Background/Aims: Ischemic stroke is still one of the leading debilitating diseases with high morbidity and mortality. NADPH oxidase (NOX)-derived reactive oxygen species (ROS) play an important role in cerebral ischemia/reperfusion (I/R) injury. However, the mechanism underlying the regulation of ROS generation is still not fully elucidated. This study aims to explore the role of transforming growth beta (TGF-β) signals in ROS generation. Methods: Sprague–Dawley rats were subjected to I/R injury, and PC-12 cells were challenged by hypoxia/reoxygenation (H/R) and/or treated with activin receptor-like kinase (ALK5) inhibitor Sb505124 or siRNA against ALK5. Brain damage was evaluated using neurological scoring, triphenyl tetrazolium chloride staining, hematoxylin and eosin staining, infarct volume measurement, TUNEL staining, and caspase-3 activity measurement. Expression of TGF-β and oxidative stress-related genes was analyzed by real-time polymerase chain reaction and Western blot; NOX activity and ROS level were measured using spectrophotometry and fluorescence microscopy, respectively. Results: I/R contributed to severe brain damage (impaired neurological function, brain infarction, tissue edema, apoptosis), TGF-β signaling activation (upregulation of ALK5, phosphorylation of SMAD2/3) and oxidative stress (upregulation of NOX2/4, rapid release of ROS [oxidative burst]). However, Sb505124 significantly reversed these alterations and protected rats against I/R injury. As in the animal results, H/R also contributed to TGF-β signaling activation and oxidative stress. Likewise, the inhibition of ALK5 or ALK5 knockdown significantly reversed these alterations in PC-12 cells. Other than ALK5 knockdown, ALK5 inhibition had no effect on the expression of ALK5 in PC-12 cells. Conclusions: Our studies demonstrated that TGF-β signaling activation is involved in the regulation of NOX2/NOX4 expression and exacerbates cerebral I/R injury.

Recent studies have uncovered that accumulation of glutamate after ischaemic stroke is closely associated with the down-regulation of glutamate transporter-1 (GLT-1) expression, suggesting that GLT-1 expression critically controls glutamate accumulation and the abnormal glutamate transport-elicited neuronal cell excitotoxicity in patients with ischaemic stroke. However, it remains unknown how GLT-1 expression is regulated under ischaemic stroke conditions. In the present study, we screened the expression of nine brain-specific or brain-enriched miRNAs in a focal cerebral ischaemia/reperfusion (I/R) injury rat model, which showed glutamate accumulation and down-regulated GLT-1 expression as expected, and revealed that the miR-107 level was elevated in both brain tissue and plasma in the model. Next, we examined the functional relationship of miR-107 with GLT-1 expression in a nerve cell hypoxia/reoxygenation (H/R) injury model. H/R treatment increased apoptosis of the nerve cells concomitant with glutamate accumulation, miR-107 elevation and suppressed GLT-1 expression, mimicking our in vivo findings in the cerebral I/R injury rat model in vitro. Co-treating the cells with an miR-107 inhibitor blocked all of the effects, demonstrating that miR-107 functions to inhibit GLT-1 expression and elevate glutamate accumulation. To extend these animal and cell-based studies to clinical patients, we measured the plasma levels of miR-107 and glutamate, and observed that both miR-107 and glutamate were elevated in patients with ischaemic stroke. On the basis of these observations, we conclude that elevated miR-107 expression after ischaemic stroke accounts, at least partially, for glutamate accumulation through suppression of GLT-1 expression. Our findings also highlight that the plasma level of miR-107 may serve as a novel biomarker for monitoring excitotoxicity in patients with ischaemic stroke.

The involvement of Nrf2 in the protective effects of (-)-Epigallocatechin-3-gallate (EGCG) on NaAsO 2 -induced hepatotoxicity The involvement of Nrf2 in the protective effects of (-)-Epigallocatechin-3-gallate (EGCG) on NaAsO2-induced hepatotoxicity

Recent studies show that resveratrol exerts beneficial effects on prevention of pulmonary hypertension. This study is performed to explore the effects of trimethoxystilbene, a novel resveratrol analog, on rat pulmonary vascular remodeling and right ventricular hypertrophy in hypoxia-induced pulmonary arterial hypertension (PAH) and the underlying mechanisms. Sprague–Dawley rats were placed in a chamber and exposed to 10% O₂ continuously for 4 weeks to induce PAH. The effects of trimethoxystilbene (5 or 10 mg/kg per day, intragastric [i.g.]) and resveratrol (as a positive control, 25 mg/kg per day, i.g.) on hypoxia-induced PAH vascular remodeling and right ventricle hypertrophy were evaluated. At the end of experiments, the index for pulmonary vascular remodeling and right ventricle hypertrophy, inflammatory cell infiltration in lung tissue, the plasma levels and lung tissue contents of hydrogen peroxide (H₂O₂), the mRNA and protein levels for NADPH oxidases (NOX2, NOX4) and vascular peroxidase 1 (VPO1) in pulmonary artery or right ventricle were measured. The results showed that trimethoxystilbene treatment significantly attenuated hypoxia-induced pulmonary vascular remodeling (such as decrease in the ratio of wall thickness to vessel external diameter) and right ventricle hypertrophy (such as decrease in the ratio of right ventricle weight to the length of the tibia), accompanied by downregulation of NOX2, NOX4, and VPO1 expression in pulmonary artery or right ventricle, decrease in H₂O₂ production and inflammatory cell infiltration in lung tissue. Trimethoxystilbene is able to prevent pulmonary vascular remodeling and right ventricle hypertrophy in hypoxia-induced rat model of PAH, which is related to inhibition of the NOX/VPO1 pathway-mediated oxidative stress and the inflammatory reaction.

Ischemic stroke is a devastating central nervous disease associated with oxidative stress and NOX2 is the main source of ROS responsible for brain tissue. miRNAs are a class of negative regulator of genes in mammals and involves the pathogenesis of ischemic stroke. This study aims to observe the role of target miRNA(miR-652) of NOX2 in ischemic stroke. A rat cerebral ischemia/reperfusion (CI/R) injury model and an SH-SY5Y cell hypoxia/reoxygenation(H/R) model were used to simulate ischemic stroke, and corresponding gene expression, biochemical indicators and pathophysiological indicators were measured to observe the role of miR-652. NOX2 significantly increased in brain tissues subjected to I/R or in SH-SY5Y cells subjected to H/R, while the expression level of miR-652(potential target of NOX2) significantly decreased in both brain tissues and plasma. Overexpression of miR-652 significantly suppressed NOX2 expression and ROS generation in H/R treated SH-SY5Y cells and reduced the relative luciferase activity of cells transfected with plasmid NOX2-WT (reporter gene plasmid). MiR-652 agomir significantly decreased the expression of NOX2 and ROS generation in brain tissues of CIR rats, as well as tissue injury. These data indicated that miR-652 protected rats from cerebral ischemia reperfusion injury by directly targeting NOX2, is a novel target for ischemic stroke therapy.