Abstract WP151: Ischemic Injury in Diabetes Augments Cerebral Microvascular Matrix Metalloproteinase Activation in Female Rats

Abstract

Introduction: Diabetic female patients are at greater risk of stroke and worsened recovery. We have previously shown that matrix metalloprotease 3 (MMP3) exacerbates hemorrhagic transformation (HT) in hyperglycemic stroke in male rats while high glucose and hypoxia increases MMP3 activity in brain microvascular endothelial cells (BMVECs). Furthermore, diabetic female rats develop greater HT. Current study was designed to test the hypothesis that ischemic stroke increases the cerebral microvascular MMP3 activity in diabetic female animals. Methods: Female rats after 7-8 weeks of diabetes (high fat diet and low dose of streptozotocin) were subjected to 60 min MCAO. After 3 days, adhesive removal time (ART) and behavioral composite score were measured and cerebral microvessels were isolated. Human female BMVECs were grown under normal (5.5mM glucose) or diabetic condition (25mM glucose plus 100μM sodium palmitate) for 48 hours, followed by hypoxia insult for 12 hours (200μM cobalt chloride). Cell lysate, media and microvascular homogenates were analyzed for MMP profiling by immunoblotting, zymography and MMP3 enzyme activity by fluorescence resonance energy transfer (FRET) assay. Results: MMP3 protein expression and activity were significantly increased in the ischemic microvasculature and this was accompanied by a worse neurobehavioral score (Table 1). In vitro, expression of MMP3, 2 and 9 proteins in cell lysate were not different between the groups. However, MMP3 and MMP9 enzyme activities were significantly increased in the diabetes condition while, there was no additional increase in enzyme activity with hypoxia. MMP2 activity was significantly increased with hypoxia in diabetes condition (Table 1). Conclusions: In female rats, diabetes and hypoxia increase MMP activity in the microvasculature and endothelial cells. These observations provide initial evidence to further explore the role of MMPs in the increased HT seen in females with diabetes.