heart disease (x ± s).

Parameter	β	SE	Walds	P	OR	95% CI
Systolic blood pressure	0.663	0.304	4.756	0.043	1.941	1.069	3.521
Fasting blood glucose	0.527	0.381	1.913	0.195	1.694	0.803	3.574
TG	0.297	0.184	2.605	0.114	1.346	0.938	1.930
Number of diseased branches	0.377	0.157	5.766	0.024	1.458	1.072	1.983
CTRP9	−0.486	0.185	6.901	0.007	0.615	0.428	0.884
SAA	0.559	0.374	2.234	0.170	1.749	0.840	3.640
Hcy	0.464	0.175	7.030	0.001	1.590	1.129	2.241

Figure 1

A VH-IVUS image showing the formation of thin cap atherosclerotic plaque in a 72-year-old man with left coronary artery disease.

Figure 2

A VH-IVUS image showing pathological intimal thickening in a 66-year-old man after undergoing left anterior descending branch surgery.

4. Result and Discussion

Atherosclerotic plaques are composed of lipid-rich aneurysmal substances covered with fibrous caps, which are important for plaque stability because unstable plaques are prone to rupture. Early clinical identification and prediction of unstable plaques are of great significance for risk stratification and prognosis evaluation of patients with coronary heart disease. Traditional plaque characteristic detection methods have their limitations in that they are mostly invasive and have low patient acceptance, thereby limiting their application. In this study, changes in serum CTRP9, SAA, and Hcy levels are evaluated for their potential as noninvasive serum markers for coronary plaque stability.

CTRP9 is a paralogous protein whose structure is closest to the adiponectin structure, and it reduces lipid accumulation in hepatocytes, regulates basal and insulin-mediated glucose uptake, reduces normal or insulin-resistant blood glucose levels, and improves metabolism. CTRP9 can reduce the proliferation and chemotaxis of vascular smooth muscle cells induced by platelet-derived growth factors, inhibit the phosphorylation of extracellular signal-regulated kinase stimulated by platelet-derived growth factors, and reduce platelet production. Thus, CTRP9 may be of great significance for evaluating the stability of coronary artery plaques.

SAA is a sensitive acute-phase reaction protein that can quickly combine with high-density lipoprotein to inhibit the activity of lecithin cholesterol peptidyl transferase, slow down the outflow and removal of cholesterol lipids, increase the deposition of lipid plasma, and increase the susceptibility of plaques. SAA is involved in the occurrence and development of coronary heart disease and has certain value in the diagnosis, treatment, and prognosis of the disease. Hcy is the product of methionine demethylation in cells and a sulfur-containing amino acid, the concentration of which is affected by environmental and genetic factors.

This study compared systolic blood pressure, fasting blood glucose and TG levels, and the number of diseased coronary arteries between patients with and without fibrous cap atherosclerosis and found that they differed significantly. The simple linear correlation method is used for analysis. The thickness of thin fibrous cap atherosclerotic plaques is significantly positively correlated with serum TC, TG, and LDL-C levels and significantly negatively correlated with HDL-C levels, thus demonstrating that the formation of thin fibrous cap atherosclerosis may be related to systolic blood pressure, fasting blood glucose and TG levels, and the number of diseased coronary arteries. The percentage of plaque necrosis core volume is significantly higher in group A than in group B, and SAA and Hcy levels are significantly higher in patients with fibrous cap atherosclerosis than in those without fibrous cap atherosclerosis. This finding could be because severe inflammatory reactions occurred in patients with fibrous cap atherosclerosis and because increased SAA levels are significantly associated with enhanced high-density lipoprotein binding ability. Hcy can cause coronary heart disease by damaging the intima and promoting the infiltration of inflammatory cells and lipids and by lipid peroxidation, vascular inflammation, smooth muscle cell proliferation, and platelet activation. The inflammatory reaction in vulnerable plaques can lead to an increase in Hcy levels, thereby predicting coronary plaque stability. CTRP9 levels are significantly lower in patients without thin-cap fibrous atherosclerosis, possibly because it produces an antiatherosclerosis effect by inhibiting inflammatory response, regulating lipid metabolism, and improving endothelial function. Logistic regression analysis revealed that the increase in systolic blood pressure, Hcy levels, and number of diseased coronary arteries and the decrease in CTRP9 levels are independent risk factors for coronary heart disease patients with thin-cap atherosclerosis. This finding is based on the fact that the increase in systolic blood pressure leads to a sharp decrease in coronary perfusion time. This results in insufficient coronary blood supply and an increase in the number of coronary lesions, leading to insufficient muscle blood supply and oxygen supply disorder. The continuous increase in coronary artery tension can damage vascular internalization, lead to lipid deposition and middle smooth muscle growth, eventually developing into atherosclerotic plaques.

5. Conclusion and Future Work

The decrease in CTRP9 levels and the increase in Hcy levels are independent risk factors for thin-cap fibrous atherosclerosis in patients with coronary heart disease.

Many prospective studies have revealed that several inflammatory markers are associated with cardiovascular events and that the degree of inflammatory response is closely related to the stability of coronary atherosclerotic plaques. However, few studies have focused on changes in CTRP9, SAA, and Hcy levels. The current study demonstrated the use of the markers to predict the stability of coronary atherosclerotic plaques and prevent the occurrence of clinical cardiovascular events reliably better. However, this study had limitations such as the small sample size, the lack of detection tools, the lack of third-party analysis of IVUS data, and the short follow-up time. Thus, further studies are needed to evaluate and confirm these findings.