Serum homocysteine as an independent risk factor in young patients of ischemic stroke

 

 

Abstract               Background: Abnormal Homocysteine (HCY) level appear to contribute to atherosclerosis by direct toxic effect that damages the arterial linings, interfering with the clotting factors and oxidation of LDL. Elevated homocysteine appears to be associated with an increased risk of the large artery subtype of ischemic stroke, and possibly to the small artery subtype. Objective: This case control study was designed to study the Homocysteine levels in young patients with Ischemic Stroke and to carry out statistical analysis to evaluate Homocysteine as an independent risk factor for Stroke. Methods: The mean serum Homocysteine in patients of Stroke and control was calculated and its association with Total cholesterol, LDL cholesterol and HDL cholesterol. Results: The mean serum Homocysteine in case and control was 28.14 µmol / L and 11.18 µmol / L respectively with S.D of 7.88 µmol / L and 1.82 µmol / L respectively with a significant p value of 0.0001. The difference in Homocysteine levels observed between patients with LDL-C ≥ 100 mg% and those with LDL-C < 100 mg% was not significant. The difference in Homocysteine levels observed between patients with HDL-C ≥ 40 mg% and those with HDL-C < 40 mg% was not significant. The difference in Homocysteine levels observed between patients with S. Cholesterol ≥ 200 mg% and those with S. Cholesterol < 200 mg% was not significant.

Conclusion: From the above findings, in this study the low levels of LDL-C and high levels of HDL-C did notprotect the patients against the Homocysteine induced ischemic stroke. Also there was not significance found between high total cholesterol and Homocysteine induced stroke thereby making homocysteine as an independent risk factor for ischemic stroke

Key Word: HDL, Homocysteine (HCY), Ischemic Stroke, LDL,

 

 

 

INTRODUCTION

Stroke is classified into two major types ¹Brain ischemia due to thrombosis, embolism, or systemic hypoperfusion.² Brain hemorrhage due to intracerebral hemorrhage or subarachnoid haemorrhage Blood disorders are an uncommon primary cause of stroke¹. Sickle cell anemia, Polycythemia vera, Essential thrombocytosis, Heparin induced thrombocytopenia, Protein C or S deficiency, acquired or congenital, Leiden factor (resistance to activated protein C), Antithrombin III deficiency, Antiphospholipid antibody, Lupus anticoagulant, Hyperhomocysteinemia. The rates of reported medical complications of stroke are high. The most common serious medical complications were pneumonia ⁵ percent, gastrointestinal bleeding ³percent, congestive heart failure ³percent, and cardiac arrest 2 percent The management of patients who have had an ischemic stroke involves several phases. The goals in the initial phase when patients first present with symptoms that are consistent with an ischemic event include:

• Insuring medical stability
• Quickly reversing any conditions that are contributing to the patient's symptoms
• Moving toward uncovering the pathophysiologic basis of the patient's neurologic symptoms
• Screening for potential contraindications to thrombolysis

Homocysteine, a sulphur containing amino acid was first described by Vigneaud in 1931. Elevated plasma levels of Homocysteine have been associated with vascular disease. The hallmarks of homocystinuria² are Ectopic lentis, Marfanoid appearance, vascular manifestations, Musculo- skeletal and CNS manifestations. In blood only about 1% of total Homocysteine is free reduced form. The major part of Homocysteine in plasma is oxidized and either colvalently bound to proteins or occur as disulfides. The determinants of total Homocysteine in plasma include several genetic enzyme defects especially cystathionine beta synthase and MTHFR polymorphism^3,4, age and gender, drugs, vitamin status, diabetes, steroid hormones, thyroid disease etc. Homocysteine has been found to induce vascular injury by multiple mechanisms: - Homocysteine promotes leucocyte recruitment by up regulating monocytechemoattractant protein –I and interleukin-8 expression and secretion⁵. It also causes oxidation of LDL which has lipid peroxidation effect. Homocysteine increases smooth muscle cell proliferation and enhances collagen production⁶. And it also causes direct endothelial injury⁷. Prothrombotic effects of Homocysteine, which have been demonstrated in patients with acute coronary syndromes and stroke, include attenuation of endothelial cell tissue plasminogen activator binding sites, activation of factor VIIa and V, inhibition of protein C and heparin sulphate, increased fibrinopeptide A and prothrombin fragments 1 and 2, increased blood viscosity, and decreased endothelial antithrombotic activity due to changes in thrombomodulin functions. Also prolonged exposure of endothelial cells to Homocysteine reduces the activity of dimethylargininedimethylaminohydrolase, the enzyme that degrades asymmetric dimethylarginine, an endogenous inhibitor of nitric oxide synthase; this impairs the production of nitric oxide. This may contribute to impaired endothelium dependent vasodilatation of both conduit and resistance vessels. Several studies^8,9 have showed that’s there is a clear dose response relationship between Homocysteine concentrations and cardiovascular mortality in patients with confirmed coronary artery disease. Normal Homocysteine concentrations range between 5 and 15 µmol/L and hyperhomocysteinemia has been classified as follows; moderate- 15–30 µmol / L, intermediate- 30–100 µmol /L, and severe- more than 100 µmol /L. In our study the difference in the homocysteine levels between the case and the control groups was highly significant (p=0.0001) suggesting homocysteine as an important predictor of Ischemic stroke. The difference in homocysteine levels observed between patient with LDL-C >100mg% and those with LDL-C < 100mg% was not statistically significant (p=0.2005). Hence in this study, the low levels of LDL -C did not protect the patients against the homocysteine induced Ischemic Stroke. The difference in homocysteine levels observed between patients with HDL-C ≥ 40mg% and those with HDL-C < 40 mg% was not statistically significant (p=0.1519) suggesting that high levels of HDL-C did not protect the patients against the homocysteine induced Stroke. Also the difference in homocysteine levels observed between the patients with S. Cholesterol ≥ 200mg% and those with S. cholesterol < 200mg% was statistically not significant (p=0.9492) suggesting that in patients irrespective of levels of Serum Cholesterol, the higher levels of serum homocysteineindependently triggered the Ischemic Stroke.

 

METHODOLOGY

The Study Population: The study population consisted of 30 patients having Ischemic Stroke and diagnosed by clinical signs and symptoms, and CT scan. Control group included 30 young individuals without Ischemic Stroke.

Inclusion Criteria:

All patients aged between 20 – 40 years.

For ischemic stroke

• Clinical signs and symptoms including weakness (Monoparesis/ monoplegia/ hemiparesis/ hemiplegia), vomiting, giddiness, altered sensorium, visual disturbances, speech disturbances, asymmetry of face etc.
• CT scan findings suggestive of ischemic stroke.
• After admission detailed history and clinical examination was carried out and CT scan was done in every patient of stroke. Patients were treated with conventional heparin, aspirin, clopidogrel, blood pressure management, ionotropic support if required, Urine examination, Lipid Profile, Serum Homocysteine were carried out in all the patients and controls.

 

Sample collection and Preparations¹⁰: The Serum Homocysteine levels and Lipid Profiles were measured 8 to 12 weeks after the attack of Ischemic stroke. Serum Homocysteine levels and Lipid Profiles were measured in a 12 hour fasting blood sample in both the case and control groups. A patient, who has had a serious illness such as MI or stroke, should not be tested for 8 to 12 weeks after the event because there is a potential for the Homocysteine level to be abnormally high during that time period. The reason for this has not been elucidated. 5 ml of blood was collected in a plain bulb for the estimation of Homocysteine and Lipid profile. 2 ml of blood was collected in Fluoride bulb for blood sugar estimation. 10 to 12 hours of fasting prior to sample collection is required. EDTA plasma or serum is preferred. Blood should be centrifuged within 30 minutes or kept on ice until centrifugation.

Methods for different Parameters: Serum cholesterol was estimated by using Cholesterol Oxidase peroxidase (CHOD-PAP) method. Serum triglycerides estimated by GPO method. Serum HDL-C was estimated by using Phosphotungstic Acid (PTA) method, whereas Serum LDL-C was calculated by using Friedwald’s Equation. Serum Homocysteine was measured by Fluorescence Polarization Immunoassay method by Ax SYM Assay system using the principle of conversion of oxidized Homocysteine to reduced form and then converting into S-adenosyl-L-Homocysteine (SAH)

Data Analysis: The data were obtained and plotted using a bar diagram. Four important data were obtained indicating the comparison of serum Homocysteine in case and control group as well as comparing serum Homocysteine with LDL-C, HDL-C, and cholesterol in case groups. Statistical analysis was done and p value was calculated using t-test.
RESULTS AND OBSERVATIONS

Table 1: s. Homocysteine levels in case and control groups with ischemic stroke

The table 1 shows that the mean serum homocysteine in case and control groups was 28.14 µmol/L and 11.18 µmol/L, respectively. The standard deviation in case and control groups was7.88 µmol/L and 1.82 µmol/L, respectively

 

Table 2: Relation of Homocysteine to LDL–C in Case Group with Ischemic Stroke

The table 2 shows that the mean homocysteine level in patients with LDL-C > 100 mg % was 26.06 µmol/L and in patients with LDL-C < 100 mg % was 30.22 µmol/L. The standard deviation in case and control groups was 8.42µmol/L and 8.95 µmol/L, respectively

 

Table 3: Relation of Homocysteine to HDL–C in Case Group with Ischemic Stroke

The table 3 shows that the mean homocysteine level in patients with HDL-C > 40 mg % was 28.98 µmol/L and in patients with HDL-C < 40 mg % was 22.67 µmol/.

 

Table 4: Relation of Homocysteine to Total Cholesterol in Case Group with Ischemic Stroke

The table 4 shows that the mean homocysteine level in patients with S. Cholesterol > 200 mg % was 28.3µmol/L and in patients with S. Cholesterol < 200 mg % was 28.07 µmol/L.

DISCUSSION

JAMA, 2002 reports a meta-analysis¹¹ in that evaluated data from 30 prospective and retrospective studies, involving 1113 stroke, provides the best analysis of the varied data. Stronger associations between the blood homocysteine concentration and cerebrovascular events were noted in retrospective compared with prospective studies. After adjustment for known risk factors, a 25 percent lower homocysteine concentration (about 3 µmol/L) in the prospective studies was associated with a 19% lower stroke risk (odds ratio 0.81, 95 percent CI 0.69 to 0.95).Jun-Hyun yoo et al.¹², reports that the prevalence of moderate hyperhomocystenemia was 3.3 fold higher among patients with cerebral infarction compared with normal controls. Kim NK et al.¹³, reports that mean plasma homocysteine level were significantly higher (p<0.05) in patients with silent brain infarction than in normal healthy people. Brattstrom et al¹⁴, shows that the prevalence of hyperhomocystenemia was 6.6 folds higher among patients with cerebral infarction compared with normal controls. The difference between the 2 groups in our study was highly significant (p value=0.0001) suggesting homocysteine as an important predictor of cerebral infarction.

Relation of homocysteine to LDL–C in case group with ischemic stroke: The difference observedbetween the 2 groups in MI was not significant in our study. (p value = 0.2005). Hence, in this study, the low levels of LDL -C did not protect the patients against the homocysteine induced stroke.

Relation of homocysteine to HDL–C in case group with ischemic stroke: The difference observedin the 2 groups in MI was not statistically significant. (p value = 0.1519). Hence in our study, the higher levels of HDL-C in did not protect the patients against the homocysteine induced stroke.

Relation of homocysteine to total cholesterol in case group with ischemic stroke: The difference observed in the 2 groups of MI was statistically not significant (p value =0.9492) suggesting no significant correlation between serum cholesterol and serum Hcy in patients of ischemic stroke.

 

CONCLUSION

From this study it can be concluded that in young patients with ischemic stroke the low levels of LDL-C and high levels of HDL-C did not protect against the homocysteine induced Ischemic stroke and homocysteine is an important and independent risk factor of ischemic stroke.

 

REFERENCES

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