Neuroprotective effects of 1-O-hexyl-2,3,5-trimethylhydroquinone on ischaemia/reperfusion-induced neuronal injury by activating the Nrf2/HO-1 pathway

Abstract

1-O-Hexyl-2,3,5-trimethylhydroquinone (HTHQ), a lipophilic phenolic agent, has an antioxidant activity and reactive oxygen species (ROS) scavenging property. However, the role of HTHQ on cerebral ischaemic/reperfusion (I/R) injury and the underlying mechanisms remain poorly understood. In the present study, we demonstrated that HTHQ treatment ameliorated cerebral I/R injury in vivo, as demonstrated by the decreased infarct volume ration, neurological deficits, oxidative stress and neuronal apoptosis. HTHQ treatment increased the levels of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream antioxidant protein, haeme oxygenase-1 (HO-1). In addition, HTHQ treatment decreases oxidative stress and neuronal apoptosis of PC12 cells following hypoxia and reperfusion (H/R) in vitro. Moreover, we provided evidence that PC12 cells were more vulnerable to H/R-induced oxidative stress after si-Nrf2 transfection, and the HTHQ-mediated protection was lost in PC12 cells transfected with siNrf2. In conclusion, these results suggested that HTHQ possesses neuroprotective effects against oxidative stress and apoptosis after cerebral I/R injury via activation of the Nrf2/HO-1 pathway.

Keywords: 1-O-Hexyl-2,3,5-trimethylhydroquinone; HO-1; Nrf2; PC12 cells; cerebral ischaemic/reperfusion.

FIGURE 1

HTHQ treatment ameliorates cerebral I/R injury in vivo. A, Representative brain sections of TTC staining. Pale areas represent infarcted brain. B, Quantification results of infarct volume ration (n = 8). C, Quantification results of neurological deficit scores (n = 8). D, Quantification results of brain water content (n = 8). **P < .01 vs the Sham group; ^# P < .05 and ^## P < .01 vs the MCAO group

FIGURE 2

HTHQ treatment attenuates oxidative stress following cerebral I/R. The levels of superoxide dismutase (SOD) (A), catalase (CAT) (B), malondialdehyde (MDA) (C) and glutathione (GSH) (D) in the brain of mice (n = 6). **P < .01 vs the Sham group; ^# P < .05 and ^## P < .01 vs the MCAO group

FIGURE 3

HTHQ treatment decreases neuronal apoptosis following cerebral I/R (A) Images and quantifications of apoptotic cell in the ischaemic cortex of mice (n = 5). (B) Western blots showing the level of Bax, Bcl‐2 and cleaved caspase‐3 in mice following cerebral I/R (n = 4). **P < .01 vs the Sham group; ^# P < .05 and ^## P < .01 vs the MCAO group

FIGURE 4

HTHQ against cerebral I/R injury via Nrf2 antioxidant pathway. A, Western blots showing the level of Nrf2 and HO‐1 in mice following cerebral I/R (n = 4). B, Western blots showing the level of nuclear Nrf2 and cytosolic Nrf2 in mice following cerebral I/R (n = 4). **P < .01 vs the MCAO group

FIGURE 5

HTHQ treatment attenuates oxidative stress in PC12 cells following H/R. The levels of superoxide dismutase (SOD) (A), catalase (CAT) (B), malondialdehyde (MDA) (C) and glutathione (GSH) (D) in PC12 cells (n = 6). **P < .01 vs the control group; ^# P < .05 and ^## P < .01 vs the H/R group

FIGURE 6

HTHQ treatment decreases apoptosis of PC12 cells following H/R. A, Flow cytometry analysis of cell apoptosis following H/R (n = 6). B, Images and quantifications of TUNEL‐positive cells following H/R (n = 5). **P < .01 vs the control group; ^## P < .01 vs the H/R group

FIGURE 7

The antioxidant activity of HTHQ involves the Nrf2/HO‐1 Pathway. The levels of superoxide dismutase (SOD) (A), catalase (CAT) (B), malondialdehyde (MDA) (C) and glutathione (GSH) (D) in PC12 cells after si‐Nrf2 transfection (n = 6). **P < .01 vs the control group; ^# P < .05 and ^## P < .01 vs the H/R group

FIGURE 8

The anti‐apoptosis ability of HTHQ involves the Nrf2/HO‐1 Pathway. A, Flow cytometry analysis of cell apoptosis after si‐Nrf2 transfection (n = 6). B, Images and quantifications of TUNEL‐positive cells after si‐Nrf2 transfection (n = 5). **P < .01 vs the control group; ^## P < .01 vs the H/R group