Elevated C-reactive protein levels predict worsening prognosis in Chinese patients with first-onset stroke

Abstract

The role of high sensitivity C-reactive protein (hsCRP) in predicting prognosis after stroke in the Asian population has not been investigated. We hypothesized that elevated levels of hsCRP were associated with worsening prognosis after stroke in Chinese patients. Two hundred and ninety consecutive patients with first-onset stroke and 290 age- and gender-matched control subjects without any cerebrovascular disease were enrolled for study. Plasma hsCRP level was detected and subsequent vascular events and death were recorded in both groups over a 5-year period. Compared to control group, patients presenting with stroke had higher plasma hsCRP level (3.3 ± 3.8 vs 1.3 ± 2.2 mg/L, P< 0.01). Furthermore, in the group of patients with stroke, the mean plasma hsCRP level was higher in patients who developed subsequent vascular diseases or died as compared with the patients without further complications (4.4 ± 4.3 vs 2.7 ± 3.3 mg/L, P< 0.01). Compared to the lowest tertile of hsCRP level, the relative risk for vascular events or death in stroke patients was 2.91 in the highest tertile of hsCRP (95% CI, 1.54–5.50, P = 0.001). This increase in relative risk for vascular events or death in stroke patients continued after adjustment for age, sex and other cardiovascular risk factors such as hypertension and diabetes (OR: 2.771, 95% CI: 1.367–5.617, P = 0.005). These findings indicate that increased hsCRP level is associated with worsening prognosis after stroke in Chinese patients and suggests that inflammation is correlated with stroke outcome.

Keywords

C-reactive protein / inflammation / stroke

Introduction

Inflammation is an important contributor to cardiovascular disease [1]. Several inflammatory markers such as soluble cell adhesion molecules, cytokines, chemokines and acute phase reactants are associated with cardiovascular events [2,3]. In particular, the acute phase reactant, C-reactive protein (CRP), which is a nonspecific inflammatory marker, has been correlated with cardiovascular outcomes including stroke [4–8]. Increased plasma high sensitivity CRP (hsCRP) level is observed in patients presenting with acute coronary syndromes and is associated with a poorer prognosis [9–11]. However, the predictive value of plasma hsCRP level in predicting the outcomes of patients presenting with stroke is unknown. Given that stroke is a major cause of morbidity and mortality in Asian patients, having a marker correlated with prognosis following stroke would allow for better risk stratification and care of these patients.

Subjects and methods

Study population

This is a prospective, multi-center case control study to assess which risk factors or markers determine prognosis of patients presenting with stroke. The study protocol was reviewed and approved by the Ministry of Public Health, Ministry of Science and Technology and Tongji Medical College, China. Informed consents were obtained from all the study subjects.

All stroke subjects were recruited consecutively from four medical centers in Hubei Province. Diagnosis of strokes was made based on the results of strict clinical medical history, neurological examination, computerized tomography (CT), or magnetic resonance imaging (MRI) according to the International Classification of Diseases, 9th version. Stroke included ischemic and intracerebral hemorrhagic strokes diagnosed using CT or MRI. Patients with other types of stroke, including transient ischemic attack, subarachnoid hemorrhage, embolic brain infarction, brain tumors and cerebrovascular malformation and severe systemic diseases such as rheumatologic disease, endocrine and metabolic disease (except diabetes mellitus), inflammation, liver, neoplastic, or renal diseases, were excluded. Controls were selected from inpatients (10%) with minor illness from the departments of ophthalmology, gastroenterology, otorhinolaryngology and orthopedics, and from community-based inhabitants (90%) free of neurological diseases following the same exclusion criteria as cases. At each local participating center, both men and women were divided into groups by age varying from 38 to 85 years (5-year range for each group), and when 1 stroke case was enrolled, a control subject was randomly selected from the corresponding center.

Two hundred and ninety patients with stroke were recruited between November, 2000 to July, 2001, including 197 patients with ischemic stroke and 93 patients with hemorrhagic stroke. Two hundred and ninety age- and sex-matched control subjects without cerebrovascular disease were collected at the same time from the same institution. All the patients and control subjects provided their medical history pertaining to hypertension, diabetes and other cardiovascular risk factors like cigarette smoking and blood pressure, fasting blood glucose, triglyceride, total cholesterol and low-density lipoprotein (LDL) and underwent routine physical examination by trained physicians.

hsCRP determination and biochemical variables

The blood samples of subjects were collected in ethylenediamine tetra acetic acid (EDTA)-Na tubes. After centrifugation at 3000 r/min for 15 min, the plasma was segregated from the blood and stored at -80°C until use. The hsCRP level in the plasma was detected using an enzyme-linked immunosorbent assay. This high sensitivity assay is a colorimetric competitive immunoassay that uses purified protein and polyclonal anti-CRP antibodies. All detections were performed in duplicate with an inter-assay variation of < 5%. Other plasma biochemical values were measured with a standard automatic analyzer (Hitachi 7060, Hitachi, Japan).

Follow-up and clinical end points

Home visits or telephone follow-up was performed every year during 5 years after the blood samples were collected. Subjects were evaluated for the following indices: self-care ability, blood pressure control, recurrent of stroke, cardiovascular events and drug therapy. The primary end point was acute myocardial infarction, recurrent stroke or death.

Statistical analysis

Analyses were performed with SPSS statistical package, version 10.0. Statistical significance for inter-group differences was analyzed by the Student t-test for continuous variables and the x² test for categorical variables. The Kaplan-Meier method (log-rank test) was used for survival analysis. The multivariate logistic regression analysis was applied to determine the odds ratio (OR) for primary endpoint during the 5-year follow-up associated with increased tertile distribution of hsCRP, with the lowest tertile as the reference, adjusted for sex, age, blood pressure and other vascular risk factors.

Results

Selected baseline characteristics of the participants

The baseline characteristics of the study subjects are shown in Table 1. There was no significant difference in age, sex, body mass index and history of hyperlipidemia and current cigarette usage between stroke patients and control subjects. However, there were more patients with a history of hypertension and diabetes in the stroke group compared to the control group (63.8% vs 41.4% and 11.7% vs 4.8%, respectively). In stroke patients, the plasma hsCRP levels were higher compared to that of control subjects (3.3 ± 3.8 mg/L vs 1.3 ± 2.2 mg/L, P < 0.01). The association between elevated hsCRP and the occurrence of stroke remained even after adjustment for age, sex and other risk factors including hypertension and diabetes (P < 0.01).

To determine whether hsCRP correlated with the type of stroke, all the stroke patients were divided into three groups based upon clinical history and imaging studies: atherothrombosis, lacunar and hemorrhagic. Interestingly, higher hsCRP levels were observed for hemorrhagic strokes compared to ischemic strokes (Fig. 1). The hsCRP levels for atherothrombotic, lacunar and hemorrhagic strokes were 2.6 ± 3.1, 2.7 ± 2.9, 4.9 ± 4.8 mg/L, respectively (P < 0.05 for all groups compared to the control).

Characteristics of the stroke population after 5-year follow-up

During the 5-year follow-up period, 97 patients (33.4% of stroke group) had a primary endpoint event. Sixty six patients died: 52 (78.8%) died of a vascular event (fatal stroke in 27, sequelae from stroke in 15, and cardiac death in 10) and 14 (21.2%) died of nonvascular causes (cancer in 7, renal failure in 3, respiratory failure with emphysema in 1, head trauma in 1, uncontrolled hyperglycemia in diabetes in 1 and severe anemia in 1). Thirty-six patients experienced new vascular events: recurrent non-fatal stroke in 33 and non-fatal myocardial infarction in 3. In contrast, 24 subjects (8.3%) in the control group had a primary endpoint event. Nineteen patients died: 8 (42.1%) died of vascular causes (fatal stroke in 3 and myocardial infarction in 5) and 11 (57.9%) died of nonvascular death (cancer in 6, respiratory failure in 3, and uncontrolled diabetes in 1). Five patients experienced new vascular events: non-fatal stroke in 4 and non-fatal myocardial infarction in 1.

hsCRP predicts primary endpoint events during the 5-year follow-up

Of the patients with stroke, the mean plasma hsCRP level in patients with primary endpoint events was significantly higher than in those patients without primary endpoint events (4.4 ± 4.3 vs. 2.7 ± 3.3 mg/L, P < 0.01). When stroke patients were stratified into tertiles based upon hsCRP levels, an increased probability of death or new vascular events was found in patients with increased levels of hsCRP (lowest to highest tertile: 23%, 32% and 46%; P < 0.002) (Fig. 2).

The analyses of primary endpoints and the distribution of the patients in different tertiles based on hsCRP level show that the relative risk of death or vascular events was 2.913 (95% CI, 1.544 to 5.495, P = 0.001) in patients with highest tertile of hsCRP level compared to those with the lowest tertile of hsCRP during the 5 year follow-up (Table 2). To determine whether hsCRP is independent of other risk factors in predicting primary endpoint events in stroke patients, we performed multiple logistic regression analyses and found that increased hsCRP level was still associated with increased risk of death and vascular events even after having adjusted for the age, sex and other cardiovascular risk factors (OR, 2.771, 95% CI 1.367–5.617, P=0.005) (Table 3 and Fig. 3). In control subjects, the mean hsCRP level was not different between those with and without primary endpoint events (1.3 ± 1.6 vs 1.2 ± 2.2 mg/L, P > 0.05). When control subjects were stratified to tertiles of hsCRP levels, there was no significant correlation between increased probability of death or vascular events and increased tertiles of hsCRP level (5.1%, 8.3% and 11.5%, P = 0.29) (Fig. 4).

Discussion

This is a case-control prospective study in the Chinese population showing that plasma concentration of hsCRP was higher in patients who presented with stroke, especially intracerebral hemorrhagic strokes, correlated with worsening prognosis in these patients. Most of the adverse outcomes, which were associated with higher hsCRP levels, were recorded as primary endpoint events, including recurrent strokes, myocardial infarction and death. The adjusted odds ratio of incurring a primary endpoint event after a 5-year follow-up period was 2.771 (95% CI, 1.367–5.617, P = 0.005). These findings suggest that hsCRP may be a useful prognostic index in Chinese patients presenting with stroke.

Inflammation plays a key role in the pathogenesis of atherosclerosis [1,12] and is correlated with increased incidence of cardiovascular disease, including strokes [13]. Several inflammatory markers have been studied, but the most widely studied is hsCRP. Elevated hsCRP level is associated with increased risk of cardiovascular disease, and in some cases, is associated with cardiovascular outcomes [5,13–15]. In particular, higher CRP levels were correlated with worsening prognosis after ischemic stroke in Caucasian patients [16,17]. However, in other studies, CRP appeared to be more correlated with stroke severity rather than recurrent events [18]. Thus, the role of CRP as a prognostic marker following stroke remains to be determined.

Recent studies suggest that CRP is more than a prognostic marker, but a mediator of vascular disease. For example, CRP is found in the arterial wall during atherogenesis, colocalizing with the terminal complement complex and foam cells [19,20]. Furthermore, CRP binds to the oxidized low-density lipoprotein (LDL) [21], up-regulates expressions of endothelial adhesion molecule and plasminogen activator inhibitor-1 [22,23], mediates LDL uptake by macrophages [24], and decreases endothelial nitric oxide synthase (eNOS) expression [25]. These studies suggest that hsCRP is not only a marker of inflammation, but also has complex modulatory functions that may contribute to the development and evolution of vascular disease such as stroke. Indeed, Napoli and his colleagues investigated the 1-year prognostic influences of CRP levels on the prognosis of 128 patients with first-ever ischemic stroke, and found that increased levels of CRP were associated with a worse outcome in patients with ischemic stroke independent on other risk factors [16].

Interestingly, we found that higher hsCRP levels were associated with hemorrhagic stroke, which tended to have poorer outcomes. Although the precise mechanism is not known, hemorrhagic stroke induces a strong inflammatory response, including upregulation of cytokines, proinflammatory transcription factors such as nuclear factor-κB, microglia activation, astrogliosis, and complement activation [26]. These inflammatory responses may, in turn, further exacerbate brain injury [27]. In addition, hypertension, which is an important risk factor for hemorrhagic stroke [28,29], may have an inflammatory component since inflammatory biomarkers such as CRP and IL-6 are elevated in patients with hypertension [30,31]. Thus, it is unclear from our study whether the higher levels of hsCRP observed in hemorrhagic stroke reflect its risk factor or its consequences. Nevertheless, this study suggests that the inflammatory response after first-onset strokes is not only a good predictor of future strokes, but also, future coronary morbidity and mortality.

In summary, this is a case-control study showing that the increased plasma hsCRP level correlates with long-term prognosis after first-ever stroke in Chinese patients. These findings suggest that inflammatory responses as assessed by plasma hsCRP levels may be a useful marker for risk stratification in stroke patients. It remains to be determined whether elevated CRP causes play critical roles in the occurrence of first-onset stroke and secondary cardiovascular events in long-term follow-up after strokes and if reduction in hsCRP level and inflammatory response using statins could help reduce the morbidity of strokes as well as ameliorate the prognosis of strokes as secondary prevention.

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