Comparison of retinal vascular geometry in obese and non-obese children
Comparison of retinal vascular geometry in obese and non-obese children
Evelyn Li Min Tai 0 1
Yee Cheng Kueh 0
Wan-Hazabbah Wan Hitam 0 1
Tien Yin Wong 0 2
Ismail Shatriah 0 1
0 Editor: Jerome W. Breslin, USF Health Morsani College of Medicine , UNITED STATES
1 Department of Ophthalmology, School of Medical Sciences, Health Campus, Universiti Sains Malaysia , Kubang Kerian, Kelantan , Malaysia , 2 Hospital Universiti Sains Malaysia , Kubang Kerian, Kelantan , Malaysia , 3 Unit of Biostatistics & Research Methodology, School of Medical Sciences, Health Campus, Universiti Sains Malaysia , Kubang Kerian, Kelantan , Malaysia , 4 Singapore Eye Research Institute, Singapore National Eye Centre, Singapore, Singapore, 5 DUKE-NUS Medical School, Singapore , Singapore
2 Centre for Eye Research Australia, University of Melbourne , Melbourne , Australia
Obese children have abnormal retinal vascular geometry. These findings suggest that
childhood obesity is characterized by early microvascular abnormalities that precede
development of overt disease. Further research is warranted to determine if these parameters
represent viable biomarkers for risk stratification in obesity.
Data Availability Statement: All relevant data are
within the paper and its Supporting Information
Funding: This work was supported by Universiti
Sains Malaysia Research Grants (http://www.
research.usm.my/); 304/PPSP/61313063 awarded
to ELMT, and 304/PPSP/61313097 awarded to
WHWH. The funder had no role in study design,
data collection and analysis, preparation of the
manuscript or decision to publish.
Competing interests: The authors have declared
that no competing interests exist.
The prevalence of childhood obesity has been increasing worldwide [
]. Childhood obesity
has adverse long-term health implications, especially for cardiometabolic conditions like
hypertension, diabetes and ischaemic heart disease . The underlying pathology in these
diseases occurs at the level of the microvasculature [
Obesity-related endothelial function has been observed in both animal and human studies.
Among obese rats, obesity has been found to increase endothelial susceptibility to
hyperglycaemia-induced oxidative stress [
]. Interestingly, animal models have shown that this protein
kinase C-mediated endothelial damage is also central to the pathogenesis of diabetic
microvascular complications [
] such as retinopathy [
] and nephropathy . This pathway is
likewise operative in humans, in whom protein kinase C blockade reduces endothelial dysfunction
secondary to hyperglycaemia, both in diabetic [
] and non-diabetic subjects [
Although obesity is often linked with metabolic disease, increased body weight per se is
independently associated with impaired endothelium-related coronary vessel function [
Healthy obese adults have been noted to have poorer endothelial function than their
nonobese counterparts [14±16]. Similarly, endothelial dysfunction has been demonstrated in
obese children [
]. In these children, compensatory elevation of circulating endothelial
progenitor cell counts suggest that in early life, obesity-mediated endothelial dysfunction may
still be reversible .
Microvasculature abnormalities in obesity may be visualized non-invasively via digital
retinal vessel analysis [
]. Based on the optimum design principle, the architecture of the
human microvasculature is designed to be energy-efficient, i.e, to provide adequate
perfusion with the minimum of energy expenditure [
]. Retinal vascular geometry not only
reflects the complexity of the vascular tree, but also provides insight into the `optimality' of
the microcirculation [
]. Retinal vascular caliber has previously been shown to be
abnormal in obese adults [
]. These changes begin even in childhood, with an inverse association
between body mass index and arteriolar caliber [
]. Newer retinal vascular parameters,
such as fractal dimension, branching coefficient, and tortuosity have been associated with
obesity-linked microvascular diseases including hypertension , diabetes [
] and stroke
]. However, few studies have examined the effect of obesity per se on these novel vessel
]. Our study thus aimed to compare retinal vascular geometry between
obese and non-obese children.
Materials and methods
This was a cross-sectional prospective study conducted in the Eye Clinic of Hospital Universiti
Sains Malaysia between January 2015 and March 2016. A total of 166 children aged 6 to 12
years old were recruited. The study was approved by the Human Research Ethics Committee
of Universiti Sains Malaysia. The conduct of the study followed the tenets of the declaration of
Children who fulfilled the inclusion and exclusion criteria were invited to participate. The
inclusion criteria was age between 6 and 12 years old at the time of examination, a best
corrected visual acuity better than 6/12, and a normal eye examination. Exclusion criteria was
strabismus, amblyopia, optic nerve abnormalities, high refractive errors (based on spherical
equivalent of ±4.0 diopters], history of ocular trauma, ocular pathology, developmental delay,
and systemic illnesses like diabetes or hypertension. Informed written consent was obtained
from at least one parent, as well as verbal assent from the child.
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Ocular examination, refraction and axial length measurement
Upon arrival at the eye clinic, distance visual acuity was assessed monocularly using a Snellen
chart for distance (Reichert; NY) at six meters. A comprehensive eye examination including
pupillary examination, anterior segment examination and complete retinal evaluation was
performed. An autokeratorefractometer (Model RK5; Canon Inc, Tokyo, Japan) was used to
obtain three consecutive readings of sphere and cylinder, with a maximum acceptable
difference of 0.25 diopters between the lowest and highest readings. Spherical equivalent was
calculated as the value of the sphere plus half of the value of the cylinder. The right eye axial length
was measured using a non-contact partial coherence interferometer (IOL Master, Carl Zeiss
Meditec; Jena, Germany). The mean axial length was derived from a mean of five consecutive
readings. An acceptable reading had a signal-to-noise ratio of more than 2 mm, and a
difference of 0.05 mm or less between the lowest and highest reading.
Blood pressure was measured in the sitting position after 5 minutes of rest, using a digital
automated sphygmomanometer (Model SEM-1 [HEM-7051-C12], Omron Healthcare Co., Ltd.;
Kyoto, Japan) with an appropriate-sized cuff. The average systolic and diastolic blood pressure
was obtained from two readings. A third blood pressure reading would be obtained if the
difference between the first two readings were greater than 10 mm Hg in systolic blood pressure
(SBP) and/or 5 mm Hg in diastolic blood pressure (DBP).
Height and weight were measured with a height and weight measuring scale (Model 220,
Seca; Hamburg, Germany) according to standard protocols. Height was recorded to the
nearest 1 mm. Weight was recorded to the nearest 0.1 kg. Body mass index (BMI) was calculated as
weight divided by the height squared (kg per meter squared). Obesity was classified as BMI
of > 2 SD (standard deviation) above the mean, based on World Health Organization age and
sex-specific growth charts.
Retinal examination and vascular analyses
Pupil dilation was achieved with a single drop of topical phenylephrine 2.5% and tropicamide
1%, after which 45 degree optic disc-centered retinal photographs were taken of both eyes
using a digital fundus camera (Model VX-10, Kowa; Tokyo, Japan). If both images were of
equivalent quality, the image from the right eye was selected. A single grader, masked to
participant identity, performed the retinal vessel analysis using a validated semi-automated
computed software, SIVA (Singapore I Vessel Assessment; National University of Singapore,
SIVA is a semi-automated program in which all retinal vessels greater than 25 um in
diameter located between one-half to two disc diameters from the optic disc margin are outlined and
their edges marked using a pixel density histogram. Retinal vascular parameters are estimated
based on measurements of the biggest six arterioles and venules in this area (zone C) (Fig 1).
Based on the Knudtson-Parr-Hubbard formula, the average retinal arteriolar and venular
caliber were calculated and summarized as the central retinal arteriolar equivalent (CRAE) and
central retinal venular equivalent (CRVE) [
]. The software also automatically provided their
ratio (arteriovenous ratio, AVR). Correction for the effect of ocular magnification on vessel
sizes was performed using the Bengtsson formula [
Retinal fractal dimension (Df) is calculated from the outlined retinal vessels using the
`boxcounting method'. In this method, the digital retinal image is compartmentalized into
equallysized squares (i.e. boxes), then the number of squares containing the skeletonized (i.e outlined)
segments of retinal vessels is calculated [
]. The process is then repeated with squares of
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Fig 1. Fundus photo centered on the optic disc in Singapore I Vessel Assessment (SIVA) software. Panel A shows
an example of a digital fundus photo pre-processing. Panel B shows a screenshot of the SIVA system. All retinal vessels
greater than 25 um in diameter located between one-half to two disc diameters from the optic disc margin (i.e. in Zone
C) are outlined and their edges marked using a pixel density histogram. The retinal arterioles are outlined in red, while
the venules are outlined in blue. In Panel C, the green lines overlying the segment of a vessel are referred to as `covers'.
A minimum of 5 covers are needed; based on these, the software will then provide an average of the mean retinal
arteriolar and venular calibers.
differing sizes. The fractal dimension is the gradient of the logarithm of the number of squares
through which the vessel outline passes against the logarithm of the size of the square. Larger
values represent a more complex branching pattern.
Branching coefficient (BC) is a method of estimating the ratio between the diameters of the
main vessel and the diameters of its branches, or `daughter vessels'. It is given by the area ratio:
S21S2S22, where S is the root, or main segment of vessel, and S1 and S2 are its branches [
A higher BC reflects similarly sized vessel diameters between the main vessel and its branch,
while a lower BC is related to a decrease in the diameters of the branches compared to the
main vessel (Fig 2).
Retinal tortuosity is an index, represented as simple tortuosity (sTORT) and curvature
tortuosity (cTORT). STORT is calculated by the actual length of vessel divided by the Euclidean
distance between the first and last points of that vessel (i.e., the length of the straight line
connecting the two points) [
]. CTORT is defined as the integral of curvature square along the
path of the vessel divided by the total arc length [
]. A lower tortuosity index represents
Statistical analyses were performed using IBM SPSS Statistics for Windows, Version 24.0 (IBM
Corp, Armonk, NY). Chi-square test was used to determine the association between obese,
non-obese children and gender. Independent t-test was used to determine the mean
differences of numerical variables between obese and non-obese children. Analysis of covariance
(ANCOVA) was performed to determine the mean differences in dependent variables (i.e.,
Fig 2. Diagrammatic illustration of branching coefficient. A portion of the retinal arteriole nasal to the disc is
shown, pre (panel A) and post-processing (panel B) by SIVA. Branching coefficient (BC) is calculated as BC
where S is the root, or main segment of vessel, and S1 and S2 are its branches. Note the relative thicknesses of the main
segment of the vessel compared to its branches, as artificially illustrated in panel C and D. A higher BC reflects
similarly sized vessel diameters between the main vessel and its branches (panel C), while a lower BC is related to a
decrease in the diameters of the branches compared to the main vessel (panel D).
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caliber, Df, AVR, BC, sTORT, cTORT) between obese and non-obese children, with
adjustment for possible confounding variables (i.e., age, gender, SBP, DBP and axial length) [
Several models were tested for each dependent variable with correction for confounding
variables in ANCOVA; Model 1 was adjusted for age and gender, Model 2 was adjusted for age,
gender, SBP and DBP, and Model 3 was adjusted for age, gender, SBP, DBP, and axial length.
P values of <0.05 were considered statistically significant.
A total of 166 Malay children were included in this study. Their mean age was 9.58 years.
Approximately 50% were female. 51.2% were categorized as obese. Other systemic
demographics and retinal vascular parameters are summarized in Table 1. Obese children had a
significantly lower retinal arteriolar caliber and arteriovenous ratio than non-obese children.
Conversely, the venular Df, BC and cTORT were significantly higher in obese children than in
The differences in the arteriolar caliber, AVR, venular Df and venular cTORT between
obese and non-obese children remained significant after adjusting for age, gender, SBP, DBP
and axial length (Table 2). After multivariable adjustment, there was no significant difference
in venular BC between obese and non-obese children.
Childhood obesity is a risk factor for various diseases, notably diabetes [
]. Diabetic patients
have previously been observed to have abnormal retinal vascular geometry [
SBP, systolic blood pressure; DBP, diastolic blood pressure; SD, standard deviation
bChi-square test and its p-value.
Statistical difference (p < 0.05) between the obese and non-obese group
(n = 81)
Mean ± SD
9.64 ± 1.75
107.53 ± 11.55
65.27 ± 8.39
23.01 ± 0.75
172.39 ± 13.25
250.35 ± 16.22
1.22 ± 0.05
1.19 ± 0.05
1.43 ± 0.27
1.24 ± 0.25
1.09 ± 0.02
1.10 ± 0.02
6.24 ± 1.11
6.43 ± 0.71
(df = 164)
CI, confidence interval; Model-1, adjusted for age and gender; Model-2, adjusted for age, gender, SBP, DBP; Model-3, adjusted for age, gender, SBP, DBP and axial
Statistical difference (p < 0.05) between the obese and non-obese group
PLOS ONE | https://doi.org/10.1371/journal.pone.0191434
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among obese individuals, data regarding these vessel parameters is scarce. Retinal vessel
analysis in obese children may identify early changes prior to the development of obesity-linked
microvascular disease. Our study demonstrates unique differences in retinal vascular
parameters between obese and non-obese primary school children.
We found that obese children had significantly narrower retinal arterioles than non-obese
children. Among studies which have evaluated the effect of BMI on retinal vessels, significant
differences in retinal arteriolar caliber have been noted between subjects in the lowest and
highest BMI quartiles [
]. Although obese children showed a trend towards wider venules,
this difference was not statistically significant, which is consistent with the literature [
BMI may have an indirect effect on the microvasculature via its association with increased
blood pressure [
], but our finding that the differences in retinal arteriolar caliber between
obese and non-obese children persisted after adjustment for blood pressure supports the
hypothesis that the arteriolar narrowing in obese children represents impaired vasodilatory
function . These results are substantiated in animal studies; Frisbee et al demonstrated that
in obese Zucker rats, there is enhancement of vasoconstrictor processes and impairment of
endothelium-dependent vasodilator responses [
]. The latter process may be mediated by
nitric oxide (NO) [
], as evidenced by studies showing NO-dependent improvement in
microvascular function after therapeutic interventions in obese rats [
We observed that venular Df was significantly higher in obese children than in non-obese
children. These results differ from those of Gopinath et al, who found that fractal dimension
was not significantly associated with body mass index [
]. However, the authors later noted
that carbohydrate intake and a high-glycaemic index diet were associated with greater retinal Df
in girls [
]. As nutrition and body mass index are inextricably intertwined, obese children may
have greater Df due to a complex interplay between these factors and the microvasculature. Df
is a proxy for the geometric complexity of the retinal branching pattern, and is associated with
] and lacunar stroke [
]. Diabetic patients have been observed to have higher
Df than controls [
]. We postulate that the higher Df in obese children may represent
microvascular alterations preceding the development of diabetes and its associated complications.
Retinal venular BC was higher in obese children than among non-obese children, but after
multivariable adjustment, the significance of these associations disappeared. To our
knowledge, no previous study has explored the relationship between obesity and BC. Venular BC
increases when the area of the branch venules increases disproportionately to that of the main
vessel. The selective effect on these venules is attributed to the fact that unlike in arterioles,
where wall shear stress is lower in second-order arterioles, shear stress, such as occurs with
elevated blood pressure, has been found to be similar in first and second-order venules [
thinner walls of second-order venules may less resistant to stress than first-order venules,
resulting in endothelial inflammation. In rat models, inflammation-induced vasodilation
predominantly affects venules, substantiating our hypothesis [
]. Endothelial inflammation also
disrupts the delicate balance between reactive oxygen species and antioxidant defenses,
resulting in damaged endothelial cells [
]. As subclinical endothelial dysfunction is present in
obese children, the underlying pathogenesis may be as discussed above [
Suboptimal BC is not only an indirect measure of endothelial dysfunction; it is also
associated with altered shear stress across the retinal vascular network, thus propagating a vicious
cycle of inflammation and further injury [
]. The increased stress on the vasculature may be
compounded by various systemic factors, which may explain the lack of statistical significance
in BC after adjustment for confounders. Suboptimal BC has been linked to impairment in
general cognitive ability and verbal fluency [
]. Although the association of BC with cognition
has not been specifically assessed in obese children, obese individuals have been found to have
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poorer cognitive ability than controls [
]. Further studies are required to determine whether
these findings are reflective of the suboptimal BC in obese patients.
Although our STORT values were similar in both groups, we found a significantly higher
venular cTORT in obese than non-obese children. These results differed from those of
Sasongko et al, in which no association of body mass index with tortuosity was observed [
As sTORT cannot distinguish between true tortuosity (multiple points of inflection) and
increased length of the vessel due to bowing, cTORT may be a more accurate measure of vessel
]. Increased retinal venular tortuosity is associated with diabetic retinopathy [
and cognitive impairment [
]. Vessels become more tortuous in response to increased
transmural pressure [
], which may explain the association of tortuosity with blood pressure [
The selective increase in venular tortuosity may be attributed to the relative paucity of smooth
muscle in venular walls, making them more vulnerable to distortion than arterioles, which
have a more developed tunica media. Furthermore, retinal arteriolar autoregulation in
response to various factors such as pressure, shear stress and metabolic demand may also
explain the observed lack of association of arteriolar parameters with obesity [
Evaluation of retinal vascular geometry in a cohort of obese children free of other systemic
disease enables non-invasive identification of early retinal microvascular alterations prior to the
development of overt disease. Our study confirms the previous observations of retinal arteriolar
narrowing in obesity, and identifies novel abnormalities in venular Df and cTORT among obese
children. The strengths of this study include its objective quantification of retinal vascular
geometry via a semi-automated, validated computer program, its sampling of subjects from a single
ethnic group, adjustment for multiple confounders and the use of vessel indices that are independent
of magnification error and cardiac cycle [
]. However, the cross-sectional nature of this study
limits our ability to make inferences of a temporal nature, and body mass index merely acts as a
substitute for obesity. Combining body mass index with other adiposity-related measures such as
fat mass by skin-fold thickness may strengthen the clinical significance of these findings [
There is also a need for prospective, longitudinal studies to demonstrate the sequential changes of
the microvasculature which occur in the development of obesity-related disease.
Obese children have abnormal retinal vascular geometry. These findings suggest that the
microvascular abnormalities observed in obesity-related diseases like diabetes have their
origins in childhood. Retinal vascular geometry may thus represent a biomarker for risk
stratification, as well as a potential therapeutic target for obesity interventions.
S1 Table. Systemic, ocular and retinal vascular parameters of study subjects.
Conceptualization: Evelyn Li Min Tai, Ismail Shatriah.
Formal analysis: Evelyn Li Min Tai, Yee Cheng Kueh.
Funding acquisition: Evelyn Li Min Tai, Wan-Hazabbah Wan Hitam.
Investigation: Evelyn Li Min Tai.
Methodology: Evelyn Li Min Tai, Yee Cheng Kueh, Ismail Shatriah.
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Resources: Evelyn Li Min Tai.
Software: Tien Yin Wong.
Supervision: Wan-Hazabbah Wan Hitam, Ismail Shatriah.
Validation: Tien Yin Wong.
Visualization: Wan-Hazabbah Wan Hitam, Ismail Shatriah.
Writing ± original draft: Evelyn Li Min Tai.
Writing ± review & editing: Evelyn Li Min Tai, Yee Cheng Kueh, Tien Yin Wong.
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