Stroke is defined as an "acute neurologic dysfunction of vascular origin with sudden (within seconds) or at least rapid (within hours) occurrence of symptoms and signs corresponding to the involvement of focal areas in the brain". The two main types of strokes are ischemic and hemorrhagic which are due to a number of different pathological mechanisms. ^[2] Hemorrhagic stroke occurs when blood from a ruptured blood vessel compresses and damages normal functioning brain tissue. In an ischemic stroke, a blocked artery prevents blood carrying oxygen and other nutrients from reaching a portion of the brain, leading to dysfunction and death of that brain tissue. ^[3,4] 

Stroke is the 3rd leading cause accounting for 10% of all death in the world (WHO, 2024). It is a major global health issue that prematurely claims millions of otherwise healthy and productive lives each year. ^[5]  According to WHO, Annually, 15 million people worldwide suffer a stroke. Of these, 5 million die and another 5 million are left permanently disabled, placing a burden on family and community. ^[6] Strokes are rare in younger people and become increasingly common with older age but at the age of 45 years, the chances of having stroke in the next 20 years is 1 in 30. There has been a definite increase in the prevalence and incidence of stroke disorder in India over the last 30 years. ^[7]

Stroke represented 1.2% of total deaths in India. The average annual incidence rate of stroke in India currently is 145 per 100,000 population, which is higher than the western nations and Indians may also be genetically prone for stroke due to high prevalence of metabolic syndrome. ^[8] Dysfunction of the brain manifests itself by various neurological deficits by various signs and symptoms that are related to the extent and site of the area of brain involved. ^[9] These include coma, hemiplegia, paraplegia, monoplegia, multiple paralysis, speech disturbances, nerve paresis, sensory impairment etc. Ischemic and hemorrhagic stroke accounts for approximately 85% and 15%, respectively. ^[10]

Oxidative stress is a mechanism involved in nerve damage caused by stroke and is the result of an imbalance between the production of free radicals (reactive oxygen species) and the antioxidant defense system. ^[11] This phenomenon leads to cellular damage, cell death and acceleration of degenerative diseases associated with aging such as cancer, cardiovascular disease, diabetes, and pulmonary and neural degenerative diseases. ^[12]

Increased production of free radicals and other chemical species has been confirmed both in ischemic and hemorrhagic strokes, and oxidative stress was introduced as a fundamental mecha nism in brain damage under these conditions. ^[13] Free radicals change the structure and function of target molecules by taking their electrons. Oxidants also effect cell membranes and genetic material such as DNA and RNA, and various enzymatic events, causing cell damage during ischemia and reperfusion. ^[14]

This is a Prospective, Observational and Single center  conducted in the Department of Biochemistry and Neurology, Index Medical College Hospital & Research Center

In this study, total 220 subjects were included; of which 110 newly diagnosed ischemic stroke patients of either sex, who  clinically diagnosed in the department of Neurology and 110 newly diagnosed haemorrhagic stroke.

Patient Selection:

After clinical examination and confirmed diagnosis by Physician, 220 patients of either sex, who meet the inclusion and exclusion criteria selected for the study from January 2022 to July 2024.

Ischemic stroke patients

• Magnetic resonance imaging (MRI) proven ischemic stroke patients attending the neurology Department
• For the assessment of stroke subtypes the classification of the trial of ORG 10172 in acute stroke treatment (TOAST)
• A detailed medical history on risk factors namely systemic hypertension (systolic blood pressure (SBP)> 140 mm Hg and diastolic blood pressure (DBP)>90 mm Hg, diabetes (Fasting blood sugar > 106 mg/dl, 2h post prandial blood sugar > 200 mg/dl or as oral hypoglycemic or insulin therapy, dyslipidemia, smoking, alcohol consumption, tobacco chewing, family history of stroke
• Patients were then finally grouped in to 3 groups: large artery disease, small vessel disease group and a group of strokes with other determined and unknown etiology.
• Due to their small sample size, we have only included large artery and small vessel disease group for performing statistical analysis of subgroups of ischemic stroke.

Patients were included on the basis of inclusion and exclusion criteria as follows:

Inclusion criteria:

• MRI proven Ischemic stroke
• Age above 40 years
• Place of birth and ethnicity - North India.

Exclusion criteria:

The patients with following disorders were excluded

• Cardio embolic stroke
• Liver and kidney failure
• Thyroid disorder and
• Cerebral venous sinus thrombosis

Hemorrhagic stroke patients

• Patients with computerized tomography (CT) proven intracerebral hemorrhage examined by the neurologist to confirm the diagnosis and  included in the study.
• CT scans were reviewed for the location of hematoma.
• A detailed medical history reviewed on blood pressure, diabetes mellitus, smoking, alcohol consumption, tobacco chewing and family history of stroke. Magnetic resonance angiography (MRA) or digital subtraction angiography (DSA)  done to exclude the vascular malformations.
• Patients were grouped in to 2 major groups namely; lobar and non lobar hemorrhages.

Patients were included on the basis of inclusion and exclusion criteria as follows:

Inclusion criteria:

• CT proven Hemorrhage,
• Above 40 years of age
• Place of birth and ethnicity- North India

Exclusion criteria: patients with

• Head injury
• Vascular malformation,
• Tumour bleed
• Vasculopathy
• Subarachnoid hemorrhage

We have excluded subarachnoid hemorrhage cases under hemorrhagic group as to maintain biological plausibility of hemorrhagic stroke studied in our case control analysis because subarachnoid hemorrhage is so different regarding pathophysiology, clinical picture and management that is often discussed separately.

Healthy Controls:

Healthy control group comprised of age and sex- matched healthy volunteers free of any neurological disorder. We have excluded the volunteers who had any sort of cardiac disease, diabetes, and hypertension.

Inclusion criteria

• Absence of stroke
• Free from any neurological disorder, cardiac disease
• Age, gender and ethnicity matched Exclusion criteria
• Anybody suffering from following
• Hypertension
• Diabetes
• No family history of stroke
• Any other chronic debilitating disease

       Full examination (general and neurological) including evaluation of stroke severity on admission using: The National Institute of Health Stroke Scale (NIHSS) was performed for initial evaluation of stroke severity, within the first 24 h on admission, in this study, a score of (0) is measured for no stroke symptoms, (1-4) is measured for minor, (5-15) is measured for moderate, (16-20) is measured for moderate to severe, and (21-42) measured for severe.

       Follow-up assessment of the patients: Modified rankin scale (mRS) was performed after 3 months post stroke to assess short-term outcome and dis- ability of stroke. Modified rankin scale scores the disability of patients into points from 0-6 according to the degree of disability. Patients with score < 2 were considered had favorable outcome, while patients with score 2-5 were considered had unfavor- able outcome and dead patients had score 6, in this study a score of (6) is measured for dead, (2-5) is measured for unfavorable outcome and (< 2) is measured for favorable outcome.

Collection of blood samples:

All the volunteer patients and normal healthy individuals (Control group) were told to fast (overnight) 12 – 14 hours, and then with all aseptic precautions the venous blood withdrawn from the anterior cubital vein, in fluoride, plain bulb and heparinised tubes for biochemical measurements. One & half hours after the meal, a second blood samples were withdrawn in a similar way for measurement of post prandial blood glucose level. For both samples, blood allowed to clot at room temperature for about 30 minutes and then centrifuged at 3000 rpm for 10 minutes. The separated

STATISTICAL ANALYSIS

At the end of the data collection, all statistical analysis done using window-based software SPSS, Version 29. Results were expressed as arithmetic means ± SD (Standard deviation) Before and after treatment levels difference between each group were evaluated using, paired student ‟t‟ test and considered statistically significant at probability values less than 0.05 (P ≤ 0.05). One-Way analysis of variance (ANOVA) done to assess the significance within and between groups.

Table 1:  Distribution of Age of the cases

In table 1, 51-60 age group shows the highest frequency for both ischemic (39.0%) and hemorrhagic strokes (43.6%). The 31-40 age group has the lowest frequencies for both types of stroke. There is a notable decline in hemorrhagic stroke cases in the 81-90 age group (2.72%).

Table 2: Distribution of the Malondialdehyde (MDA) among Ischemic stroke group and Haemorrhagic stroke

In table 2, In our study, we observed that mean Ischemic Stroke: MDA levels are significantly higher, with a mean of 2.83 ± 0.43 nmol/mL. Hemorrhagic Stroke: MDA levels are lower, with a mean of 2.29 ± 0.39 nmol/mL. This suggests that oxidative stress, as indicated by MDA levels, may differ between the two types of strokes. Higher MDA levels in ischemic stroke could reflect increased lipid peroxidation and oxidative damage.

Table 3: Distribution of the Nitric oxide (NO) among Ischemic stroke group and Haemorrhagic stroke

In table 3, Ischemic Stroke: NO levels are higher, with a mean of 5.02 ± 0.41 µg/mL. Hemorrhagic Stroke: NO levels are slightly lower, with a mean of 4.83 ± 0.51 µg/mL. The difference in NO levels may indicate variations in vascular responses or inflammatory processes between the two types of strokes. There is an increase in Nitric oxide in ischemic stroke (ISPs) and Haemorrhagic stroke patients when compared to control subjects. This indicates that lipid peroxidation is significantly increased in Ischemic stroke than Haemorrhagic stroke patients.

Table 4: Distribution of the Glutathione peroxidase (GPX) among Ischemic stroke group and Haemorrhagic stroke

In table 4, Ischemic Stroke: GPX levels are higher, with a mean of 4.39 ± 0.41 μmol/mL. Hemorrhagic Stroke: GPX levels are lower, with a mean of 4.21 ± 0.29 μmol/mL. The difference in GPX levels suggests variations in antioxidant defense mechanisms between the two types of strokes. Higher GPX levels in ischemic stroke may reflect a compensatory response to increased oxidative stress.

Table 5: Distribution of the Uric acid (mg/dl) among Ischemic stroke group and Haemorrhagic stroke

In table 5, Ischemic Stroke: Uric acid levels are higher, with a mean of 7.91 ± 0.73 mg/dL. Hemorrhagic Stroke: Uric acid levels are lower, with a mean of 7.01 ± 0.83 mg/dL. The elevated uric acid levels in ischemic stroke may indicate increased oxidative stress or inflammatory responses, as uric acid can act as both an antioxidant and a pro-oxidant depending on the context.

Table 6: Distribution of the Superoxide dismutase (SOD) (U/mg) among Ischemic stroke group and Haemorrhagic stroke

In table 6, Ischemic Stroke: SOD levels are higher, with a mean of 9.89 ± 0.93 U/mg. Hemorrhagic Stroke: SOD levels are lower, with a mean of 9.01 ± 0.93 U/mg. The increased SOD levels in ischemic stroke may reflect a heightened antioxidant response to counteract oxidative stress, while the lower levels in hemorrhagic stroke might indicate a different balance of oxidative stress and antioxidant defenses.

Table 7: Distribution of the Catalase (IU/mg) among Ischemic stroke group and Haemorrhagic stroke

In table 7, Ischemic Stroke: Catalase levels are lower, with a mean of 9.01 ± 0.73 IU/mg. Hemorrhagic Stroke: Catalase levels are higher, with a mean of 9.89 ± 0.83 IU/mg. This indicates that individuals with hemorrhagic stroke may have a stronger catalase response to oxidative stress, while those with ischemic stroke show lower levels, which might suggest a different oxidative stress profile or antioxidant capacity in the two conditions.

Table 8: Distribution of the Total Antioxidant capacity (TAC) (mmol/L) among Ischemic stroke group and Haemorrhagic stroke

In table 8, Ischemic Stroke: TAC levels are higher, with a mean of 1.89 ± 0.41 mmol/L. Hemorrhagic Stroke: TAC levels are lower, with a mean of 1.21 ± 0.31 mmol/L. The elevated TAC in ischemic stroke suggests a greater overall capacity to counteract oxidative stress compared to hemorrhagic stroke, which may have lower antioxidant defenses. This difference could reflect distinct metabolic and inflammatory responses in the two types of strokes.

Table 9: Distribution of the Total Antioxidant capacity (TAC) (mmol/L) among Ischemic stroke group and Haemorrhagic stroke

In Table 9, Ischemic Stroke: Thiol group levels are higher, with a mean of 332.35 ± 31.64 mmol/L. Hemorrhagic Stroke: Thiol group levels are lower, with a mean of 318.12 ± 28.55 mmol/L. The higher levels of thiol groups in ischemic stroke may indicate a greater presence of these important antioxidants, which can help protect against oxidative damage. In contrast, the lower levels in hemorrhagic stroke might suggest a reduced capacity for antioxidant defense in that condition.

In our study, 51-60 age group shows the highest frequency for both ischemic (39.0%) and hemorrhagic strokes (43.6%). The 31-40 age group has the lowest frequencies for both types of strokes. There is a notable decline in hemorrhagic stroke cases in the 81-90 age group (2.72%). These findings matched with most of the previous studies. ^[15] 

In our study we observed that mean Ischemic Stroke: MDA levels are significantly higher, with a mean of 2.83 ± 0.43 nmol/mL. Hemorrhagic Stroke: MDA levels are lower, with a mean of 2.29 ± 0.39 nmol/mL. This suggests that oxidative stress, as indicated by MDA levels, may differ between the two types of strokes. Higher MDA levels in ischemic stroke could reflect increased lipid peroxidation and oxidative damage. InimioaraM et al. ^[6] observed significantly higher concentration of MDA in stroke patients compared with controls, similar to our study and suggested that increased level of lipid peroxides may be due to oxidation of blood or neural lipids by ischemia.

We found Ischemic Stroke: NO levels are higher, with a mean of 5.02 ± 0.41 µg/mL. Hemorrhagic Stroke: NO levels are slightly lower, with a mean of 4.83 ± 0.51 µg/mL. The difference in NO levels may indicate variations in vascular responses or inflammatory processes between the two types of strokes. There is an increase in Nitric oxide in ischemic stroke (ISPs) and Haemorrhagic stroke patients when compared to control subjects. This indicates that lipid peroxidation is significantly increased in Ischemic stroke than Haemorrhagic stroke patients.

We noticed, Ischemic Stroke: GPX levels are higher, with a mean of 4.39 ± 0.41 μmol/mL. Hemorrhagic Stroke: GPX levels are lower, with a mean of 4.21 ± 0.29 μmol/mL. The difference in GPX levels suggests variations in antioxidant defense mechanisms between the two types of strokes. Higher GPX levels in ischemic stroke may reflect a compensatory response to increased oxidative stress.

Several GPx isoforms are known, differing in their location and subunit structure. There are GPx1-4, selenoperoxidases with selenocysteine in the active site, cytosolic glutathione peroxidase (GPx1), gastrointestinal peroxidase (GPx2), extracellular peroxidase located in the serum (GPx3) and glutathione peroxidase of phospholipid and cholesterol peroxides (GPx4). In addition, there is Gpx5, which contains cysteine instead of selenocysteine, acts as a secretory enzyme of the epididymis, GPx6, a selenoprotein created by the olfactory epithelium, and the low-activity peroxidases GPx7 and GPx8. ^[17]

In this study, Ischemic Stroke: Uric acid levels are higher, with a mean of 7.91 ± 0.73 mg/dL. Hemorrhagic Stroke: Uric acid levels are lower, with a mean of 7.01 ± 0.83 mg/dL. The elevated uric acid levels in ischemic stroke may indicate increased oxidative stress or inflammatory responses, as uric acid can act as both an antioxidant and a pro-oxidant depending on the context.

We found Ischemic Stroke: SOD levels are higher, with a mean of 9.89 ± 0.93 U/mg. Hemorrhagic Stroke: SOD levels are lower, with a mean of 9.01 ± 0.93 U/mg. The increased SOD levels in ischemic stroke may reflect a heightened antioxidant response to counteract oxidative stress, while the lower levels in hemorrhagic stroke might indicate a different balance of oxidative stress and antioxidant defenses.

Srikrishna R et al. ^[18] observed reduced SOD in cases 4.04±0.03U/ml as compared to 9.01±1.04 U/ml in controls. Spranger M et al. ^[19] found that mean serum levels of SOD in cases of both ischemic and hemorrhagic stroke were significantly lower as compared to controls suggesting that antioxidants are depleted as a consequence of an excessive production of oxygen free radicals very early after the onset of stroke.

Cherubini et al. ^[20] found that antioxidants including SOD are reduced immediately after an acute stroke, possibly as a consequence of increased oxidative stress and A specific antioxidant profile is associated with a poor early outcome .Thus our findings of serum SOD levels matched with previous studies. Hence increasing the anti oxidative capacity in serum within the first day after the onset of symptoms might be a therapeutic option to minimize the oxidative injury caused by oxygen free radicals until the endogenous free radical scavenging systems recovers.

We found that Ischemic Stroke: Catalase levels are lower, with a mean of 9.01 ± 0.73 IU/mg. Hemorrhagic Stroke: Catalase levels are higher, with a mean of 9.89 ± 0.83 IU/mg. This indicates that individuals with hemorrhagic stroke may have a stronger catalase response to oxidative stress, while those with ischemic stroke show lower levels, which might suggest a different oxidative stress profile or antioxidant capacity in the two conditions.

In conclusion, we determined that diabetes mellitus brings an additive oxidative stress load to acute ischemic stroke patients. These patients need to be managed carefully with regard to their poor prognosis. We consider that high TAC levels in diabetic stroke patients render the antioxidant supplementation useless at least for the acute-phase (24 hours) treatment of stroke. Oxidative stress and TAC in the later periods of acute ischemic stroke need to be explored in further studies.

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