Research Paper Volume 12, Issue 24 pp 26121—26139

Subanesthetic isoflurane abates ROS-activated MAPK/NF-κB signaling to repress ischemia-induced microglia inflammation and brain injury

Figure 3. Sub-anesthetic ISO post-conditioning abates OGD-induced COX2/PGE2 and iNOS/NO generation in microglial cells in co-cultures. Co-cultures with or without 3 h-OGD stimulation were exposed to RA with or without 0.7% ISO for 30 min. And co-cultures were continuously cultured under normal conditions for 6, 12, or 24 h after OGD treatment. Then, microglial cells were harvested to measure some indexes. (A) qPCR analysis of COX2 and iNOS mRNA levels at 6 h after OGD treatment. GAPDH was used as the endogenous control. (B) Western blot analysis of COX2 and iNOS protein levels at 12 h after OGD treatment. β-actin was used as the internal control. (C) RIA analysis of PGE2 levels at 24 h after OGD treatment. (D) The iNOS activity was assessed by monitoring the conversion of arginine to citrulline at 12 h after OGD treatment. (E) Colorimetric estimation of NO levels with Griess reagent at 24 h after OGD treatment. Representative data are from three independent experiments and expressed as mean ± SD. Statistical significance: *P < 0.05 vs. Ctrl groups; #P < 0.05 vs. OGD + RA group. Ctrl: control; ISO: isoflurane; OGD: oxygen and glucose deprivation; RA: room air.