bluuebunny/crossref_metadata_2025_split huggingface dataset:
Beta and Angiotensin Blockades Are Associated With Improved 10‐Year Survival in Renal Transplant Recipients
Authors: Aftab Waqas , Varadarajan Padmini , Rasool Shuja , Kore Arputharaj , Pai Ramdas G. 
Published: May 2013

Background
Mortality in allograft kidney transplant recipients is high, and cardiovascular disease is the leading cause of death in these patients. They have heightened activity of sympathetic and renin–angiotensin systems. We tested the hypothesis that blockade of sympathetic and renin–angiotensin systems in these patients may offer a survival benefit using a large cohort of patients with long‐term follow up.

Methods and Results

Medical records of 321 consecutive patients from our institution who had received renal transplantation between 1995 and 2003 were abstracted. Survival was analyzed as a function of pharmacological therapies adjusted for age, sex, and comorbidities. The characteristics of the 321 patients were as follows: age at transplant, 44±13 years; 40% male; 89% with hypertension; 36% with diabetes, and mean left ventricular ejection fraction of 60%. Over a follow‐up of 10±4 years, there were 119 deaths. Adjusted for age, sex, diabetes, and coronary artery disease, use of a beta‐blocker therapy (
P
=0.04) and angiotensin‐converting enzyme inhibitor or receptor blocker (
P
=0.03) was associated with better survival. This treatment effect was seen across all major clinical subgroups and was supported by propensity score analysis. The propensity score–adjusted 10‐year survival was 95% in those taking both groups of medications, 72% in those taking either of them, and 64% in those taking neither (
P
=0.004).

Conclusions
Use of beta‐blocker and angiotensin blocking therapies is associated with higher survival after renal transplantation, indicating their potential protective role in this high‐risk population.
Periaortic Adipose Tissue and Aortic Dimensions in the Framingham Heart Study
Authors: Thanassoulis George , Massaro Joseph M. , Corsini Erin , Rogers Ian , Schlett Christopher L. , Meigs James B. , Hoffmann Udo , O'Donnell Christopher J. , Fox Caroline S. 
Published: May 2013

Background
Periaortic fat, because of its contiguity with the aorta, may promote vascular remodeling and aortic dilatation. However, the relations between perioartic fat depots and aortic dimensions have not been previously described.

Methods and Results

A total of 3001 individuals (mean age 50±10 years, 49% women) from the
F
ramingham
O
ffspring and
T
hird
G
eneration cohorts underwent computed tomography for quantification of periaortic fat and aortic dimensions. We estimated the association between quantitative periaortic and visceral adipose tissue volumes (per standard deviation [
SD
] increment of volume) with aortic dimensions in both the thorax and abdomen. Thoracic periaortic fat was associated with higher thoracic aortic dimensions (β coefficient per SD of fat volume 0.67 mm, 95% confidence interval 0.58 to 0.76 mm;
P
<0.001). The association persisted after adjustment for age, sex, and cardiovascular risk factors including body mass index and visceral adipose tissue volume. Results for the association of periaortic fat and abdominal aortic dimensions were similar. Further adjustment for adipokines (resistin and adiponectin) had no significant impact on these associations.

Conclusions
Periaortic fat volume was associated with aortic dimensions in both the thorax and abdomen, supporting the notion that local fat depots may contribute to aortic remodeling. Further work to understand the mechanisms underlying this association is warranted.
Prospective Relationship of Low Cardiovascular Risk Factor Profile at Younger Ages to Ankle‐Brachial Index: 39‐Year Follow‐Up—The Chicago Healthy Aging Study
Authors: Vu Thanh‐Huyen T. , Stamler Jeremiah , Liu Kiang , McDermott Mary M. , Lloyd‐Jones Donald M. , Pirzada Amber , Garside Daniel B. , Daviglus Martha L. 
Published: May 2013

Background

Data are sparse regarding the long‐term association of favorable levels of all major cardiovascular disease risk factors (RFs) (ie, low risk [
LR
]) with ankle‐brachial index (
ABI
).

Methods and Results

In 2007–2010, the
C
hicago
H
ealthy
A
ging
S
tudy reexamined a subset of participants aged 65 to 84 years from the
C
hicago
H
eart
A
ssociation
D
etection
P
roject in
I
ndustry cohort (baseline examination, 1967–1973).
RF
groups were defined as
LR
(untreated blood pressure ≤120/≤80 mm Hg, untreated serum cholesterol <200 mg/dL, body mass index <25 kg/m
2
, not smoking, no diabetes) or as 0
RF
s, 1
RF
, or 2+
RF
s based on the presence of blood pressure ≥140/≥90 mm Hg or receiving treatment, serum cholesterol ≥240 mg/dL or receiving treatment, body mass index ≥30 kg/m
2
, smoking, or diabetes.
ABI
at follow‐up was categorized as indicating
PAD
present (≤0.90), as borderline
PAD
(0.91 to 0.99), or as normal (1.00 to 1.40). We included 1346 participants with
ABI
≤1.40. After multivariable adjustment, the presence of fewer baseline
RF
s was associated with a lower likelihood of
PAD
at 39‐year follow‐up (
P
for trend is <0.001). Odds ratios (95% CIs) for
PAD
in persons with
LR
, 0
RF
s, or 1
RF
compared with those with 2+
RF
s were 0.14 (0.05 to 0.44), 0.28 (0.13 to 0.59), and 0.33 (0.16 to 0.65), respectively; findings were similar for borderline
PAD
(
P
for trend is 0.005). The association was mainly due to baseline smoking status, cholesterol, and diabetes. Remaining free of adverse
RF
s or improving
RF
status over time was also associated with
PAD
.

Conclusions

LR
profile in younger adulthood (ages 25 to 45) is associated with the lowest prevalence of
PAD
and borderline
PAD
39 years later.
Relation Between Serum Total Cholesterol Level and Cardiovascular Disease Stratified by Sex and Age Group: A Pooled Analysis of 65 594 Individuals From 10 Cohort Studies in Japan
Authors: Nagasawa Sin‐ya , Okamura Tomonori , Iso Hiroyasu , Tamakoshi Akiko , Yamada Michiko , Watanabe Makoto , Murakami Yoshitaka , Miura Katsuyuki , Ueshima Hirotsugu ,   for the Evidence for Cardiovascular Prevention from Observational Cohorts in Japan (EPOCH‐JAPAN) Research Group
Published: May 2013

Background

The relation between serum total cholesterol (
TC
) and cardiovascular disease in women and in the elderly is unclear, especially in
A
sian populations.

Methods and Results

We examined this relation in the largest‐scale pooled analysis of the
J
apanese population, the Evidence for Cardiovascular Prevention from Observational Cohorts in Japan (
EPOCH
‐
JAPAN
) study. A total of 65 594 participants who were 40 to 89 years of age and did not have a past history of cardiovascular disease were examined.
C
ox proportional‐hazards models were used to estimate hazard ratios for death from total stroke, cerebral infarction, intracranial cerebral hemorrhage, or coronary heart disease. The mean follow‐up period was 10.1 years, with the number of deaths from total stroke, cerebral infarction, cerebral hemorrhage, and coronary heart disease being 875, 457, 212, and 374, respectively. The participants were divided into 2 age groups: middle‐aged (40 to 69 years; mean age 55 years) and elderly (70 to 89 years; mean age 75 years). In men, the multivariate‐adjusted hazard ratios for coronary heart disease in the highest
TC
category (≥6.21 mmol/L) compared with the lowest category (<4.14 mmol/L) were 2.52 (95% confidence interval: 1.15–5.07) in middle‐aged participants and 2.77 (1.09–7.03) in elderly participants. In women, the hazard ratios of the highest
TC
category (≥6.72 mmol/L) compared with the lowest category (<4.66 mmol/L) were 3.20 (1.44–7.09) in middle‐aged participants and 1.02 (0.42–2.49) in elderly participants.
TC
levels were not associated with cerebral infarction in any age or sex group and were associated negatively with total stroke and cerebral hemorrhage.

Conclusion

High serum
TC
levels are associated with coronary heart disease in middle‐aged
J
apanese men and women, but evidence in elderly
J
apanese individuals is still limited.
Microvascular Coronary Artery Spasm Presents Distinctive Clinical Features With Endothelial Dysfunction as Nonobstructive Coronary Artery Disease
Authors: Ohba Keisuke , Sugiyama Seigo , Sumida Hitoshi , Nozaki Toshimitsu , Matsubara Junichi , Matsuzawa Yasushi , Konishi Masaaki , Akiyama Eiichi , Kurokawa Hirofumi , Maeda Hirofumi , Sugamura Koichi , Nagayoshi Yasuhiro , Morihisa Kenji , Sakamoto Kenji , Tsujita Kenichi , Yamamoto Eiichiro , Yamamuro Megumi , Kojima Sunao , Kaikita Koichi , Tayama Shinji , Hokimoto Seiji , Matsui Kunihiko , Sakamoto Tomohiro , Ogawa Hisao 
Published: May 2013

Background

Angina without significant stenosis, or nonobstructive coronary artery disease, attracts clinical attention. Microvascular coronary artery spasm (microvascular
CAS
) can cause nonobstructive coronary artery disease. We investigated the clinical features of microvascular
CAS
and the therapeutic efficacy of calcium channel blockers.

Methods and Results

Three hundred seventy consecutive, stable patients with suspected angina presenting nonobstructive coronary arteries (<50% diameter) in coronary angiography were investigated with the intracoronary acetylcholine provocation test, with simultaneous measurements of transcardiac lactate production and of changes in the quantitative coronary blood flow. We diagnosed microvascular
CAS
according to lactate production and a decrease in coronary blood flow without epicardial vasospasm during the acetylcholine provocation test. We prospectively followed up the patients with calcium channel blockers for microvascular coronary artery disease. We identified 50 patients with microvascular
CAS
who demonstrated significant impairment of the endothelium‐dependent vascular response, which was assessed by coronary blood flow during the acetylcholine provocation test. Administration of isosorbide dinitrate normalized the abnormal coronary flow pattern in the patients with microvascular
CAS
. Multivariate logistic regression analysis indicated that female sex, a lower body mass index, minor–borderline ischemic electrocardiogram findings at rest, limited–baseline diastolic‐to‐systolic velocity ratio, and attenuated adenosine triphosphate–induced coronary flow reserve were independently correlated with the presence of microvascular
CAS
. Receiver‐operating characteristics curve analysis revealed that the aforementioned 5‐variable model showed good correlation with the presence of microvascular
CAS
(area under the curve: 0.820). No patients with microvascular
CAS
treated with calcium channel blockers developed cardiovascular events over 47.8±27.5 months.

Conclusions

Microvascular
CAS
causes distinctive clinical features and endothelial dysfunction that are important to recognize as nonobstructive coronary artery disease so that optimal care with calcium channel blockers can be provided.

Clinical Trial Registration

URL:
www.umin.ac.jp/ctr
. Unique identifier: UMIN000003839.
Premature Cardiac Contractions and Risk of Incident Ischemic Stroke
Authors: Ofoma Uchenna , He Fan , Shaffer Michele L. , Naccarelli Gerard V. , Liao Duanping 
Published: May 2013

Background

The etiologies of ischemic stroke remain undetermined in 15% to 40% of patients. Apart from atrial fibrillation, other arrhythmias are less well‐characterized as risk factors. Premature cardiac contractions are known to confer long‐term cardiovascular risks, like myocardial infarction. Ischemic stroke as cardiovascular risk outcome remains a topic of interest. We examined the prospective relationships in the Atherosclerosis Risk in Communities (
ARIC
) study, to determine whether premature atrial (
PAC
) or ventricular (
PVC
) contractions are associated with increased risk for incident ischemic stroke.

Methods and Results

We analyzed 14 493 baseline stroke‐free middle‐aged individuals in the
ARIC
public‐use data. The presence of PAC or PVC at baseline was assessed from 2‐minute electrocardiogram. A physician‐panel confirmed and classified all stroke cases. Average follow‐up time was 13 years. Proportional hazards models assessed associations between premature contractions and incident stroke.
PAC
s and
PVC
s were identified in 717 (4.9%) and 793 (5.5%) participants, respectively. In all, 509(3.5%) participants developed ischemic stroke. The hazard ratio (
HR
) (95% confidence interval [
CI
]) associated with
PVC
was 1.77 (1.30, 2.41), attenuated to 1.25 (0.91, 1.71) after adjusting for baseline stroke risk factors. The interaction between
PVC
and baseline hypertension was marginally significant (
P
=0.08). Among normotensives, having
PVC
s was associated with nearly 2‐fold increase in the rate of incident ischemic stroke (
HR
1.69; 95%
CI
1.02, 2.78), adjusting for stroke risk factors. The adjusted risk of ischemic stroke associated with
PAC
s was 1.30 (95%
CI
0.92, 1.83).

Conclusions

Presence of
PVC
s may indicate an increased risk of ischemic stroke, especially in normotensives. This risk approximates risk of stroke from being black, male, or obese in normotensives from this cohort.
Growth Differentiation Factor‐15 Deficiency Inhibits Atherosclerosis Progression by Regulating Interleukin‐6–Dependent Inflammatory Response to Vascular Injury
Authors: Bonaterra Gabriel A. , Zügel Stefanie , Thogersen Joel , Walter Sabrina A. , Haberkorn Uwe , Strelau Jens , Kinscherf Ralf 
Published: May 2013

Background

Growth differentiation factor (
GDF
)‐15 is a distant and divergent member of the transforming growth factor‐β superfamily (TGF‐β) . There is growing evidence indicating the involvement of
GDF
‐15 in various pathologies. Expression of
GDF
‐15 is induced under conditions of inflammation and increased
GDF
‐15 serum levels are suggested as a risk factor for cardiovascular diseases.

Methods and Results

We show here that
GDF
‐15 and proinflammatory cytokine interleukin (
IL
)‐6 levels are highly increased (5‐fold) in cultured oxidized low‐density lipoproteins–stimulated peritoneal macrophages derived from
GDF
‐15
+/+
/apolipoprotein (apo)
E
−/−
, mice. Notably,
IL
‐6 induction on oxidized low‐density lipoproteins stimulation is completely abolished in the absence of
GDF
‐15. Consistent with our in vitro data
GDF
‐15
mRNA
expression and protein levels are upregulated (2.5‐ to 6‐fold) in the atherosclerotic vessel wall of
GDF
‐15
+/+
/apo
E
−/−
mice after a cholesterol‐enriched diet.
GDF
‐15 deficiency inhibits lumen stenosis (52%) and
18
FDG
uptake (34%) in the aortic arch despite increased serum triglyceride/cholesterol levels and elevated body weight. Immunohistomorphometric investigations of atherosclerotic lesions reveal a decreased percentage of inflammatory
CD
11b
+
(57%) or
IL
‐6
+
, leukocytes, and apoptotic cells (74%) after 20 weeks. However, the total number of macrophages and cell density in atherosclerotic lesions of the innominate artery are increased in
GDF
‐15
−/−
/apo
E
−/−
mice.

Conclusions

Our data suggest that
GDF
‐15 is involved in orchestrating atherosclerotic lesion progression by regulating apoptotic cell death and
IL
‐6–dependent inflammatory responses to vascular injury.
Relation of Carotid Artery Diameter With Cardiac Geometry and Mechanics in Heart Failure With Preserved Ejection Fraction
Authors: Liao Zhen‐Yu , Peng Ming‐Cheng , Yun Chun‐Ho , Lai Yau‐Huei , Po Helen L. , Hou Charles Jia‐Yin , Kuo Jen‐Yuan , Hung Chung‐Lieh , Wu Yih‐Jer , Bulwer Bernard E. , Yeh Hung‐I , Tsai Cheng‐Ho 
Published: May 2013

Background
Central artery dilation and remodeling are associated with higher heart failure and cardiovascular risks. However, data regarding carotid artery diameter from hypertension to heart failure have remained elusive. We sought to investigate this issue by examining the association between carotid artery diameter and surrogates of ventricular dysfunction.

Methods and Results

Two hundred thirteen consecutive patients including 49 with heart failure and preserved ejection fraction (
HFpEF
), 116 with hypertension, and an additional 48 healthy participants underwent comprehensive echocardiography and tissue
D
oppler imaging. Ultrasonography of the common carotid arteries was performed for measurement of intima‐media thickness and diameter (
CCAD
). Cardiac mechanics, including
LV
twist, were assessed by novel speckle‐tracking software. A substantial graded enlargement of
CCAD
was observed across all 3 groups (6.8±0.6, 7.7±0.73, and 8.7±0.95 mm for normal, hypertension, and
HFpEF
groups, respectively;
ANOVA
P
<0.001) and correlated with serum brain natriuretic peptide level (
R
2
=0.31,
P
<0.001). Multivariable models showed that
CCAD
was associated with increased
LV
mass,
LV
mass‐to‐volume ratio (β‐coefficient=10.9 and 0.11, both
P
<0.001), reduced
LV
longitudinal and radial strain (β‐coeffficient=0.81 and −3.1, both
P
<0.05), and twist (β‐coefficient=−0.84,
P
<0.05).
CCAD
set at 8.07 mm as a cut‐off had a 77.6% sensitivity, 82.3% specificity, and area under the receiver operating characteristic curves (
AUROC
) of 0.86 (95% CI 0.80 to 0.92) in discriminating
HFpEF
. In addition,
CCAD
superimposed on myocardial deformation significantly expanded
AUROC
(for longitudinal strain, from 0.84 to 0.90,
P
of Δ
AUROC
=0.02) in heart failure discrimination models.

Conclusions

Increased carotid artery diameter is associated with worse LV geometry, higher brain natriuretic peptide level, and reduced contractile mechanics in individuals with
HFpEF
.
Dipeptidyl Peptidase‐4 Inhibitors Attenuate Endothelial Function as Evaluated by Flow‐Mediated Vasodilatation in Type 2 Diabetic Patients
Authors: Ayaori Makoto , Iwakami Naotsugu , Uto‐Kondo Harumi , Sato Hiroki , Sasaki Makoto , Komatsu Tomohiro , Iizuka Maki , Takiguchi Shunichi , Yakushiji Emi , Nakaya Kazuhiro , Yogo Makiko , Ogura Masatsune , Takase Bonpei , Murakami Takehiko , Ikewaki Katsunori 
Published: May 2013

Background

Endothelial dysfunction is an independent predictor for cardiovascular events in patients with type 2 diabetes (
T2DM
). Glucagon like peptide‐1 (
GLP
‐1) reportedly exerts vasodilatory actions, and inhibitors of dipeptidyl peptidase‐4 (
DPP
‐4), an enzyme‐degrading
GLP
‐1, are widely used to treat
T2DM
. We therefore hypothesized that
DPP
‐4 inhibitors (
DPP
‐4Is) improve endothelial function in
T2DM
patients and performed 2 prospective, randomized crossover trials to compare the
DPP
‐4I sitagliptin and an α‐glucosidase inhibitor, voglibose (in study 1) and the
DPP
‐4Is sitagliptin and alogliptin (in study 2).

Methods and Results

In study 1, 24 men with
T2DM
(46±5 years) were randomized to sitagliptin or voglibose for 6 weeks without washout periods. Surprisingly, sitagliptin significantly reduced flow‐mediated vasodilatation (
FMD
; −51% compared with baseline,
P
<0.05) of the brachial artery despite improved diabetic status. In contrast, voglibose did not affect
FMD
. To confirm this result and determine whether it is a class effect, we conducted another trial (study 2) to compare sitagliptin and alogliptin in 42
T2DM
patients (66±8 years) for 6 weeks with 4‐week washout periods. Both
DPP
‐4Is improved glycemic control but significantly attenuated
FMD
(7.2/4.3%,
P
<0.001, before/after sitagliptin; 7.0/4.8%,
P
<0.001, before/after alogliptin, respectively). Interestingly,
FMD
reduction was less evident in subjects who were on statins or whose
LDL
cholesterol levels were reduced by them, but this was not correlated with parameters including
DPP
‐4 activity and
GLP
‐1 levels or diabetic parameters.

Conclusions

Our 2 independent trials demonstrated that
DPP
‐4 inhibition attenuated endothelial function as evaluated by
FMD
in
T2DM
patients. This unexpected unfavorable effect may be a class effect of
DPP
‐4Is.

Clinical Trial Registration

URL:
http://center.umin.ac.jp
, Unique Identifiers: UMIN000005682 (sitagliptin versus voglibose) and UMIN000005681 (sitagliptin versus alogliptin).
Detection of Aortic Arch Calcification in Apolipoprotein E‐Null Mice Using Carbon Nanotube–Based Micro‐CT System
Authors: Wait John M. S. , Tomita Hirofumi , Burk Laurel M. , Lu Jianping , Zhou Otto Z. , Maeda Nobuyo , Lee Yueh Z. 
Published: May 2013

Background

We performed in vivo micro‐computed tomography (micro‐
CT
) imaging using a novel carbon nanotube (
CNT
)–based x‐ray source to detect calcification in the aortic arch of apolipoprotein
E
(apoE)–null mice.

Methods and Results

We measured calcification volume of aortic arch plaques using
CNT
‐based micro‐
CT
in 16‐ to 18‐month‐old males on 129S6/SvEvTac and C57BL/6J genetic backgrounds (129‐apoE
KO
and B6‐apoE
KO
). Cardiac and respiratory gated images were acquired in each mouse under anesthesia. Images obtained using a
CNT
micro‐
CT
had less motion blur and better spatial resolution for aortic calcification than those using conventional micro‐
CT
, evaluated by edge sharpness (slope of the normalized attenuation units, 1.6±0.3 versus 0.8±0.2) and contrast‐to‐noise ratio of the calcifications (118±34 versus 10±2); both
P
<0.05, n=6. Calcification volume in the arch inner curvature was 4 times bigger in the 129‐apoE
KO
than in the B6‐apoE
KO
mice (0.90±0.18 versus 0.22±0.10 mm
3
,
P
<0.01, n=7 and 5, respectively), whereas plaque areas in the inner curvature measured in dissected aorta were only twice as great in the 129‐apoE
KO
than in the B6‐apoE
KO
mice (6.1±0.6 versus 3.7±0.4 mm
2
,
P
<0.05). Consistent with this, histological calcification area in the plaques was significantly higher in the 129‐apoE
KO
than in the B6‐apoE
KO
mice (16.9±2.0 versus 9.6±0.8%,
P
<0.05, 3 animals for each).

Conclusions

A novel
CNT
‐based micro‐
CT
is a useful tool to evaluate vascular calcifications in living mice. Quantification from acquired images suggests higher susceptibility to calcification of the aortic arch plaques in 129‐apoE
KO
than in B6‐apoE
KO
mice.
Multiple Risk Factor Intervention Trial Revisited: A New Perspective Based on Nonfatal and Fatal Composite Endpoints, Coronary and Cardiovascular, During the Trial
Authors: Stamler Jeremiah , Neaton James D. , Cohen Jerome D. , Cutler Jeffrey , Eberly Lynn , Grandits Gregory , Kuller Lewis H. , Ockene Judith , Prineas Ronald ,   The MRFIT Research Group
Published: May 2013

Background

The
M
ultiple
R
isk
F
actor
I
ntervention
T
rial evaluated a multifactor intervention on coronary heart disease (
CHD
) in 12 866 men. A priori defined endpoints (
CHD
death,
CHD
death or nonfatal myocardial infarction, cardiovascular disease [
CVD
] death, and all‐cause death) did not differ significantly between the special intervention (
SI
) and usual care (
UC
) groups over an average follow‐up period of 7 years. Event rates were lower than anticipated, reducing power. Other nonfatal
CVD
outcomes were prespecified but not considered in composite outcomes comparing
SI
with
UC
.

Methods and Results

Post‐trial
CVD
mortality risks associated with nonfatal
CVD
events occurring during the trial were determined with
C
ox regression. Nonfatal outcomes associated with >2‐fold risk of
CVD
death over the subsequent 20 years were combined with during‐trial deaths to create 2 new composite outcomes.
SI
/
UC
hazard ratios and 95% confidence intervals were estimated for each composite outcome. Of 10 during‐trial nonfatal events, 6 were associated (
P
<0.001) with >2‐fold risk of
CVD
death. A
CHD
composite outcome (
CHD
death, myocardial infarction [clinical or serial
ECG
change],
CHF
, or coronary artery surgery) was experienced by 520
SI
and 602
UC
men (
SI
/
UC
hazard ratio = 0.86; 95% confidence interval, 0.76–0.97;
P
=0.01). A
CVD
composite outcome (
CHD
[as above], other
CVD
deaths, stroke, or renal impairment) was experienced by 581
SI
and 652
UC
men (hazard ratio = 0.89; 95% confidence interval, 0.79–0.99;
P
=0.04).

Conclusions

In post hoc analyses, composite fatal/nonfatal
CHD
and
CVD
rates over 7 years were significantly lower for
SI
than for
UC
. These findings reinforce recommendations for improved dietary/lifestyle practices, with pharmacological therapy as needed, to prevent and control major
CVD
risk factors.