Haiping Zhao1,2, Guangwen Li1,2, Rongliang Wang1,2, Zhen Tao1,2, Qingfeng Ma1, Sijia Zhang1, Ziping Han1, Feng Yan1,2, Fangfang Li1, Ping Liu1, Shubei Ma1, Xunming Ji1, Yumin Luo1,2,3. 1. Institute of Cerebrovascular Disease Research and Department of Neurology, Xuanwu Hospital of Capital Medical University, Beijing, China. 2. Beijing Geriatric Medical Research Center and Beijing Key Laboratory of Translational Medicine for Cerebrovascular Diseases, Beijing, China. 3. Beijing Institute for Brain Disorders, Capital Medical University, Beijing, China.
Abstract
BACKGROUND AND PURPOSE: T helper cell 1 (Th1)-skewed neurotoxicity contributes to the poor outcome of stroke in rodents. Here, we have elucidated the mechanism of the Th1/Th2 shift in acute ischaemic stroke (AIS) patients at hyperacute phase and have looked for a miRNA-based therapeutic target. EXPERIMENTAL APPROACH: MiR-494 levels in blood from AIS patients and controls were measured by real-time PCR. C57BL/6J mice were subjected to transient middle cerebral artery occlusion, and cortical neurons were subjected to oxygen-glucose deprivation. Luciferase reporter system, chromatin immunoprecipitation sequencing (ChIP-Seq), and ChIP-PCR were used to uncover possible mechanisms. KEY RESULTS: In lymphocytes from AIS patients, there was a Th1/Th2 shift and histone deacetylase 2 (HDAC2) was markedly down-regulated. ChIP-seq showed that HDAC2 binding sites were enriched in regulation of Th1 cytokine production, and ChIP-PCR confirmed that HDAC2 binding was changed at the intron of STAT4 and the promoter of T-box transcription factor 21 (T-bet) in lymphocytes from AIS patients. MiR-494 was the most significantly increased miRNA in lymphocytes from AIS patients, and miR-494-3p directly targeted HDAC2. A strong association existed between miR-494 and Th1 cytokines, and neurological deficit as measured by the National Institute of Health Stroke Scale (NIHSS) in AIS patients. In vitro and in vivo experiments showed that antagomir-494 reduced Th1 shift-mediated neuronal and sensorimotor functional damage in the mouse model of ischaemic stroke, via the HDAC2-STAT4 pathway. CONCLUSION AND IMPLICATIONS: We demonstrated that miR-494 inhibition prevented Th1-skewed neurotoxicity through regulation of the HDAC2-STAT4 cascade.
BACKGROUND AND PURPOSE: T helper cell 1 (Th1)-skewed neurotoxicity contributes to the poor outcome of stroke in rodents. Here, we have elucidated the mechanism of the Th1/Th2 shift in acute ischaemic stroke (AIS) patients at hyperacute phase and have looked for a miRNA-based therapeutic target. EXPERIMENTAL APPROACH: MiR-494 levels in blood from AISpatients and controls were measured by real-time PCR. C57BL/6J mice were subjected to transient middle cerebral artery occlusion, and cortical neurons were subjected to oxygen-glucose deprivation. Luciferase reporter system, chromatin immunoprecipitation sequencing (ChIP-Seq), and ChIP-PCR were used to uncover possible mechanisms. KEY RESULTS: In lymphocytes from AISpatients, there was a Th1/Th2 shift and histone deacetylase 2 (HDAC2) was markedly down-regulated. ChIP-seq showed that HDAC2 binding sites were enriched in regulation of Th1 cytokine production, and ChIP-PCR confirmed that HDAC2 binding was changed at the intron of STAT4 and the promoter of T-box transcription factor 21 (T-bet) in lymphocytes from AISpatients. MiR-494 was the most significantly increased miRNA in lymphocytes from AISpatients, and miR-494-3p directly targeted HDAC2. A strong association existed between miR-494 and Th1 cytokines, and neurological deficit as measured by the National Institute of Health Stroke Scale (NIHSS) in AISpatients. In vitro and in vivo experiments showed that antagomir-494 reduced Th1 shift-mediated neuronal and sensorimotor functional damage in the mouse model of ischaemic stroke, via the HDAC2-STAT4 pathway. CONCLUSION AND IMPLICATIONS: We demonstrated that miR-494 inhibition prevented Th1-skewed neurotoxicity through regulation of the HDAC2-STAT4 cascade.
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