Total Flavones of <i>Rhododendron</i> Protect Against Ischemic Cerebral Injury by Regulating the Phosphorylation of the RhoA-ROCK<sub>2</sub> Pathway via Endothelial-Derived H<sub>2</sub>S
This study aims to investigate the mechanism by which the total flavones of <i>Rhododendron</i> (TFR) protect against cerebral ischemic injury through the endothelial-derived H<sub>2</sub>S-mediated regulation of RhoA phosphorylation at the Ser188 and Rho kinase 2 (ROCK<su...
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| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
| Published: |
MDPI AG
2025-07-01
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| Series: | Current Issues in Molecular Biology |
| Subjects: | |
| Online Access: | https://www.mdpi.com/1467-3045/47/7/513 |
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| Summary: | This study aims to investigate the mechanism by which the total flavones of <i>Rhododendron</i> (TFR) protect against cerebral ischemic injury through the endothelial-derived H<sub>2</sub>S-mediated regulation of RhoA phosphorylation at the Ser188 and Rho kinase 2 (ROCK<sub>2</sub>) phosphorylation at Thr436. For experimental design, mouse or rat cerebrovascular endothelial cells (ECs) were cultured with or without neurons and subjected to hypoxia/reoxygenation (H/R) injury. The vasodilation of the cerebral basilar artery was assessed. Cerebral ischemia/reperfusion (I/R) injury was induced in mice by bilateral carotid artery ligation, followed by Morris water maze and open field behavioral assessments. The protein levels of cystathionine-γ-lyase (CSE), 3-mercaptopyruvate sulfurtransferase (3-MST), RhoA, ROCK<sub>2</sub>, p-RhoA (RhoA phosphorylated at Ser188), and p-ROCK<sub>2</sub> (ROCK<sub>2</sub> phosphorylated at Thr436) were quantified. Additionally, the activities of RhoA and ROCK<sub>2</sub> were measured. Notably, TFR significantly inhibited H/R-induced H<sub>2</sub>S reduction and suppressed the increased expression and activity of RhoA and ROCK<sub>2</sub> in ECs, effects attenuated by CSE or 3-MST knockout. Moreover, TFR-mediated cerebrovascular dilation was reduced by RhoA or ROCK<sub>2</sub> inhibitors, while the protective effect of TFR against cerebral I/R injury in mice was markedly attenuated by the heterozygous knockout of ROCK<sub>2</sub>. In the ECs-co-cultured neurons, the inhibition of TFR on H/R-induced neuronal injury and decrease in H<sub>2</sub>S level in the co-culture was attenuated by the knockout of CSE or 3-MST in the ECs. TFR notably inhibited the H/R-induced upregulation of neuronal RhoA, ROCK<sub>2</sub>, and p-ROCK<sub>2</sub> protein levels, as well as the activities of RhoA and ROCK<sub>2</sub>, while reversing the decrease in p-RhoA. However, the knockout of CSE or 3-MST in the ECs significantly attenuated the inhibition of TFR on these increases. Furthermore, 3-MST knockout in ECs attenuated the TFR-mediated suppression of p-RhoA reduction. Additionally, CSE or 3-MST knockout in ECs exacerbated H/R-induced neuronal injury, reduced H<sub>2</sub>S level in the co-culture system, and increased RhoA activity and ROCK<sub>2</sub> expression in neurons. In summary, TFR protected against ischemic cerebral injury by endothelial-derived H<sub>2</sub>S promoting the phosphorylation of RhoA at Ser188 but inhibited the phosphorylation of ROCK<sub>2</sub> at Thr436 to inhibit the RhoA-ROCK<sub>2</sub> pathway in neurons. |
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| ISSN: | 1467-3037 1467-3045 |