# Proinflammation and Hypertension: A Population-Based Study

Authors: Vanhala Mauno; Kautiainen Hannu; Kumpusalo Esko
Journal: Mediators of Inflammation
 (2008)

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## Abstract

There is evidence that proinflammation may be linked to the development of hypertension (HT). We examined the association of both the interleukin-1 beta (IL-1β) and the interleukin 1-receptor antagonist (IL-1ra) with future blood pressure (BP) and HT occurrence (BP≥140/90 mmHg, or antihypertensive drug) in a population-based prospective study. Our study consisted of 396 (147 men and 249 women) middle-aged, baseline apparently healthy, normotensive subjects participating in a 6.5-year follow-up study. Subjects with high-sensitivity CRP (hs-CRP)<10 mg/L were excluded at the initial visit.  At follow-up, the occurrence of HT was 32%. The levels of baseline IL-1β and IL-1ra were significantly higher for subjects who developed HT during the follow-up than for those who did not (IL-1β; 0.67 ± 0.62 pg/mL versus 0.56 ± 0.32 pg/mL, P=.020 and  IL-1ra; 184 ± 132 pg/mL versus 154 ± 89 pg/mL, P=.007). After adjustments for age, follow-up time, sex, baseline systolic BP, and BMI, our results confirm a statistically significant (P=.036) linear association between the quartiles of IL-1β and change of systolic BP during the study. After adjustments for age, follow-up time, sex, and BMI, our results also show a linear association between incident HT and the quartiles of IL-1ra. (P=.026).  These results provide evidence that proinflammation may precede BP elevation and HT.


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## Body

## 1. Introduction

There exists a growing body of evidence describing the role of cytokines
in the context of hypertension (HT) [1–5].
Interleukin-1beta (IL-1β) is a crucial mediator of inflammatory response.
Animal experiments that examined the vascular wall of hypertensive rats showed
an increase in mRNA expression levels of IL-6, IL-1β, and TNF-alpha [6].
Previous human work has shown increased levels of IL-1β in the systemic
circulation of patients with essential HT. Most IL-1β is secreted by peripheral
blood monocytes instead of adipocytes in hypertensive subjects, and angiotensin
II (Ang II) may be directly involved in the process of monocyte activation [7].
IL-1β and Ang II together may also regulate sympathetic nervous system (SNS)
activity through negative feedback by modulating the expression of nitric oxide
(NO) in the brain [8–10]. IL-1β could also be an additional risk factor
for atherogenesis in hypertensive patients [11].The production and effects of IL-1β are controlled at many levels, a critical
one being the inhibition of its activities by the secreted form of the
interleukin-1 receptor antagonist (IL-1ra) [12].
Elevated concentrations of this substance have been connected to insulin
resistance and type 2 diabetes [13]. Moreover, elevated levels of IL-1ra have been found in subjects with essential HT
[14]. Even if proinflammation is linked to cardiovascular disease (CVD), there
currently exists no evidence showing that IL-1ra could have an independent role in the
etiology of CVD [15].IL-1β also stimulates the synthesis of hs-CRP and other inflammatory mediators [16–18]. It has been speculated that
inflammation plays an important role in triggering vascular fibrosis, which is
an important aspect of extracellular matrix remodeling in hypertension [19]. Another research has
proven that CRP predicts future cardiovascular events, at least in women [20].Baseline blood pressure (BP), obesity, use of alcohol, and lack of
physical activity have traditionally been suggested as precursors of HT that works via mechanisms including
proinflammation and insulin resistance [21, 22].Despite several reliable cross-sectional studies showing the connection
of IL-1β and IL-1ra
to hypertension, researchers have not yet conclusively linked these
proinflammatory markers to blood pressure in a longitudinal study. Accordingly,
we examined the two aforementioned proinflammatory markers, lifestyle factors,
and other traditional risk factors in a normotensive, apparently healthy
population. Our goal was to understand their roles in both future BP changes
and HT in follow-up examinations.


## 2. Materials and Methods

### 2.1. Subjects

All subjects (N=1294) were born in 1942, 1947, 1952, 1957, and 1962 in
Pieksämäki, a town in eastern Finland. The patients were invited to participate
in a general health examination in 1997-1998. A total of 923 of
the 1294 subjects (71.3%) participated in an initial examination in 1997-1998, and 690 of those
underwent a second checkup in 2003-2004. The mean
follow-up time was 6.45 (range 5.3–7.4) years. There
were no additional examinations between these two checkups. All participants
gave informed written consent. The study protocol was approved by the Ethics
Committee of the Kuopio University Hospital and the University of Kuopio.All participants completed a standard questionnaire in the beginning and
at the end of the study. It included questions about smoking habits, use of
alcohol, and physical activity. Those who smoked daily were classified as
smokers. Use of alcohol was measured using a scale with ten graded levels: 0 =
never used, 1 = not used alcohol in the last year, 3 = used less than 1 unit (standard drinks)/week, 4 = 1-2 units/week, 5 = 3–5 units/week, 6 = 6–9 units/week, 7 = 10–14 units/week, 8
= 15–21 units/week, 9 = 22–28 units/week, and 10 = more than 29 units/week. A subject was classified as drinking more
than the recommended 1-2 doses/day if he/she consumed 10 or more alcohol units/week. Physical activity was measured
using a scale that comprised six graded levels. The patients were asked to
define physical exercise as sufficiently strenuous exertion that results in
perspiration and breathlessness. 1 = daily exercise, 2 = three or more
times/week, 3 = twice/week, 4 = about once/week, 5 = once/month, and 6 = less
than once/month. A subject was classified as physically active if he/she
exercised three or more times per week.All patients with hs-CRP > 10 mg/L at the initial
examination were excluded  due to the fact that they might have had an active
infection. Since certain cardiovascular drugs can affect BP and cytokines, all
patients taking medication for another cardiovascular disease were excluded in the beginning of the
study.


### 2.2. Clinical and Laboratory Methods

Height and weight were measured to the nearest 0.5 cm and 0.1 kg,
respectively. Body mass index (BMI) was calculated as weight (kg) divided by
height (m) squared. The waist was measured to the nearest 1.0 cm at the
midpoint between the lateral iliac crest and the lowest rib. Trained nurses
measured BP twice with subjects in a seated position using a mercury
sphygmomanometer. The mean of two measurements was used for subsequent
statistical analyses. Measurements were performed at the initial and final
visits. Hypertension was determined to be present if subjects had a BP ≥140/90 mmHg or if subjects were taking antihypertensive medication [23].Fasting blood samples were drawn after 12 hours of fasting. C-reactive
protein (CRP) was measured with an Immulite (R) and a DPC high-sensitivity CRP assay (hs-CRP). Plasma concentrations
of IL-β and IL-1ra
were measured using assay kits from R&D Systems (Minneapolis, Minn, USA; for
IL-1β, resolution power was 0.125–8 pg/mL and
sensitivity 0.1 pg/mL; for IL-1ra, resolution power was 46.9–3000 pg/mL and
sensitivity 22 pg/mL). Cytokines were measured during the initial examination.


### 2.3. Statistical Analysis

The results were expressed as means, standard deviations (SDs), ranges,
and 95 percent confidence intervals (95% CI). IL-1β and IL-1ra exhibited skewed
distributions and were, therefore, logarithmically transformed for statistical
analysis. The t-test and chi-squared test were used to derive
statistical comparisons between
genders or between groups of hypertensive and normotensive subjects in
respect of baseline characteristics and other subgroup parameters. The
hypotheses of linearity between quartiles of plasma cytokine levels and changes
in systolic BP or occurrence of HT were analyzed with two general linear models
known as the analysis of covariance (ANCOVA) and with logistical models. All
models were adjusted for gender, age, baseline BMI, follow-up time, and
baseline systolic BP when appropriate. All statistical tests were two sided, withan alpha-level of
0.05.


## 2.1. Subjects

All subjects (N=1294) were born in 1942, 1947, 1952, 1957, and 1962 in
Pieksämäki, a town in eastern Finland. The patients were invited to participate
in a general health examination in 1997-1998. A total of 923 of
the 1294 subjects (71.3%) participated in an initial examination in 1997-1998, and 690 of those
underwent a second checkup in 2003-2004. The mean
follow-up time was 6.45 (range 5.3–7.4) years. There
were no additional examinations between these two checkups. All participants
gave informed written consent. The study protocol was approved by the Ethics
Committee of the Kuopio University Hospital and the University of Kuopio.All participants completed a standard questionnaire in the beginning and
at the end of the study. It included questions about smoking habits, use of
alcohol, and physical activity. Those who smoked daily were classified as
smokers. Use of alcohol was measured using a scale with ten graded levels: 0 =
never used, 1 = not used alcohol in the last year, 3 = used less than 1 unit (standard drinks)/week, 4 = 1-2 units/week, 5 = 3–5 units/week, 6 = 6–9 units/week, 7 = 10–14 units/week, 8
= 15–21 units/week, 9 = 22–28 units/week, and 10 = more than 29 units/week. A subject was classified as drinking more
than the recommended 1-2 doses/day if he/she consumed 10 or more alcohol units/week. Physical activity was measured
using a scale that comprised six graded levels. The patients were asked to
define physical exercise as sufficiently strenuous exertion that results in
perspiration and breathlessness. 1 = daily exercise, 2 = three or more
times/week, 3 = twice/week, 4 = about once/week, 5 = once/month, and 6 = less
than once/month. A subject was classified as physically active if he/she
exercised three or more times per week.All patients with hs-CRP > 10 mg/L at the initial
examination were excluded  due to the fact that they might have had an active
infection. Since certain cardiovascular drugs can affect BP and cytokines, all
patients taking medication for another cardiovascular disease were excluded in the beginning of the
study.


## 2.2. Clinical and Laboratory Methods

Height and weight were measured to the nearest 0.5 cm and 0.1 kg,
respectively. Body mass index (BMI) was calculated as weight (kg) divided by
height (m) squared. The waist was measured to the nearest 1.0 cm at the
midpoint between the lateral iliac crest and the lowest rib. Trained nurses
measured BP twice with subjects in a seated position using a mercury
sphygmomanometer. The mean of two measurements was used for subsequent
statistical analyses. Measurements were performed at the initial and final
visits. Hypertension was determined to be present if subjects had a BP ≥140/90 mmHg or if subjects were taking antihypertensive medication [23].Fasting blood samples were drawn after 12 hours of fasting. C-reactive
protein (CRP) was measured with an Immulite (R) and a DPC high-sensitivity CRP assay (hs-CRP). Plasma concentrations
of IL-β and IL-1ra
were measured using assay kits from R&D Systems (Minneapolis, Minn, USA; for
IL-1β, resolution power was 0.125–8 pg/mL and
sensitivity 0.1 pg/mL; for IL-1ra, resolution power was 46.9–3000 pg/mL and
sensitivity 22 pg/mL). Cytokines were measured during the initial examination.


## 2.3. Statistical Analysis

The results were expressed as means, standard deviations (SDs), ranges,
and 95 percent confidence intervals (95% CI). IL-1β and IL-1ra exhibited skewed
distributions and were, therefore, logarithmically transformed for statistical
analysis. The t-test and chi-squared test were used to derive
statistical comparisons between
genders or between groups of hypertensive and normotensive subjects in
respect of baseline characteristics and other subgroup parameters. The
hypotheses of linearity between quartiles of plasma cytokine levels and changes
in systolic BP or occurrence of HT were analyzed with two general linear models
known as the analysis of covariance (ANCOVA) and with logistical models. All
models were adjusted for gender, age, baseline BMI, follow-up time, and
baseline systolic BP when appropriate. All statistical tests were two sided, withan alpha-level of
0.05.


## 3. Results

Of the 233 nonparticipant subjects, 48% (n=112) were men and 52% (n=121)
were women. Our
data shows no statistical difference in BP, BMI, age, or lifestyle factors in
nonparticipants as compared with the participants. Of the 690 participants, 406
subjects (59%) had a BP < 140/90 mmHg and were not undergoing antihypertensive
drug treatment at the time of the baseline checkup. Six of these patients had hs-CRP > 10 mg/L, and
four subjects were taking a certain drug for another CVD; all of such participants
were excluded. Thus, the final study population consisted of 396 apparently
healthy subjects (147 men and 249 women).At the initial checkup, mean systolic BP was 126 ± 8 mmHg in male patients
and 122 ± 10 mmHg in female subjects (P between sexes < .001). Mean
diastolic BP was, respectively, 77 ± 6 mmHg and 75 ± 7 mmHg (P<.001). BMI
in men was higher than in women (25.9±3.1 kg/m2 versus 25.1±3.6 kg/m2, P between sexes = .029). Twelve percent of men consumed 10 or more units
of alcohol each week, and this was statistically significantly higher than the
2% of women who reported the same alcohol consumption (P between sexes
< .001). There was no sex-related difference in physical activity. The
percentages of physically active subjects were 30% in men and 28% in women.The levels of cytokines recorded at the baseline checkup were statistically
similar for both sexes. The level of IL-1β was 0.59 pg/mL ± 0.43 pg/mL in men and
0.58 pg/mL ± 0.43 pg/mL in women. The corresponding figures for IL-1ra were 157 pg/mL ± 92 pg/mL in men and 168 pg/mL ± 102 pg/mL in women. There was no statistically
significant correlation between these two cytokines.Between the initial and follow-up examinations, smoking decreased from
28% to 23% in men and from 21% to 17% in women, and consumption of 10 or more
alcohol units/week increased from 12% to 18% in men and from 2% to 4% in women.
The percentage of physically active subjects did not change.Between examinations, the systolic BP increased 9 ± 12 mmHg and the
diastolic BP increased 4 ± 8 mmHg. In total, 128 (32.3%) of the 396 subjects, 59
men (41%) and 69 women (27%,P=.005 between sexes), developed HT. Five
of those 59 men and 14 of those 69 women were prescribed antihypertensive
medication between the initial and the subsequent visit.At the end of the follow-up, six (4%) men and seven (3%) women were
receiving drug treatment for another cardiovascular condition. The use of
antilipolytic medication increased among the men from four (3%) to 28 (19%)
users, and in women from two (1%) to 27 (11%) users over the duration of our
study. In those subjects treated with lipid-lowering drugs at the end of the
study, the mean systolic BP by the end of our study was 138 ± 16 mmHg, as
compared to 131 ± 14 mmHg in the nontreated subjects (P<.001 between
groups). The corresponding figures for diastolic BP were 82 ± 9 mmHg and 80 ± 9 mmHg, P=.060.Table1 shows that the mean level of baseline IL-1β was significantly
higher among those subjects who had developed hypertension between the initial
and the subsequent visits
as compared to those subjects who were normotensive at the time of the second
checkup (0.56 pg/mL versus 0.67 pg/mL, P=.020). The same phenomenon was
present in the levels of IL-1ra
(154 pg/mL versus 184 pg/mL, P=.007).Table 1
Baseline characteristics of 396 baseline normotensive subjects according tohypertension status at the end of the follow-up period. HT−: normotensive at the end of study, HT+: hypertensive at the end of
study (blood pressure ≥140/90 mmHg and/or antihypertensive drug use), and CVD: cardiovascular
disease.
HT−HT+Pn=268n=128Number of female = 249/396 (63%)180 (67%)69 (54%)—Age (years)50.6±6.153.0 ± 51<.001Follow-up
time (years)6.4±0.46.4±0.4.091BMI (kg/m2)25.1±3.626.0±3.611IL-1β (pg/mL)0.56±0.320.67±0.62.020IL-1ra (pg/mL)154 ±  89184 ± 1327Lipid-lowering drug at the beginning of the study1%3%.091Lipid-lowering drug at the end of the study9%26%<.001Drug for some other CVD at the end of the study3%6%.398Smoking30%26%.807Alcohol > 10
units/week%5%6%.807Physically active25%33%.086The systolic BP increased 6 ± 14 mmHg from the lowest to the highest
quartile of IL-1β. This difference was statistically significant (P=.008).
The BP values sorted by quartile of IL-1β concentration were 129 ± 13 mmHg,
132 ± 16 mmHg, 130 ± 14 mmHg, and 135 ± 15 mmHg (P for linearity = .019).
Figure 1 shows that after adjusting
gender, age, follow-up time, BMI, and baseline systolic BP, a
statistically significant (P=.036) linear association was evidenced
between the quartiles of IL-1β and the change in the systolic BP during the
follow-up. Further adjustment for
smoking, use of alcohol, and physical exercise at baseline did not change the
results. Taking into account lipid-lowering medication or drugs for other
cardiovascular disease did not change these results.Change of systolic blood
pressure (mmHg) by IL-1β and IL-1ra
quartiles among 377 baseline normotensive subjects without antihypertensive
drug treatment at the end of the 6.5-year prospective study. Adjusted for gender, age, baseline BMI, baseline
systolic blood pressure, and follow-up time.
(a)(b)The prevalence rates of incident HT, from the lowest to the highest
quartile of IL-1ra, were as follows: 26.5%, 23.2%, 34%, and 45.5% (crudeP=.003
for linearity). The increase in HT occurrence from the lowest to the highest
quartile of IL-1ra was
19% (P=.05
after adjustments for gender, age, baseline BMI, and follow-up time). Table 2
and Figure 2 show that there was also a linear association between HT incidence
and quartiles of IL-1ra
(P=.026) after adjustments as previously described. Further adjustment
for smoking, use of alcohol, and physical exercise at baseline did not change
the results. Taking into account lipid-lowering medication or drugs for other
cardiovascular disease did not change these results.Table 2
Odds ratios of hypertension (≥140/≥90 mmHg or drug treatment for
hypertension) according to quartiles of plasma levels of baseline IL-1β and IL-1ra.
VariableQuartile of plasma levelP for linearity*IIIIIIIVIL-1βMedian (range), pg/mL0.25 (0.088–0.378)0.48 (0.379–0.566)0.63 (0.567–0.696)0.82 (0.700–5.540)Odds ratio (95% CI)1.001.47 (0.79 to 2.73)0.83 (0.43 to 1.58)1.55 (0.84 to 2.89).46IL-1raMedian (range), pg/mL84 (46–98)117 (99–139)163 (140–192)244 (193–1134)Odds ratio (95% CI)1.000.61 (0.31 to 1.20)1.07 (0.56 to 2.04)1.82 (0.95 to 3.46).026*P-values were calculated by using a logistic
model, adjusted for gender, age, follow-up time, and baseline body mass index.Occurrence of hypertension (RR ≥ 140/90 mmHg or drug treatment for hypertension)
over a 6.5-year timeframe according to quartiles of baseline IL-1β and IL-1ra. Adjusted for gender, age, baseline BMI, and follow-up time.
(a)(b)


## 4. Discussion

To the best of our knowledge, this study is the first prospective,
population-based study providing evidence that high levels of proinflammatory
cytokine IL-1β precede future changes in systolic BP and in levels of its
specific antagonist IL-1ra
in the context of HT. These associations were present after adjustments for
sex, age, BMI, and follow-up time; they also remained for analyses that only examined
BP from baseline.Earlier findings suggest that the pressor effect of IL-1β is mediated by
AT1 receptors and depends on the stimulation and the release of Angiotensin II
(Ang II). [20, 21] Ang II raises BP via its peripheral and central effects. In
the brain, ANG II downregulates neuronal NOS (nNOS) and leads to the decrease of NO
production and the increase
of SNS activity [8, 9, 18, 24]. There
are also observations that IL-1β downregulates SNS activity by increased local
expression of NOS mRNA as well as that Ang II-mediated increases in arterial BP
can be caused by inhibition of the expression of IL-1β and nNOS at the brain
level [10]. ANG II and IL-1β may thus act in a negative feedback loop in the
central BP control system [8]. It seems possible that central IL-1β is also
involved in stress-induced HT [25].The unbalanced production of IL-1β, and of its natural specific inhibitor IL-1ra,
plays an important role in chronic/sterile inflammation [12]. IL-1ra is an acute phase reactant for IL-1β, since it reflects inflammatory
reactions, acts in an antagonistic manner, and serves as a natural compensatory
mechanism for the IL-1-induced disease process. IL-1ra concentrations increase during both insulin
resistance and the metabolic syndrome [13]. A human experiment has shown that
IL-1β induces the secretion of IL-1ra, and that induction of IL-1β precedes secretion of IL-1ra.In our study, IL-1β is independently associated (even after taking into
account age, sex, baseline BMI, and certain lifestyle factors) with changes in
systolic BP that may reflect an incipient change in arterial stiffness. High
levels of IL-1ra seem
to be linked to occurrence of HT, and this can most likely be explained by the
fact that those subjects already in the initial examination exhibited elevated
BP subsequently developed stable HT (defined as ≥140/90 mmHg and/or prescribed
antihypertensive drugs). These findings are in line with previous data
suggesting that proinflammation precedes HT and that it may be a trigger for
arterial wall stiffness [20].Lipid-lowering medications and some drugs used to treat cardiovascular
diseases other than HT may have BP lowering effects [26]. Evidence also
suggests that lipid-lowering medications can affect the levels of cytokines
[27]. Therefore, we excluded subjects who were using this kind of medication at
the beginning of the study. Previous work has shown that lifestyle factors like
exercise, smoking, and consumption of alcohol can affect both BP and HT [21, 22]. Therefore,
we used the baseline information to correct our analyses for these potential
confounding factors.During the course of the study, 59 men (41%) and 69 women (27%,P=.005
between sexes) developed HT. This is consistent with both earlier results [22]
and the knowledge that middle-aged men are at greater risk for all
cardiovascular diseases than women. Our study also showed that lipid-lowering
medications were prescribed more frequently for men; this finding is also
consistent with the present recommendations [26]. Antilipolytic medication use
increased during the study, especially among those subjects who developed HT.
Therefore, the increase occurred in subjects with a high risk for having HT and
dyslipidemia simultaneously.One strength of this study lies in its population-based, prospective
design, which includes both men and women and evaluates many traditional risk
factors simultaneously. We also took into account the possibility that
lifestyle factors and some drug treatments may have an effect on the results by
correcting our analysis for these confounding factors. Lifestyle factors are
very difficult to evaluate accurately [28], and one shortcoming in our study is
that these factors were self-reported. We also did not measure sympathetic
nervous activity or endothelial function at the initial visit or levels of
cytokines during the subjects’ subsequent visit. Future studies should evaluate
the possible association of sympathetic overflow and cytokines with respect to
HT.We conclude that our report provides important evidence regarding the
relationship between cytokines and future BP or HT. This work shows that IL-1β
and IL-1ra are
markers for future BP and HT, and thus for prehypertension, in the
normotensive, apparently healthy population. We suggest that both IL-1β and IL-1ra reflect the same phenomenon, namely, vascular
inflammation and arterial stiffness. The clinical importance of this study lies
in the fact that both Il-1β and IL-1ra are linked to increased cardiovascular risk in
hypertensive subjects. This increased risk may reflect the presence of
extended-duration proinflammatory vasculature stress.


## 5. Summary

What is known about this topic?(i)
There
is evidence that proinflammation may be linked to the development of
hypertension.(ii)
Increased levels of IL-1β
and IL-1ra have
been shown to be linked with essential hypertension in cross-sectional
studies.
What this study adds?(i)
This
longitudinal study of apparently healthy middle-aged men and women shows that
high levels of IL-1β and IL-1ra
are risk factors for subsequent hypertension.



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