Oxidative stress and post-ischemic inflammatory response are the key pathogenic mechanisms of the neuronal injury caused by ischemic stroke (IS). On the other hand, Diabetes is a major risk factor for the development of stroke, increasing the susceptibility to atherosclerosis and the prevalence of atherogenic risk factors, including hypertension, obesity, and abnormal blood lipids. The aim of current study was to analyze the functional activity of oxidant-antioxidant system and post-ischemic inflammatory response in IS patients complicated and noncomplicated with diabetes mellitus type 2 (DM2). ELISA, photochemiluminescent and spectrophotometric methods were used to analyze the serum samples of IS patients complicated and noncomplicated with DM2, DM2 patients, as well as healthy subjects. The results obtained suggest that IS complicated with diabetes is characterized by higher intensity of the lipid peroxidation process as compared to IS noncomplicated with diabetes that, probably, is one of the determining factors responsible for more severe clinical course of IS patients complicated with DM2 compared to those noncomplicated with DM2. It is also shown that mechanisms of the compensatory response to oxidative stress on the level of antioxidants in IS patients complicated with diabetes differ from those detected in IS noncomplicated with diabetes. A significant increase of the levels of the five analyzed cytokines in both groups of IS patients were detected. Based on the results obtained we suggest that metabolic, molecular, and cellular level alterations are typical for long-term DM2 impair compensatory mechanisms protecting the body from oxidative stress, and that in IS complicated with DM2 the systemic inflammatory reactions and oxidative stress are more intense than in case of IS noncomplicated with DM2.
Stroke is the third leading cause of death and disability worldwide after heart disease and cancer and is the major cause of morbidity, particularly in the middle aged and elderly population [
Oxidative stress (OS) and postischemic inflammatory response are the key pathogenic factors leading to uncontrolled cell damage and death, which badly influences stroke progression and outcome [
The present study aimed to reveal the molecular mechanisms responsible for the development of OS and inflammatory reactions in human IS on the systemic level and to identify the molecular components involved in the above mentioned processes. To assess the peculiarities in the development of systemic OS we measured the blood levels of the oxidized derivatives of lipids. To evaluate the functional state of antioxidant system we determined the total capacity of low-molecular non-enzymatic water-soluble antioxidants (TAC) and ferroxidase activity of ceruloplasmin (FAC) in the blood of studied groups. Then, to study the inflammatory reactions we assessed the state of cytokine network in acute ischemic stroke and comparison of functional activities of proinflammatory and chemotactic cytokines, including IL-1b, IL-6, TNF-a, MCP-1 and CXCL1, in IS complicated and noncomplicated with DM2 in postischemic inflammatory response progression.
In total 90 IS patients complicated with DM2 (mean age ± SD: 65 ± 10 years, females/males: 39/51), 120 IS patients noncomplicated with DM2 (mean age ± SD: 65 ± 13 years, females/males: 66/54), 110 patients with DM2 (mean age ± SD: 59 ± 8 years, females/males: 63/47) and 116 healthy subjects (mean age ± SD: 60 ± 12 years, females/males: 69/47) were enrolled in this study (
Patients with large vessel atherothromboembolic stroke (n = 171) and cardioembolic stroke (n = 39) were selected for this study; those with lacunar stroke syndromes were excluded from the study group. Severity of neurological deficit was defined using the National Institutes of Health Stroke Scale (NIHSS). In the present study, patients with a moderate to severe impairment (average NIHSS score 17) were involved. Healthy subjects (controls) were volunteers from the institutes of NAS RA reported no personal or family history of IS, myocardial infarction, and any other cerebrovascular or cardiovascular disease. They had no serious medical disorder or treatment during the past 12 months. Exclusion criteria for all subjects included chronic inflammation or/and infectious diseases, present or past history of metabolic (diabetes mellitus, etc), neuropsychiatric, immune system and oncological disorders, myocardial infarction or any other serious medical conditions. All subjects or their
Study Groups | IS complicated with DM2 | IS non complicated with DM2 | DM2 | HS |
---|---|---|---|---|
n | 90 | 120 | 110 | 116 |
Age (М ± SD), years | 65 ± 10 | 65 ± 13 | 59 ± 8 | 60 ± 12 |
Female/male | 39/51 | 66/54 | 63/47 | 69/47 |
p values are given in the text.
legal representatives gave their informed consent to participate in the study, which was approved by the Ethical Committee of the Institute of Molecular Biology NAS RA (IRB #00004079).
Practically fasting blood samples were collected by venipuncture in appropriate tubes and kept on ice for 60 min. After that the coagulated blood was centrifuged at 3000 g for 15 min at 4˚C to separate serum from blood corpuscles. The obtained serum samples were stored in aliquots at −30˚C and thawed immediately prior to use.
The contents of lipid hydroperoxides and the FAC in the blood serum samples were determined by spectrophotometric assays published earlier [
Data statistics was performed by Mann-Whitney’s U-test using the “Graphpad Prism” (GraphPad Software Inc., USA) software package. Values of p < 0.05 were accepted as statistically significant.
The results obtained for the peculiarities in the development of systemic OS showed significantly increased levels of lipid hydroperoxides in the serum of all three patients groups as compared to the group of HS (
The results obtained for the functional state of antioxidant system (
Concerning the FAC significant differences were found only in the blood serum of IS patients complicated with DM2 (
The results obtained in the study of cytokine system showed that the levels of all 5 cytokines measured (IL-1b, IL-6, TNF-a, MCP-1 and CXCL1) were higher in diseased groups compared to HS (
With regard to IL-6, its levels were 3 and 2 times significantly higher in the blood serum of IS patients complicated and noncomplicated with DM2, respectively, compared to HS (p < 0.05), while the levels of IL-6 in DM2 patients (М ± SD: 19.2 ± 3.8 pg/ml) did not significantly differ from those of HS (М ± SD: 18.4 ± 3.1 pg/ml). Though the levels of IL-6 were found to be 1.51 times significantly higher in IS patients complicated with DM2 compared to IS patients noncomplicated with DM2 (p < 0.05)
According to the results obtained for TNF-a, its levels were 5.2 and 3.7 times significantly increased in IS
Study Groups | Content of lipid hydroperoxides, А480 | TAC, mmol/L | FAC, Umol/L/min |
---|---|---|---|
IS complicated with DM2 | 0.12 ± 0.04 | 2.8 ± 0.9 | 574 ± 129 |
IS noncomplicated with DM2 | 0.09 ± 0.03 | 3.4 ± 1.1 | 475 ± 107 |
DM2 | 0.09 ± 0.03 | 2.6 ± 0.8 | 483 ± 111 |
HS | 0.03 ± 0.01 | 2.5 ± 0.8 | 429 ± 94 |
p values are given in the text.
Study Groups | [IL-1b] | [IL-6] | [TNF-a] | [MCP-1] | [CXCL1] |
---|---|---|---|---|---|
IS complicated with DM2 | 45.4 ± 9.1 | 55.1 ± 10.8 | 69.00 ± 13.7 | 855 ± 140.4 | 180 ± 31.6 |
IS noncomplicated with DM2 | 28.8 ± 4.8 | 36.5 ± 6.7 | 50.00 ± 9.6 | 600 ± 101.7 | 120 ± 20.7 |
DM2 | 6.0 ± 1.1 | 19.2 ± 3.8 | 15.00 ± 2.9 | 280 ± 49.3 | 78 ± 13.2 |
HS | 5.1 ± 0.8 | 18.4 ± 3.1 | 13.37 ± 2.5 | 254 ± 46.2 | 74 ± 12.9 |
p values are given in the text.
patients both complicated and noncomplicated with DM2, respectively, compared to HS (p < 0.05). However, we again did not found any significant differences between its levels in DM2 patients (М ± SD: 15.00 ± 2.9 pg/ml) and HS (М ± SD: 13.37 ± 2.5 pg/ml), but we found significantly high levels (1.38 times) of TNF-a between IS patients complicated and noncomplicated with DM2 (p < 0.05).
The data obtained for MCP-1 showed significantly high levels (p < 0.05) in IS complicated and noncomplicated with DM2 (3.4 and 2.4 times, respectively), and no difference between the DM2 (М ± SD: 280 ± 49.3 pg/ml) and HS (М ± SD: 254 ± 46.2 pg/ml). However, the levels of MCP-1 were 1.43 times higher in IS complicated with DM2 than in IS noncomplicated with DM2 (p < 0.05).
Finally, the levels of CXCL1 were 2.43 and 1.62 times significantly increased in IS patients complicated and noncomplicated with DM2 (p < 0.05), while they did not differ in DM2 patients (М ± SD: 180 ± 31.6 pg/ml) and HS (М ± SD: 120 ± 20.7 pg/ml). Moreover, the levels of CXCL1 were 1.5 times significantly increased in IS complicated with DM2 than in IS noncomplicated with DM2 (p < 0.05).
Based on the results obtained we suggest that metabolic, molecular, and cellular level alterations are typical for long-term DM2 impair compensatory mechanisms protecting the body from OS, and that in IS complicated with DM2 the systemic inflammatory reactions and OS are more intense than in case of IS noncomplicated with DM2.
The authors express thanks to administration and physicians of Medical Centers of the Ministry of Health of the Republic of Armenia as well as all healthy and diseased subjects involved in this study. This study was supported by the State Committee of Science of Republic of Armenia and the National Foundation of Science and Advanced Technologies.
Gohar Tsakanova,Elina Arakelova,Armine Soghoyan,Violetta Ayvazyan, (2015) Oxidative Stress and Post-Ischemic Inflammatory Response in Ischemic Stroke Complicated with Diabetes Mellitus Type 2. Journal of Biosciences and Medicines,03,94-98. doi: 10.4236/jbm.2015.33014