Mechanism of Mitochondrial Dysfunction in Brain Vasculature during Ischemic Stroke: Role of Tetrahydrocurcumin

Abstract

Cerebral ischemic/reperfusion (I/R) injury is a major cause of stroke and is associated with the formation of leaky mitochondria leading to mitochondrial dysfunction. Tetrahydrocurcumin (THC), a major herbal antioxidant and anti‐inflammatory compound, has shown to protect the brain against injury. The objectives of this study are to identify the mechanism of mitochondrial dysfunction during I/R and the therapeutic potential of THC to mitigate mitophagy in an experimental stroke model. In our study, 8–10 weeks old male C57BL/6 wild‐type mice were subjected to middle cerebral artery occlusion (MCAO) for 40 min, followed by reperfusion for 72 hours. THC (25 mg/kg‐BW/day) was injected intraperitoneally (I.P.) once daily for 3 days after 40 minutes of ischemia. The experimental groups were: (i) sham (control), (ii) I/R (stroke) and (iii) I/R+THC. We found that infarct size, brain edema, and permeability were increased in ischemic mice as compared to sham‐operated control group; however THC treatment in I/R mice almost normalized these changes. Plasma levels of tHcy were significantly increased in the I/R mice compared to the sham group, but THC treatment restored the tHcy to control level in I/R+THC group. Western blot analysis also revealed that the protein levels of CBS, CSE, and MTHFR were significantly increased while the level of SAHH was significantly decreased in I/R mice compared to sham. THC treatment ameliorates these changes to control levels. The levels of mitochondrial p47phox, gp91phox (oxidants) were increased, and levels of Trx‐2, SOD‐2 (anti‐oxidants) were decreased, respectively, in the ischemic mitochondrial fraction. Treatment with THC mitigated these levels. Interestingly, matrix extracted from isolated mitochondria of the ischemic brain also showed an increase in MMP‐9 activity in gelatin‐gel zympography. We also observed an increase in TIMP‐1 and a decrease in TIMP‐2 protein expression in the ischemic brain as compared to sham. THC treatments reversed all these changes to normal levels. Further, we identified that the activation of MMP‐9 is associated with reduced expression of mitochondrial tight junction proteins and collagen IV/elastion degradation leading to mitophagy. To determine whether occludin and zona occluden‐1 (ZO1) are MMP target, we determined TJP protein expression by Western analysis. The results showed downregulation of occludin and ZO1 in the mitochondria from the ischemic brain, which was mitigated by THC treatment. Experimental data also revealed a robust degradation of elastin and accumulation of collagen IV in the ischemic brain as compared to sham. To further support the claim of mitophagy occurrence in ischemic brains, the levels of mitophagy markers (conversion of LC3‐I to LC3‐II in mitochondria) Mfn‐1(fusion protein) and DRP‐1 (fission protein) were measured by Western Blot. The LC3‐II was increased, whereas Mfn‐1 and DRP‐1 protein expression were decreased in ischemic mitochondria. THC treatment inhibited the conversion of LC3‐Ito LC‐3‐II and enhanced the expression of MFN‐1 and DRP‐1. In conclusion, this study suggests a potential therapeutic role of dietary THC to repair mitochondrial damage during ischemia stroke.Support or Funding InformationThis work is financially supported by National Institute of health grant AR‐067667 and HL‐107640‐NT are greatly acknowledgedThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.