CRP in Patients with Acute Ischemic Stroke in Bangladesh

Objectives: This aim of the study was to evaluate CRP and the risk factors in ischaemic stroke. Methods: This case control study was carried out from January 2006 to December 2007 in the department of Neurology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka in all the OPD and admitted stroke patients meeting the inclusion and exclusion criteria. Results: Out of 30 patients with ischaemic stroke 28(93.3%) patients had positive C-reactive protein value. 37% stroke patients were smoker. Mean total cholesterol, triglyceride and LDL level were higher and mean HDL level was lower among stroke patients in comparison to controls. 17% of stroke patients were suffering from diabetes mellitus & 80% of stroke patients were hypertensive. Conclusion: C-reactive protein is elevated in the acute phase of ischa emic stroke and could present a prognostic marker.


Introduction
Stroke is the commonest neurological disorder of adult life and remains third leading cause of death after heart disease and cancer, after the age of 40. 1 Of the hundreds of thousands of stroke survivors each year, approximately 30 percent require assistance for activities of daily living, 20 percent require assistance with ambulation and 16 percent require institutional care. 2 In general, its incidence is 150 to 300 per 100,000 populations and prevalence is 600/100,000 population.Of all strokes, 85 percent are due to ischaemic infarction and 15 percent are due to haemorrhage.The highest incidence of stroke seen is 7/1000 at Novosibriska, Russia, and lowest is 2.4/1000 at Dijon, France. 3 In Bangladesh the incidence of stroke is 700,000 per year.Stroke causing about 4 million deaths per year) and is the most important single cause of severe disability in people living in their own home. 4ovel plasma marker of atherothrombotic disease. 6urthermore, elevated plasma levels of C-reactive protein are not disease specific but are sensitive markers produced in response to tissue injury, infectious agents, immunologic stimuli, and inflammation. 7Elevated blood levels of Creactive protein (CRP) are associated with an increased risk of atherosclerotic vascular disease including stroke. 8owever the role of CRP in the etiology and prognosis of ischemic stroke remains to be clearly defined. 9One hypothesis is that CRP plays a direct causal role in the pathogenesis of atherosclerosis by promoting endothelial cell adhesion molecule expression, monocyte recruitment or complement activation, or by mediating low density lipoprotein cholesterol uptake by macrophages. 10evated blood concentrations of CRP during the acute phase of ischemic stroke event primarily reflect the extent of cerebral ischemic injury and its complications. 11Higher values of C-reactive protein (CRP) are significantly associated with large infarct size and worst outcome. 12Use of plasma CRP levels may aid in identifying a potentially large number of men and women who are at risk for cerebrovascular events. 7The purpose of this study is To evaluate C-reactive protein in patients with acute ischaemic stroke, To evaluate the risk factors in ischaemic stroke and To observe the elevation of C-reactive protein in patients with acute ischaemic stroke.

Materials and Methods
This was a case control study which was carried out in the department of Neurology in collaboration with the department of Microbiology and Immunology, Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka.The study was done from January 2006 to December 2007 in all the patients with stroke attending the outpatient department (OPD) and also admitted into the indoor of the neurology department, meeting the inclusion and exclusion criteria, were .included in the study.Patient of more than 30 yrs age and presented with a acute ischaemic stroke (<4 weeks duration) proved clinically and by neuroimaging (CT/MRI of brain) were defined as case and Apparently healthy and age and sex matched with the case were control.Age below 30 years, Hemorrhagic stroke proved by neuroimaging, history of acute myocardial infarction, venous thrombosis, and myeloproliferative diseases, history of rheumatological and connective tissue diseases, history of malignancy,History of recent surgery (within one month) and history of recent infection (within one month) were excluded from the study.
After taking the detail history and physical examination a total 73 patients were included for neuroimaging (CT Scan/ MRI of brain) study.Among them 23 patients were excluded due to the evidence of hemorrhage in neuroimaging.Among the remaining 50 patients 20 patients were excluded from the study group due to the evidence of infection, rheumatological disorder and history of acute myocardial infarction.Ultimately a total of 30 patients were included in the study group whom met the inclusion and exclusion criteria.Out of 30 patients CT scan of brain was done in 25 patients and MRI of brain was done in 5 patients.
Detail history was taken about age, hypertension, diabetes, smoking habit, cardiac disease and oral contraceptive pills.They were also asked about febrile infection or productive cough, diarrhoea and dysuria.They were clinically evaluated for the presence of sinusitis, diabetes, hypertension, and tuberculosis, pneumonia, bronchitis.For detection of lymphadenopathy cervical, supraclavicular, axillary and inguinal groups of lymph nodes were searched for.They were also clinically evaluated for presence of any valvular and ischemic heart disease.
All of them were investigated for serum C-reactive protein, complete blood count, serum creatinine, random blood sugar RBS, serum lipid profile, ECG and chest X-ray PA view.Written consent was taken from the patient or relatives after explaining the study procedure and aims of the study A total of 30 age and sex matched and apparently healthy persons were taken as controls.Controls never suffered from stroke and came in neurology out patient department for nonspecific disorders and symptoms like anxiety neurosis, myalgia, burning limbs, tingling numbness, headache and somatoform disorders and had no clinically apparent infection or inflammation or sign of neurological deficit.All the obtained data were recorded in a pre-structured case record form.

C-reactive protein in serum was estimated by Nephelometric
System by using commercial kit (DADE BEHRING BN 100, USA) in the department of Microbiology and Immunology of Bangabandhu Sheikh Mujib Medical University.Under aseptic precaution 5ml of random venous blood was collected in a test tube.It was kept for 5-10 minutes for clotting.It was then centrifuged at the rate of 5,000-10,000 rpm for 10 minutes.
In this way serum was separated.200µl serum was taken into sample cup.Then reagent was taken into predefined reagent cup.The Behring Nephelometer 100 analyzer took reaction buffer, diluent, distilled water automatically.Quantitative measurement of C-reactive protein was estimated from the serum following automated analyzer program directed in computer.C-reactive protein level was expressed in mg/L.Random blood sugar was also estimated by kit method in Biochemistry department of BSMMU.Fasting lipid profile was also estimated by kit method in Biochemistry department of BSMMU.Total count of WBC, differential count of WBC, Hb% and ESR was done in clinical pathology department of BSMMU.Urine routine and microscopic examination were done in clinical pathology department of BSMMU to exclude urinary tract infection.X-ray chest P/A view was done in Radiology and imaging Department of BSMMU.Neuroimaging (CT Scan of brain/MRI of brain) was done in Radiology and Imaging department of BSMMU and also from some private diagnostic centers.Electrocardiogram [ECG] -A 12 lead standard ECG was done in the department of Cardiology of BSMMU.
Blood pressure was measured by Mercury Sphygmomanometer.It was measured from the mid-arm of the left side in lying position.Data analysis was done by Computer based software; statistical package for social sciences (SPSS) was used to analyze collected data.Appropriate analysis, such as Unpaired Student's't' test, Chi-square test, Z-test, etc, were carried out.P value<0.05 was taken as minimum level of significance.

Results
This was a case control study carried out to evaluate the serum C reactive protein in patients with acute ischaemic stroke.A total 30 patients diagnosed as ischaemic stroke clinically and confirmed by neuroimaging were evaluated.Age and sex matched apparently healthy person of same number were also taken as control.Majority (63%) of study subjects were in between 51 to 80 years of age.The study showed 21(70%) patients were male and 9(30%) patients were female.Among control 25(83.3%)were male and 5(16.7%) were female.
It was found 28(93.3%)patients had positive C-reactive protein value and only 2(6.7%) patients had negative Creactive protein value.Among control group all had negative C-reactive protein value.Seventy (70) percent of the study subjects were male.Male to female ratio was 2.33:1.A study in 1995 showed that stroke affects male 1.5 times more often than female 14 .This difference found in the study may be due to the attitude of our society that the females are less frequently brought to the hospitals for treatment.
In this study serum C-reactive protein level was found to be elevated in 28 (93.3%)patients out of 30.Mean C-reactive protein value was 42.06 (SD±21.26)whereas among the control it was 4.30 (SD±0.72).These findings are consistent with several studies 5,16,17,18 The possible explanations are: 1. CRP concentration may reflect the degree of stroke severity, correlating with the degree of inflammation directly consequent to cerebral infarction.2. CRP concentration may indicate underlying unstable atherosclerotic disease.3. CRP may be raised as a consequence of secondary complications of stroke at the time of sampling. 17We have found 2 stroke patients were negative for C-reactive protein value.Grau, 1997 found about 25% of patients with first ever ischaemic stroke had normal levels of CRP after stroke.It might be due to the variability of the degree of inflammatory response to ischaemic stroke (Gabay et al., 1999).In 1999 had shown high CRP values in their study compared with much lower levels predictive of cardiovascular morbidity in epidemiological studies that could suggest a vigorous acute phase response rather than a chronic inflammatory low grade response 16 .The inflammatory process represented by CRP elevation could also result from underlying atherosclerotic plaque itself.
The study revealed 37% stroke patients were smoker.In a study in BSMMU showed 59.84% smoker among their stroke patients. 19This difference may be due to the variation in the number of cases and anti smoking campaign.The contribution of cigarette smoking to an increased risk of stroke is controversial.Although it may have an indirect effect. 20One study in 1998 showed an ordered relationship between increasing exposure to cigarette smoke and the prevalence of cerebral infarction. 21ghty (80) percent of stroke patients were hypertensive.In this study we have also found that total count of WBC was higher in case group along with neutrophilic leucocytosis.Marquardt et al., 2005 showed in their study that leucocyte count was followed a time course compatible with the pattern of an acute phase reaction showing initial peak and a rapid decline after stroke.Emsley et al., 2003 observed a persistent elevation of leucocyte count along with other factors until 3 months after the event.ESR was found to be a bit higher in the study group but there was no significant relationship between ESR and CRP in case and controls.
The potential role of viral infection in atherogenesis has received much attention in ischaemic heart disease but has yet to be found relevant to carotid artery disease.Inflammatory cell infiltration, especially by macrophages has been associated with carotid plaque rupture in a manner analogous to coronary artery disease, and concentration of both circulating and locally synthesized pro inflammatory cytokines are elevated in patients with carotid atherosclerosis.In the majority of patients CRP may therefore be a useful marker of atherosclerotic instability. 23 coronary artery disease two recent studies seem to demonstrate that therapy with statins and reducing levels of CRP could reduce progression of atherosclerosis. 23,24 cerebrovascular disease the influence of some categories of drugs on atherosclerotic process modifying the acute phase of inflammation was studied.It was suggested that statins and Angiotensin converting enzyme inhibitors could be more effective in the presence of high CRP levels and that the efficacy of anti platelet therapy, such as aspirin or clopidogrel, in secondary prevention could be related to the levels of inflammatory markers. 25mitations of the study The study was not prospectively designed to assess the effect of C-reactive protein on outcome of ischaemic stroke.
Only a single blood level was checked.The number of cases in this study was small.

Conclusion
Stroke is actually one of the main problems of health worldwide because of its high incidence, prevalence, mortality, and residual disability.It is of utmost importance to select patients with high risk of negative outcomes and to address them through a most intensive pharmacological and rehabilitative approach.Our study confirms that C-reactive protein is elevated in the acute phase of ischaemic stroke and could present a prognostic marker.Further prospective studies are necessary to demonstrate this hypothesis.

Table - I
Age distribution of the study subjects nsChi square test/Unpaired Student's 't' test ns = Not significant

Table - VI
Lipid profile (SD±8.38) mmHg between cases and control groups respectively.This table showing Systolic Blood Pressure and Diastolic Blood Pressure were significantly higher in the case group (p<0.001).CRP +ve and CRP -ve was not significant.Among control group all are CRP -ve.Comparison of lipid profile between case CRP -ve and control CRP -ve also didn't show any significant difference 22  Napoli et al., 2001in their study of 1020 stroke patients had shown two third patients were hypertensive.22Sheuli,2006 found 63.3 % stroke patients were hypertensive in a study.