JIACM 2012; 13(3): 198-202

ORIGINAL ARTICLE

Study of platelet aggregation in subjects with

early glucose intolerance (EGI)
Rupali Malik*, Arun Gogna**, Sumita Saluja***

Abstract
Background: Role of platelet abnormalities like increased aggregation preceding cardiovascular complications in frank diabetic
patients is well established. It was thus intended to see if such a relationship also exists in pre-diabetic stage like early glucose
intolerance, so that vascular complications could be dealt with at the earliest.
Objectives: To study platelet aggregation in subjects with EGI and to see if any correlation of platelet aggregation existed with
parameters of metabolic syndrome.
Methodology: We studied 75 patients, 25 each with EGI, frank diabetes and normal glucose tolerance based on oral glucose tolerance
test (OGTT) with 75 gm glucose.
All patients underwent detailed history and clinical examination. Measurements of various anthropometric parameters like height,
weight, waist circumference (WC), BMI, WC/height ratio were also taken.
Routine lab examination like complete blood count, peripheral blood smear, fasting lipid profile, KFT, and ECG were also done.
Finally, all subjects underwent optical platelet aggregation test with agonist like ADP in 5 gm concentration in a resting, fasting,
and non-smoking state.
Results: We found statistically significant increase in platelet aggregation in subjects with DM as well as EGI compared to normal.
40% subjects in EGI group and 52% in frank diabetic group had increased aggregation with ADP in 5 gm concentration.
No correlation of platelet aggregation was seen with various parameters of metabolic syndrome like weight, WC, BMI, fasting
blood glucose, fasting triglyceride, total cholesterol, HDL and LDC cholesterol.
Conclusion: Abnormal platelet behaviour like increased aggregation which precede the onset of cardio-vascular disease in subjects
with DM starts in the latent diabetic stage like EGI itself. Abnormal glucose tolerance is the most important predictor of abnormal
platelet behaviour and the anthropometric parameters of metabolic syndrome were not found to be correlated.
Key words: Diabetes mellitus, early glucose intolerance, platelet aggregation.

Introduction Moreover, in diabetic patients, the clock for macrovascular

The worldwide prevalence of diabetes mellitus (DM) has
risen dramatically over the past 2 decades. The native
Asian Indians lead the globe in DM prevalence with more
than 50 million cases with possibly an equal number of
pre-diabetics1. Pre-diabetes classically included impaired
fasting glucose (IFG) and impaired glucose tolerance (IGT).
But in the American Diabetes Association (ADA)
diagnostic criteria presently used for their diagnosis, no
mention is made about plasma glucose response
intermediate to fasting and 2-hour during an OGTT. But
many non-diabetic subjects show isolated abnormal
intermediate values > 160 mg/dl, with fasting plasma
glucose (FPG) < 100 mg/dl and 2-hour post-prandial
glucose (PPG) < 140 mg/dl1. Thus, EGI is defined as 1-hour
post-glucose load > 160 with normal FPG and 2-hour PPG.
So, if only fasting and 2-hour post-glucose testing is done,
subjects with EGI are missed out.
complications starts ticking early, almost 1 - 2 decades
before the diagnosis when the patient is in a state of
glucose intolerance but not classical IGT 1 . The
hyperglycaemia peaks at this stage and causes atheroma,
endothelial thinning, stunning, endothelial dysfunction,
and causes irreversible vascular damage.
Also, studies like DCCT-EDIC and UKPDS have proved the
legacy effect of early intensive glucose control on long-
term rate of vascular complications in DM2. Thus, it is
important to intervene at the earliest possible stage for
better vascular outcomes in diabetic patients.
Also, Asians have a higher rate of DM and CAD despite
being lean1. While a part of this is explained by increased
rates of insulin resistance, these factors do not fully explain
the excess CAD risk in this asthenic group. Platelet
aggregation has been considered an important

* Resident, ** Consultant and Associate Professor, Department of Medicine, *** Consultant and Associate Professor,
Department of Haematology, Vardhman Mahavir Medical College (VMMC) and Safdarjung Hospital,
New Delhi - 110 029.
pathogenic event which predisposes to CAD. Studies in
the western population have shown platelet aggregation
to be higher in diabetic subjects3 suggesting a key role of
platelets in diabetic vascular occlusive disorders. But little
information is available on platelet behaviour in pre-
diabetics, especially in high-risk Asian Indians. Therefore
this study was undertaken to study platelet aggregation
in subjects with EGI. Moreover, as impaired glucose
tolerance is an integral part of the metabolic syndrome, it
was also intended to study any correlation of aggregation
with various parameters of the metabolic syndrome.
patients reported high aggregation.
In group 3 (NGT) out of 25 patients, 11 had low, and 14
had normal platelet aggregation. No patient in this group
high platelet aggregation.
This result (Fig. 1 and Table I) was found to be highly
significant statistically on comparing aggregation
between group 1 and 3 (p value = 0.001) and between
group 2 and 3 (p value = 0.0001) by both chi-square and
Fisher exact test.

16 ADP-5
Material and methods
14
This study was conducted at Vardhman Mahavir Medical 14 13
College (VMMC) and Safdarjung Hospital, New Delhi, on 12
12 11
patients attending the medicine OPD and diabetes clinic. 10
10
Based on OGTT with 75 gm glucose, the patients were
8
divided into 3 groups: EGI, frank diabetics, and those with
6 6
normal glucose tolerance, with 25 patients in each group. 6
Patients were excluded based on criteria like past h/o 4 3
hypertension, chronic renal failure, ischaemic heart 2
disease. Also excluded were patients on drugs like statins, 0
antiplatelet drugs (clopidogrel, aspirin, ticlopidine), 0
NSAIDs, h/o stroke within 5 years, Hb < 8 gm%, platelet Low Normal High
count < 1,50,000 or > 4,50,000, haematological disorders, Platelet aggregation
family or personal h/o bleeding disorders or surgical EGI DM N
procedure within 1 week before enrolment.
Fig. 1: Graph showing platelet aggregation with ADP 5g.
Statistical analysis
Table I: Table showing platelet aggregation with ADP
Observed data was presented in terms of minimum, g.
5
maximum, and standard deviation for a descriptive
analysis. Further analysis done using chi-square test and Group 1 Group 2 Group 3
Fisher exact test. Correlation was done by Karl Pearson EGI25 DM25 normal25
test. Low aggregation 3 6 11

The level of statistical significance was taken as p < 0.05. Normal aggregation 12 6 14
The data was analysed using statistical software. High aggregation 10 13 0
P= 0.001 (Group 1 and 3); P = 0.0001 (Group 2 and 3).
Results
Based on Karl Pearson test, no correlation of platelet
Subjects in all these groups had similar age distribution aggregation was seen with various parameters of
and demographic profile. metabolic syndrome like weight, WC, BMI, fasting blood
In optical platelet aggregation testing it was found:- glucose, fasting triglyceride, total cholesterol, HDL and LDL
cholesterol.
In group 1 (EGI) out of 25 patients, 3 patients had low
aggregation, 12 had normal, and 10 had high aggregation. Also, no correlation of platelet aggregation was found with
Thus 40% patients in this group had reported increased the sex of the patient.
platelet aggregation.
Discussion
In group 2 (DM) out of 25 patients, 6 had low, 6 had
normal, and 13 had high aggregation. Thus, a total of 52% Abnormalities of platelet aggregation and haemostatic

Journal, Indian Academy of Clinical Medicine l Vol. 13, No. 3 l July-September, 2012 199
balance before the clinical evidence of diabetic vascular
disease suggest that this defect may be acquired early in
the natural history of diabetes, and could underlie the
vascular disease onset. While long-term prospective
studies in vivo and in vitro are required to show whether
abnormal platelet aggregation is causally related to
vascular disease, a consequence of it. or simply a
characteristic of the diabetic state regardless of whether
vascular disease is present or not, the present study is in
accord with the hypothesis that abnormal platelet
aggregation is integral to vascular disease onset.
In spite of leading the globe in diabetes prevalence, the
Indian subcontinent is lacking in studies on platelet
behaviour. One study done by Mohan et al4 in 2004 on
the subjects of ongoing Chennai Urban Population Study
(CUPS-13) found increased platelet activation by flow
cytometry in subjects of DM and CAD.
In our study, we studied platelet aggregation in subjects
with EGI and DM, and it was compared to that in subjects
with normal glucose tolerance. We targeted EGI because
it is a stage which lies much earlier in the spectrum of
diabetes, and as reported by various studies like Decode
study5 and others5-13, post-prandial hyperglycaemia is an
independent factor for CAD. Also, the Honolulu heart
study 8 reported 1-hour post-prandial glucose as a
predictor of CVD in patients of DM.
We found in our study that platelet aggregation to ADP
in concentration of 5g was found to be significantly
increased in subjects with EGI and frank diabetes as
compared to subjects with normal glucose tolerance. With
ADP 5g concentration, 40% subjects with EGI and 52%
subjects with frank diabetics had increased aggregation.
Though platelet aggregation was significantly higher in
EGI group than normal, it was less than that in patients
with frank diabetes.
Similarly, Sagel et al14 found increased platelet aggregation
tendency, determined as an increase in light transmission
after 4 min, using ADP, adrenaline, and collagen in platelet-
rich plasma from patients with frank diabetes, unrelated
to presence of vascular disease. They also studied platelet
aggregation in subjects with latent diabetes and pre-
diabetes. Though they found increased second phase
aggregation to all concentrations of ADP (0.125M, 0.25
M, 0.5M and 1M) in patients with frank diabetes, in
latent diabetics the aggregation was found to be high only
with ADP concentration of 0.5M and 1M.
In contrast, Colwell et al15 in one of there studies reported
increased second phase platelet aggregation induced
by ADP and epinephrine in frank diabetics and latent
diabetics, but not in pre-diabetics. In their study, pre-
200 Journal, Indian Academy of Clinical Medicine
diabetics were defined as the subjects with normal
glucose tolerance, and both parents unequivocal
diabetics. And latent diabetics were patients with post-
prandial glucose values of 185 (1 hr), 160 (1.5 hrs) and
140 (2 hrs). Thus it could be concluded that platelet
aggregation is affected only when glucose tolerance is
abnormal.
Heath et al16 also found increased platelet aggregation
tendency in vitro defined by broader amplitude after
adding ADP in low concentration (1M, 2M) and by
decrease in disaggregation of platelets in patients with
severe diabetic retinopathy.
Hassanein et al17 also found decreased disaggregation of
platelets after addition of ADP in patient with diabetes.
OMalley et al18 showed increased sensitivity of platelets
from diabetic patients with peripheral neuropathy to ADP
and epinephrine-induced aggregation.
Leone et al19 had shown that patients of diabetes with
retinopathy, peripheral vascular thrombosis, or abnormal
ECG had increased platelet aggregation to ADP. They also
found similar findings in some patients without apparent
vascular disease.
Breddin et al 20 and Wu Hoak et al 21 found increased
tendency to formation of spontaneous platelet
aggregates in vivo and in vitro.
In contrast to all these studies, Kutti et al22 reported that
there was no increase in in vitro ADP sensitivity of platelets
from diabetic patients. The reason for this contrasting
result could be the fact that all patients in their study were
insulin dependent diabetics. And as described by Harrison
et al23, chronic use of insulin administration (8 days), and
not acute use, would restore prostacyclin PGI2 production
in platelets of diabetic animals leading to a decrease in
their aggregation tendency.
In our study, we found that there was no correlation of
platelet aggregation with various parameters of metabolic
syndrome like waist circumference, BMI, fasting blood
glucose, fasting triglycerides level, total cholesterol level,
HDL cholesterol, and LDL cholesterol.
Similarly, Sagel et al14 found no correlation between
fasting blood glucose level and platelet aggregation
tendency.
Kwaan et al24 who showed that ADP-induced platelet
aggregation of normal platelet-rich plasma was increased
by adding plasma from patients with diabetes, especially
those with vascular complications, suggesting a key role
of plasma factor in increased platelet aggregation in
diabetics; also found no correlation of increased
aggregation tendency and age, weight, level of blood
l Vol. 13, No. 3 l July-September, 2012
glucose, triglycerides, cholesterol, free fatty acids or
albuminuria.
In contrast, Hassanein et al17 reported that decrease in
this aggregation of platelets after addition of ADP in
diabetics was marked in patients with high cholesterol
levels.
We also studied the correlation between platelet
aggregation and waist circumference/height ratio. This
parameter was taken into consideration as it is
considered a strong predictor of intraabdominal fat25 and
has been proposed as a better indicator of cardiovascular
risk25 and weight management26 than the classic BMI. But
no correlation was found between aggregation and
waist circumference/height ratio. Thus it is very clear that
glucose intolerance is the only parameter which is
related to increased platelet aggregation tendency.
We also found that the incidence of increased platelet
aggregation was similar between male and female
patients in both EGI and diabetics.
This is in contrast with the findings of a study done by
Kueh et al27 who reported incidence of increased platelet
aggregation in female diabetics was significantly higher
than male diabetics. No reason was found for this
observation. None of the females in their study was on
oral contraceptive pills or hormone replacement therapy.
But all the females had undergone platelet aggregation
study in the first week of their menstrual cycle. But stage
of menstrual cycle was not taken into consideration in
our study. This might be the reason for the contrast found
between the two studies.
The limitation of our study is the small sample size. In this
context, more number of long-term prospective studies
on platelet behaviour are required.
Our study supports the observations made by previous
studies and also extends the abnormal behaviour of
diabetic platelets to the latent diabetic phase. And it seems
logical to postulate that the abnormal behaviour of
platelets may be involved in the genesis of occlusive
lesions in the micro-circulation of diabetics, thus leading
to vascular complications.
Conclusion
Our study found statistically significant increase in platelet
aggregation in subjects with EGI and DM compared to
subjects with normal glucose tolerance. The increase in
platelet aggregation in subjects with EGI was slightly less
than that in subjects with DM. With ADP 5g, high
aggregation was seen in 40% subjects with EGI, and 52%
with frank diabetics.
Journal, Indian Academy of Clinical Medicine l Vol. 13, No. 3
In our study, we found that there was no correlation of
platelet aggregation with various parameters of metabolic
syndrome like waist circumference, BMI, fasting blood
glucose, fasting triglycerides level, total cholesterol level,
HDL cholesterol, and LDL cholesterol.
Even waist circumference/height ratio, which is a better
predictor of cardio-vascular disease risk, was found to have
no correlation with increased platelet aggregation
tendency.
Also, no correlation existed between the increased platelet
aggregation and sex of the patient. The prevalence of
increased aggregation was found to be similar between
males and the females.
So, it could be concluded that the platelet abnormality
which precedes the onset of cardio-vascular disease in
subjects with DM, starts in the latent diabetic stage itself,
like in EGI. Therefore, it seems important for the clinicians
to target these subjects with EGI at the earliest possible
so that macro-vascular complication of diabetes could be
dealt with even before their onset.
Thus we conclude that abnormal glucose tolerance is the
most important predictor of the abnormal platelet
behaviour and so subjects with abnormal glucose
tolerance like EGI should be screened as early as possible.
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202 Journal, Indian Academy of Clinical Medicine l Vol. 13, No. 3 l July-September, 2012