# Efficacy of Clopidrogel on Reperfusion and High-Sensitivity C-Reactive Protein in Patients with Acute Myocardial Infarction

Authors: Mehmet Akbulut; Makbule Kutlu; Yılmaz Ozbay; Veli Polat; Mehmet Nail Bilen; Adil Baydas; Yakup Altas
Journal: Mediators of Inflammation
 (2009)

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## Abstract

We investigated the effects of clopidogrel on reperfusion and inflammatory process in STEMI. A total of 175 STEMI patients with similar clinical characteristics were included to this study. One was the standard pharmacological reperfusion therapy group (group 1,n : 90), who received 300 mg aspirin, 70 U/kg bolus, and 12 U/kg/hr continuous infusion of  unfractioned heparin and accelerated t-PA. Clopidogrel 450 mg loading and 75 mg/d thereafter was added to standard reperfusion therapy in the other group (group 2, n : 85). The ST-segment resolution, CK-MB, and high-sensitive CRP (hs-CRP) parameters were measured. Complete ST resolution was observed in 32 patients (36.8%) in group 1 and 53 patients (63.8%) in group 2 (P<.001). Also in the first 24 hours, the CK-MB levels of patients in group 1 were significantly higher than those of group 2 (P=.001). The hs-CRP values were  greater in group 1 than group 2 at 48th hour (gruop 1: 9.4±0.1 mg/L, group 2: 3.7±1.4 mg/L; P=.000). We concluded that adding clopidogrel to standard treatment in STEMI patients provided early reperfusion and suppression of inflammatory response.


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## Body

## 1. Introduction

Thromboxane and ADP, which are among the mediators of platelet activation and aggregation, play
a key role in initiating and propagating coronary thrombosis and are raised
during myocardial infarction. Simultaneous inhibition of both of these pathways
with the combination of clopidogrel and aspirin produces
greater antiplatelet effects than either agent alone [1, 2]. In recent reports,
dual antiplatelet therapy with aspirin and clopidogrel showed significant
improvements on prognosis in ST-elevating myocardial infarction (STEMI) [3–5].
Moreover, this beneficial effect is not only limited to the acute phase, but
also extended to a 1-year follow-up period [6]. But substantial uncertainty
remained regarding the net effects of adding clopidogrel to aspirin upon the
reperfusion, acute inflammatory response, and ischemic events in this setting.
In this study, we evaluated the effect of the addition of clopidogrel to
standard reperfusion protocol on reperfusion and acute inflammatory response.


## 2. Material
and Methods

### 2.1. Patient Population

From December 2006 to January 2008, consecutive patients were enrolled who presented
with ischemic discomfort lasting >20 minutes at rest within 12 hours, before
randomization pain lasting >20 minutes, ST-segment elevation of at least 0.2 mV in at least two contiguous
precordial leads. The study was limited to only anteriorly located myocardial
infarction to homogenize the investigated population. The exclusion criteria
were conduction or rhythm abnormalities (bundle branch block, idioventricular
rhythm, etc.); any contraindication to thrombolytics [7]; early coronary
angiography (within first 48 hours) due to recurrent ischemia or failed
thrombolysis; those patients under the treatment of aspirin or thienopyridines; any contraindication to aspirin or
clopidogrel; past history of MI or coronary revascularization; presence of clinically
assessed heart failure (Killip II/III) or cardiogenic shock; hepatic failure;
renal failure (serum creatinine >2.5 mg/dL); thrombocytopenia (<100.000/mm3) and patients older than 70 years old were excluded.The study protocol was approved by
institutional review board, and written informed consent was taken from all
patients.


### 2.2. Study Design

A total of 175 patients were included in the present
study and were allocated in 2 groups. One was the standard pharmacological
reperfusion therapy group (group 1,n:90), consisting of patients who received
300 mg aspirin at first then 150 mg/d thereafter, 70 U/kg (maximum 5000 U)
bolus, and 12 U/kg/hr continuous infusion of unfractioned heparin and accelerated t-PA therapy (15 mg intravenous
bolus followed by an infusion of 0.75 mg/kg (maximum 50 mg) over 30 minutes,
followed by an infusion of 0.5 mg/kg (maximum 35 mg) over 60 minutes).
Clopidogrel 450 mg loading and 75 mg/d thereafter was added to standard
reperfusion therapy in the other group (group 2, n:85). The ST-segment
resolution within 120 minutes, CK-MB parameters within first 24 hours, and high-sensitive
C-reactive protein (hs-CRP) parameters within 48 hours were measured. Also patients
were examined for inhospital major and minor bleeding complications. All data
were evaluated by two investigators who are blinded to treatment.


#### 2.2.1. Electrocardiographic Analysis

Standard 12-lead electrocardiograms were
obtained at baseline and at 30-minute periods during the first120 minutes after
initiation of fibrinolytic therapy. ST-segment elevation was analyzed manually
with lens-intensified calipers to the nearest of 0.025 mV 20 milliseconds after the end of QRS complex with the PR segment as the reference
baseline from leads I, aVL, and V1 through V6 for
anterior infarction. The sum of ST deviation was measured at baseline and at
30, 60, 90, 120 minutes using previously described methods [8]. The percent
resolution of ST deviation from baseline to 30, 60, 90, 120 minutes was
calculated and categorized as complete (≥70%), partial (30% to 70%), or no
resolution (≤30%).


#### 2.2.2. Blood Sampling

Blood samples were obtained for CK-MB and
hs-CRP analysis at 0, 4, 8, 12, 24 and 0, 24, 48 hours of admission, respectively.
Biochemical and haematological parameters were measured by an Olympus AU 600
autoanalyzer (Olympus Optical Co., Ltd., Schimatsu-Mishima, Japan) and Bayer Advia 120 Cell CBC Counter Hematologia
autoanalyzer (Bayer Advia 120 CBC counter, NJ, USA). Hs-C-reactive protein was measured by
the high-sensitivity nephelometric method (Dade Behring, Marburg, Germany).


### 2.3. Statistical Analysis

The SPSS for windows program (version 15.0) was
used for all data analysis. Study data were expressed as mean value ± standard
deviation or percent values. Numerical variables were compared by Mann-WhineyU-test,
whereas nonnumerical ones were compared by Chi-squere test. Results with a P value less than .05 were considered statistically significant.


## 2.1. Patient Population

From December 2006 to January 2008, consecutive patients were enrolled who presented
with ischemic discomfort lasting >20 minutes at rest within 12 hours, before
randomization pain lasting >20 minutes, ST-segment elevation of at least 0.2 mV in at least two contiguous
precordial leads. The study was limited to only anteriorly located myocardial
infarction to homogenize the investigated population. The exclusion criteria
were conduction or rhythm abnormalities (bundle branch block, idioventricular
rhythm, etc.); any contraindication to thrombolytics [7]; early coronary
angiography (within first 48 hours) due to recurrent ischemia or failed
thrombolysis; those patients under the treatment of aspirin or thienopyridines; any contraindication to aspirin or
clopidogrel; past history of MI or coronary revascularization; presence of clinically
assessed heart failure (Killip II/III) or cardiogenic shock; hepatic failure;
renal failure (serum creatinine >2.5 mg/dL); thrombocytopenia (<100.000/mm3) and patients older than 70 years old were excluded.The study protocol was approved by
institutional review board, and written informed consent was taken from all
patients.


## 2.2. Study Design

A total of 175 patients were included in the present
study and were allocated in 2 groups. One was the standard pharmacological
reperfusion therapy group (group 1,n:90), consisting of patients who received
300 mg aspirin at first then 150 mg/d thereafter, 70 U/kg (maximum 5000 U)
bolus, and 12 U/kg/hr continuous infusion of unfractioned heparin and accelerated t-PA therapy (15 mg intravenous
bolus followed by an infusion of 0.75 mg/kg (maximum 50 mg) over 30 minutes,
followed by an infusion of 0.5 mg/kg (maximum 35 mg) over 60 minutes).
Clopidogrel 450 mg loading and 75 mg/d thereafter was added to standard
reperfusion therapy in the other group (group 2, n:85). The ST-segment
resolution within 120 minutes, CK-MB parameters within first 24 hours, and high-sensitive
C-reactive protein (hs-CRP) parameters within 48 hours were measured. Also patients
were examined for inhospital major and minor bleeding complications. All data
were evaluated by two investigators who are blinded to treatment.


### 2.2.1. Electrocardiographic Analysis

Standard 12-lead electrocardiograms were
obtained at baseline and at 30-minute periods during the first120 minutes after
initiation of fibrinolytic therapy. ST-segment elevation was analyzed manually
with lens-intensified calipers to the nearest of 0.025 mV 20 milliseconds after the end of QRS complex with the PR segment as the reference
baseline from leads I, aVL, and V1 through V6 for
anterior infarction. The sum of ST deviation was measured at baseline and at
30, 60, 90, 120 minutes using previously described methods [8]. The percent
resolution of ST deviation from baseline to 30, 60, 90, 120 minutes was
calculated and categorized as complete (≥70%), partial (30% to 70%), or no
resolution (≤30%).


### 2.2.2. Blood Sampling

Blood samples were obtained for CK-MB and
hs-CRP analysis at 0, 4, 8, 12, 24 and 0, 24, 48 hours of admission, respectively.
Biochemical and haematological parameters were measured by an Olympus AU 600
autoanalyzer (Olympus Optical Co., Ltd., Schimatsu-Mishima, Japan) and Bayer Advia 120 Cell CBC Counter Hematologia
autoanalyzer (Bayer Advia 120 CBC counter, NJ, USA). Hs-C-reactive protein was measured by
the high-sensitivity nephelometric method (Dade Behring, Marburg, Germany).


## 2.2.1. Electrocardiographic Analysis

Standard 12-lead electrocardiograms were
obtained at baseline and at 30-minute periods during the first120 minutes after
initiation of fibrinolytic therapy. ST-segment elevation was analyzed manually
with lens-intensified calipers to the nearest of 0.025 mV 20 milliseconds after the end of QRS complex with the PR segment as the reference
baseline from leads I, aVL, and V1 through V6 for
anterior infarction. The sum of ST deviation was measured at baseline and at
30, 60, 90, 120 minutes using previously described methods [8]. The percent
resolution of ST deviation from baseline to 30, 60, 90, 120 minutes was
calculated and categorized as complete (≥70%), partial (30% to 70%), or no
resolution (≤30%).


## 2.2.2. Blood Sampling

Blood samples were obtained for CK-MB and
hs-CRP analysis at 0, 4, 8, 12, 24 and 0, 24, 48 hours of admission, respectively.
Biochemical and haematological parameters were measured by an Olympus AU 600
autoanalyzer (Olympus Optical Co., Ltd., Schimatsu-Mishima, Japan) and Bayer Advia 120 Cell CBC Counter Hematologia
autoanalyzer (Bayer Advia 120 CBC counter, NJ, USA). Hs-C-reactive protein was measured by
the high-sensitivity nephelometric method (Dade Behring, Marburg, Germany).


## 2.3. Statistical Analysis

The SPSS for windows program (version 15.0) was
used for all data analysis. Study data were expressed as mean value ± standard
deviation or percent values. Numerical variables were compared by Mann-WhineyU-test,
whereas nonnumerical ones were compared by Chi-squere test. Results with a P value less than .05 were considered statistically significant.


## 3. Results

### 3.1. Baseline Characteristics

A total of 175 patients were included in the study
(group 1,n:90; group 2, n:85). Three
patients due to ischemic events in group 1 (2 failed thrombolysis, 1 heart
failure) and patients in group 2
were excluded from the study due to ischemic and hemorrhagic events (1 heart
failure, 1 gluteal hemorrhage). However, there were no important differences in
baseline clinical characteristics and initial therapy between the two treatment
groups. Clinical characteristics of the study groups were shown in Table 1.Table 1
Baseline characteristics.
CharacteristicsGroup
1(n:87)Group
2(n:83)P  valueMean age (year)57 ± 756 ± 5.31Men (%)79.380.7.48Time for symptom to lytic (h)2.7 ± 22.6 ± 1.56PresentationHeart rate (beat/min)75 ± 1474 ± 12.37Systolic blood pressure (mmHg)133 ± 14135 ± 16.31Diastolic blood pressure (mmHg)73 ± 1175±15.29History (%)Current smoker44.844.6.51Diabetes mellitus17.219.3.44Hypertension29.932.5.35Hyperlipidemia27.631.3.35High TIMI risk score (≥5)13.814.8.33Medication during hospitalization (%)β-blocker9394.50Statins9093.51ACE-I or ARB7374.43


### 3.2. ST-Segment

Complete ST resolution was achieved in 32 of
patients (36.8%) in group 1 and 53 of patients (63.8%) in group 2 (P < .001)
at the end of 120 minutes (Table 2 and Figure 1). The difference between groups
started after 60th minute
of the therapy, and it became more significant at 120th minute (Figure 2).Table 2
Number of patients with
resolution in 120th minute.
Group 1(n:87)Group 2(n:83)P value>70%36.8 (32)63.8 (53).00130–70%49.4 (43)27.7 (23).001<30%13.8 (12)8.4 (7).01Figure 1
120 minute ST-segment resolution.Figure 2
Patient number with ST-segment resolution in the course of time.


### 3.3. CK-MB

Peak CK-MB values were reached at 10–12th hours in group 1, whereas
in group 2 they were
at 8–10th hours. Peak CK-MB value
was 233.5 ± 145 U/L in group 1 and
132.5 ± 122 U/L in group 2 (P = .01), and at the end of 24 hours CK-MB value
was 138 ± 88 U/L in group 1, and 56 ± 35 U/L in group 2 (P = .001) (Table 3).Table 3
CK-MB (U/L) levels of the patients in the groups during
the first 24 hours.
0 hour4th hour8th hour12th hour24th hourGroup I29.2 ± 23106.4 ± 54193.2 ± 125233.5 ± 145138 ± 88Group II28.5 ± 2255.4 ± 35132.5 ± 122111.3 ± 2756 ± 35P value.66.006.01.001.001


### 3.4. The hs-CRP

Although baseline hs-CRP values were similar at
baseline (group 1:1.6±0.9 mg/L, group 2: 1.9±1.3 mg/L; P = .67), they were greater in
group 1 than group 2 at 48th hour (group 1: 9.4±0.1 mg/L, group 2: 3.7±1.4 mg/L; P = .000) (Figure 3).Figure 3
hs-CRP levels of the patients in the groups during the first 48 hours.


### 3.5. Other Clinical Outcome

No death was observed in both groups. Failed
thrombolysis in two patients and heart failure in one patient were seen in
group 1; one patient with heart failure and one patient with major hemorrhage
(≥2 unit blood transfusion) were seen in group 2. However, there is no
significant difference between the groups with respect to these unfavorable
events (P > .05).


## 3.1. Baseline Characteristics

A total of 175 patients were included in the study
(group 1,n:90; group 2, n:85). Three
patients due to ischemic events in group 1 (2 failed thrombolysis, 1 heart
failure) and patients in group 2
were excluded from the study due to ischemic and hemorrhagic events (1 heart
failure, 1 gluteal hemorrhage). However, there were no important differences in
baseline clinical characteristics and initial therapy between the two treatment
groups. Clinical characteristics of the study groups were shown in Table 1.Table 1
Baseline characteristics.
CharacteristicsGroup
1(n:87)Group
2(n:83)P  valueMean age (year)57 ± 756 ± 5.31Men (%)79.380.7.48Time for symptom to lytic (h)2.7 ± 22.6 ± 1.56PresentationHeart rate (beat/min)75 ± 1474 ± 12.37Systolic blood pressure (mmHg)133 ± 14135 ± 16.31Diastolic blood pressure (mmHg)73 ± 1175±15.29History (%)Current smoker44.844.6.51Diabetes mellitus17.219.3.44Hypertension29.932.5.35Hyperlipidemia27.631.3.35High TIMI risk score (≥5)13.814.8.33Medication during hospitalization (%)β-blocker9394.50Statins9093.51ACE-I or ARB7374.43


## 3.2. ST-Segment

Complete ST resolution was achieved in 32 of
patients (36.8%) in group 1 and 53 of patients (63.8%) in group 2 (P < .001)
at the end of 120 minutes (Table 2 and Figure 1). The difference between groups
started after 60th minute
of the therapy, and it became more significant at 120th minute (Figure 2).Table 2
Number of patients with
resolution in 120th minute.
Group 1(n:87)Group 2(n:83)P value>70%36.8 (32)63.8 (53).00130–70%49.4 (43)27.7 (23).001<30%13.8 (12)8.4 (7).01Figure 1
120 minute ST-segment resolution.Figure 2
Patient number with ST-segment resolution in the course of time.


## 3.3. CK-MB

Peak CK-MB values were reached at 10–12th hours in group 1, whereas
in group 2 they were
at 8–10th hours. Peak CK-MB value
was 233.5 ± 145 U/L in group 1 and
132.5 ± 122 U/L in group 2 (P = .01), and at the end of 24 hours CK-MB value
was 138 ± 88 U/L in group 1, and 56 ± 35 U/L in group 2 (P = .001) (Table 3).Table 3
CK-MB (U/L) levels of the patients in the groups during
the first 24 hours.
0 hour4th hour8th hour12th hour24th hourGroup I29.2 ± 23106.4 ± 54193.2 ± 125233.5 ± 145138 ± 88Group II28.5 ± 2255.4 ± 35132.5 ± 122111.3 ± 2756 ± 35P value.66.006.01.001.001


## 3.4. The hs-CRP

Although baseline hs-CRP values were similar at
baseline (group 1:1.6±0.9 mg/L, group 2: 1.9±1.3 mg/L; P = .67), they were greater in
group 1 than group 2 at 48th hour (group 1: 9.4±0.1 mg/L, group 2: 3.7±1.4 mg/L; P = .000) (Figure 3).Figure 3
hs-CRP levels of the patients in the groups during the first 48 hours.


## 3.5. Other Clinical Outcome

No death was observed in both groups. Failed
thrombolysis in two patients and heart failure in one patient were seen in
group 1; one patient with heart failure and one patient with major hemorrhage
(≥2 unit blood transfusion) were seen in group 2. However, there is no
significant difference between the groups with respect to these unfavorable
events (P > .05).


## 4. Discussion

Platelet activation and aggregation play a key
role initiating and propagating coronary artery thrombosis, and antiplatelet
therapy improves outcomes across the spectrum of acute coronary syndromes. Beside
this, it was thought that simultaneous inhibition of platelet activation and
aggregation pathways with specific antiplatelet combinations might be more
efficient in acute coronary syndromes [1, 2]. We observed improvements in both
reperfusion and inflammatory response with the early addition of clopidogrel to
standard reperfusion therapy in STEMI patients in this study. Moreover, these
findings supported the hypothesis of dual antiplatelet inhibition with aspirin
and clopidogrel might be more beneficial in acute coronary syndrome.Currently, there are promising developments in
the prevention of ischemic complications in sympthomatic atherothrombotic
cases. The CLARITY-TIMI 28 study demonstrated that the addition of clopidogrel
to aspirin in patients with STEMI receiving fibrinolytic therapy improved the
patency rate of the infarct-related artery after 48 hours and reduced ischemic
complications [3]. Among the patients in this trial who underwent percutaneous
coronary intervention (PCI), described in the PCI-CLARTY substudy, clopidogrel
pretreatment significantly reduced the incidence of cardiac death or ischemic
complications both before and after PCI [9]. In the COMMIT trial, the routine
addition of clopidogrel 75 mg/d to aspirin therapy for 4 weeks in patients with
acute myocardial infarction resulted in a 9% proportional reduction in death,
reinfarction, or stroke (P = .002) and a 7% proportional reduction in
death (P = .03) [5]. Even though no loading dose was used, the benefit of
clopidogrel therapy was seen almost immediately by an 11% proportional
reduction in mortality in the first 2 days after the initiation of therapy (P = .019)
[5]. The results of both CLARITY-TIMI 28 and COMMIT showed that adding
clopidogrel to aspirin and other standard treatments safely reduced mortality
and major vascular events in patients with acute myocardial infarction.Beneficial effects of clopidogrel may arise
through some different proposed mechanisms. (i) Clopidogrel may facilitate initial fibrinolysis and thereby
improve early reperfusion with a glycoprotein IIb-IIIa inhibitor [10]-like
effect. (ii) Clopidogrel may exert its effects in myocardial infarction mainly
by preventing reocclusion or by limiting microvascular effects of platelet
activation rather than enhancing fibrinolysis [5]. While most of the benefit of
clopidogrel has been attributed to inhibition of platelet activation, blockage
of the P2Y12 receptor may also have pleotropic effects with respect to
endothelial cell function, leukocyte activation, and inflammation [11].Previous studies have suggested some
anti-inflammatory properties of clopidogrel. A reduction in the number of
platelet-leukocyte interactions has been described [12, 13], and one study
reported a special benefit of the drug in reducing the augmented risk of
percutaneous interventions in patients with elevated levels of C-reactive
protein [14]. In another study, it was stated that clopidogrel pretreatment
attenuated the periprocedural increase in CRP by 65% [15]. Quinn et
al. said that clopidogrel pretreatment reduces platelet inflammatory marker expression in patients undergoing
PCI [16], and they also concluded that it might be very important for clinical
events. All these findings constituted an important clue for us to investigate
the limiting effect of clopidogrel in increment of high-sensitive C-reactive protein in STEMI and to verify
this. Moreover, we could not see another study in literature about this topic.Activated
platelets have been clearly implicated in the tendency for microembolization
during acute coronary syndrome [17, 18] and play an important role in the
inflammatory response. Studies have shown that activated platelets express the
CD40 ligand, a potent stimulus of vascular inflammation [19, 20]. The CD40
ligand promotes platelet-leukocyte interactions [19] and induces tissue factor
expression [21], thus providing a link among inflammation, platelet activation,
embolization, and coagulation. The inhibition of subcellular platelet CD40
ligand by clopidogrel treatment may explain the mechanism of reduction in
inflammation as expressed by an attenuation in C-reactive protein increase
after acute STEMI. By the way, we think that large extensive clinical trials
are needed to exactly clarify the mechanisms of beneficial effects of
clopidogrel on clinical results in previous ones and that is what we have
determined in our study because the impressive improvement in clinical results
with added clopidogrel on aspirin requires  some other mechanisms except for a
potent antiplatelet effect.


## 5. Limitations

The first and most important limitation is the small
study population and the lack of a placebo control group. Only the anteriorly
located STEMI patients were included to homogenize the study population, and a
fibrinolytic agent only rt-PA was used and this condition was an important
limiting factor for our study. Clopidogrel loading dose (450 mg) may be a
limitation. We think that different
dosages (300–600 mg) should be
applied for both evaluations
of safety and efficacy because there is a correlation between loading dose and
time to reach maximal antiplatelet effect [22]. Another limiting factor of our
study was the restriction of the study with 48 hours. A long follow-up period
would be more useful to exactly realize all the efficiency and safety of
clopidogrel (major and minor hemorrhages
included) treatment. So that with long follow-up period, difference among the
groups of medical treatment, PCI and coronary artery bypass grefting in the direction of angiographic findings could
be clearly understood.


## 6. Conclusions

In conclusion, we found that addition of
clopidogrel to medical reperfusion therapy in STEMI had favorable effects on
reperfusion and suppression of hs-CRP. We think that these favorable clinical
and laboratory effects of dual antiplatelet therapy with clopidogrel are not only limited to
its antiplatelet effect, but also there may be some pleiotropic effects of
clopidogrel; those were not explained clearly. Large extensive studies are
needed to explain the complete effect of clopidogrel in acute STEMI.



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*Source: 932515-2009-04-12.xml*