Journal of the American Society of Hypertension 12(3) (2018) 230–237

Research Article
Common variants at somatostatin are significantly
associated with hypertension incidence in smoking
and drinking populations
Hui Zhu, BSa, Lijun Zhu, MSa, Zhengmei Fang, BSa, Song Yang, MSb,
Yanchun Chen, MSb, Yuelong Jin, MSa, Xianghai Zhao, MSb,
Chong Shen, PhDc,*, and Yingshui Yao, PhDa,**
a
Department of Epidemiology and Biostatistics, School of Public Health, Wannan Medical College, Wuhu, China;
b
Department of Cardiology, Affiliated Yixing People’s Hospital of Jiangsu University, People’s Hospital of Yixing City, Yixing, China; and
c
Department of Epidemiology, School of Public Health, Nanjing Medical University, Nanjing, China
Manuscript received September 1, 2017 and accepted December 21, 2017

Abstract

Somatostatin (SST) and growth hormone–releasing hormone (GHRH) are involved in the development of hypertension. This study
aimed to evaluate whether SST and GHRH contribute to genetic susceptibility to hypertension. A case-control study consisting of
2012 hypertensive patients and 2210 matched control individuals was performed, and three tagging single-nucleotide polymorphisms
were genotyped. The association of these single-nucleotide polymorphisms with hypertension and ischemic stroke was further eval-
uated among 4098 participants in a follow-up study. Hazard ratio (HR) and 95% confidence interval were estimated by Cox propor-
tional hazards regression. The follow-up study indicated that in smoking population, variants at SST presented significant association
with hypertension incidence; the adjusted HR of rs3755792 (GA þ AA vs. GG) was 0.634 (P ¼ .037), and the adjusted HR of
rs7624906 (TC þ CC vs. TT) was 1.803 (P ¼ .005). In drinking population, rs3755792 at SST was associated with hypertension inci-
dence, and the adjusted HR was 0.580 (P ¼ .009). Moreover, rs6032470 at GHRH had a statistical association with ischemic stroke
incidence in smoking population, and the adjusted HR of the additive model was 1.625 (P ¼ .049). These results suggested that SST
and GHRH harbor genetic susceptible loci with incident hypertension and ischemic stroke and that smoking and drinking might
modify the genetic effect. J Am Soc Hypertens 2018;12(3):230–237. Ó 2017 American Society of Hypertension. All rights reserved.
Keywords: GHRH; hypertension; ischemic stroke; polymorphism; SST.

Hui Zhu, Lijun Zhu, and Zhengmei Fang contributed equally to
the work.
Supplemental Material can be found at www.ashjournal.com.
Conflict of interest: None.
*Corresponding author: Chong Shen, PhD, Department of
Epidemiology, School of Public Health, Nanjing Medical Univer-
sity, 101 Longmian Avenue, Jiangning, Nanjing, 211166 China.
Tel: þ86 25 86868291; Fax: þ86 25 86527613.
**Corresponding author: Yingshui Yao, PhD, Department of
Epidemiology and Biostatistics, School of Public Health, Wannan
Medical College, No. 22, Wen Chang Road, Wuhu, 241002, An-
hui, China. Tel: þ86 553 3932651; Fax: þ86 553 3932637.
E-mails: sc@njmu.edu.cn, yingshuiyao@163.com
1933-1711/$ - see front matter Ó 2017 American Society of Hypertension. All rights reserved.
https://doi.org/10.1016/j.jash.2017.12.009
Introduction
Cardiovascular disease (CVD) has been one of the most
serious diseases threatening human health in the world.1,2 It
has also been the main public health problem contributing
to majority of deaths among Chinese adults.3 Hypertension
is known as the leading risk factor for CVDs such as stroke,
coronary heart disease, and arrhythmia, and these CVDs
exacerbate morbidity and mortality, directly impacting
about 1 billion people worldwide.4–6 In China, the number
of people with hypertension has substantially increased
over the past decades, and the estimated number might in-
crease to 352 million in 2030.7
Hypertension is a complex condition that results from
interaction of environmental exposure and genetic varia-
tions.8 Recent genome-wide association studies identified
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
that multiple genetic loci were associated with hypertension
and the level of blood pressure.8–10
Somatostatin (SST) and growth hormone–releasing hor-
mone (GHRH) are the major mediators of growth hormone
(GH).11 GH increases gap junctions and/or calcium-
activated potassium channel distribution, thereby enhancing
the endothelium-derived hyperpolarizing factor transfer,12
which hyperpolarizes vascular smooth muscle cells and
contributes to vasorelaxation.13 SST is mainly released in
the periventricular nucleus of the hypothalamus14 and reg-
ulates the negative feedback manipulated by GH on its own
secretion.12 GHRH is a hypothalamic peptide released in
the arcuate nucleus of the hypothalamus and presents as
the central regulator of GH synthesis and release.15
Furthermore, SST presents potent effects on vascular in-
flammatory changes by diminishing proinflammatory neu-
ropeptides such as calcitonin gene–related peptide and
substance P, which cause a long-lasting arteriolar vasodila-
tion.16,17 SST also inhibits vasoactive intestinal peptide,
which presents important vasodilatory function and lowers
mean arterial pressure (MAP) effectively.18 Some previous
animal studies revealed that intracerebroventricular admin-
istration of an SST analog (octreotide) raises the plasma
vasopressin concentration and MAP effectively.19–21 Two
prospective cohort studies reported that endogenous SST
precursor (N-terminal prosomatostatin) is the independent
predictor of CVDs, and it is more stable and easy to mea-
sure.22,23 Likewise, neuronostatin, another peptide encoded
by the SST gene, suppresses cardiac contractile function
and increases blood pressure via protein kinase A and
JUN NH2-terminal kinase pathways.24–26
A study in French-Canadian population showed that
rs34872250 polymorphism at SST was strongly related to
interindividual variation in blood pressure, especially
among women and overweight/obese subjects.27 Unfortu-
nately, no data of SST rs34872250 mutant allele were iden-
tified in Han Chinese population in the SNPinfo website
(https://snpinfo.niehs.nih.gov/).
Previous studies have shown limited evidence of the rela-
tionship between SST as well as GHRH and hypertension.
To determine whether polymorphisms at SST and GHRH
are associated with hypertension, we conducted a case-
control study on 2012 hypertensive patients and 2210
healthy control individuals in Han Chinese population
and the subjects were further followed up to evaluate the
genetic effects of SST and GHRH on hypertension and
ischemic stroke incidence.
Methods
Subjects
A case-control study was performed in a rural population
in the Jiangsu province, China.28 A total of 4222 partici-
pants consisting of 2012 hypertensive patients and 2210
231
age- (5 years) and sex-matched control individuals were re-
cruited from May to October 2009. Hypertension was
defined as an average systolic pressure 140 mm Hg
and/or diastolic pressure  90 mm Hg, and/or currently
receiving antihypertensive medication to lower blood
pressure.
In the follow-up study, 94 matched elderly control indi-
viduals and 30 hypertensive patients with a history of stroke
were excluded; thus, 4098 subjects were further followed
up for an average of 5.01 years to observe the incidence
of ischemic stroke, and 2116 normotensive control individ-
uals among those subjects were also prospectively followed
up to observe the incidence of hypertension (Figure 1).
Face-to-face or telephonic interviews were conducted to
ascertain the disease status. Registered disease records
from the local public health department were reviewed
and verified by a study-wide end point assessment commit-
tee at People’s Hospital of Yixing City.
This research was performed in accordance with the
ethics committee of Nanjing Medical University. Written
informed consents were obtained from all subjects during
the epidemiologic interview.
Data Collection
A questionnaire survey was conducted by trained staff to
gather demographic information including age, gender, na-
tionality, marriage, education, medical history, smoking,
and drinking status. Smokers were defined as individuals
who smoked 20 cigarettes/wk lasting for at least 3 mon/
y. Drinkers were defined as those with current or past
alcohol consumption of 2 times/wk lasting for at least
6 mon/y. Furthermore, physical examination to measure
weight (kg), height (cm), and blood pressure was also per-
formed. The blood pressure was measured three times in
the sitting position according to the standard procedures
and recommendations.29
Peripheral venous blood samples were collected after
overnight fasting (>10 hours), and the glucose (GLU),
serum total cholesterol (TCH), triglyceride (TG), low-
density lipoprotein cholesterol (LDL-C), and high-density
lipoprotein cholesterol (HDL-C) levels were measured
enzymatically on a Siemens automatic biochemistry
analyzer ADVIA 1200 (Siemens Healthcare Diagnostics
Inc., Beijing, MN).
SNP Selection
The SST gene is located on chromosome 3q27.3 (gene
ID:6750; NC_000003.12), spans 1.5 kbps, and consists of
2 exons. The GHRH gene is mapped to chromosome
20q11.23 (gene ID:2691; NC_000020.11), spans
10.7 kbps, and contains 5 exons. Beijing Han population
China gene database was downloaded from the National
Human Genome Research Institute 1000 Genome Project
232 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237

Figure 1. Flow chart for selection of participants. Inclusion of participants for evaluating the association between SST, GHRH and hy-
pertension in the case-control study, and selection of participants for the further follow-up study. Matched elderly controls (n ¼ 94) and
30 hypertension patients with a history of stroke were excluded in the further follow-up study. HT, hypertension; IS, ischemic stroke.

database (GRCh37, http://phase3browser.1000genomes. using SDS 2.32 Allelic Discrimination Software

org/index.html). Haploview software (version 4.2) was
used to screen the tagging single-nucleotide polymor-
phisms (tagSNPs), with r2  0.8 and minor allele
frequency  0.05. SNPs’ function was assessed and pre-
dicted using the SNPinfo website (snpinfo, http://snpinfo.
niehs.nih.gov/), and tagSNPs with potential function were
therefore preferred for inclusion (Supplementary Table 1).
Finally, rs6032470 (T>C) at GHRH and rs7624906
(T>C) and rs3755792 (G>A) at SST were selected as
tagSNPs to investigate the genetic association with hyper-
tension and ischemic stroke.
Genotyping
The DNA was isolated from peripheral blood leuko-
cytes by a standard phenol-chloroform method.30 Geno-
typing was conducted by using TaqMan-based allelic
discrimination assay on the platform of ABI 9700 real-
time polymerase chain reaction system. The probes,
primers, and TaqMan master mix for assays were
purchased from Nanjing BioSteed Biotechnologies Co.,
Ltd. Reaction conditions were as follows: 50 C for 2
minutes and 95 C for 10 minutes, followed by 45 cycles
at 95 C for 15 seconds and 60 C for 1 minute. The total
reaction volume was 5 mL, including 1 mL DNA
template. The assay results were displayed on the ABI
7900 fluorescence quantitative polymerase chain reaction
instrument, and the experimental data were obtained
(Applied Biosystems).
Statistical Analysis
Quantitative variables with normal distribution are pre-
sented as mean  standard deviation, and the differences be-
tween patients and control subjects were evaluated with the
unpaired Student t test. Qualitative variables are presented as
absolute frequencies (n) and relative frequencies (%). Qualita-
tive variables and the allele and genotype frequency distribu-
tions of patients and control subjects were compared using the
chi-square (c2) test. Hardy-Weinberg equilibrium was esti-
mated with a Fisher exact test in the control group. General
linear model was applied to compare blood pressure levels
(mean  standard deviation) between genotypes among pa-
tients (divided into treated and untreated groups) and controls,
after adjustment for age, gender, TCH, TG, LDL-C, HDL-C,
body mass index (BMI), GLU, smoking, and drinking status.
Additive, dominant, and recessive models were tested for
adaptive genetic effect using multiple logistic regression anal-
ysis and Cox proportional hazards regression analysis. The
best-fitting genetic model was defined as the one with the
smallest P value in the analysis and that was further evaluated
by Bonferroni correction. Multiple logistic regression analysis
was conducted to evaluate the association between the
tagSNPs and hypertension with odds ratio and 95% confidence
interval (95% CI) in the case-control study and adjusted for
age, gender, TCH, TG, LDL-C, HDL-C, BMI, GLU, smoking,
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237 233

and drinking status. Cox proportional hazards regression anal-
ysis was performed to estimate the risk of SSTand GHRH var-
iants for ischemic stroke incidence with hazard ratios (HR)
and 95% CI, adjusted for covariates in the follow-up study.
Proportional hazards assumption over time was assessed visu-
ally using the Kaplan-Meier survival plots and log-minus-log
plots. A two-tailed P < .05 was defined to be statistically sig-
nificant. Bonferroni correction was applied to adjust P value
by multiplying 3 times (the number of SNPs tested). All of
the statistical analysis was performed with Statistical Product
and Service Solutions 18.0 (SPSS; SPSS Inc, Chicago, IL).
Results
Demographic and Clinical Characteristics of
Study Population
The demographic and clinical characteristics of the par-
ticipants in the case-control study are described as previ-
ously reported (Table 1).28 The hypertensive patients were
slightly older than control subjects by 3.42 years
(P < .05), although the control subjects were matched by
age (5 years). No significant differences in sex, HDL-C
level, smoking, and drinking status were found between
the two groups. The hypertensive patients presented a
higher systolic pressure, diastolic pressure, BMI, TCH,
TG, LDL-C, and GLU levels than control subjects.
Demographic and clinical characteristics of follow-up
population at baseline are shown in Supplementary
Table 2. During an average follow-up of 5.01 years, a total
of 613 (29.0%) and 171 (4.2%) subjects developed hyper-
tension and ischemic stroke, respectively, with an incidence
density of 65.68 and 8.05 per 1000 person-years. Among
613 subjects with incident hypertension, self-report of
taking an antihypertensive medication for the treatment of
hypertension accounted for 162 (26.4%), and their disease
records were further reviewed and verified. The other hy-
pertensive patients (n ¼ 451) were diagnosed by standard
blood pressure measurements.
Association Analyses of Case-Control
Population
The genotypes of all three SNPs in the control subjects
were in Hardy-Weinberg equilibrium (P > .05). There was
no significant association between the three SNPs and hyper-
tension (Supplementary Table 3), even after adjustment for
age, gender, TCH, TG, LDL-C, HDL-C, BMI, GLU, smok-
ing, and drinking status (Supplementary Table 4). Mean-
while, further stratification analyses by age, gender,
smoking, and drinking showed that none of the three SNPs
was correlated with hypertension even after adjustment for
covariates (Supplementary Tables 7 and 8). Haplotype anal-
ysis also did not show significant haplotype of rs3755792
and rs7624906 for hypertension (Supplementary Table 5).
Quantitative trait analysis with general linear model sug-
gested that no statistical difference in blood pressure was
observed among the different genotypes in hypertensive pa-
tients (including the treated and untreated group) and con-
trol subjects, even after adjustment for age, gender, TCH,
TG, LDL-C, HDL-C, BMI, GLU, smoking, and drinking
status (Supplementary Table 6).
Follow-up Analysis
No major violations of the proportional hazards assump-
tion were found (Supplementary Figures 1–6). Cox propor-
tional hazards regression analysis showed that all three

Table 1
Demographic and clinical characteristics of the participants in the case-control study
Characteristics Group Control (n ¼ 2210) Case (n ¼ 2012) t/c2 P
Gender Male 884 (40.0%) 829 (41.2%) 0.63 .427
Female 1326 (60.0%) 1183 (58.8%)
Age (y) 58.93  10.45 62.35  10.73 10.48 <.001
GLU (mmol/L) 5.46  1.61 5.83  2.05 6.68 <.001
TCH (mmol/L) 4.79  1.01 4.93  1.05 4.57 <.001
TG (mmol/L) 1.54  1.21 1.86  1.58 7.62 <.001
HDL-C (mmol/L) 1.36  0.33 1.37  0.33 0.18 .861
LDLC (mmol/L) 2.65  0.73 2.80  0.89 6.23 <.001
BMI (kg/m2) 23.64  3.2 24.76  3.51 10.84 <.001
Blood pressure (mm Hg) SBP 124.24  11.36 142.86  14.3 47.02 <.001
DBP 79.08  6.51 87.53  8.54 36.37 <.001
Smoking Yes 533 (24.1%) 480 (23.9%) 0.04 .843
No 1677 (75.9%) 1532 (76.1%)
Drinking Yes 476 (21.5%) 423 (21.0%) 0.17 .683
No 1734 (88.5%) 1589 (79.0%)
BMI, body mass index; DBP, diastolic blood pressure; GLU, glucose; HDL-C, high-density lipoprotein cholesterol; LDL-C, low-density
lipoprotein cholesterol; SBP, systolic blood pressure; TCH, total cholesterol; TG, triglyceride.
234 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237

SNPs were not associated with hypertension and ischemic Discussion

stroke incidence in the follow-up population, even after
adjustment for age, gender, TCH, TG, LDL-C, HDL-C,
BMI, hypertension, type 2 diabetes mellitus (T2DM), smok-
ing, and drinking status (Supplementary Tables 9 and 10).
Further stratification analyses by age, gender, smoking,
and drinking status showed that rs3755792 variation at SST
was significantly associated with a decreased risk of hyper-
tension in drinking population, and the adjusted HR of the ad-
ditive model was 0.580 (P ¼ .009) after adjustment for
covariates. The association remained statistically significant
even after Bonferroni correction (corrected P ¼ .027). In
smoking population, rs3755792 (GA þ AA vs. GG) and
rs7624906 (TC þ CC vs. TT) variations at SST displayed sig-
nificant associations with incident hypertension; the adjusted
HRs (95% CI) were 0.634 (0.413–0.973) and 1.803 (1.198–
2.712), with P values of .037 and .005, respectively
(Table 2). After Bonferroni correction, the association of
rs7624906 with hypertension in the smoking group was still
significant (corrected P ¼ .015).
Furthermore, the smoking population with rs6032470C
allele presented an increased risk of ischemic stroke inci-
dence, and the adjusted HR (95% CI) of the additive model
was 1.625 (1.002–2.635) with P ¼ .049 (Table 3), after
adjustment for age, gender, TCH, TG, LDL-C, HDL-C,
BMI, drinking, T2DM, and hypertension. However, the as-
sociation was not significant after Bonferroni correction
(corrected P ¼ .147).
In French-Canadian population, rs34872250 polymor-
phism at SST significantly increased the risk of hypertension
incidence, especially among women and overweight/obese
subjects.24 However, no valid data of rs34872250 were prob-
able for linkage disequilibrium analysis with rs3755792 in
Caucasian and Chinese population. In this study, we observed
that rs3755792 at SST was significantly associated with a
decreased risk of hypertension incidence in drinking and
smoking individuals, and rs7624906 at SST was associated
with the increased risk of hypertension incidence in smoking
individuals. The results indicated that the association of SST
and hypertension varied based on ethnicity.
Previous studies showed that long-term exposure to ciga-
rette smoking could modulate SST expression.31–33 Cigarette
smoking exerted anti-inflammatory effect in ulcerative coli-
tis by upregulating SST gene expression in interleukin-10–
deficient mice.32 Long-term nicotine exposure could in-
crease the levels of somatostatin-28 significantly,33 and the
elevated levels of somatostatin-28 in the central nervous sys-
tem could result in a rise in MAP effectively.19 Interestingly,
a cohort study observed that circulating concentration of
endogenous SST precursor (NT-proSST), a fragment of the
somatostatin precursor, was positively associated with smok-
ing status, use of antihypertensive medication, and higher
TCH level.22 On the other hand, ethanol ingestion may
modify endocrine function by influencing the expression of

Table 2
Stratification analysis of SNPs with incident hypertension in the follow-up study
SNP Stratum Genotype n Person-years Incidence Density HR (95% CI)
(/100 Person-years)
Additive Dominant Recessive
rs3755792 Smoking GG 133 1757.02 7.57 0.684 (0.456–1.027) 0.634 (0.413–0.973) 1.802 (0.431–7.539)
GA 24 447.69 5.36 P ¼ .067 P ¼ .037 P ¼ .420
AA 2 27.41 7.3
No smoking GG 344 5142.4 6.69 0.992 (0.821–1.198) 0.978 (0.789–1.213) 1.105 (0.606–2.015)
GA 99 1484.43 6.67 P ¼ .935 P ¼ .839 P ¼ .745
AA 11 126.89 8.67
Drinking GG 128 1507.02 8.49 0.580 (0.385–0.874) 0.558 (0.365–0.854) 0.753 (0.103–5.496)
GA 26 451.32 5.76 P ¼ .009 P ¼ .007 P ¼ .780
AA 1 25.19 3.97
No drinking GG 349 5392.4 6.47 1.019 (0.844–1.231) 0.999 (0.804–1.240) 1.225 (0.689–2.181)
GA 97 1480.8 6.55 P ¼ .844 P ¼ .990 P ¼ .489
AA 12 129.11 9.29
rs7624906 Smoking TT 129 1929.05 6.69 1.660 (1.125–2.449) 1.803 (1.198–2.712) –
TC 30 289.99 10.35 P ¼ .011 P ¼ .005 –
CC – 13.08 –
No smoking TT 404 5954.41 6.78 0.917 (0.681–1.234) 0.917 (0.681–1.234) –
TC 50 797.97 6.27 P ¼ .566 P ¼ .566 –
CC – – –
CI, confidence interval; HR, hazard ratio; SNP, single-nucleotide polymorphism.
Smoking stratification was adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, drinking status, and T2DM.
Drinking stratification was adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, smoking status, and T2DM.
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
Table 3
Stratification analyses of rs6032470 with incident ischemic stroke in the follow-up study
Stratum Genotype n Person-year Incidence Density
(/1000 Person-year)
Smoking TT 23 3408.88 6.75
TC 15 1605.48 9.34
CC 5 217.48 22.99
No smoking TT 75 9665.38 7.76
TC 47 5578.9 8.43
CC 6 726.46 8.26
CI, confidence interval; HR, hazard ratio.
Smoking stratification was adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, drinking, T2DM, and hypertension.
neurotransmitter receptors.34–36 Mice with long-term ethanol
consumption presented a downregulated expression of
SSTR4 receptor,34 and the decreased receptors may stimulate
SST expression.35 Furthermore, a test for heterogeneity
showed that the genotype distribution of rs3755792 and
rs7624906 presented moderate-to-high heterogeneity in
smoking and drinking subgroups. These results suggested
that cigarette smoking and alcohol drinking may modify
the genetic effect of SST on hypertension by influencing the
SST gene expression.
In this study, we observed that the smoking population
with CC genotype of rs6032470 presented an increased
risk of ischemic stroke, although statistical significance
did not remain after Bonferroni correction (P ¼ .147).
Previous studies suggested that GHRH could increase
the expression of inducible nitric oxide synthase and
neuronal nitric oxide synthase.37,38 It had been proved
that inducible nitric oxide synthase and neuronal nitric ox-
ide synthase expressed in endothelial cells and infiltrating
leukocytes exacerbated ischemic brain injury.39,40 This
was the first study to observe the association of GHRH
rs6032470 polymorphisms with ischemic stroke in Han
Chinese population, and further functional study is
warranted.
Several limitations of our study should be noted. First,
owing to a lack of adequate selection of normotensive
subjects as controls, hypertensive patients were slightly
older than control subjects, although controls were
matched to cases according to 5-year age strata. Thus,
age was included as a covariate in the subsequent ana-
lyses, and age-stratified analyses were also conducted.
Second, we did not detect the plasma SST and GHRH
or conduct further functional research tests to prove the
role of SST and GHRH genetic variations in vascular
endothelial function and blood pressure. Third, potential
bias including recall bias of smoking and drinking habits
might exist, but the significant association would not be
distorted in a subgroup population. As we defined the
235
HR (95% CI)
Additive Dominant Recessive
1.625 (1.002–2.635) 1.775 (0.939–3.354) 2.185 (0.734–6.504)
P ¼ .049 P ¼ .077 P ¼ .160
0.887 (0.657–1.199) 0.968 (0.674–1.389) 0.442 (0.164–1.196)
P ¼ .436 P ¼ .86 P ¼ .108
smoking and drinking status according to the current
and/or the past habits, the estimated association might
be weakened but not distorted by prevalence versus inci-
dence differences. Fourth, in the cohort study, loss to
follow-up might weaken the efficacy, particularly in the
subgroups categorized by smoking and drinking status.
Regardless of these limitations, this is the first study to
report the association of SST and GHRH polymorphisms
with hypertension and ischemic stroke in Han Chinese
population. Furthermore, we adjusted for potential con-
founders including age, gender, TCH, TG, LDL-C,
HDL-C, BMI, GLU, T2DM, smoking, and drinking
status to reduce the false-positive rate and evaluated
the association by Bonferroni correction. The findings
of our study provide a new insight into susceptibility
gene (GHRH and SST) to hypertension and ischemic
stroke.
In conclusion, the findings of this study indicated that
rs3755792 and rs7624906 at SST were associated with hy-
pertension incidence, and rs6032470 at GHRH contributed
to the genetic susceptibility to ischemic stroke. Smoking
and drinking might modify the genetic effect of SST and
GHRH on hypertension and ischemic stroke. How SST
and GHRH promote the genetic susceptibility to the
CVDs merits further investigation.
Acknowledgment
This work was supported by the National Natural Sci-
ence Foundation of China (grant nos. 81541071,
81573232, and 81673266), Anhui Provincial Natural Sci-
ence Foundation (1308085MH135), the Priority Academic
Program for the Development of Jiangsu Higher Education
Institutions (Public Health and Preventive Medicine), and
the Flagship Major Development of Jiangsu Higher Educa-
tion Institutions. The funders had no role in the study
design, data collection and analysis, decision to publish,
or preparation of the manuscript.
236 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
Supplementary data
Supplementary data related to this article can be found at
https://doi.org/10.1016/j.jash.2017.12.009.
References
1. Wu L, He Y, Jiang B, Sun D, Wang J, Liu M, et al.
Trends in prevalence, awareness, treatment and control
of hypertension during 2001-2010 in an Urban elderly
population of China. PLoS One 2015;10:e0132814.
2. Dregan A, Ravindrarajah R, Hazra N, Hamada S,
Jackson SH, Gulliford MC. Longitudinal trends in hy-
pertension management and mortality among octoge-
narians: prospective cohort study. Hypertension 2016;
68:97–105.
3. Guo J, Zhu YC, Chen YP, Hu Y, Tang XW, Zhang B.
The dynamics of hypertension prevalence, awareness,
treatment, control and associated factors in Chinese
adults: results from CHNS 1991-2011. J Hypertens
2015;33:1688–96.
4. Yoon SS, Gu Q, Nwankwo T, Wright JD, Hong Y,
Burt V. Trends in blood pressure among adults with hy-
pertension: United States, 2003 to 2012. Hypertension
2015;65:54–61.
5. Mills KT, Bundy JD, Kelly TN, Reed JE, Kearney PM,
Reynolds K, et al. Global disparities of hypertension
prevalence and control: a systematic analysis of
population-based studies from 90 countries. Circulation
2016;134:441–50.
6. Padwal RS, Bienek A, McAlister FA, Campbell NR.
Epidemiology of hypertension in Canada: an update.
Can J Cardiol 2016;32:687–94.
7. Li D, Lv J, Liu F, Liu P, Yang X, Feng Y, et al. Hyper-
tension burden and control in mainland China: analysis
of nationwide data 2003-2012. Int J Cardiol 2015;184:
637–44.
8. Lu X, Wang L, Lin X, Huang J, Charles Gu C, He M,
et al. Genome-wide association study in Chinese iden-
tifies novel loci for blood pressure and hypertension.
Hum Mol Genet 2015;24:865–74.
9. Levy D, Ehret GB, Rice K, Verwoert GC, Launer LJ,
Dehghan A, et al. Genome-wide association study of
blood pressure and hypertension. Nat Genet 2009;41:
677–87.
10. Adeyemo A, Gerry N, Chen G, Herbert A,
Doumatey A, Huang H, et al. A genome-wide associa-
tion study of hypertension and blood pressure in Afri-
can Americans. PLoS Genet 2009;5:e1000564.
11. Pivonello C, De Martino MC, Negri M, Cuomo G,
Cariati F, Izzo F, et al. The GH-IGF-SST system in he-
patocellular carcinoma: biological and molecular path-
ogenetic mechanisms and therapeutic targets. Infect
Agent Cancer 2014;9:27.
12. Gray C, Li M, Reynolds CM, Vickers MH. Pre-wean-
ing growth hormone treatment reverses hypertension
and endothelial dysfunction in adult male offspring of
mothers undernourished during pregnancy. PLoS One
2013;8:e53505.
13. Ozkor MA, Quyyumi AA. Endothelium-derived hyper-
polarizing factor and vascular function. Cardiol Res
Pract 2011;2011:156146.
14. Patel YC. Somatostatin and its receptor family. Front
Neuroendocrinol 1999;20:157–98.
15. Qian Y, Yan A, Lin H, Li W. Molecular characteriza-
tion of the GHRH/GHRH-R and its effect on GH syn-
thesis and release in orange-spotted grouper
(Epinephelus coioides). Comp Biochem Physiol B Bio-
chem Mol Biol 2012;163:229–37.
16. Pinter E, Helyes Z, Szolcsanyi J. Inhibitory effect of
somatostatin on inflammation and nociception. Phar-
macol Ther 2006;112:440–56.
17. Gazelius B, Brodin E, Olgart L, Panopoulos P. Evi-
dence that substance P is a mediator of antidromic
vasodilatation using somatostatin as a release inhibitor.
Acta Physiol Scand 1981;113:155–9.
18. Henning RJ, Sawmiller DR. Vasoactive intestinal pep-
tide: cardiovascular effects. Cardiovasc Res 2001;49:
27–37.
19. Brown MR. Somatostatin-28 effects on central nervous
system regulation of vasopressin secretion and blood
pressure. Neuroendocrinology 1988;47:556–62.
20. Rettig R, Geist R, Sauer U, Rohmeiss P, Unger T. Cen-
tral effects of somatostatin: pressor response, AVP
release, and sympathoinhibition. Am J Physiol 1989;
257(3 Pt 2):R588-94.
21. Gardi J, Szentirmai E, Hajdu I, Obal F Jr, Krueger JM.
The somatostatin analog, octreotide, causes accumula-
tion of growth hormone-releasing hormone and deple-
tion of angiotensin in the rat hypothalamus. Neurosci
Lett 2001;315:37–40.
22. Abbasi A, Kieneker LM, Corpeleijn E, Gansevoort RT,
Gans RO, Struck J, et al. Plasma N-terminal Prosoma-
tostatin and risk of incident cardiovascular disease and
all-cause mortality in a prospective observational
cohort: the PREVEND study. Clin Chem 2017;63:
278–87.
23. Hedback T, Almgren P, Nilsson PM, Melander O.
N-Terminal prosomatostatin as a risk marker for car-
diovascular disease and diabetes in a general popula-
tion. J Clin Endocrinol Metab 2016;101:3437–44.
24. Samson WK, Zhang JV, Avsian-Kretchmer O, Cui K,
Yosten GL, Klein C, et al. Neuronostatin encoded by
the somatostatin gene regulates neuronal, cardiovascu-
lar, and metabolic functions. J Biol Chem 2008;283:
31949–59.
25. Vainio L, Perjes A, Ryti N, Magga J, Alakoski T,
Serpi R, et al. Neuronostatin, a novel peptide encoded
by somatostatin gene, regulates cardiac contractile
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
function and cardiomyocyte survival. J Biol Chem
2012;287:4572–80.
26. Hua Y, Ma H, Samson WK, Ren J. Neuronostatin in-
hibits cardiac contractile function via a protein kinase
A- and JNK-dependent mechanism in murine hearts.
Am J Physiol Regul Integr Comp Physiol 2009;297:
R682-9.
27. Tremblay M, Brisson D, Gaudet D. Association be-
tween a polymorphic poly-T repeat sequence in the
promoter of the somatostatin gene and hypertension.
Hypertens Res 2016;39:467–74.
28. Yang S, Chen Y, Liu C, Zhao X, Zhuang Q, Sun J, et al.
Association study of Common variants in PFN1 with
hypertension in a Han Chinese population: a case-
control Study and A Follow-up study. Am J Hypertens
2017;30:1024–31.
29. Perloff D, Grim C, Flack J, Frohlich ED, Hill M,
McDonald M, et al. Human blood pressure determina-
tion by sphygmomanometry. Circulation 1993;88(5 Pt
1):2460–70.
30. Sambrook J, Russell DW. Isolation of high-molecular-
weight DNA from Mammalian cells using Proteinase K
and Phenol. CSH Protoc 2006;2006(1). https:
//doi.org/10.1101/pdb.prot4036.
31. Yang J, Liu AY, Tang B, Luo D, Lai YJ, Zhu BL, et al.
Chronic nicotine differentially affects murine transcrip-
tome profiling in isolated cortical interneurons and py-
ramidal neurons. BMC Genomics 2017;18:194.
32. Eliakim R, Fan QX, Babyatsky MW. Chronic nicotine
administration differentially alters jejunal and colonic
inflammation in interleukin-10 deficient mice. Eur J
Gastroenterol Hepatol 2002;14:607–14.
237
33. Petruzziello F, Falasca S, Andren PE, Rainer G,
Zhang X. Chronic nicotine treatment impacts the regu-
lation of opioid and non-opioid peptides in the rat dor-
sal striatum. Mol Cell Proteomics 2013;12:1553–62.
34. Jonsson S, Ericson M, Soderpalm B. Modest long-term
ethanol consumption affects expression of neurotrans-
mitter receptor genes in the rat nucleus accumbens.
Alcohol Clin Exp Res 2014;38:722–9.
35. Barrios V, Puebla-Jimenez L, del Carmen Boyano-
Adanez M, Sanz M, Soriano-Guillen L, Arilla-
Ferreiro E. Differential effects of ethanol ingestion on
somatostatin content, somatostatin receptors and ad-
enylyl cyclase activity in the frontoparietal cortex of
virgin and parturient rats. Life Sci 2005;77:1094–105.
36. Rage F, Arancibia S, Tapia-Arancibia L. Effect of
acute, but not chronic ethanol treatment on somato-
statin secretion in rat hypothalamic neurons. Neurosci
Lett 1998;245:175–9.
37. Barabutis N, Siejka A, Schally AV. Growth hormone
releasing hormone induces the expression of nitric ox-
ide synthase. J Cell Mol Med 2011;15:1148–55.
38. Tsumori M, Murakami Y, Koshimura K, Kato Y. Growth
hormone-releasing hormone and gonadotropin-releasing
hormone stimulate nitric oxide production in 17beta-
estradiol-primed rat anterior pituitary cells. Endocrine
2002;17:215–8.
39. Terpolilli NA, Moskowitz MA, Plesnila N. Nitric ox-
ide: considerations for the treatment of ischemic stroke.
J Cereb Blood Flow Metab 2012;32:1332–46.
40. Zhu J, Song W, Li L, Fan X. Endothelial nitric oxide
synthase: a potential therapeutic target for cerebrovas-
cular diseases. Mol Brain 2016;9:30.
237.e1 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237

Table S1
The biological information and functional prediction of three tagSNPs of SST and GHRH genes
No SNP Chromosome Position Allele TFBS RegPotential Conservation Nearby Gene Distance (bp)
1 rs6032470 20 35320941 C/T Y 0 0 GHRHjjMANBAL 2235jj30524
2 rs3755792 3 188872655 A/G Y NA 0 SSTjjRTP2 1760jj26086
3 rs7624906 3 188872330 C/T Y NA 0.019 SSTjjRTP2 1435jj26411
TFBS, transcription factor binding site; RegPotential, regulatory potential score; MANBAL, mannosidase beta like; RTP2, receptor
transporter protein 2.
Table S2
Characteristics of follow-up population at baseline
Characteristics Group Incident No Hypertension t/c2 P Incident Ischemic Stroke No Stroke (n ¼ 3915) t/c2 P
Hypertension (n ¼ 1503) (n ¼ 171)
(n ¼ 613)
Gender Male 266 (43.4%) 587 (39.1%) 3.405 .071 84 (49.1%) 1568 (40.1%) 7.969 .005
Female 347 (56.6%) 916 (60.9%) 87 (50.9%) 2347 (59.9%)
Age (y) 60.72  9.56 57.47  10.42 6.676 3.12  1011 71.81  9.76 59.78  10.42 14.839 2.07  1048

H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
Glu (mmol/L) 5.55  1.83 5.45  1.54 1.252 .211 5.84  2.25 5.65  1.84 1.351 .177
TCH (mmol/L) 4.86  0.97 4.76  1.03 2.221 .026 5.04  1.1 4.85  1.03 2.343 .019
TG (mmol/L) 1.59  1.11 1.52  1.26 1.15 .25 1.95  1.67 1.69  1.40 2.411 .016
HDL-C (mmol/L) 1.37  0.33 1.38  0.33 0.629 .53 1.37  0.35 1.37  0.33 0.112 .911
LDL-C (mmol/L) 2.68  0.73 2.64  0.73 0.945 .345 2.87  1.04 2.72  0.80 2.352 .019
BMI (kg/m2) 24.12  3.29 23.53  3.16 3.826 1.34  104 24.47  3.77 24.2  3.36 1.025 .305
Blood pressure (mmHg) SBP 126.72  9.73 123.36  11.89 6.189 7.25  1010 140.52  17.06 133.04  15.77 6.049 1.59  109
DBP 80.29  5.46 78.76  6.83 4.934 8.67  107 85.37  9.28 83.15  8.60 3.288 .001
Smoking Yes 159 (25.9%) 366 (24.3%) 0.588 .471 43 (25.1%) 949 (24.2%) 0.073 .785
No 454 (74.1%) 1137 (75.7%) 128 (74.9%) 2966 (75.8%)
Drinking Yes 155 (25.3%) 313 (20.8%) 5.029 .028 37 (21.6%) 843 (21.5%) 0.001 .999
No 458 (74.7%) 1190 (79.2%) 134 (78.4%) 3072 (78.5%)
Person-year 9333.13 21,231.21
Incidence density (/103 65.68 8.05
person-years)
BMI, body mass index; DBP, diastolic blood pressure; GLU, glucose; HDL-C, high-density lipoprotein cholesterol; LDL-C, low-density lipoprotein cholesterol; SBP, systolic
blood pressure; TCH, total cholesterol; TG, triglyceride.

237.e2
Table S3

237.e3
Association analysis of three SNPs at GHRH and SST genes with hypertension
Gene SNP Group WT/HT/MT Crude OR (95% CI) Allele Gene Py
Additive Dominant Recessive Major/Minor OR (95% CI)/P*
GHRH rs6032470 TT/TC/CC T/C
Case 1248/681/82 0.954 (0.86–1.059) 0.972 (0.859–1.101) 0.808 (0.603–1.083) 3177/845 0.954 (0.859–1.059) .502
Control 1352/740/110 P ¼ .378 P ¼ .660 P ¼ .154 3444/960 P ¼ .378
GG/GA/AA G/A

H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
SST rs3755792 Case 1545/434/32 1.004 (0.882–1.142) 1.007 (0.872–1.162) 0.974 (0.603–1.574) 3524/498 1.004 (0.882–1.143) .632
Control 1696/472/36 P ¼ .955 P ¼ .924 P ¼ .914 3864/544 P ¼ .955
TT/TC/CC T/C
rs7624906 Case 1763/238/8 1.018 (0.853–1.215) 1.012 (0.841–1.218) 1.255 (0.454–3.467) 3764/254 1.0180 (0.854–1.214) .580
Control 1937/260/7 P ¼ .841 P ¼ .897 P ¼ .662 4134/274 P ¼ .842
CI, confidence interval; HT, heterozygote; MT, minor homozygote; OR, odds ratio; SNP, single-nucleotide polymorphism; WT, major homozygote.
Additive model: WT vs. HT vs. MT, dominant model: WT vs. HT and MT, and recessive model: WT and HT vs. MT.
* P value of c2 test for comparison of allele frequency between the cases and controls.
y
Hardy-Weinberg equilibrium test in controls.
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237 237.e4

Table S4
Association analysis of three SNPs at GHRH and SST genes with hypertension
Gene SNP Group WT/HT/MT Genotype OR (95% CI)*
Additive Dominant Recessive
GHRH rs6032470 TT/TC/CC
Case 1248/681/82 0.970 (0.870–1.080) 0.990 (0.870–1.130) 0.810 (0.600–1.110)
Control 1352/740/110 P ¼ .549 P ¼ .881 P ¼ .188
GG/GA/AA
SST rs3755792 Case 1545/434/32 1.030 (0.900–1.180) 1.040 (0.890–1.200) 1.010 (0.610–1.670)
Control 1696/472/36 P ¼ .676 P ¼ .651 P ¼ .974
TT/TC/CC
rs7624906 Case 1763/238/8 1.030 (0.860–1.240) 1.020 (0.840–1.240) 1.440 (0.500–4.190)
Control 1937/260/7 P ¼ .757 P ¼ .842 P ¼ .498
CI, confidence interval; HT, heterozygote; MT, minor homozygote; OR, odds ratio; SNP, single-nucleotide polymorphism; WT, major
homozygote.
* Adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, GLU, smoking, and drinking status.

Table S5
Haplotype frequencies of rs3755792(G/A) and rs7624906(C/T) and association with hypertension
Gene Haplotype Genotype Case Frequency Control Frequency OR (95% CI)* P
SST rs3755792-rs7624906 G-T 0.8133 0.8143 Reference –
G-C 0.0632 0.0622 1.018 (0.841–1.195) .8454
A-T 0.1234 0.1235 1.001 (0.870–1.132) .9875
A-C 0 0 – –
CI, confidence interval; OR, odds ratio.
* Adjusted for age, gender, TCH, TG, HDL-C, LDL-C, GLU, BMI, smoking, and drinking status.
Table S6

237.e5
Quantitative trait analysis for blood pressure (mm Hg) between genotypes of the SST and GHRH genes
SNPs Genotype SBP DBP
Case Control Case Control
Treated Untreated Treated Untreated
rs6032470 TT 142.26  16.39 (632) 143.26  11.42 (616) 124.51  11.37 (1352) 87.225  9.22 (632) 87.84  7.51 (616) 79.15  6.49 (1352)
TC 143.54  16.70 (365) 142.55  11.36 (316) 123.96  11.27 (740) 87.800  9.17 (365) 87.07  8.14 (316) 78.94  6.57 (740)

H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237
CC 142.34  17.05 (40) 143.29  13.48 (42) 122.91  11.98 (110) 88.613  9.18 (40) 87.95  9.03 (42) 79.33  6.35 (110)
F 1.184 0.300 0.662 1.505 0.918 0.464
P* .306 .741 .516 .223 .400 .629
rs3755792 GG 142.82  16.80 (800) 143.23  11.65 (745) 124.06  11.45 (1696) 87.488  9.17 (800) 87.76  7.80 (745) 79.11  6.55 (1696)
GA 142.36  15.30 (221) 142.17  11.02 (213) 124.93  11.04 (472) 87.611  8.97 (221) 86.89  7.77 (213) 79.09  6.47 (472)
AA 141.94  19.55 (16) 145.25  9.66 (16) 124.36  12.05 (36) 85.313  13.12 (16) 89.47  7.50 (16) 77.9  5.14 (36)
F 0.001 0.738 1.150 0.408 0.956 0.261
P* .999 .478 .317 .665 .385 .771
rs7624906 TT 142.58  16.63 (910) 142.98  11.56 (853) 124.16  11.38 (1937) 87.443  9.30 (910) 87.58  7.76 (853) 79.02  6.48 (1937)
TC 143.42  15.83 (124) 143.42  11.24 (114) 125.02  11.29 (260) 87.702  8.48 (124) 87.84  8.06 (853) 79.56  6.72 (260)
CC 149.00  12.73 (2) 144.33  5.92 (6) 122.14  11.80 (7) 85  0.00 (2) 86.67  8.02 (6) 78.79  7.07 (7)
F 0.303 0.172 0.697 0.125 0.262 0.568
P* .738 .842 .498 .882 .77 .567
DBP, diastolic blood pressure; SBP, systolic blood pressure; SNP, single-nucleotide polymorphism.
* Adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, GLU, smoking, and drinking status.
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237 237.e6

Table S7
Stratification analysis by age, gender, smoking, and drinking status for the association between the three SNPs and hypertension
SNP Stratum Group WT/HT/MT Crude OR (95% CI)
Additive Dominant Recessive
rs6032470 TT/TC/CC
<55 y Case 321/191/21 1.024 (0.849–1.235) 1.061 (0.850–1.324) 0.858 (0.499–1.475)
Control 527/289/39 P ¼ .804 P ¼ .600 P ¼ .580
55 y Case 927/490/61 0.926 (0.815–1.051) 0.939 (0.807–1.094) 0.774 (0.545–1.098)
Control 825/451/71 P ¼ .232 P ¼ .421 P ¼ .151
Male Case 539/254/59 0.939 (0.795–1.110) 0.914 (0.750–1.114) 1.008 (0.629–1.616)
Control 556/289/59 P ¼ .462 P ¼ .376 P ¼ .974
Female Case 709/427/59 0.966 (0.845–1.105) 1.0160 (0.865–1.192) 0.707 (0.486–1.029)
Control 796/451/59 P ¼ .613 P ¼ .850 P ¼ .070
Smoking Case 313/144/22 0.979 (0.788–1.217) 0.944 (0.729–1.223) 1.169 (0.635–2.154)
Control 340/170/21 P ¼ .851 P ¼ .663 P ¼ .616
No smoking Case 935/537/60 0.946 (0.840–1.066) 0.981 (0.851–1.130) 0.725 (0.518–1.013)
Control 1012/570/89 P ¼ .362 P ¼ .786 P ¼ .059
Drinking Case 282/126/14 0.949 (0.749–1.202) 0.959 (0.727–1.266) 0.912 (0.683–1.219)
Control 313/143/19 P ¼ .664 P ¼ .769 P ¼ .535
No drinking Case 966/555/68 0.954 (0.849–1.072) 0.974 (0.847–1.120) 0.804(0.583–1.109)
Control 1039/597/91 P ¼ .426 P ¼ .711 P ¼ .183
rs3755792 GG/GA/AA
<55 y Case 409/115/9 0.977 (0.776–1.230) 0.963 (0.746–1.242) 1.114 (0.473–2.624)
Control 651/192/13 P ¼ .843 P ¼ .771 P ¼ .805
55 y Case 1136/319/23 1.024 (0.874–1.200) 1.038 (0.871–1.238) 0.911 (0.508–1.631)
Control 1045/280/23 P ¼ .771 P ¼ .676 P ¼ .753
Male Case 632/182/59 1.103 (0.898–1.355) 1.098 (0.876–1.377) 1.360 (0.614–3.014)
Control 687/183/59 P ¼ .349 P ¼ .416 P ¼ .448
Female Case 913/252/59 0.944 (0.799–1.115) 0.950 (0.789–1.144) 0.802 (0.435–1.478)
Control 1009/289/59 P ¼ .498 P ¼ .590 P ¼ .480
Smoking Case 367/104/8 1.136 (0.868–1.487) 1.144 (0.85–1.54) 1.2740 (0.458–3.54)
Control 420/105/7 P ¼ .353 P ¼ .373 P ¼ .642
No smoking Case 1178/330/24 0.967 (0.834–1.121) 0.968 (0.822–1.141) 0.9020 (0.523–1.556)
Control 1276/367/29 P ¼ .654 P ¼ .700 P ¼ .710
Drinking Case 330/88/4 0.912 (0.683–1.219) 0.925 (0.676–1.267) 0.640 (0.186–2.201)
Control 365/103/7 P ¼ .535 P ¼ .627 P ¼ .479
No drinking Case 1215/346/28 1.028 (0.889–1.187) 1.029 (0.876–1.209) 1.0510(0.623–1.775)
Control 1331/369/29 P ¼ .713 P ¼ .724 P ¼ .851
rs7624906 TT/TC/CC
<55 y Case 472/58/3 1.064 (0.767–1.475) 1.023 (0.728–1.439) 4.840 (0.502–46.647)
Control 760/95/1 P ¼ .710 P ¼ .895 P ¼ .173
55 y Case 1291/180/5 0.979 (0.792–1.21) 0.986 (0.790–1.232) 0.760 (0.231–2.497)
Control 1177/165/6 P ¼ .844 P ¼ .904 P ¼ .651
Male Case 719/103/59 0.961 (0.738–1.249) 0.943 (0.714–1.247) 1.334 (0.357–4.983)
Control 760/117/59 P ¼ .764 P ¼ .682 P ¼ .669
Female Case 1044/135/59 1.064 (0.838–1.352) 1.066 (0.832–1.364) 1.12(0.226–5.558)
Control 1177/143/59 P ¼ .611 P ¼ .614 P ¼ .890
Smoking Case 406/70/3 1.059 (0.765–1.465) 1.079 (0.762–1.528) 0.832 (0.185–3.736)
Control 456/72/4 P ¼ .729 P ¼ .669 P ¼ .810
Nosmoking Case 1357/168/5 1.003 (0.812–1.239) 0.989 (0.795–1.230) 1.824 (0.435–7.645)
Control 1481/188/3 P ¼ .979 P ¼ .918 P ¼ .411
Drinking Case 365/54/3 0.978 (0.685–1.394) 0.968 (0.661–1.417) 1.126 (0.226–5.611)
Control 409/63/3 P ¼ .901 P ¼ .866 P ¼ .884
No drinking Case 1398/184/5 1.034 (0.843–1.268) 1.028 (0.832–1.27) 1.363 (0.365–5.085)
Control 1528/197/4 P ¼ .751 P ¼ .800 P ¼ .645
CI, confidence interval; HT, heterozygote; MT, minor homozygote; OR, odds ratio; SNP, single-nucleotide polymorphism; WT, major
homozygote.
237.e7 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237

Table S8
Stratification analysis by age, gender, smoking, and drinking status for the association between the three SNPs and hypertension
SNP Stratum Group WT/HT/MT Adjusted OR (95% CI)
Additive Dominant Recessive
rs6032470 TT/TC/CC
<55 y Case 321/191/21 1.049 (0.863–1.274) 1.089 (0.865–1.370) 0.898 (0.515–1.565)
Control 527/289/39 P ¼ .633 P ¼ .469 P ¼ .704
55 y Case 927/490/61 0.933 (0.818–1.063) 0.945 (0.808–1.105) 0.793 (0.553–1.137)
Control 825/451/71 P ¼ .295 P ¼ .481 P ¼ .206
Male Case 539/254/59 0.988 (0.832–1.174) 0.969 (0.790–1.190) 1.085 (0.666–1.765)
Control 556/289/59 P ¼ .892 P ¼ .765 P ¼ .744
Female Case 709/427/59 0.953 (0.827–1.097) 1.006 (0.850–1.192) 0.666 (0.448–0.991)
Control 796/451/59 P ¼ .502 P ¼ .941 P ¼ .045
Smoking Case 313/144/22 1.039 (0.827–1.305) 1.009 (0.768–1.325) 1.285 (0.678–2.434)
Control 340/170/21 P ¼ .743 P ¼ .949 P ¼ .442
No smoking Case 935/537/60 0.950 (0.839–1.076) 0.989 (0.852–1.147) 0.716 (0.504–1.016)
Control 1012/570/89 P ¼ .421 P ¼ .879 P ¼ .061
Drinking Case 282/126/14 0.989 (0.772–1.267) 1.026 (0.767–1.372) 0.776 (0.373–1.616)
Control 313/143/19 P ¼ .931 P ¼ .865 P ¼ .499
No drinking Case 966/555/68 0.961 (0.851–1.086) 0.982 (0.848–1.136) 0.818 (0.583–1.146)
Control 1039/597/91 P ¼ .527 P ¼ .804 P ¼ .243
rs3755792 GG/GA/AA
<55 y Case 409/115/9 0.975 (0.768–1.238) 0.956 (0.735–1.243) 1.184 (0.482–2.905)
Control 651/192/13 P ¼ .833 P ¼ .736 P ¼ .713
55 y Case 1136/319/23 1.056 (0.897–1.243) 1.078 (0.900–1.292) 0.916 (0.504–1.666)
Control 1045/280/23 P ¼ .511 P ¼ .414 P ¼ .774
Male Case 632/182/59 1.170 (0.945–1.447) 1.166 (0.923–1.474) 1.545 (0.679–3.513)
Control 687/183/59 P ¼ .149 P ¼ .198 P ¼ .300
Female Case 913/252/59 0.939 (0.787–1.120) 0.949 (0.780–1.153) 0.759 (0.394–1.459)
Control 1009/289/59 P ¼ .483 P ¼ .597 P ¼ .408
Smoking Case 367/104/8 1.211 (0.912–1.609) 1.223 (0.894–1.673) 1.460 (0.495–4.300)
Control 420/105/7 P ¼ .186 P ¼ .208 P ¼ .493
No smoking Case 1178/330/24 0.980 (0.84–1.1430) 0.985 (0.830–1.168) 0.896 (0.504–1.593)
Control 1276/367/29 P ¼ .796 P ¼ .861 P ¼ .708
Drinking Case 330/88/4 0.96 (0.707–1.302) 0.969 (0.697–1.348) 0.774 (0.210–2.852)
Control 365/103/7 P ¼ .792 P ¼ .853 P ¼ .701
No drinking Case 1215/346/28 1.044 (0.897–1.215) 1.047 (0.884–1.239) 1.086 (0.626–1.887)
Control 1331/369/29 P ¼ .576 P ¼ .595 P ¼ .768
rs7624906 TT/TC/CC
<55 y Case 472/58/3 1.055 (0.743–1.499) 5.537 (0.526–58.235) 5.537 (0.526–58.235)
Control 760/95/1 P ¼ .764 P ¼ .154 P ¼ .154
55 y Case 1291/180/5 0.986 (0.792–1.226) 0.989 (0.787–1.243) 0.877 (0.258–2.979)
Control 1177/165/6 P ¼ .897 P ¼ .923 P ¼ .834
Male Case 719/103/59 0.999 (0.762–1.310) 0.989 (0.742–1.319) 1.238 (0.319–4.803)
Control 760/117/59 P ¼ .994 P ¼ .941 P ¼ .757
Female Case 1044/135/59 1.057 (0.821–1.361) 1.044 (0.804–1.354) 2.002 (0.376–10.659)
Control 1177/143/59 P ¼ .666 P ¼ .748 P ¼ .416
Smoking Case 406/70/3 1.125 (0.801–1.580) 1.156 (0.803–1.664) 0.857 (0.181–4.043)
Control 456/72/4 P ¼ .496 P ¼ .436 P ¼ .845
No smoking Case 1357/168/5 0.997 (0.800–1.243) 0.974 (0.776–1.223) 2.688 (0.594–12.163)
Control 1481/188/3 P ¼ .980 P ¼ .819 P ¼ .199
Drinking Case 365/54/3 0.986 (0.680–1.428) 0.989 (0.664–1.474) 0.911 (0.174–4.777)
Control 409/63/3 P ¼ .940 P ¼ .957 P ¼ .912
No drinking Case 1398/184/5 1.038 (0.839–1.285) 1.022 (0.820–1.275) 2.025 (0.510–8.030)
Control 1528/197/4 P ¼ .730 P ¼ .845 P ¼ .316
CI, confidence interval; HT, heterozygote; MT, minor homozygote; OR, odds ratio; SNP, single-nucleotide polymorphism; WT, major homozygote.
Age stratification was adjusted for gender, TCH, TG, LDL-C, HDL-C, BMI, GLU, smoking, and drinking status. Gender stratification was
adjusted for age, TCH, TG, LDL-C, HDL-C, BMI, GLU, smoking, and drinking status. Smoking stratification was adjusted for age, gender,
TCH, TG, LDL-C, HDL-C, BMI, GLU, and drinking status. Drinking stratification was adjusted for age, gender, TCH, TG, LDL-C, HDL-C,
BMI, GLU, and smoking status
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237 237.e8

Table S9
Association analysis of three SNPs at GHRH and SST genes with incident hypertension
SNP Genotype Case Person-year Incidence Density Adjusted HR (95% CI)*
(/100 Person years)
Additive Dominant Recessive
rs6032470 TT 382 5555.38 6.88 1.004 (0.876–1.151) 1.014 (0.860–1.195) 0.961 (0.66–1.401)
TC 202 2986.43 6.76 P ¼ .952 P ¼ .869 P ¼ .837
CC 29 440.92 6.58
rs3755792 GG 477 6899.42 6.91 0.928 (0.782–1.100) 0.897 (0.741–1.086) 1.142 (0.658–1.982)
GA 123 1932.12 6.37 P ¼ .389 P ¼ .266 P ¼ .638
AA 13 154.3 8.43
rs7624906 TT 533 7883.46 6.76 1.069 (0.847–1.349) 1.089 (0.859–1.380) –
TC 80 1087.96 7.35 P ¼ .574 P ¼ .481 –
CC – 13.08 –
CI, confidence interval; HR, hazard ratio; SNP, single-nucleotide polymorphism.
* Adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, T2DM, smoking, and drinking status.

Table S10
Association analysis of three SNPs in GHRH and SST with incident ischemic stroke
SNP Genotype Case Person-year Incidence Density HR (95% CI)*
(/103 Person-years)
Additive Dominant Recessive
rs6032470 TT 98 13,074.26 7.50 1.02 (0.793–1.312) 1.103 (0.808–1.506) 0.732 (0.36–1.489)
TC 62 7184.38 8.63 P ¼ .875 P ¼ .537 P ¼ .389
CC 11 943.93 11.65
rs3755792 GG 130 16,284.38 7.98 1.021 (0.734–1.419) 1.071 (0.749–1.531) 0.425 (0.059–3.047)
GA 40 4581.31 8.73 P ¼ .904 P ¼ .706 P ¼ .395
AA 1 343.1 2.92
rs7624906 TT 149 18,646.54 7.99 1.226 (0.795–1.889) 1.212 (0.768–1.911) 2.153 (0.297–15.582)
TC 21 2488.61 8.44 P ¼ .357 P ¼ .409 P ¼ .448
CC 1 62.29 16.05
CI, confidence interval; HR, hazard ratio; SNP, single-nucleotide polymorphism.
* Adjusted for gender, age, TCH, TG, LDL-C, HDL-C, BMI, smoking, drinking, T2DM, and hypertension.
237.e9 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237

Table S11
Stratification analysis by age, gender, smoking, and drinking for the association between the three SNPs and incident hypertension
SNP Group WT/HT/MT n Person-years Incidence HR (95% CI)
Density
Additive Dominant Recessive
(/102 Person-
years)
rs6032470 <55 y TT 117 2305.91 5.07 1.002 (0.774–1.298) 0.972 (0.715–1.321) 1.188 (0.596–2.369)
TC 57 1238.02 4.6 P ¼ .987 P ¼ .854 P ¼ .624
CC 9 169.75 5.3
55 y TT 265 3249.47 8.16 1.003 (0.852–1.181) 1.021 (0.838–1.244) 0.920 (0.584–1.449)
TC 145 1748.41 8.29 P ¼ .972 P ¼ .838 P ¼ .718
CC 20 271.17 7.38
Male TT 172 2283.16 7.53 1.037 (0.840–1.281) 1.103 (0.853–1.425) 0.779 (0.407–1.490)
TC 84 1143.61 7.35 P ¼ .736 P ¼ .455 P ¼ .450
CC 10 152.95 6.54
Female TT 210 3272.22 6.42 0.989 (0.826–1.184) 0.970 (0.781–1.205) 1.072 (0.672–1.709)
TC 118 1842.81 6.4 P ¼ .903 P ¼ .782 P ¼ .772
CC 19 287.97 6.6
Smoking TT 108 1421.32 7.6 0.876 (0.650–1.180) 0.874 (0.62–1.233) 0.742 (0.293–1.882)
TC 46 717.28 6.41 P ¼ .383 P ¼ .443 P ¼ .530
CC 5 93.52 5.35
No smoking TT 274 4134.06 6.63 1.051 (0.900–1.228) 1.067 (0.883–1.29) 1.044 (0.689–1.581)
TC 156 2269.15 6.87 P ¼ .527 P ¼ .500 P ¼ .840
CC 24 347.4 6.91
Drinking TT 105 1322.77 7.94 0.830 (0.624–1.104) 0.929 (0.656–1.316) 0.283 (0.099–0.811)
TC 46 580.19 7.93 P ¼ .200 P ¼ .679 P ¼ .019
CC 4 80.58 4.96
No drinking TT 277 4232.62 6.54 1.036 (0.886–1.213) 1.019 (0.843–1.231) 1.173 (0.782–1.762)
TC 156 2406.24 6.48 P ¼ .657 P ¼ .846 P ¼ .441
CC 25 360.34 6.94
rs3755792 <55 y GG 139 2823.76 4.92 0.965 (0.706–1.319) 0.953 (0.675–1.344) 1.064 (0.336–3.366)
GA 41 832.93 4.92 P ¼ .823 P ¼ .782 P ¼ .916
AA 3 60.1 4.99
55 y GG 338 4075.66 8.29 0.916 (0.747–1.124) 0.882 (0.700–1.112) 1.111 (0.590–2.089)
GA 82 1099.19 7.46 P ¼ .402 P ¼ .288 P ¼ .745
AA 10 94.2 10.62
Male GG 213 2783.66 7.65 0.873 (0.657–1.161) 0.855 (0.630–1.16) 1.011 (0.320–3.195)
GA 50 750.54 6.66 P ¼ .351 P ¼ .313 P ¼ .985
AA 3 45.51 6.59
Female GG 264 4115.75 6.41 0.933 (0.752–1.157) 0.897 (0.699–1.15) 1.121 (0.595–2.11)
GA 73 1181.57 6.18 P ¼ .528 P ¼ .392 P ¼ .724
AA 10 108.79 9.19
Smoking GG 133 1757.02 7.57 0.684 (0.456–1.027) 0.634 (0.413–0.973) 1.802 (0.431–7.539)
GA 24 447.69 5.36 P ¼ .067 P ¼ .037 P ¼ .420
AA 2 27.41 7.3
No smoking GG 344 5142.4 6.69 0.992 (0.821–1.198) 0.978 (0.789–1.213) 1.105 (0.606–2.015)
GA 99 1484.43 6.67 P ¼ .935 P ¼ .839 P ¼ .745
AA 11 126.89 8.67
Drinking GG 128 1507.02 8.49 0.580 (0.385–0.874) 0.558 (0.365–0.854) 0.753 (0.103–5.496)
GA 26 451.32 5.76 P ¼ .009 P ¼ .007 P ¼ .780
AA 1 25.19 3.97
No drinking GG 349 5392.4 6.47 1.019 (0.844–1.231) 0.999 (0.804–1.240) 1.225 (0.689–2.181)
GA 97 1480.8 6.55 P ¼ .844 P ¼ .990 P ¼ .489
AA 12 129.11 9.29
rs7624906 <55 y TT 157 3284.31 4.78 1.062 (0.697–1.620) 1.064 (0.697–1.623) –
TC 26 429.38 6.06 P ¼ .779 P ¼ .774 –
CC – 3.09 –
(continued on next page)
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237 237.e10

Table S11 (continued )

SNP Group WT/HT/MT n Person-years Incidence HR (95% CI)
Density
Additive Dominant Recessive
(/102 Person-
years)
55 y TT 376 4599.15 8.18 1.016 (0.765–1.348) 1.045 (0.782–1.397) –
TC 54 658.57 8.2 P ¼ .913 P ¼ .767 –
CC – 9.99 –
Male TT 221 3088.91 7.15 1.311 (0.957–1.796) 1.375 (0.993–1.905) –
TC 45 476.39 9.45 P ¼ .092 P ¼ .056 –
CC – 13.08 –
Female TT 312 4794.55 6.51 0.862 (0.605–1.226) 0.862 (0.605–1.226) –
TC 35 611.56 5.72 P ¼ .408 P ¼ .408 –
CC – – –
Smoking TT 129 1929.05 6.69 1.660 (1.125–2.449) 1.803 (1.198–2.712) –
TC 30 289.99 10.35 P ¼ .011 P ¼ .005 –
CC – 13.08 –
No smoking TT 404 5954.41 6.78 0.917 (0.681–1.234) 0.917 (0.681–1.234) –
TC 50 797.97 6.27 P ¼ .566 P ¼ .566 –
CC – – –
Drinking TT 127 1694.91 7.49 1.083 (0.719–1.632) 1.125 (0.735–1.720) –
TC 28 277.29 10.1 P ¼ .702 P ¼ .588 –
CC – 9.99 –
No drinking TT 406 6188.55 6.56 1.039 (0.778–1.388) 1.040 (0.778–1.390) –
TC 52 810.66 6.41 P ¼ .795 P ¼ .791 –
CC – 3.09
CI, confidence interval; HR, hazard ratio; HT, heterozygote; MT, minor homozygote; SNP, single-nucleotide polymorphism; WT, major
homozygote.
Age stratification was adjusted for gender, TCH, TG, LDL-C, HDL-C, BMI, smoking, drinking, and T2DM. Gender stratification was
adjusted for age, TCH, TG, LDL-C, HDL-C, BMI, smoking, drinking, and T2DM. Smoking stratification was adjusted for age, gender,
TCH, TG, LDL-C, HDL-C, BMI, drinking, and T2DM. Drinking stratification was adjusted for age, gender, TCH, TG, LDL-C, HDL-
C, BMI, smoking, and T2DM.
237.e11 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237

Table S12
Stratification analysis by age, gender, smoking, and drinking for the association between the three SNPs and incident ischemic stroke
SNP Group Genotype n Person-year Incidence HR (95% CI)
Density
Additive Dominant Recessive
(/103 Person-
years)
rs6032470 <55 y TT 7 4694.07 1.49 0.92 (0.362–2.342) 0.988 (0.271–3.601) 0.637 (0.062–6.529)
TC 3 2662.16 1.13 P ¼ .861 P ¼ .986 P ¼ .704
CC 2 335.71 5.96
55 y TT 91 8380.19 10.86 1.066 (0.816–1.392) 1.112 (0.805–1.536) 0.935 (0.446–1.96)
TC 59 4522.21 13.05 P ¼ .64 P ¼ .52 P ¼ .859
CC 9 608.22 14.8
Male TT 49 5572.53 8.79 1.148 (0.804–1.639) 1.237 (0.791–1.933) 1.012 (0.397–2.581)
TC 30 2729.63 10.99 P ¼ .447 P ¼ .351 P ¼ .98
CC 8 363.61 22
Female TT 49 7501.73 6.53 0.913 (0.632–1.318) 0.98 (0.629–1.525) 0.527 (0.159–1.743)
TC 32 4454.75 7.18 P ¼ .628 P ¼ .927 P ¼ .294
CC 3 580.32 5.17
Smoking TT 23 3408.88 6.75 1.625 (1.002–2.635) 1.775 (0.939–3.354) 2.185 (0.734–6.504)
TC 15 1605.48 9.34 P ¼ .049 P ¼ .077 P ¼ .160
CC 5 217.48 22.99
No smoking TT 75 9665.38 7.76 0.887 (0.657–1.199) 0.968 (0.674–1.389) 0.442 (0.164–1.196)
TC 47 5578.9 8.43 P ¼ .436 P ¼ .86 P ¼ .108
CC 6 726.46 8.26
Drinking TT 21 3098.34 6.78 0.869 (0.481–1.571) 1.116 (0.541–2.304) 0.159 (0.02–1.235)
TC 13 1401.2 9.28 P ¼ .642 P ¼ .767 P ¼ .079
CC 3 174.97 17.15
No drinking TT 77 9975.92 7.72 1.062 (0.798–1.414) 1.119 (0.789–1.587) 0.893 (0.4–1.992)
TC 49 5783.18 8.47 P ¼ .679 P ¼ .529 P ¼ .782
CC 8 768.97 10.4
rs3755792 <55 y GG 6 5869.25 1.02 1.298 (0.356–4.731) 1.488 (0.353–6.274) –
GA 6 1706.1 3.52 P ¼ .692 P ¼ .588 –
AA – 122.81 –
55 y GG 124 10,415.13 11.91 1.005 (0.711–1.421) 1.065 (0.728–1.558) 0.396 (0.055–2.84)
GA 34 2875.21 11.83 P ¼ .978 P ¼ .745 P ¼ .357
AA 1 220.29 4.54
Male GG 64 6682.88 9.58 1.134 (0.717–1.793) 1.234 (0.754–2.018) –
GA 23 1843.36 12.48 P ¼ .592 P ¼ .403 –
AA – 139.53 –
Female GG 66 9601.5 6.87 0.924 (0.569–1.502) 0.928 (0.545–1.579) 0.775 (0.106–5.681)
GA 17 2737.94 6.21 P ¼ .751 P ¼ .782 P ¼ .802
AA 1 203.57 4.91
Smoking GG 32 4055.58 7.89 0.94 (0.463–1.908) 1.016 (0.474–2.179) –
GA 11 1091.37 10.08 P ¼ .863 P ¼ .967 –
AA – 84.89 –
No smoking GG 98 12,228.8 8.01 1.022 (0.697–1.499) 1.066 (0.703–1.618) 0.54 (0.075–3.896)
GA 29 3489.94 8.31 P ¼ .911 P ¼ .763 P ¼ .542
AA 1 258.21 3.87
Drinking GG 28 3609.6 7.76 1.129 (0.523–2.44) 1.229 (0.537–2.814) –
GA 9 1000.85 8.99 P ¼ .757 P ¼ .625 –
AA – 64.05 –
No drinking GG 102 12,674.78 8.05 0.97 (0.666–1.413) 1.004 (0.668–1.509) 0.514 (0.072–3.689)
GA 31 3580.45 8.66 P ¼ .873 P ¼ .986 P ¼ .508
AA 1 279.05 3.58
rs7624906 <55 y TT 11 6829.47 1.61 0.988 (0.124–7.89) 1.031 (0.118–9.008) –
TC 1 844.07 1.19 P ¼ .991 P ¼ .978 –
CC – 24.62 –
55 y TT 138 11,817.07 11.68 1.144 (0.739–1.773) 1.138 (0.715–1.812) 1.582 (0.218–11.482)
(continued on next page)
 H. Zhu et al. / Journal of the American Society of Hypertension 12(3) (2018) 230–237 237.e12

Table S12 (continued )

SNP Group Genotype n Person-year Incidence HR (95% CI)
Density
Additive Dominant Recessive
(/103 Person-
years)
TC 20 1644.54 12.16 P ¼ .546 P ¼ .586 P ¼ .65
CC 1 37.67 26.55
Male TT 74 7501.68 9.86 1.387 (0.798–2.413) 1.38 (0.751–2.537) 2.394 (0.327–17.497)
TC 12 1114.09 10.77 P ¼ .246 P ¼ .299 P ¼ .39
CC 1 43.66 22.9
Female TT 75 11,144.86 6.73 1.069 (0.534–2.137) 1.077 (0.536–2.163) –
TC 9 1374.52 6.55 P ¼ .851 P ¼ .836 –
CC – 18.63 –
Smoking TT 35 4463.42 7.84 1.168 (0.532–2.563) 1.21 (0.538–2.721) –
TC 8 733.8 10.9 P ¼ .699 P ¼ .644 –
CC – 34.63 –
No smoking TT 114 14,183.13 8.04 1.297 (0.765–2.199) 1.25 (0.712–2.194) 3.657 (0.495–27.011)
TC 13 1754.81 7.41 P ¼ .335 P ¼ .437 P ¼ .204
CC 1 27.66 36.15
Drinking TT 29 4037.93 7.18 1.964 (0.945–4.085) 1.914 (0.826–4.432) 5.807 (0.756–44.582)
TC 7 608.91 11.5 P ¼ .071 P ¼ .130 P ¼ .091
CC 1 26.33 37.98
No drinking TT 120 14,608.62 8.21 0.991 (0.572–1.718) 1.013 (0.578–1.776) –
TC 14 1879.7 7.45 P ¼ .975 P ¼ .964 –
CC – 35.96 –
CI, confidence interval; HR, hazard ratio; SNP, single-nucleotide polymorphism.
Age stratification was adjusted for gender, TCH, TG, LDL-C, HDL-C, BMI, smoking, drinking, T2DM, and hypertension. Gender strat-
ification was adjusted for age, TCH, TG, LDL-C, HDL-C, BMI, smoking, drinking, T2DM, and hypertension. Smoking stratification was
adjusted for age, gender, TCH, TG, LDL-C, HDL-C, BMI, drinking, T2DM, and hypertension. Drinking stratification was adjusted for age,
gender, TCH, TG, LDL-C, HDL-C, BMI, smoking, T2DM, and hypertension.