4-benzyl-2-methyl-1-2-4-thiadiazolidine-3-5-dione and Brain-Ischemia

Diabetes aggravates brain injury after cerebral ischemia/reperfusion (I/R).. To investigate whether limb I/R causes cerebral injury in a rat diabetes model and whether glycogen synthase kinase-3β (GSK-3β) is involved.. Male adult Sprague-Dawley rats were assigned into streptozotocin-induced diabetes (n = 30; blood glucose ≥16.7 mmol/L) or control (n = 20) groups, further subdivided into diabetes I/R (3-hour femoral artery/vein clamping), diabetes-I/R + TDZD-8 (I/R plus GSK-3β inhibitor), diabetes-sham, control-sham and control-I/R groups (n = 10 each). Cortical and hippocampal morphology (hematoxylin/eosin); hippocampal CA1 apoptosis (TUNEL assay); cleaved caspase-3 (apoptosis), and Iba1 (microglial activation) protein expression (immunohistochemistry); phosphorylated/total GSK-3β and nuclear factor-κB (NF-κB) protein levels (Western blotting); and serum and brain tissue tumor necrosis factor (TNF)-α levels (enzyme-linked immunosorbent assay) were analyzed.. The diabetes-I/R group showed greater cortical and hippocampal injury, apoptosis, cleaved caspase-3 expression and Iba1 expression than the control-I/R group; TDZD-8 reduced injury/apoptosis and cleaved caspase-3/Iba1 expressions. The diabetes-I/R group had lower p-GSK-3β and p-NF-κBp65 expression than the control-I/R group (P < 0.05); TDZD-8 increased p-GSK-3β expression but decreased p-NF-κBp65 expression (P < 0.05). The diabetes-I/R group showed higher elevation of serum and brain tissue TNF-α than the control-I/R group (P < 0.05); TDZD-8 reduced TNF-α production.. Diabetes exacerbates limb I/R-induced cerebral damage and activates NF-κB and GSK-3β.

Topics: Animals; Apoptosis; Brain Ischemia; Cerebral Cortex; Diabetes Mellitus, Experimental; Diabetic Angiopathies; Extremities; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hippocampus; Male; NF-kappa B; Rats, Sprague-Dawley; Reperfusion Injury; Thiadiazoles

There is evidence that insulin reduces brain injury evoked by ischemia/reperfusion (I/R). However, the molecular mechanisms underlying the protective effects of insulin remain unknown. Insulin is a well-known inhibitor of glycogen synthase kinase-3beta (GSK-3beta). Here, we investigate the role of GSK-3beta inhibition on I/R-induced cerebral injury in a rat model of insulinopenic diabetes.. Rats with streptozotocin-induced diabetes were subjected to 30-min occlusion of common carotid arteries followed by 1 or 24 h of reperfusion. Insulin (2-12 IU/kg i.v.) or the selective GSK-3beta inhibitor TDZD-8 (0.2-3 mg/kg i.v.) was administered during reperfusion.. Insulin or TDZD-8 dramatically reduced infarct volume and levels of S100B protein, a marker of cerebral injury. Both drugs induced phosphorylation of the Ser9 residue, thereby inactivating GSK-3beta in the rat hippocampus. Insulin, but not TDZD-8, lowered blood glucose. The hippocampi of the drug-treated animals displayed reduced oxidative stress at 1 h of reperfusion as shown by the decreased generation of reactive oxygen species and lipid peroxidation. I/R-induced activation of nuclear factor-kappaB was attenuated by both drug treatments. At 24 h of reperfusion, TDZD-8 and insulin significantly reduced plasma levels of tumor necrosis factor-alpha; neutrophil infiltration, measured as myeloperoxidase activity and intercellular-adhesion-molecule-1 expression; and cyclooxygenase-2 and inducible-NO-synthase expression.. Acute administration of insulin or TDZD-8 reduced cerebral I/R injury in diabetic rats. We propose that the inhibitory effect on the activity of GSK-3beta contributes to the protective effect of insulin independently of any effects on blood glucose.

Topics: Animals; Blood Glucose; Blotting, Western; Body Weight; Brain Ischemia; Diabetes Mellitus, Experimental; Enzyme-Linked Immunosorbent Assay; Gene Expression; Glycogen Synthase Kinase 3; Hippocampus; Hormones; Hypoglycemic Agents; Insulin; Lipid Peroxidation; Male; NF-kappa B; Oxidative Stress; Phosphorylation; Rats; Rats, Wistar; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Thiadiazoles; Tumor Necrosis Factor-alpha

The serine/threonine glycogen synthase kinase 3beta (GSK-3beta) is abundant in the central nervous system, particularly in the hippocampus, and plays a pivotal role in the pathophysiology of a number of diseases, including neurodegeneration. This study was designed to investigate the effects of GSK-3beta inhibition against I/R injury in the rat hippocampus. Transient cerebral ischemia (30 min) followed by 1 h of reperfusion significantly increased generation of reactive oxygen species and modulated superoxide dismutase activity; 24 h of reperfusion evoked apoptosis (determined as mitochondrial cytochrome c release and Bcl-2 and caspase-9 expression), resulted in high plasma levels of TNF-alpha and increased expression of cyclooxygenase-2, inducible nitric oxide synthase, and intercellular adhesion molecule-1. The selective GSK-3beta inhibitor, 4-benzyl-2-methyl-1,2,4-thiadiazolidine-3,5-dione (TDZD-8), was administered before and after ischemia or during reperfusion alone to assess its potential as prophylactic or therapeutic strategy. Prophylactic or therapeutic administration of TDZD-8 caused the phosphorylation (Ser(9)) and hence inactivation of GSK-3beta. Infarct volume and levels of S100B protein, a marker of cerebral injury, were reduced by TDZD-8. This was associated with a significant reduction in markers of oxidative stress, apoptosis, and the inflammatory response resulting from cerebral I/R. These beneficial effects were associated with a reduction of I/R-induced activation of the mitogen-activated protein kinases JNK1/2 and p38 and nuclear factor-kappaB. The present study demonstrates that TDZD-8 protects the brain against I/R injury by inhibiting GSK-3beta activity. Collectively, our data may contribute to focus the role of GSK-3beta in cerebral I/R.

Topics: Animals; Antioxidants; Brain; Brain Ischemia; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hippocampus; Male; Mitochondria; Models, Biological; Nerve Growth Factors; Phosphorylation; Rats; Rats, Wistar; Reperfusion Injury; S100 Calcium Binding Protein beta Subunit; S100 Proteins; Thiadiazoles