Abstract
Background
Studies have confirmed a correlation between angiotensin-converting enzyme 2 (ACE2) gene polymorphisms and the risk of hypertension; however, its correlation with congenital heart disease (CHD)-related-pulmonary arterial hypertension (PAH) risk in neonates has not been reported.
Methods
The study enrolled 321 Han Chinese neonates, comprising 113 healthy controls and 208 patients with left-to-right shunt CHD. Among the CHD patients, 98 cases were classified as the PAH subtype [CHD PAH (+)]. Tag SNP genotyping was performed using Sanger sequencing. Associations between three ACE2 SNPs (rs2074192, rs2285666, and rs2106809) and CHD PAH (+) neonates were assessed via sex-stratified logistic regression. Differences in circulating ACE2 and angiotensin1-7 [Ang(1–7)] levels across ACE2 haplotypes were compared using analysis of variance (ANOVA).
Results
No significant associations were observed between the three ACE2 SNPs and susceptibility to CHD or the risk of PAH in either univariable or multivariable analyses. In females, the CCT haplotype showed a nominally suggestive association with CHD-PAH in both the univariable model (OR = 0.216, 95% CI: 0.047–0.740; P = 0.025; FDR_P = 0.074) and the multivariable model (OR = 0.187, 95% CI: 0.039–0.670; P = 0.018; FDR_P = 0.053). A nominal difference in circulating Ang-(1–7) levels was also observed across haplotypes among females, with higher levels in CCT haplotype carriers than in those carrying the CTC or TCT haplotypes (160.16 ± 19.24 pg/mL vs. 140.54 ± 28.40 pg/mL and 139.77 ± 29.85 pg/mL, respectively; P = 0.037). However, this difference did not survive FDR correction (FDR_P = 0.074).
Conclusions
Our study showed that there was no significant association between ACE2 SNPs or haplotypes and the risk of CHD-PAH in neonates.
Trial registration
Our study is an observational study. According to the International Committee of Medical Journal Editors (ICMJE), purely observational studies (in which the allocation of medical interventions is not under the investigator’s discretion) do not require registration.
Supplementary Information
The online version contains supplementary material available at https://doi.org/10.1186/s12872-026-06368-6.
Keywords: Angiotensin-converting enzyme 2, Congenital heart disease, Neonate, Pulmonary arterial hypertension, Single nucleotide polymorphism
Background
Congenital heart disease (CHD) is the most common congenital malformation worldwide, and a leading cause of neonatal and child mortality [1, 2]. Left-to-right shunt CHD [such as ventricular septal defect (VSD), atrial septal defect (ASD), and patent ductus arteriosus (PDA), etc.] is an important cause of neonatal pulmonary arterial hypertension (PAH) [3, 4]. Its main pathological and anatomical basis includes abnormal shunting leading to increased pulmonary blood flow, pulmonary vascular hyperperfusion and endothelial injury, pulmonary vascular remodeling, and potential genetic susceptibility [4]. PAH is associated with increased mortality, and early identification and intervention are particularly important [5, 6].
The renin-angiotensin system (RAS), present in the respiratory system lung tissue, regulates pulmonary circulation, blood pressure, and lung development and function [7, 8]. Angiotensin-converting enzyme 2 (ACE2) is one of the main regulators of RAS and has a counterbalancing role in blood pressure regulation [9]. ACE2 has one active enzymatic site and degrades angiotensin II (Ang II) into the vasodilator angiotensin 1–7 [Ang (1–7)] [10–12]. Ang (1–7) has a high affinity for Mas receptors (MasR) and has some beneficial effects on the cardiovascular system including a protective role in essential hypertension (EH) and PAH [13, 14]. ACE2 is a type one integral membrane glycoprotein found in most tissues and is highly expressed in the pulmonary vascular endothelium [9, 15]. The Leeds Family Study found that genetic factors accounted for 67% of the phenotypic variation in circulating ACE2 [16], which has a vasoprotective function in patients with CHD-PAH [17, 18].
The human ACE2 gene maps to chromosome Xp22 and spans 39.98 kb of genomic DNA, containing 18 exons and 20 introns. The ACE2 gene is polymorphic. According to The National Center for Biotechnology Information (https://www.ncbi.nlm.nih.gov/snp), the ACE2 gene has 21,754 variants, including 19,797 SNPs; however, some SNPs have a functional effect on the expression of ACE2 [19–22].
Studies investigating ACE2 SNPs correlated it with EH risk in the Chinese population [23], showing differences in the region (south and north) [24–29], ethnicity (Han and non-Han populations) [28, 29], and sex (female and male) [20, 27, 28]. Therefore, effects differ among populations, and environmental and genetic factors are important. There is limited research on the relationship between ACE2 SNPs and CHD-PAH [30], and studies on neonates have yet to be done. In this study, we therefore aimed to investigate the correlation between ACE2 gene variations and CHD-PAH in Chinese Han neonates residing in southern Fujian.
Methods
Study participants
This was a hospital-based case-control study as we previously reported [31]. A total of 321 Chinese Han neonates from the Neonatology Department of Quanzhou First Hospital affiliated of Fujian Medical University and Reproductive Medicine Center of the First Affiliated Hospital of Xiamen University were enrolled from September 2019 to December 2022. They included 113 healthy neonates (control group, 60 males and 53 females) and 208 patients (CHD group, 111 males and 97 females) with left-to-right shunt CHD. According to the guidelines for the diagnosis and treatment of PAH in China (2021 edition) [32], the CHD group was divided into 110 neonates without PAH (60 males and 50 females) [CHD PAH (-)] and 98 neonates with CHD and PAH (51 males and 47 females) [CHD PAH (+)]. The inclusion criteria for the CHD PAH (-) group were as follows: congenital heart disease with a left-to-right shunt, confirmed via echocardiography, and normal pulmonary arterial pressure, as indicated using echocardiography. Two senior cardiologists performed measurements using a double-blind approach, the inclusion criteria for the CHD-PAH (+) group were as follows: congenital heart disease with a left-to-right shunt confirmed via echocardiography; mean pulmonary artery pressure at rest ≥ 25 mmHg, as indicated using echocardiography. The echocardiographic parameters used for grading the probability of PAH include the main parameter and other parameters. If the main parameter, maximum velocity of tricuspid regurgitation (TRVmax) measured by resting state is > 3.4 m/s then the echocardiographic probability of PAH is high. If the TRVmax is 2.9–3.4 m/s, then other echocardiographic parameters suggesting PAH must be used to assign the probability of PAH. These parameters are split into three categories [A: the ventricles; B: the pulmonary artery; C: the inferior vena cava (IVC) and right atrium]. Parameters from at least two different categories are needed to determine the probability of PAH [32]. Patients with concomitant bronchial dysplasia, congenital diaphragmatic hernia, or inherited metabolic disease were excluded. Written informed consent was obtained from all participants and from the parents or legal guardians of participants under 16 years old. This study was reviewed and approved by the Medical Ethics Committee of Quanzhou Medical College [Quanzhou Medical College Ethics Censorship: 2019002].
Clinical parameters
The clinical parameters of all participants were recorded, including sex, gestational week at delivery, birth height and weight, mode of delivery, and clinical classification of CHD.
Collection of blood samples
Two whole blood samples were collected from all patients using EDTA anticoagulant tubes, one was subpackaged in EP tubes and stored in a refrigerator (-80 ℃) for subsequent DNA extraction, and the other sent to the laboratory for centrifugation (2–8 ℃, 3500 rpm, 5 min). The separated plasma was quickly divided into EP tubes, labeled with subject groups and quantities, and stored in the refrigerator (-80 ℃) for subsequent ACE2 and Ang (1–7) measurements.
Tag SNP selection and haplotype reconstruction
Tag SNPs were identified with literature obtained from the PubMed and Chinese HapMap databases (http://hapmap.ncbi.nlm.nih.gov/). Haploview 4.2 software searched for the tag SNPs using a pairwise r2 value and minor allele frequency (MAF) of ≥ 0.8 and 1%, respectively. According to the selection criteria, three ACE2 SNPs (rs2074192, rs2285666, and rs2106809) were identified because of their correlation with EH risk in the population, especially in the Chinese Han population. We used the SHEsisPlus program (http://shesisplus.bio-x.cn/SHEsis.html) to reconstruct the haplotypes [33, 34].
Deoxyribonucleic acid (DNA) extraction and genotyping
Genomic DNA was extracted from the blood cells using the Sangon Biotech DNA Extraction Kit (Shanghai, China). Sanger sequencing was performed for tag SNP genotyping, as previously described [31]. PCR primer sequences and amplification parameters were shown in Supplementary Tables 1, and image of PCR amplification products by agarose gel electrophoresis for ACE2 gene three different SNPs: rs2074192, rs2285666, and rs2106809 were shown in Supplementary Fig. 1.
Measurement of plasma ACE2 and Ang (1–7) levels
Circulating ACE2 and Ang (1–7) levels were measured in neonates with CHD using commercial enzyme-linked immunosorbent assay (ELISA) kits (Human ACE2 ELISA Kit, E-EL-H0281, Elabscience; Human Ang (1–7) ELISA Kit, E-EL-H5518, Elabscience, Wuhan, China). The ELISA kit for the measurement of ACE2 and Ang (1–7) employed the sandwich enzyme immunoassay technique and the competitive inhibition enzyme immunoassay, respectively. All procedures were performed in strict accordance with the manufacturer’s instructions. We conducted batches of tests every six months, with each sample measured in triplicate, and the mean value used for subsequent analysis.
Statistical analysis
The Hardy–Weinberg equilibrium was assessed only for females, owing to the location of ACE2 on the X chromosome, and all male and female data were analyzed separately. Haplotype distribution was analyzed using the SHEsis software (http://analysis.bio-x.cn/SHEsisMain.htm) [33, 34]. Continuous variables were normally distributed and expressed as mean ± standard deviation (x ± s). Significant differences for continuous variables were analyzed via one-way analysis of variance (ANOVA) or the independent sample t-test. The count data was expressed as frequency (%), and the ratio between the two groups was compared using the chi-squared (χ2) test. Univariable and multivariable logistic regression analyses were conducted to assess the associations of three ACE2 single-nucleotide polymorphisms (SNPs) with CHD and PAH risk. Haplotype-based logistic regression analysis was further performed to evaluate the association with PAH risk. Multivariable models were adjusted for gestational age, birth height, birth weight and delivery mode. The results were presented as Odds ratios (ORs) with 95% confidence intervals (CIs). All analyses were performed using SPSS statistical software (version 26.0; SPSS, Chicago, IL, USA) or R software (Version 4.2.0). A 2-tailed P < 0.05 was considered statistically significant. P values were adjusted for multiple testing using the Benjamini–Hochberg procedure to control the false discovery rate. Post hoc power analyses were additionally conducted for sex-stratified SNP and haplotype analyses to assess the impact of the reduced sample size and genotype/haplotype frequencies. After sex stratification, statistical power was limited: approximately 80% power was achieved only for relatively large effects (ORs of approximately 2.3 in the additive SNP model and 2.4 in the haplotype analyses), whereas the dominant SNP model did not achieve 80% power within the tested range. Thus, the sex-stratified and haplotype analyses were not adequately powered to detect small-to-moderate genetic effects.
Results
Comparison of baseline information
The results of comparing neonates with CHD to healthy controls, stratified by sex, are shown in Table 1. No statistically significant differences were observed between the two groups regarding sex, gestational week at delivery, birth height and weight, or mode of delivery (all P > 0 0.05). We next compared the CHD PAH (+) and CHD PAH (-) groups within each sex (Table 2) and similarly found no statistically significant differences in baseline characteristics (all P > 0 0.05).
Table 1.
Baseline data of healthy control and CHD groups
| Factors | Males | Females | ||||||
|---|---|---|---|---|---|---|---|---|
| Control (N = 60) |
CHD (N = 111) |
χ2 /t | P | Control (N = 53) |
CHD (N = 97) |
χ2 /t | P | |
|
Gestational age (weeks) |
39.75 ± 0.58 | 39.60 ± 0.61 | 1.552 | 0.123 | 39.84 ± 0.56 | 39.72 ± 0.68 | 1.103 | 0.272 |
|
Birth height (cm) |
50.38 ± 2.27 | 50.27 ± 2.00 | 0.336 | 0.737 | 49.30 ± 2.24 | 49.40 ± 2.13 | -0.270 | 0.787 |
|
Birth weight (g) |
3299.20 ± 357.76 | 3229.48 ± 272.61 | 1.426 | 0.156 | 3213.11 ± 311.68 | 3265.80 ± 303.57 | -1.007 | 0.316 |
|
Delivery mode (Eutocia/Cesarean) |
45/15 | 85/26 | 0.053 | 0.852 | 40/13 | 74/23 | 0.013 | 1.000 |
CHD Congenital heart disease
Table 2.
Baseline information of neonates in the CHD PAH(+) and CHD PAH(-) groups
| Characteristics | Males | Females | ||||
|---|---|---|---|---|---|---|
| CHD PAH(-) (N = 60) |
CHD PAH(+) (N = 51) |
P | CHD PAH(-) (N = 50) |
CHD PAH(+) (N = 47) |
P | |
| Gestational age | 39.63 ± 0.53 | 39.57 ± 0.69 | 0.579 | 39.75 ± 0.65 | 39.69 ± 0.71 | 0.659 |
| (weeks) | 50.50 ± 2.11 | 50.00 ± 1.85 | 0.192 | 49.18 ± 2.26 | 49.64 ± 1.98 | 0.292 |
| Birth height | ||||||
| (cm) | 3254.78 ± 266.61 | 3199.71 ± 279.20 | 0.291 | 3257.20 ± 315.51 | 3274.96 ± 293.46 | 0.775 |
| Birth weight | ||||||
| (g) | 47/13 | 38/13 | 0.659 | 37/13 | 37/10 | 0.639 |
| Delivery mode | ||||||
| (Eutocia/Cesarean) | 24 | 20 | 0.194 | 20 | 24 | 0.312 |
| Ventricular septal defect | ||||||
| Atrial septal defect | 31 | 26 | 1.000 | 27 | 22 | 0.545 |
| Patent ductus arteriosus* | 5 | 5 | 1.000 | 3 | 1 | 0.618 |
CHD PAH(-) Congenital heart disease without pulmonary arterial hypertension, CHD PAH(+) Congenital heart disease with pulmonary arterial hypertension
*Differences between CHD PAH(-) and CHD PAH(+) were tested by Fisher’s exact test
Hardy-Weinberg equilibrium
The genotyping distribution of the three SNPs was consistent with the Hardy-Weinberg genetic equilibrium in the control (rs2074192: χ2 = 1.397, P = 0.237; rs2285666: χ2 = 2.184, P = 0.139; and rs2106809: χ2 = 3.182, P = 0.074) and CHD group (rs2074192: χ2 = 0.267, P = 0.606; rs2285666: χ2 = 1.721, P = 0.190; and rs2106809: χ2 = 0.792, P = 0.373) after the Hardy-Weinberg genetic equilibrium analysis in females.
Sex-stratified association of ACE2 gene polymorphisms with neonatal CHD and CHD-PAH risk
The genotype and allele frequencies of ACE2 gene polymorphisms are summarized in Tables 3 and 4, Supplementary Tables 2 and 4 for female participants, and in Supplementary Tables 3 and 5 for male participants. Sex-stratified logistic regression analyses were conducted to assess the associations of the three ACE2 SNPs with the risks of CHD and CHD-PAH. Both univariable and multivariable models were used, with the latter adjusted for available potential confounders. No significant associations were observed between the three SNPs and susceptibility to CHD or the risk of PAH in either univariable or multivariable analyses among patients with CHD. After correction for multiple comparisons, none of the ACE2 variants showed a robust association with the risk of CHD or PAH.
Table 3.
Genotype distribution and allele frequency of ACE2 polymorphism between healthy control and CHD groups in female
| SNPs | Genotype /Allele | Cases (%) | Univariable model# | Multivariable model* | |||||
|---|---|---|---|---|---|---|---|---|---|
| Control (N = 53) | CHD (N = 97) |
OR (95%CI) | P | FDR_P | OR (95%CI) | P | FDR_P | ||
| rs2074192 | CC | 18 (33.96) | 33 (34.02) | 1.000 | 1.000 | ||||
| CT + TT | 35 (66.04) | 64 (65.98) | 0.997(0.492–2.022) | 0.994 | 0.994 | 1.000(0.490–2.042) | 1.000 | 1.000 | |
| C | 58 (54.72) | 111 (57.22) | 1.000 | 1.000 | |||||
| T | 48 (45.28) | 83 (42.78) | 0.911(0.577–1.438) | 0.69 | 0.898 | 0.927(0.585–1.469) | 0.746 | 0.898 | |
| rs2285666 | CC | 14 (26.42) | 29 (29.90) | 1.000 | 1.000 | ||||
| CT + TT | 39 (73.58) | 68 (70.10) | 0.842(0.398–1.781) | 0.652 | 0.898 | 0.799(0.374–1.705) | 0.561 | 0.898 | |
| C | 49 (46.23) | 100 (51.55) | 1.000 | 1.000 | |||||
| T | 57 (53.77) | 94 (48.45) | 0.832(0.535–1.294) | 0.414 | 0.898 | 0.816(0.523–1.275) | 0.372 | 0.898 | |
| rs2106809 | CC | 17 (32.08) | 29 (29.90) | 1.000 | 1.000 | ||||
| CT + TT | 36 (67.92) | 68 (70.10) | 1.107(0.538–2.280) | 0.782 | 0.898 | 1.102(0.532–2.283) | 0.794 | 0.898 | |
| C | 54 (50.94) | 102 (52.58) | 1.000 | 1.000 | |||||
| T | 52 (49.06) | 92 (47.42) | 0.944(0.607–1.471) | 0.800 | 0.898 | 0.948(0.607–1.481) | 0.814 | 0.898 | |
CHD Congenital heart disease
FDR indicates false discovery rate-adjusted P values (Benjamini-Hochberg method)
# Univariable logistic regression
* Multivariable logistic regression adjusting for gestational age, birth height, birth weight and delivery mode
Table 4.
Genotype distribution and allele frequency of ACE2 polymorphism between CHD PAH (−) and CHD PAH (+) groups in female
| SNPs | Genotype /Allele | CHD Cases (%) | Univariable model# | Multivariable model* | |||||
|---|---|---|---|---|---|---|---|---|---|
| PAH(-) (N = 50) | PAH(+) (N = 47) | OR (95%CI) | P | FDR_P | OR (95%CI) | P | FDR_P | ||
| rs2074192 | CC | 17 (34.00) | 16 (34.04) | 1.000 | 1.000 | ||||
| CT + TT | 33 (66.00) | 31 (65.96) | 0.998(0.431–2.313) | 0.996 | 0.996 | 1.023(0.435–2.407) | 0.958 | 0.989 | |
| C | 58 (58.00) | 53 (56.38) | 1.000 | 1.000 | |||||
| T | 42 (42.00) | 41 (43.62) | 1.065(0.611–1.855) | 0.825 | 0.996 | 1.100(0.626–1.932) | 0.741 | 0.988 | |
| rs2285666 | CC | 16 (32.00) | 13 (27.66) | 1.000 | 1.000 | ||||
| CT + TT | 34 (68.00) | 34 (72.34) | 1.231(0.514–2.946) | 0.641 | 0.996 | 1.192(0.490–2.899) | 0.699 | 0.988 | |
| C | 55 (55.00) | 45 (47.87) | 1.000 | 1.000 | |||||
| T | 45 (45.00) | 49 (52.13) | 1.288(0.756–2.196) | 0.352 | 0.946 | 1.259(0.730–2.171) | 0.407 | 0.988 | |
| rs2106809 | CC | 13 (26.00) | 16 (34.04) | 1.000 | 1.000 | ||||
| CT + TT | 37 (74.00) | 31 (65.96) | 0.681(0.284–1.631) | 0.388 | 0.946 | 0.698(0.287–1.696) | 0.427 | 0.988 | |
| C | 49 (49.00) | 53 (56.38) | 1.000 | 1.000 | |||||
| T | 51 (51.00) | 41 (43.62) | 0.761(0.441–1.312) | 0.326 | 0.946 | 0.778(0.446–1.357) | 0.376 | 0.988 | |
CHD PAH(-) Congenital heart disease without pulmonary arterial hypertension, CHD PAH(+) Congenital heart disease with pulmonary arterial hypertension
FDR indicates false discovery rate-adjusted P values (Benjamini-Hochberg method)
# Univariable logistic regression
* Multivariable logistic regression adjusting for gestational age, birth height, birth weight and delivery mode
Linkage disequilibrium (LD) and haplotype analysis
LD analysis using the SHEsisplus software showed that the three selected SNPs might fall within the same linkage area. In females, rs2074192 and rs2285666: D’= 0.97, R2 = 0.66; rs2285666 and rs2106809: D’= 1, R2 = 0.84; rs2074192 and rs2106809: D’= 0.88, R2 = 0.64. In males, rs2074192 and rs2285666: D’= 0.96, R2 = 0.60; rs2285666 and rs2106809: D’= 0.98, R2 = 0.68; rs2074192 and rs2106809: D’= 0.93, R2 = 0.66.
Haplotype analysis was performed based on the three ACE2 SNPs, rs2074192, rs2285666, and rs2106809. Three common haplotypes with an estimated frequency of ≥ 3% were identified, namely CTC, TCT, and CCT. The distributions of these haplotypes in CHD-PAH(+) and CHD-PAH(−) are presented separately for males and females in Table 5. We further evaluated whether these ACE2 haplotypes were associated with the risk of PAH among patients with CHD.
Table 5.
The relationship between the haplotype of ACE2 gene and PAH in male and female CHD neonates
| Gender | Haplotype | Haplotype Counts (%) | Univariable model# | Multivariable model* | |||||
|---|---|---|---|---|---|---|---|---|---|
| CHD PAH(-) | CHD PAH(+) | OR (95%CI) | P | FDR_P | OR (95%CI) | P | FDR_P | ||
| Female | CTC | 43 (43.00) | 48 (51.00) | 1.214 (0.516–2.888) | 0.657 | 0.84 | 1.175 (0.491–2.841) | 0.717 | 0.726 |
| TCT | 42 (42.00) | 39 (41.49) | 1.090 (0.472–2.529) | 0.84 | 0.84 | 1.164 (0.498–2.757) | 0.726 | 0.726 | |
| CCT | 12 (12.00) | 3 (3.10) | 0.216 (0.047–0.740) | 0.025 | 0.074 | 0.187 (0.039–0.670) | 0.018 | 0.053 | |
| Male | CTC | 28 (46.67) | 25 (47.06) | 0.688 (0.135–2.950) | 0.620 | 0.805 | 0.670 (0.125–3.060) | 0.611 | 0.805 |
| TCT | 24 (40.00) | 19 (37.25) | 1.099 (0.520–2.327) | 0.805 | 0.805 | 1.102 (0.509–2.391) | 0.805 | 0.805 | |
| CCT | 5 (8.33) | 3 (5.88) | 0.891 (0.411–1.917) | 0.767 | 0.805 | 0.902 (0.411–1.965) | 0.795 | 0.805 | |
CHD PAH(-) Congenital heart disease without pulmonary arterial hypertension, CHD PAH(+) Congenital heart disease with pulmonary arterial hypertension
FDR indicates false discovery rate-adjusted P values (Benjamini-Hochberg method)
# Univariable logistic regression
* Multivariable logistic regression adjusting for gestational age, birth height, birth weight and delivery mode
In the sex-stratified analyses, associations between the three common haplotypes and CHD-PAH risk were assessed using both univariable and multivariable logistic regression models. Among female patients, neither the CTC nor the TCT haplotype showed a significant association with the risk of CHD-PAH in either the crude or adjusted models. The CCT haplotype showed a nominal association with a reduced risk of CHD-PAH in the univariable analysis (OR = 0.216, 95% CI: 0.047–0.740; P = 0.025), and this association remained nominally significant after adjustment for available potential confounders in the multivariable model (OR = 0.187, 95% CI: 0.039–0.670; P = 0.018). However, this finding did not remain statistically significant after correction for multiple comparisons (all FDR_P > 0.05). In addition, the number of the CCT haplotype counts in female was very small, which may have resulted in unstable effect estimates and limited precision of the confidence intervals. Therefore, this nominal association should be interpreted with caution. Among male patients, none of the three common ACE2 haplotypes was significantly associated with the risk of PAH in either univariable or multivariable analyses. Overall, no ACE2 haplotype showed a robust association with PAH risk among patients with CHD after correction for multiple testing.
Comparisons of ACE2 haplotypes with circulating ACE2 and Ang (1–7) levels in neonates with CHD
As shown in Table 6, ANOVA indicated a difference in circulating Ang-(1–7) levels across ACE2 haplotypes among female neonates. Specifically, females carrying the CCT haplotype had higher circulating Ang-(1–7) concentrations than those carrying the CTC or TCT haplotypes (160.16 ± 19.24 pg/mL vs. 140.54 ± 28.40 pg/mL and 139.77 ± 29.85 pg/mL, respectively; P = 0.037). In contrast, no significant difference in Ang-(1–7) levels across haplotypes was observed among male neonates. However, the association observed in females did not remain statistically significant after correction for multiple comparisons (FDR_ P = 0.074). In the present study, circulating ACE2 levels did not differ significantly among the three common ACE2 haplotypes in either females or males.
Table 6.
Comparisons of circulating ACE2 and Ang (1–7) levels with ACE2 haplotypes in CHD neonates
| Haplotype | Female | Male | ||||
|---|---|---|---|---|---|---|
| Cases | ACE2 (ng/ml) | Ang(1–7) (pg/ml) | Cases | ACE2 (ng/ml) | Ang(1–7) (pg/ml) | |
| CTC | 66 | 11.38 ± 2.41 | 140.54 ± 28.40 | 53 | 12.19 ± 3.84 | 143.63 ± 31.35 |
| TCT | 63 | 11.24 ± 2.53 | 139.77 ± 29.85 | 43 | 12.20 ± 3.42 | 139.78 ± 29.7 |
| CCT | 15 | 12.16 ± 1.49 | 160.16 ± 19.24 | 8 | 13.08 ± 2.96 | 142.81 ± 32.89 |
| P-value | 0.401 | 0.037 | 0.800 | 0.828 | ||
| FDR_P | 0.401 | 0.074 | 0.828 | 0.828 | ||
Significant differences for continuous variables were analyzed via one-way analysis of variance (ANOVA)
ACE2 Angiotensin-converting enzyme 2, Ang (1–7) Angiotensin (1–7)
FDR indicates false discovery rate-adjusted P values (Benjamini-Hochberg method)
Further carrier-based analysis showed that female patients carrying one copy of the CCT haplotype had higher circulating Ang-(1–7) levels than non-carriers of this haplotype (160.16 ± 19.24 pg/mL vs. 137.11 ± 29.09 pg/mL; P = 0.016). No comparable association was observed for the CTC or TCT haplotypes. Nevertheless, this association also lost statistical significance after FDR correction (FDR_P = 0.096; Table 7), suggesting that the observed difference should be interpreted cautiously. Consistently, no significant differences in circulating ACE2 levels were detected across haplotypes or carrier groups (all P > 0.05).
Table 7.
Comparison of circulating ACE2 and Ang (1–7) levels among three haplotypes of ACE2 in female CHD neonates
| Haplotype | Indicator | Cases | Subtypes | P | FDR_P | ||
|---|---|---|---|---|---|---|---|
| None | Heterozygote | Homozygote | |||||
| CTC | ACE2 (ng/ml) | 31/41/25 | 11.48 ± 2.26 | 11.35 ± 2.74 | 11.42 ± 1.80 | 0.973 | 0.973 |
| Ang(1–7) (pg/ml) | 140.96 ± 30.56 | 144.12 ± 29.35 | 134.69 ± 26.30 | 0.442 | 0.663 | ||
| TCT | ACE2 (ng/ml) | 34/45/18 | 11.73 ± 1.99 | 11.31 ± 2.60 | 11.05 ± 2.40 | 0.575 | 0.690 |
| Ang(1–7) (pg/ml) | 142.35 ± 27.55 | 142.82 ± 29.72 | 132.16 ± 29.62 | 0.387 | 0.663 | ||
| CCT | ACE2 (ng/ml) | 82/15/0 | 11.27 ± 2.46 | 12.16 ± 1.49 | 0.405 | 0.663 | |
| Ang(1–7) (pg/ml) | 137.11 ± 29.09 | 160.16 ± 19.24 | 0.016 | 0.096 | |||
Significant differences for continuous variables were analyzed via one-way analysis of variance (ANOVA)
ACE2 Angiotensin-converting enzyme 2, Ang (1–7) Angiotensin (1–7)
FDR indicates false discovery rate-adjusted P values (Benjamini-Hochberg method)
Heterozygote: patients with two different haplotypes
Homozygote: patients with two identical haplotypes
Discussion
The prevalent types of left-to-right shunt CHD include VSD, ASD, and PDA. Left-to-right shunt CHD causes increased pulmonary arterial pressure and/or blood flow, which is the main pathological and anatomical cause of PAH, and may induce endothelial dysfunction and vascular remodeling, increase the mortality [3–6]. Therefore, PAH caused by left-to-right shunt CHD is a gradual pathological process from “high pulmonary blood flow” to “vascular remodeling”. CHD-PAH is the most common cause of PAH in China, and many CHD patients lose the opportunity for surgery due to the complication of PAH [32]. Previous studies of PAH have mostly focused on adults, in order to reduce the influence of vascular remodeling and environmental factors on pulmonary artery pressure, we chose newborns as the research subjects in this study. Due to the influence of geographic area and ethnicity on the frequency of genetic variants, the parents of all patients in this study are Han Chinese residents who have been living in southern Fujian for a long time.
Owing to limited research on the association between ACE2 SNPs and PAH risk, the correlation between ACE2 SNPs and susceptibility to EH was used to help select tag SNPs. The reports on the association between ACE2 SNPs and EH risk are mostly on rs2285666 and rs2106809 [23, 24, 27, 35–41]. Among them, a meta-analysis involving 14,122 Asians and Caucasians confirmed that rs2285666 and rs2106809 polymorphisms were correlated in both males and females with the risk of hypertension, and may have important roles in the etiology of EH [35]. Three studies investigated the association between ACE2 rs2074192 and the risk of EH; however, the results of these studies were all positive [24, 38, 42]. There are few reports on other SNPs including ACE2 rs4646155 [24, 38, 43], rs879922 [24, 43], and rs1978124 [26, 35, 39]; however, the results from these studies were inconsistent. Therefore, rs2074192, rs2285666, and rs2106809 are correlated with the risk of EH in the population, especially in the Chinese Han population. The functional SNP remains unknown. According to the Ensemble project, ACE2 rs2285666 is expected to be a functional SNP owing to its location in the splice region of ACE2. In addition, the intronic ACE2 SNP, rs2106809, may create an intronic exonic splicing enhancer (ESE) site to influence the splicing efficiency of the ACE2 gene [27] and may stay in a microRNA-binding site or gene, thereby possibly regulating gene expression by altering the miRNA– mRNA interaction [44]. The distribution frequencies of the alleles at these three loci of the ACE2 gene in the East Asian population, which we searched on the NCBI website (https://www.ncbi.nlm.nih.gov/snp/rs2074192?vertical_tab=true#publications), are as follows: rs2074192: C = 0.575, T = 0.425; rs2285666: C = 0.438, T = 0.562; rs2106809: T = 0.39, C = 0.61. Moreover, rs2074192 is in the same linkage disequilibrium (LD) block as rs2106809; however, it is in strong LD with many other ACE2 SNPs [40]. There are both similarities and significant differences in the pathogenesis of CHD-PAH and EH. Our study relied on SNPs previously studied in EH, the rationale for limiting the analysis to only three tag SNPs remained weak, and the study design did not capture broader ACE2 variation. In the future, we will expand the sample size and consider using high-throughput sequencing to screen SNPs associated with neonatal CHD.
Sex-stratified logistic regression analyses were performed to evaluate the associations between the three ACE2 SNPs and the risks of CHD and CHD-PAH. Both univariable and multivariable models were applied, with the multivariable models adjusted for available potential confounders. From the results, no robust association was identified between these ACE2 variants and CHD or PAH risk after correction for multiple comparisons (female: Tables 3 and 4 and Supplementary Tables 2 and 4, male: Supplementary Tables 3 and 5). For CHD-PAH susceptibility, our results are similar to those of a previous relation study on CHD-PAH and ACE2 gene polymorphism in Chinese Han patients aged ≥ 14 years old [30].
Haplotype analysis revealed three major haplotypes - CTC, TCT, and CCT in the study region. It is noteworthy that CCT haplotype in females showed a nominal association with CHD-PAH in univariable and multivariable models (both P < 0.05), and this association did not remain statistically significant after FDR correction (both FDR_P > 0.05), suggesting that the finding should be interpreted cautiously. No haplotype showed an association with CHD-PAH risk in males (Table 5). ACE2 gene had special sex-based specificity, previous research on the correlation between ACE2 SNPs and PAH risk was limited. A similar study showed that females with CHD (≥ 14 years old) carrying the CCA or CCG haplotypes (rs2074192-rs2285666-rs2106809) had a lower risk of elevated pulmonary artery pressure than did those without the haplotypes [30], and so far, studies on neonates have yet to be done.
Comparisons of ACE2 haplotypes with circulating levels of ACE2 and Ang (1–7) revealed that either carrying CCT haplotype or with CCT heterozygotes in female patients had higher circulating Ang (1–7) levels than did those without (both P < 0.05). However, further FDR correction suggested that these associations did not remain statistically significant(both FDR_P > 0.05). Similar results among the haplotypes were not found for ACE2 levels in females. Neither ACE2 nor Ang(1–7) levels were associated with haplotypes in males (Table 6). A recent study showed no statistically significant differences between ACE2 levels and ACE2 SNPs in patients with CHD-PAH [30].
Several limitations should be considered. First, the study was underpowered for sex-stratified SNP and haplotype analyses. This limitation was particularly relevant in female CHD-PAH risk analyses, in which only 3 CHD PAH(+) carriers of the CCT haplotype were available. The post hoc analyses showed that approximately 80% power was achieved only for relatively large effects (ORs of approximately 2.3 in the additive SNP model and 2.4 in the haplotype analyses); the dominant SNP model did not achieve 80% power within the tested range. Accordingly, small-to-moderate genetic effects may have been missed, and subgroup estimates may be unstable. The nominal associations, particularly those from female subgroup and haplotype analyses, should therefore be interpreted as exploratory. No SNP- or haplotype-level association remained statistically significant after correction for multiple comparisons, and replication in larger, adequately powered independent cohorts is required. Second, the restricted candidate-SNP strategy did not provide comprehensive coverage of genetic variation within the genes of interest or other loci potentially relevant to CHD-PAH; rare variants and broader gene-gene interactions were not assessed. Third, PAH was analyzed as a binary outcome because detailed hemodynamic data and biomarker-based severity indicators were unavailable. We were therefore unable to evaluate genotype-phenotype relationships across the spectrum of PAH severity, potentially obscuring more subtle associations. Finally, although multivariable logistic regression was adjusted for available covariates, CHD severity, heart failure status, and oxygenation status were unavailable for adjustment. Residual confounding may therefore remain. Future studies with comprehensive genetic coverage and detailed longitudinal clinical phenotyping are needed to validate and refine these findings.
Conclusions
In conclusion, the present study found no significant associations between ACE2 rs2074192, rs2285666, or rs2106809, either individually at the allele and genotype levels or jointly as haplotypes, and the risk of CHD-PAH in neonates of either sex. These findings should be considered exploratory and require validation in larger, adequately powered, independent cohorts.
Supplementary Information
Acknowledgements
We would like to thank the editors and reviewers for their suggestions and Editage (https://www.editage.cn) for English language editing.
Abbreviations
- CHD
Congenital heart disease
- CHD-PAH
Congenital heart disease-related pulmonary arterial hypertension
- CHD PAH (+)
Congenital heart disease with pulmonary arterial hypertension
- CHD PAH (-)
Congenital heart disease without pulmonary arterial hypertension
- PAH
Pulmonary arterial hypertension
- EH
Essential hypertension
- SNPs
Single nucleotide polymorphisms
- ACE2
Angiotensin-converting enzyme 2
- Ang(1–7)
Angiotensin1-7
- Ang II
Angiotensin II
- RAS
Renin-angiotensin system
- OR
Odds ratio
- CI
Confidence interval
Authors’ contributions
YFC and LL conceived and designed the trial, and reviewed and edited the manuscript. CLW, QFL, JC and LFD contributed to curate and analyze the data. YFC and CLW wrote the manuscript. CLW, SML, and YSL contributed to the implementation and monitoring of the trial. All authors had read and approved the fnal version of the manuscript, and agree with the order of presentation of the authors.
Funding
This study was supported by grants from the Key Science and Technology Project of Quanzhou Medical College, China (XJK2204A), Natural Science Foundation project of Fujian Province, China (2023J011781), and the Quanzhou City Science and Technology Program, China (2019C081R).
Data availability
The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.
Declarations
Ethics approval and consent to participate
This study adhered to the Helsinki Declaration and was reviewed and approved by the Medical Ethics Committee of Quanzhou Medical College [Quanzhou Medical College Ethics Censorship: 2019002]. Written informed consent was obtained from all participants and from the parents or legal guardians of participants under 16 years old.
Consent for publication
Not applicable.
Competing interests
The authors declare no competing interests.
Footnotes
Publisher’s note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Youfang Chen and Cuiling Wang contributed equally to this article.
References
- 1.Verdurmen KMJ, Eijsvoogel NB, Lempersz C, Vullings R, Schroer C, van Laar JOEH, et al. A systematic review of prenatal screening for congenital heart disease by fetal electrocardiography. Int J Gynaecol Obstet. 2016;135(2):129–34. [DOI] [PubMed] [Google Scholar]
- 2.Zhang Y, Wang J, Zhao J, Huang G, Liu K, Pan W, et al. Current status and challenges in prenatal and neonatal screening, diagnosis, and management of congenital heart disease in China. Lancet Child Adolesc Health. 2023;7(7):479–89. [DOI] [PubMed] [Google Scholar]
- 3.Wacker J, Beghetti M. Pulmonary hypertension in pediatrics: from clinical suspicion to management. Eur J Pediatr. 2025;184(5):288. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Jumoboyev IJ. Congenital heart disease and pulmonary hypertension in pediatrics. Sci Innov. 2025;4(7):172–84. [Google Scholar]
- 5.Kolte D, Lakshmanan S, Jankowich MD, Brittain EL, Maron BA, Choudhary G. Mild Pulmonary Hypertension Is Associated With Increased Mortality: A Systematic Review and Meta-Analysis. J Am Heart Assoc. 2018;7(18):e009729. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Constantine A, Dimopoulos K, Opotowsky AR. Congenital Heart Disease and Pulmonary Hypertension. Cardiol Clin. 2020;38:445–56. [DOI] [PubMed] [Google Scholar]
- 7.Marshall RP. The pulmonary renin-angiotensin system. Curr Pharm Des. 2003;9(9):715–22. [DOI] [PubMed] [Google Scholar]
- 8.Tan WSD, Liao W, Zhou S, Mei D, Wong W-SF. Targeting the renin-angiotensin system as novel therapeutic strategy for pulmonary diseases. Curr Opin Pharmacol. 2018;40:9–17. [DOI] [PubMed] [Google Scholar]
- 9.Gintoni I, Adamopoulou M, Yapijakis C. The Impact of ACE and ACE2 Gene Polymorphisms in Pulmonary Diseases Including COVID-19. Vivo. 2022;36:13–29. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Donoghue M, Hsieh F, Baronas E, Godbout K, Gosselin M, Stagliano N, et al. A novel angiotensin-converting enzyme-related carboxypeptidase (ACE2) converts angiotensin I to angiotensin 1–9. Circ Res. 2000;87:E1–9. [DOI] [PubMed] [Google Scholar]
- 11.Tipnis SR, Hooper NM, Hyde R, Karran E, Christie G, Turner AJ. A human homolog of angiotensin-converting enzyme. Cloning and functional expression as a captopril-insensitive carboxypeptidase. J Biol Chem. 2000;275:33238–43. [DOI] [PubMed] [Google Scholar]
- 12.Varagic J, Ahmad S, Nagata S, Ferrario CM. ACE2: angiotensin II/angiotensin-(1–7) balance in cardiac and renal injury. Curr Hypertens Rep. 2014;16:420. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Burrell LM, Harrap SB, Velkoska E, Patel SK. The ACE2 gene: its potential as a functional candidate for cardiovascular disease. Clin Sci (Lond). 2013;124:65–76. [DOI] [PubMed] [Google Scholar]
- 14.Papavassiliou KA, Gogou VA, Papavassiliou AG. Angiotensin-Converting Enzyme 2 (ACE2) Signaling in Pulmonary Arterial Hypertension: Underpinning Mechanisms and Potential Targeting Strategies. IJMS. 2023;24:17441. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 15.Saravi B, Li Z, Lang CN, Schmid B, Lang FK, Grad S, et al. The Tissue Renin-Angiotensin System and Its Role in the Pathogenesis of Major Human Diseases. Quo Vadis? Cells. 2021;10:650. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Rice GI, Jones AL, Grant PJ, Carter AM, Turner AJ, Hooper NM. Circulating Activities of Angiotensin-Converting Enzyme, Its Homolog, Angiotensin-Converting Enzyme 2, and Neprilysin in a Family Study. Hypertension. 2006;48:914–20. [DOI] [PubMed] [Google Scholar]
- 17.Dai H, Guo Y, Guang X, Xiao Z, Zhang M, Yin X. The Changes of Serum Angiotensin-Converting Enzyme 2 in Patients with Pulmonary Arterial Hypertension due to Congenital Heart Disease. Cardiology. 2013;124:208–12. [DOI] [PubMed] [Google Scholar]
- 18.Sandoval J, Del Valle-Mondragón L, Masso F, Zayas N, Pulido T, Teijeiro R, et al. Angiotensin converting enzyme 2 and angiotensin (1–7) axis in pulmonary arterial hypertension. Eur Respir J. 2020;56:1902416. [DOI] [PubMed] [Google Scholar]
- 19.Wu Y-H, Li J-Y, Wang C, Zhang L-M, Qiao H. The ACE2 G8790A Polymorphism: Involvement in Type 2 Diabetes Mellitus Combined with Cerebral Stroke. J Clin Lab Anal. 2017;31:e22033. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 20.Zhang Q, Cong M, Wang N, Li X, Zhang H, Zhang K, et al. Association of angiotensin-converting enzyme 2 gene polymorphism and enzymatic activity with essential hypertension in different gender: A case–control study. Medicine. 2018;97:e12917. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 21.Asselta R, Paraboschi EM, Mantovani A, Duga S. ACE2 and TMPRSS2 variants and expression as candidates to sex and country differences in COVID-19 severity in Italy. Aging (Albany NY). 2020 Jun 5;12(11):10087–98. [DOI] [PMC free article] [PubMed]
- 22.Khayat AS, De Assumpção PP, Meireles Khayat BC, Thomaz Araújo TM, Batista-Gomes JA, Imbiriba LC et al. ACE2 polymorphisms as potential players in COVID-19 outcome. Pershouse MA, editor. PLoS ONE. 2020;15:e0243887. [DOI] [PMC free article] [PubMed]
- 23.Lu N, Yang Y, Wang Y, Liu Y, Fu G, Chen D, et al. ACE2 gene polymorphism and essential hypertension: an updated meta-analysis involving 11,051 subjects. Mol Biol Rep. 2012;39:6581–9. [DOI] [PubMed] [Google Scholar]
- 24.Luo Y, Liu C, Guan T, Li Y, Lai Y, Li F, et al. Association of ACE2 genetic polymorphisms with hypertension-related target organ damages in south Xinjiang. Hypertens Res. 2019;42:681–9. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 25.Pan Y, Wang T, Li Y, Guan T, Lai Y, Shen Y, et al. Association of ACE2 polymorphisms with susceptibility to essential hypertension and dyslipidemia in Xinjiang, China. Lipids Health Dis. 2018;17:241. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 26.Niu W, Qi Y, Hou S, Zhou W, Qiu C. Correlation of angiotensin-converting enzyme 2 gene polymorphisms with stage 2 hypertension in Han Chinese. Transl Res. 2007;150:374–80. [DOI] [PubMed] [Google Scholar]
- 27.Chen YY, Liu D, Zhang P, Zhong JC, Zhang CJ, Wu SL, et al. Impact of ACE2 gene polymorphism on antihypertensive efficacy of ACE inhibitors. J Hum Hypertens. 2016;30:766–71. [DOI] [PubMed] [Google Scholar]
- 28.Yi L, Gu Y, Wang X, An L, Xie X, Shao W, et al. Association of ACE, ACE2 and UTS2 Polymorphisms with Essential Hypertension in Han and Dongxiang Populations from North-western China. J Int Med Res. 2006;34:272–83. [DOI] [PubMed] [Google Scholar]
- 29.Fan X, Wang Y, Wang H, Sun K, Zhang W, Song X, et al. Polymorphisms of angiotensin-converting enzyme (ACE) and ACE2 are not associated with orthostatic blood pressure dysregulation in hypertensive patients. Acta Pharmacol Sin. 2009;30:1237–44. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 30.Zi-yang Y, Nanshan X, Dongling L, Tao H, Yigao H, Danqing Y, et al. ACE2 gene polymorphisms are associated with elevated pulmonary artery pressure in congenital heart diseases. Gene. 2023;882:147642. [DOI] [PubMed] [Google Scholar]
- 31.Lin Q-F, Rao J-H, Luo S-M, Wang Q-M, Deng L-F, Chen X, et al. Relation between endothelial nitric oxide synthase genetic polymorphisms and pulmonary arterial hypertension in newborns with congenital heart disease. Clin Exp Hypertens. 2022;44:567–72. [DOI] [PubMed] [Google Scholar]
- 32.Pulmonary Embolism and Pulmonary Vascular Disease Group of the Respiratory Disease Branch of the Chinese Medical Association, Chinese Guidelines for Diagnosis and Treatment of Pulmonary Hypertension. (2021 Edition). Zhonghua Yi Xue Za Zhi. 2021;101.1:11–51.
- 33.Shi YY, He L. SHEsis, a powerful software platform for analyses of linkage disequilibrium, haplotype construction, and genetic association at polymorphism loci. Cell Res. 2005;15:97–8. [DOI] [PubMed] [Google Scholar]
- 34.Li Z, Zhang Z, He Z, Tang W, Li T, Zeng Z, et al. A partition-ligation-combination-subdivision EM algorithm for haplotype inference with multiallelic markers: update of the SHEsis. Cell Res. 2009;19:519–23. http://analysis.bio-x.cn. [DOI] [PubMed] [Google Scholar]
- 35.Yang M, Zhao J, Xing L, Shi L. The association between angiotensin-converting enzyme 2 polymorphisms and essential hypertension risk: A meta-analysis involving 14,122 patients. J Renin Angiotensin Aldosterone Syst. 2015;16:1240–4. [DOI] [PubMed] [Google Scholar]
- 36.Zhang X, Wang Y, Zheng Y, Yuan J, Tong J, Xu J, et al. Effect of ACE, ACE2 and CYP11B2 gene polymorphisms and noise on essential hypertension among steelworkers in China: a case–control study. BMC Med Genomics. 2022;15:22. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 37.Pinheiro DS, Santos RS, Jardim PCBV, Silva EG, Reis AAS, Pedrino GR et al. The combination of ACE I/D and ACE2 G8790A polymorphisms revels susceptibility to hypertension: A genetic association study in Brazilian patients. Ciccacci C, editor. PLoS ONE. 2019;14:e0221248. [DOI] [PMC free article] [PubMed]
- 38.Fan Z, Wu G, Yue M, Ye J, Chen Y, Xu B, et al. Hypertension and hypertensive left ventricular hypertrophy are associated with ACE2 genetic polymorphism. Life Sci. 2019;225:39–45. [DOI] [PubMed] [Google Scholar]
- 39.Li J, Feng M, Wang Y, Li Y, Zhang Y, Li L, et al. The relationship between three X-linked genes and the risk for hypertension among northeastern Han Chinese. J Renin Angiotensin Aldosterone Syst. 2015;16:1321–8. [DOI] [PubMed] [Google Scholar]
- 40.He Y, Yang W, Liu S, Gan L, Zhang F, Mu C, et al. Interactions between angiotensin-converting enzyme-2 polymorphisms and high salt intake increase the risk of hypertension in the Chinese Wa population. Int J Clin Exp Pathol. 2017;10:11159–68. [PMC free article] [PubMed] [Google Scholar]
- 41.Al-Eitan L, Al-Khaldi S, Ibdah RK. ACE gene polymorphism and susceptibility to hypertension in a Jordanian adult population. PLoS ONE. 2024;19:e0304271. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 42.Hamet P, Pausova Z, Attaoua R, Hishmih C, Haloui M, Shin J, et al. SARS-CoV-2 Receptor ACE2 Gene Is Associated with Hypertension and Severity of COVID 19: Interaction with Sex, Obesity, and Smoking. Am J Hypertens. 2021;34:367–76. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 43.Liu D, Chen Y, Zhang P, Zhong J, Jin L, Zhang C, et al. Association between circulating levels of ACE2-Ang-(1–7)-MAS axis and ACE2 gene polymorphisms in hypertensive patients. Medicine. 2016;95:e3876. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 44.Bhaumik P, Gopalakrishnan C, Kamaraj B, Purohit R. Single nucleotide polymorphisms in microRNA binding sites: implications in colorectal cancer. ScientificWorldJournal. 2014;2014:547154. [DOI] [PMC free article] [PubMed] [Google Scholar]
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Supplementary Materials
Data Availability Statement
The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.