DOI: 10.1111/j.1365-2362.2008.01952.

Infection
M. Odermarsky
recurrence
et al.and atherosclerosis in diabetes
ORIGINAL ARTICLE
Blackwellrecurrence
ORIGINAL
Infection Publishing
ARTICLE:
andLtd
atherosclerosis
DIABETES MELLITUS
in diabetes

Respiratory infection recurrence and passive

smoking in early atherosclerosis in children
and adolescents with type 1 diabetes
M. Odermarsky*, S. Andersson†, E. Pesonen*, S. Sjöblad‡, S. Ylä-Herttuala§ and P. Liuba*
*
Paediatric Cardiology, and ‡Endocrinology, Department of Paediatrics, Lund University Hospital, Lund, Sweden, †Department
of Paediatrics, Helsinki University Hospital, Helsinki, Finland, §A. I. Virtanen Institute, University of Kuopio, Kuopio, Finland

ABSTRACT
Background Optimal glucose control in juvenile type 1 diabetes mellitus is necessary but not sufficient to reduce
the burden of cardiovascular events in later life. This emphasizes the importance of searching for other possible
risk factors associated with diabetes. We investigated whether recurrent episodes of acute respiratory infections
and exposure to tobacco smoke could influence vascular phenotypes for early atherosclerosis in children and
adolescents with type 1 diabetes.
Materials and methods Common carotid artery elasticity and intima-media thickness along with circulating
markers of lipid, inflammatory and glycaemic profiles were investigated in up to 98 children and adolescents
with type 1 diabetes. The number of clinically manifest acute respiratory tract infections (RTI) during the past
year, and the degree of exposure to environmental tobacco smoke (ETS), were assessed by separate
questionnaires.
Results Carotid artery compliance (CAC) was decreased in patients with high (≥ 4/year; n = 22) recurrence of
RTI compared to the remaining patients (n = 40; P < 0·05). In a multivariate analysis, the number of RTI during
the past year and HbA1C were independently associated with decreased CAC (P < 0·05 for both). The inverse
relationship between RTI recurrence and CAC was strengthened by frequent exposure to ETS.
Conclusions High recurrence of respiratory infections in young type 1 diabetics is associated with increased
stiffening of the carotid artery particularly in those often exposed to tobacco smoke.
Keywords Atherosclerosis, infection, type 1 diabetes.
Eur J Clin Invest 2008; 38 (6): 381–388

Introduction
Individuals with type 1 diabetes have a 2 to 3 fold increased
risk for cardiovascular disease compared to non-diabetic subjects
[1]. Accelerated atherosclerosis appears to play an important role
in the excess cardiovascular morbidity associated with diabetes.
Accumulating evidence suggests that non-invasive measures of
early subclinical atherosclerosis in peripheral arteries of young
type 1 diabetics could have prognostic value in cardiovascular
risk prediction [2–4].
What exactly drives the atherosclerotic process in type 1 diabetes
remains unclear. Close to normal glycaemic control is not sufficient
to lower the cardiovascular risk at levels comparable to those in
healthy individuals [5]. Much of the current data support the view
that atherosclerotic vasculopathy in diabetes has a multifactorial
aetiology.
Infection has been hypothesized to contribute to atherosclerosis
particularly in conjunction with other cardiovascular risk factors
[6]. Prior studies suggested that type 1 diabetes may increase
the propensity for both chronic and acute infections in part by
weakening the immune mechanisms [7,8]. Of note, both diabetes
and passive smoke appear to predispose to recurrent respiratory
infections [7,9]. Although earlier studies in hypercholesterolaemic
animals suggested increased predisposition for atherosclerosis
with increasing number of pathogen inoculation [10,11], a possible
association between infection recurrence and atherosclerosis in
type 1 diabetes has not been studied.
In the present study, we investigated whether recurrent episodes
of respiratory tract infections (RTI) in children with type 1 diabetes
could pose cumulative atherogenic effects, and whether these

European Journal of Clinical Investigation Vol 38 381

 M. ODERMARSKY ET AL.
effects could be amplified by exposure to environmental tobacco
smoke (ETS). High-resolution ultrasound was used to investigate
several indices of subclinical atherosclerosis in the common carotid
artery, while the frequency of RTI experienced during the past year
and the exposure to ETS were estimated via questionnaire.
Materials and methods
Study population
Ninety-eight children and adolescents (54 male and 44 female)
aged between seven and 22 (mean 15) years with type 1
diabetes mellitus for at least six months were recruited from
the paediatric outpatient diabetes clinic at the Lund University
Hospital. Exclusion criteria were: family history for other major
cardiovascular risk factors (primary hypercholesterolaemia,
hypertension, premature coronary and cerebrovascular disease),
active smoking, systemic hypertension, asthma and allergy.
Body weight, height and arterial blood pressure (systolic
and diastolic) were measured upon the ultrasound visit.
Data on demographic information, parental education and
current occupation were collected from questionnaires. Data on
diabetes duration were obtained from the registry of outpatient
diabetes clinic.
Detailed infection history (type and number of acute infections
during the past year; type, length and severity of symptoms,
medication if any, and time elapsed since last infection prior to
the ultrasound visit) was also determined by questionnaire.
Acute infection was defined by clinical symptoms. Patients were
grouped as follows in relation to the number of infections during
the past year: group 1 = 0–1 infections (low frequency);
group 2 = 2 to 3 infections (moderate frequency), and
group 3 = ≥ 4 infections (high frequency).
Exposure to ETS was assessed through a confidential
questionnaire, and defined as occasional or regular cigarette
smoking in the presence of study participants in or outside the
home (e.g. private or public transportation, in or around school,
playground or other public places). Thus, patients were divided
into three groups: group 1 = no exposure during the past year;
group 2 = occasional exposure, i.e. presence in a smoky
environment less than once a week; and group 3 = weekly to daily
exposure. In addition, household exposure to ETS was categorized
in relation to the average number of cigarettes smoked per day in
or around the home by the patient’s cohabitants. The number of
household smokers was also obtained.
Both questionnaires were filled out by patients and their
guardians upon the ultrasound visit, but were reviewed
after the study’s completion. The study was performed between
January 2004 and October 2005. A new law banning all smoking
in public areas came to force in July 2005.
The parental social status was deduced based on the
level (secondary/high school = low, university or professional
382 © 2008 The Authors. Journal Compilation © 2008 Blackwell Publishing Ltd
www.ejci-online.com
qualification = high) and number of years of education of the
mother and the mother’s partner.
The study was approved by the ethics committee for human
research at the Lund University. Written consent was obtained
from all participants over 18 years or from their guardians if
under 18 years of age. All participants gave oral consent.
Biochemical analyses
All blood samples were taken at the time of the ultrasound
examination, and stored at –80 °C until final analysis.
High-density lipoprotein (HDL-C), low-density lipoprotein
(LDL-C) and total cholesterol (TC) were analysed from lithium
heparin plasma by enzymatic method (Roche/Hitachi 912, Roche
Diagnostics, Mannheim, Germany). Plasma high-sensitivity
C-reactive protein (hsCRP) was measured from lithium heparin
plasma by enzyme-linked immunoassay using polyclonal
antibodies (DACO Diagnostics, Glostrup, Denmark). Plasma
fibrinogen was assessed by automated coagulation analyser
from sodium citrate plasma (Sysmex CA-7000, Sysmex
Corporation, Mundelein, IL, USA). Plasma tumour necrosis
factor-alpha (TNF-alpha) was detected using chemiluminescent
immunometric assay from serum (Immulite 1000 LKNF1,
Siemens Medical Solutions Diagnostics, Llanberis, UK). Plasma
cyclic guanosine monophosphate (cGMP) was measured by an
enzyme immunoassay (Amersham Pharmacia Biotech UK,
Buckinghamshire, UK) according to the manufacturer’s
recommendations [12]. In brief, 125 μL of EDTA plasma was
precipitated with 500 μL of ice cold ethanol. After centrifugation
of this suspension (4000 r.p.m. for 5 min), the supernatant was
removed and the remaining precipitate was washed with 500 μL
of ethanol, centrifuged and dried at 56 °C. The dried extract was
dissolved in 500 μL of assay buffer for analysis. The detection limit
was 0·02 pmol mL–1. For measurement of nitrite-nitrate, 200 μL
plasma samples were first deproteinized by adding 400 μL ZnSO4
and 500 μL NaOH. The samples were mixed and after 10 min
centrifuged at 1000 ×/g at 4 °C. Nitrate (NO3) in supernatants was
reduced to nitrite (NO2) with copper-coated granules of cadmium.
The concentration of nitrite was then determined after reaction
with a Greiss reagent using an Elx808 Bio-Tek microtitreplate
instrument reader [13]. A modified enzyme-linked
immunosorbent assay (ELISA) was used to determine antibodies
against oxidized LDL-C (oxLDL-C) in serum. The technique is
described in details elsewhere [14]. The data are expressed as
the ratio of binding to oxLDL-C to binding to native LDL-C.
Carotid artery ultrasound protocol
A high-resolution ultrasound system (Acuson Sequoia C256,
Siemens AG, Germany) equipped with a 15 MHz probe was
used. The imaging protocol was described in detail previously [15].
In short, longitudinal scans in bi-dimensional mode of the 1 cm
long distal end of the left common carotid artery were imaged
 INFECTION RECURRENCE AND ATHEROSCLEROSIS IN DIABETES
so that the lumen-intima and media–adventitia interfaces
were distinguishable. All images corresponded to the R-wave
on electrocardiogram (ECG). Four scans obtained from each
individual were recorded on videotape for offline analysis
of the carotid artery compliance (CAC), stiffness index (SI)
and intima-media thickness (IMT). The ultrasound scans were
taken by two sonographers unaware of the infection and smoke
exposure status. The mean carotid IMT was measured from each
scan manually. Mean IMT obtained from all scans from the same
subject were averaged and the resulting mean IMT was used for
statistical analyses.
CAC and SI were calculated according to the following
formulas: CAC = [(Ds – Dd)Dd]/(Ps – Pd), and SI = ln(Ps/Pd)/
[(Ds – Dd)/Dd], where Ds is systolic diameter, Dd is diastolic
diameter, Ps is systolic blood pressure, and Pd is diastolic blood
pressure. CAC reflects the ability of arteries to expand in
response to the pulse pressure caused by cardiac contraction
and relaxation. SI was introduced to reduce the impact of the
curvilinear pressure-stiffness relationship on arterial stiffness,
and is therefore considered to be relatively independent of blood
pressure [16].
In a prior reproducibility study on 10 patients, the coefficient
of interobserver variability (MO and AM) was 6·2%.
Statistical analysis
Differences in the studied variables were assessed by anova.
Eventual correlations between the hypothesised predictor
variables and the dependent variable were assessed by simple
regression. A multiple regression model was used to identify
independent factors affecting IMT, CAC and SI. hsCRP was
logarithmically transformed due to its skewed distribution. All
other numeric variables showed a Gaussian distribution. All data
are expressed as mean ± SE unless otherwise specified. Statistical
significance was accepted when P was less or equal to 0·05. All
statistical analyses were performed with StatView for Windows
(version 5·0, SAS Institute, USA).
Results
Infection, demographic and metabolic characteristics
The infection questionnaire was answered by 72 (41 male and 31
female) patients (73%). Fifteen patients experienced none or 1
episode of RTI during the past year (group 1), 33 patients had
2–3 RTI (group 2), while the remaining (24) patients had ≥ 4 RTI
(group 3). No other types of clinically manifest acute or chronic
infections were identified through the questionnaire.
There was a significant inverse correlation between the number
of RTI during the past year and the time elapsed from the last
infection (P = 0·003, r = –0·45). By anova there was however no
difference in this regard between those with a moderate or high
frequency of RTI (P = 0·70), but both groups differed significantly
from that with a low RTI recurrence (P < 0·01). There was no
significant difference between the infection groups in terms of
age, BMI, systolic and diastolic blood pressure, hsCRP, fibrinogen,
TNF-alpha, TC, HDL-C and LDL-C. HbA1C was lower in patients
with moderate (group 2) and high (group 3) frequency of RTI than
in those with low frequency (group 1, P < 0·01). Although it did
not reach statistical significance, plasma nitrate showed a stepwise
reduction with an increasing number of RTI (16·6 ± 1·7 in group
1, 14·9 ± 1·1 in group 2 and 13·0 ± 0·9 in group 3; group 3 vs.
group 1: P = 0·07). Overall, there was no association between
the frequency of RTI and oxLDL-C antibodies (P > 0·2). Patients’
demographic and metabolic characteristics in relation to the
number of infections are summarized in Table 1.
Fibrinogen (P = 0·002) but not hsCRP (P = 0·13) was higher
in girls than in boys. No other gender-related differences in the
inflammatory and metabolic variables were noted.
Effects of infection, demographic and metabolic
variables on carotid artery elasticity
The mean CAC of the study cohort was 2·80 ± 0·13%/10 mmHg
(2·69 ± 0·22 in males and 2·94 ± 0·13 in females, P = 0·34). In
univariate analysis, weak but significant inverse correlations
were observed between CAC and diabetes duration (P = 0·007,
r = –0·29), HbA1c (P = 0·004, r = –0·31), age (P = 0·004, r = –0·30),
and frequency of RTI (P = 0·01, r = –0·32; Fig. 1a). CAC was
decreased in patients with a high frequency of RTI compared to
those with low and moderate frequency (Fig. 1b). Although
diabetes duration was significantly greater in the third infection
tertile, the relationship between RTI and CAC remained significant
when diabetes duration along with other variables (age, BMI,
IMT, hsCRP and HbA1c) were entered in a multiple regression
analysis. In this model, in addition to RTI, HbA1c was also found
to independently predict the decrease in CAC (P = 0·01, r = –0·36).
Carotid SI was positively correlated with the number of RTI
frequency (P = 0·01, r = 0·31, Fig. 1c), being greater in the high
frequency group than in the remaining two groups (Fig. 1d).
The time between the last RTI and ultrasound had no impact on
the relationship between RTI recurrence and CAC or SI. Neither
plasma cGMP (P = 0·06; r = 0·22), nor nitrate (P = 0·13, r = 0·18)
showed a significant correlation with CAC.
Effects of infection, demographic and metabolic
variables on carotid artery intima-media thickness
There was no association between IMT and infection recurrence.
IMT positively correlated with diabetes duration (P = 0·01,
r = 0·27), BMI (P = 0·03, r = 0·22), LDL-C (P = 0·01, r = 0·28),
TC (P = 0·002, r = 0·35) and TC:HDL-C ratio (P = 0·05, r = 0·23).
There was no relationship between IMT and age, hsCRP,
fibrinogen or HbA1c levels. IMT correlated with TNF-alpha
in boys (P = 0·03, r = 0·34) but not in girls (P = 0·31). No
difference in IMT was observed between girls and boys (P = 0·32).
European Journal of Clinical Investigation Vol 38 383
 M. ODERMARSKY ET AL. www.ejci-online.com

Table 1 Patients demographic, biochemical and ultrasound characteristics in relation to the number of acute respiratory tract
infections (RTI) during the past year
P value
Groups based on 1 2 3
number of RTI during past year (0–1) (2–3) (≥ 4) 1 vs. 2 1 vs. 3
Number of patients (n) 15 33 24
Age (years) 15 ± 1 15 ± 1 15 ± 1 NS NS
Diabetes duration (years) 6·5 ± 0·9 5·7 ± 0·7 8·5 ± 1·0 NS NS
SBP (mmHg) 123 ± 6 122 ± 2 124 ± 4 NS NS
DBP (mmHg) 72 ± 4 69 ± 1 69 ± 2 NS NS
BMI 21·3 ± 1·1 21·3 ± 0·6 20·7 ± 0·7 NS NS
HbA1c (%) 7·8 ± 0·6 6·7 ± 0·2 6·9 ± 0·2 0·01 0·04
–1
hsCRP (mg L ) 1·5 ± 0·5 1·6 ± 0·6 1·1 ± 0·4 NS NS
–1
Fibrinogen (mg L ) 2·54 ± 0·11 2·64 ± 0·11 2·40 ± 0·08 NS NS
TC (mmol L–1) 4·44 ± 0·42 4·07 ± 0·13 4·33 ± 0·22 NS NS
–1
HDL-C (mmol L ) 1·71 ± 0·12 1·60 ± 0·07 1·62 ± 0·10 NS NS
–1
LDL-C (mmol L ) 2·43 ± 0·35 2·12 ± 0·14 2·30 ± 0·18 NS NS
TNF-alpha (ng L–1) 9·9 ± 0·8 10·3 ± 0·6 9·0 ± 0·4 NS NS
cGMP (pmol L–1) 20·8 ± 1·9 20·3 ± 0·9 21·0 ± 1·0 NS NS
–1
Nitrate (μmol L ) 16·6 ± 1·7 14·9 ± 1·1 13.0 ± 0.9 NS 0.07
oxLDL-C antibodies 1·94 ± 0·24 2·48 ± 0·28 2·19 ± 0·19 NS NS
*CAC (%/10 mmHg) 3·40 ± 0·64 2·94 ± 0·16 2·21 ± 0·19 NS 0·01
*SI 2·53 ± 0·14 2·48 ± 0·05 2·80 ± 0·09 NS 0·05
IMT (mm) 0·41 ± 0·02 0·38 ± 0·01 0·39 ± 0·01 NS NS

SBP, systolic blood pressure; DBP, diastolic blood pressure; BMI, body mass index; HbA1c , glycosylated haemoglobin; hsCRP, high sensitivity C-reactive protein;
TC, total cholesterol; HDL-C, high density lipoprotein cholesterol; LDL-C, low density lipoprotein cholesterol; TNF-alpha, tumour necrosis factor alpha; cGMP, cyclic
guanosine monophosphate; oxLDL-C antibodies, ratio of binding to oxLDL-C to binding to native LDL-C; CAC, carotid artery compliance; SI, carotid artery stiffness
index; IMT, carotid artery intima-media thickness. *n = 13, 27 and 22 in groups 1, 2, and 3, respectively. Data are expressed as mean ± SE.

Influences of smoke exposure on infection, carotid
atherosclerosis, and inflammatory and metabolic
variables
The questionnaire enquiring about ETS exposure was
answered by 93 patients (95% of study participants). Twenty
respondents (22%) indicated daily and weekly ETS exposure
(group 3), while rare exposure (group 2) was indicated by
44 patients (47%). Regarding household smoking, data on the
number of smokers per home and the average daily cigarette
consumption by patients’ cohabitants were provided by
93 patients. Non-smoking homes were indicated by
66 respondents (71%), while the remaining respondents
indicated 1 (16 respondents, 17%) and ≥ 2 smokers
per home (11 respondents, 12%). Parents’ social status
(expressed as number of years of education) was lower in
smokers than in non-smokers (P < 0·05). However, this variable
had no effect on the correlation between infection recurrence
and CAC (data not shown).
The number of RTI during the past year was higher in those
patients with daily to weekly exposure to ETS (P = 0·005).
There was also a trend toward increased recurrence of RTI
in those patients living in homes with ≥ 2 smokers compared
to those living in homes with no or 1 smoker (P = 0·09).
No direct association between the selected measures of smoke
exposure and indexes of carotid atherosclerosis (CAC, SI, IMT)
was observed (P > 0·2). However, patients with a moderate to high
prevalence of RTI (≥ 2/year) also frequently exposed to ETS had
lower CAC than those presenting either one or none of these
factors (P < 0·05; Fig. 2a; ancova after adjustment for age, diabetes
duration and HbA1C). Also, the relationship between CAC and RTI
recurrence became strongly significant (P = 0·0002, r = –0·56) in
patients with ETS exposure (2nd and 3rd tertiles) (Fig. 2b). In
patients with frequent ETS exposure (3rd tertile), but not in
the 1st (P = 0·61), and 2nd (P = 0·13) tertiles, plasma levels of
oxLDL-C antibodies were correlated with the frequency of RTI
(P = 0·04, r = 0·53). Plasma levels of cGMP were also significantly

384 © 2008 The Authors. Journal Compilation © 2008 Blackwell Publishing Ltd
 INFECTION RECURRENCE AND ATHEROSCLEROSIS IN DIABETES

Figure 1 Relationship between non-invasive

indices of carotid artery elasticity [carotid
artery compliance (CAC)]: (a) and (b), and
carotid artery stiffness index (SI): (c) and (d)
and frequency of respiratory tract infections
(RTI) during the past year (P = 0·01, r = –0·32
for CAC, and P = 0·01, r = 0·31 for SI). *denotes
P < 0·01, and **denotes P < 0·05 vs. the 1st and
3rd tertiles; n = 13 (0–1 infection), n = 27 (2–3
infections), n = 22 (≥ 4 infections). Data are
presented as mean ± SE.

Figure 2 (a) Interaction between frequency of respiratory tract infections (RTI) and exposure to environmental tobacco smoke (ETS)
on carotid artery compliance by ANCOVA (adjustment for age, diabetes duration and HbA1C). (b) Relationship between carotid artery
compliance (CAC) and frequency of respiratory tract infections (RTI) in patients with exposure to environmental tobacco smoke (ETS)
(P = 0·0002, r = –0·56). ‘RTI–’: < 2 RTI during the past year; ‘RTI+’: ≥ 2 RTI during the past year. ‘ETS+RTI+’: frequent (daily to weekly)
smoke exposure and 2 RTI during the past year. n = 12 in the 1st tertile; n = 32 in the 2nd tertile; n = 12 in the 3rd tertile.
*denotes P < 0·05. Data are presented as mean ± SE.

decreased in patients with both risk factors compared to those Discussion

presenting with either one or none (P < 0·05).
Exposure to ETS expressed as daily cigarette consumption
in and around the home was associated with LDL-C : HDL-C ratio
(P = 0·001, r = 0·36; Fig. 3). Plasma levels of both hsCRP and cGMP
showed weak, yet significant correlations with daily cigarette
consumption by patients’ cohabitants in or around the home
(P = 0·04, r = 0·22 and P = 0·009, r = –0·30, respectively). Neither
fibrinogen nor TNF-alpha were influenced by smoke (P > 0·3).
Nitrate showed no relationship with exposure to ETS (P = 0·5).
To our knowledge, the present study is the first to demonstrate
that in children and adolescents with type 1 diabetes, the number
of RTI during the past year is associated with a decrease in carotid
artery elasticity independent of other variables known to affect this
vascular index in the course of diabetes. The putative cumulative
stiffening effects of recurrent RTI seemed to be amplified in
patients with exposure to ETS. Given the important role of lipid
peroxidation in atherosclerosis [17], the significant correlation

European Journal of Clinical Investigation Vol 38 385

 M. ODERMARSKY ET AL.
Figure 3 Relationship between LDL-C:HDL-C ratio and exposure
to ETS expressed as daily cigarette consumption by cohabitants
in or around the home (P = 0·001, r = 0·36).
between RTI frequency and oxLDL-C antibodies in patients with
frequent ETS exposure is another finding that supports a synergy
of RTI frequency and passive smoking in arterial disease. Neither
the lipid nor the inflammatory indices differed in relation to the
number of RTI. LDL-C:HDL-C ratio, an established metabolic
index of atherosclerosis, rose with increasing levels of exposure
to ETS.
Various cardiovascular risk factors, including type 1 diabetes,
may cause arterial damage already in childhood, with potentially
long term consequences for atherosclerosis-related morbidity. Loss
of carotid artery elasticity was previously demonstrated in young
individuals with cardiovascular risk factors in childhood [18], and
was suggested to independently predict cardiovascular events in
high risk patients [19], supporting the view that arterial stiffening
progresses hand in hand with atherosclerosis. In type 1 diabetes,
the risk of developing accelerated atherosclerosis in the carotid
artery appears to be in part related to the duration of diabetes
and to the glycaemic level [20,21]. Using HbA1C as a measure of
glycaemic control, we observed similar trends for the carotid artery
elasticity in young diabetic patients. However, after adjustment for
potential confounders, only HbA1C and the frequency of RTI
remained associated with decreased carotid elasticity.
The exact role of infections in atherosclerosis remains in dispute.
In experimental studies, a plethora of pathogens has been
proposed to trigger or accelerate atherosclerosis through multiple
mechanisms including vascular endothelial damage,
dyslipidaemia, autoimmunity and endovascular infection [6].
In a few prior paediatric studies, detrimental effects on arterial
homeostasis were associated with clinically manifest acute
386 © 2008 The Authors. Journal Compilation © 2008 Blackwell Publishing Ltd
www.ejci-online.com
infections. Thus, impairment of the brachial artery’s endothelial
function and thickening of the carotid artery intima-media were
evidenced in otherwise healthy children but also in those with type
1 diabetes weeks to months after clinical recovery from acute
infections in the respiratory or urinary tract [22–24]. In these
studies, the magnitude of arterial damage observed after recovery
from the infectious illness was not related to the severity of the
illness, rendering conceivable the assumption that both mild and
severe infections are important in triggering vascular changes.
Flu-like infections have also been linked with an increased
incidence of major coronary events in middle-aged people [25].
During the past decade, it has become increasingly apparent
that many of the previously suggested influences of infection
on vasculature could be coupled to atherosclerosis if certain
conditions are met. Pathogen burden, infection recurrence and
coexistence with cardiovascular risk factors have been gradually
proposed as important prerequisites in order for this association
to take place [6]. Studies by Törmäkangas et al. showed that
repeated rather than single infections in mice worsened
atherosclerosis in the aorta through increased lipid accumulation
although, as in our study, the circulatory levels of cholesterol
remained unchanged [10]. Intimal accumulation of lipids with
subsequent enhanced lipid peroxidation contributes to stiffening
of the arterial wall in part via inflammatory and proliferative
changes initiated by oxidized lipoproteins [26]. Recently,
Mayr et al. showed in adults a rise in oxLDL-C antibodies with
increasing number of microbes known to cause chronic infection
[27]. Persistent co-infection with multiple pathogens predisposes
to frequent infectious relapses, and this mechanism might have
played a role in the observed association between pathogen
burden and lipid peroxidation. This hypothesis is in agreement
with the significant correlation observed in our study between
oxLDL-C antibodies and RTI frequency, even though this
association was restricted to patients regularly exposed to ETS.
In Mayr’s study, although the magnitude of atherosclerosis in the
carotid arteries increased with pathogen burden, the investigators
found no independent association between oxLDL-C antibodies
and carotid atherosclerosis, which could imply that the
infection-mediated pro-atherosclerotic effects involve multiple
pathways. Similarly, we found no association between oxLDL-C
antibodies and carotid artery elasticity.
In view of the multifactorial aetiology of atherosclerosis,
it is likely that the additional burden imposed by other vascular
risk factors is important in the development of vascular changes
following infection. This concept might have particular relevance
in type 1 diabetes. Diabetic hyperglycaemia seems to predispose
to increased recurrence of both bacterial and viral infection by
lowering the efficacy of the cell-mediated immune response
[10]. Type 1 diabetes is associated with multiple metabolic,
inflammatory and immunological abnormalities that render
possible a number of interactions at a vascular level between
 INFECTION RECURRENCE AND ATHEROSCLEROSIS IN DIABETES
both acute and chronic conditions associated with diabetes. Iwata
et al. showed that high glucose levels in vitro stimulate production
of inflammatory cytokines in lipopolysaccharide-stimulated
monocytes from diabetes patients [28]. In another study,
subcutaneous inoculation of lipopolysaccharide in diabetic mice
caused a more rapid and pronounced up-regulation of adhesion
molecules, cytokines and chemokines than in non-diabetic mice
[29].
Juonala et al. found that the number of childhood risk factors
predicts the decrease in carotid artery elasticity in middle-aged
adults [18]. ETS exerts adverse vascular effects nearly as much
as active smoking [30], and a previous prospective study
demonstrated an additive interaction between both active and
passive smoking and chronic infections on plaque progression
[31]. In healthy children and young adults, ETS exposure
correlated with the magnitude of atherogenic changes in
peripheral arteries [32,33]. Although we could not observe
any direct influence of smoke exposure on indices of carotid
atherosclerosis in the cohort of type 1 diabetics, the decrease
in CAC in patients with frequent RTI was greatest in those with
frequent exposure to ETS, suggesting a possible synergy of these
two in the early development of arterial disease. The decline in
CAC in patients with both frequent RTI and ETS exposure was
paralleled by a decrease in plasma cGMP. Previous studies have
shown an inhibitory effect of both infection [34] and smoke
exposure [35] on the vascular endothelial nitric oxide (NO)
pathway. cGMP is an intracellular messenger of NO-mediated
relaxation of vascular smooth muscle layer, and its plasma level
correlates with endothelial function [12]. Our finding could
therefore suggest that ETS and infection cause a synergistic
inhibition on the endothelial mechanisms involving NO.
This hypothesis remains to be confirmed in future studies.
ETS could also affect the immune system, especially in
the respiratory tract, due to the inhaled noxious substances
in the tobacco smoke. This could in part explain the increased
susceptibility of smoke exposed children to infectious airway
diseases [36]. A similar finding was observed in the present
study. Possible mechanisms might involve increased microbial
adherence, impaired lung and nasal clearance, and changes in
the immune response [37].
In the present study, tobacco smoke exposure expressed as
daily cigarette consumption in or around the home adversely
influenced lipid levels by lowering HDL cholesterol levels and
increasing LDL cholesterol levels. Increased oxidant burden and
release of pro-inflammatory cytokines, which have been
associated with passive smoking [38] could be possible
underlying mechanisms.
Study limitations: The size of the studied groups in relation
to infectious recurrence and smoke exposure is relatively small.
Assessment of infection history through a questionnaire might
not be highly accurate in identifying all infections an individual
is exposed to. Smoke exposure assessment through a questionnaire
might not closely reflect the daily life situation. However, earlier
studies using an identical questionnaire suggested good sensitivity
to discriminate between children’s ETS exposure levels by
demonstrating a significant correlation with biomarkers for smoke
exposure [39].
In conclusion, recurrent respiratory tract infections seem to pose
cumulative deleterious effects on the arterial elasticity in diabetic
children, particularly in those with frequent exposure to ETS.
Further prospective studies are warranted to verify whether
such effects might in time translate into adverse changes in
arterial structure.
Address
Paediatric Cardiology, Department of Paediatrics, Lund
University Hospital, Lund, Sweden (M. Odermarsky, E. Pesonen,
P. Liuba), Department of Paediatrics, Helsinki University Hospital,
Finland (S. Andersson), Paediatric Endocrinology, Department of
Paediatrics, Lund University Hospital, Lund, Sweden (S. Sjöblad)
A. I. Virtanen Institute, University of Kuopio, Kuopio, Finland
(S. Ylä-Herttuala).
Correspondence to: Petru Liuba MD, PhD, Division of Paediatric
Cardiology, Lund University Hospital, 221 85 Lund, Sweden.
Tel.: +46 46 17 82 67; fax: +46 46 17 81 50; e-mail:
petru.liuba@med.lu.se
Received 13 July 2007; accepted 17 March 2008
Acknowledgements
We thank Ms. Annica Maxedius (AM), research nurse, for the
excellent technical assistance in blood sampling and carotid artery
ultrasound. The study was supported by a clinical scientist award
(PL) from FAMRI, USA. Additional funding was received from
Lund University (PL).
References
1 Pyorala K, Laaksi M, Uusitupa M. Diabetes and atherosclerosis:
an epidemiological view. Diabetes Metab Rev 1987;3:463–524.
2 Krantz JS, Mack WJ, Hodis HN, Liu CR, Liu CH, Kaufman FR.
Early onset of subclinical atherosclerosis in young persons with
type 1diabetes. J Pediatr 2004;145:452–7.
3 Jarvisalo MJ, Jartti L, Nanto-Salonen K, Irjala K, Ronnemaa T,
Hartiala JJ et al. Increased aortic intima-media thickness: a marker
of preclinical atherosclerosis in high-risk children. Circulation
2001;104:2943–7.
4 Krebs A, Schmidt-Trucksass A, Wagner J, Krebs K, Doerfer J,
Schwab KO. Adult-like but regressive increase of intima-media
thickness and roughness in a child with type 1 diabetes.
Pediatr Diabetes 2005;6:161–4.
5 Selvin E, Coresh J, Golden SH, Brancati FL, Folsom AR, Steffes MW.
Glycemic control and coronary heart disease risk in persons with and
without diabetes: the atherosclerosis risk in communities study. Arch
Intern Med 2005;165:1910–6.
6 O’Connor S, Taylor C, Campbell LA, Epstein S, Libby P. Potential
infectious etiologies of atherosclerosis: a multifactorial perspective.
Emerg Infect Dis 2001;7:780–8.
European Journal of Clinical Investigation Vol 38 387
 M. ODERMARSKY ET AL.
7 Joshi N, Caputo GM, Weitekamp MR, Karchmer AW. Infections in
patients with diabetes mellitus. N Engl J Med 1999;341:1906–12.
8 Ojetti V, Pitocco D, Bartozzoli F, Danese S, Migneco A, Lupascu A
et al. High rate of Helicobacter pylori re-infection in patients affected
by type 1 diabetes (Letter). Diabetes Care 2002;25:1485.
9 Larsson ML, Loit HM, Meren M, Polluste J, Magnusson A,
Larsson K et al. Passive smoking and respiratory symptoms in the
FinEsS Study. Eur Respir J 2003;21:672–6.
10 Tormakangas L, Erkkila L, Korhonen T, Tiirola T, Bloigu A, Saikku
P et al. Effects of repeated Chlamydia pneumoniae inoculations on
aortic lipid accumulation and inflammatory response in C57BL/6J
mice. Infect Immun 2005;73:6458–66.
11 Liuba P, Karnani P, Pesonen E, Paakkari I, Forslid A, Johansson L
et al. Endothelial dysfunction after repeated Chlamydia pneumoniae
infection in apolipoprotein E-knockout mice. Circulation
2000;102:1039–44.
12 Turanlahti M, Pesonen E, Pohjavuori M, Lassus P, Fyhrquist F,
Andersson S. Plasma cyclic guanosine monophosphate reflecting
the severity of persistent pulmonary hypertension of the newborn.
Biol Neonate 2001;80:107–12.
13 Green LC. Wagner DA, Glogowski J, Skipper PL, Wishnok JS,
Tannenbaum SR. Analysis of nitrate, nitrite, and [15N] nitrate in
biological fluids. Anal Biochem 1982;126:131–8.
14 Mäkimattila S, Luoma JS, Ylä-Herttuala S, Bergholm R, Utriainen T,
Virkamäki A et al. Autoantibodies against oxidized LDL and
endothelium-dependent vasodilation in insulin-dependent diabetes
mellitus. Atherosclerosis 1999;147:115–22.
15 Persson J, Stavenow L, Wikstrand J, Israelsson B, Formgren J,
Berglund G. Noninvasive quantification of atherosclerosis.
Reproducibility of ultrasound measurement of arterial wall thickness
and plaque size. Arterioscler Thromb 1992;12:261–6.
16 Salomaa V, Riley W, Kark JD, Nardo C, Folsom AR. Non-insulin
dependent diabetes mellitus and fasting glucose and insulin
concentrations are associated with arterial stiffness indexes. The
ARIC Study. Atherosclerosis Risk in Communities Study.
Circulation 1995;91:1432–43.
17 Stocker R, Keaney JF Jr. Role of oxidative modifications in
atherosclerosis. Physiol Rev 2004;84:1381–478.
18 Jounala M, Järvisalo MJ, Mäki-Torkko N, Kähönen M, Viikari JSA,
Raitakari OT. Risk factors identified in childhood and decreased
carotid artery elasticity in adulthood: the Cardiovascular Risk in
Young Finns Study. Circulation 2005;112:1486–93.
19 Blacher J, Pannier B, Guerin AP, Marchais SJ, Safar ME, London GM.
Carotid arterial stiffness as a predictor of cardiovascular and
all-cause mortality in end stage renal disease. Hypertension
1998;32:570–4.
20 Jarvisalo MJ, Raitakari M, Toikka JO, Putto-Laurila A, Rontu R,
Laine S et al. Endothelial dysfunction and increased arterial
intima-media thickness in children with type 1 diabetes.
Circulation 2004;109:1750–5.
21 Jensen-Urstad KJ, Reichard PG, Rosfors JS, Lindblad LE, Jensen-
Urstad MT. Early atherosclerosis is retarded by improved
long-term blood glucose control in patients with IDDM. Diabetes
1996;45:1253–8.
22 Charakida M, Donald AE, Terese M, Leary S, Halcox JP, Ness A et al.
ALSPAC (Avon Longitudinal Study of Parents and Children) Study
Team. Endothelial dysfunction in childhood infection. Circulation
2005;111:1660–5.
23 Liuba P, Persson J, Luoma J, Ylä-Herttualla S, Pesonen E. Acute
388 © 2008 The Authors. Journal Compilation © 2008 Blackwell Publishing Ltd
www.ejci-online.com
infections in children are accompanied by oxidative modification of
LDL and decrease of HDL cholesterol, and are followed by
thickening of carotid intima-media. Eur Heart J 2003;24:515–21.
24 Aburawi EH, Liuba P, Pesonen E, Ylä-Herttuala S, Sjöblad S. Acute
respiratory viral infections aggravate arterial endothelial dysfunction
in children with type 1 diabetes. Diabetes Care 2004;27:2733–5.
25 Meier CR, Jick SS, Derby LE, Vasilakis C, Jick H. Acute respiratory-tract
infections and risk of first-time acute myocardial infarction. Lancet
1998;351:1467–71.
26 Kita T, Kume N, Minami M, Hayashida K, Murayama T, Sano H et al.
Role of oxidized LDL in atherosclerosis. Ann N Y Acad Sci
2001;947:199–205.
27 Mayr M, Kiechl S, Tsimikas S, Miller E, Sheldon J, Willeit J et al.
Oxidized low-density lipoprotein autoantibodies, chronic infections,
and carotid atherosclerosis in a population-based study. J Am Coll
Cardiol 2006;47:2436–43.
28 Iwata H, Soga Y, Meguro M, Yoshizawa S, Okada Y, Iwamoto Y et al.
High glucose up-regulates lipopolysaccharide-stimulated
inflammatory cytokine production via c-jun N-terminal kinase in
the monocytic cell line THP-1. J Endotoxin Res 2007;13:227–34.
29 Lu H, Raptis M, Black E, Stan M, Amar S, Graves DT. Influence of
diabetes on the exacerbation of an inflammatory response in
cardiovascular tissue. Endocrinology 2004;145:4934–9.
30 Barnoya J, Glantz SA. Cardiovascular effects of second hand smoke:
nearly as large as smoking. Circulation 2005;111:2684–98.
31 Kiechl S, Werner P, Egger G, Oberhollenzer F, Mayr M, Xu Q et al.
Active and passive smoking, chronic infections, and the risk of
carotid atherosclerosis: prospective results from the Bruneck Study.
Stroke 2002;33:2170–6.
32 Celermajer DS, Sorensen KE, Georgakopoulos D, Bull C, Thomas O,
Robinson J et al. Cigarette smoking is associated with dose-related
and potentially reversible impairment of endothelium-dependent
dilation in healthy young adults. Circulation 1993;88:2149–55.
33 Kallio K, Jokinen E, Raitakari OT, Hämäläinen M, Siltala M, Volanen
I et al. Tobacco smoke exposure is associated with attenuated
endothelial function in 11-year-old healthy children. Circulation
2007;115:3205–12.
34 Bouwman JJ, Visseren FL, Bevers LM, van der Vlist WE, Bouter KP,
Diepersloot RJ. Azithromycin reduces Chlamydia pneumoniae-
induced attenuation of eNOS and cGMP production by endothelial
cells. Eur J Clin Invest 2005;35:573–82.
35 Rahman MM, Laher I. Structural and functional alteration of blood
vessels caused by cigarette smoking: an overview of molecular
mechanisms. Curr Vasc Pharmacol 2007;5:276–92.
36 Robbins CS. Dawe DE, Goncharova SI, Pouladi MA, Drannik AG,
Swirski FK et al. Cigarette smoke decreases pulmonary dendritic cells
and impacts antiviral immune responsiveness. Am J Respir Cell Mol
Biol 2004;30:202–11.
37 DiFranza JR, Aligne CA, Weitzman M. Prenatal and postnatal
environmental tobacco smoke exposure and children’s health.
Pediatrics 2004;113:1007–15.
38 Zhang J, Liu Y, Shi J, Larson DF, Watson RR. Side-stream
cigarette smoke induces dose–response in systemic
inflammatory cytokine production and oxidative stress.
Exp Biol Med 2002;227:823–9.
39 Johansson A, Halling A, Hermansson G, Ludvigsson J. Assessment
of passive smoking behaviour in the home and their influence on
children’s passive smoking: development of questionnaire. Ann
Epidemiol 2005;15:453–9.