Yun-Liang Xie1, Bo Zhang2, Ling Jing3. 1. a Medical Department , The Affiliated Hospital of Bei Hua University , Jilin , China. 2. b Health Care Department , The Affiliated Hospital of Bei Hua University , Jilin , China. 3. c College of Pharmacy , Jilin University , Changchun , China.
Abstract
OBJECTIVE: To explore the potential effect of miR-125b on p53-mediated regulation of Bax/Cytochrome C/Caspase-3 apoptotic signaling pathway in rats with cerebral ischemia-reperfusion (CIR) injury. METHODS: Sprague-Dawley (SD) rats were used to conduct CIR injury and injected with miR-125b mimic/inhibitor or p53 inhibitor (Pifithrin-α, PFT-α). Dual-luciferase reporter gene assay was used to analyze the targeting relationship between miR-125b and p53. Longa scoring and Triphenyl tetrazolinm chloride (TTC) staining were used to test the neurologic function and determine infarct size, respectively. Hematoxylin-eosin (HE) and Nissl's stainings were conducted to observe the morphology of cortical neurons. Neuronal nuclei (NeuN) expression was detected by immunohistochemical staining. QRT-PCR was performed to detect the expressions of miR-125b and p53. TUNEL staining and Western blotting was used to determine neuronal apoptosis and expressions of Bax/Cytochrome C/Caspase-3 signaling pathway-related proteins, respectively. RESULTS: Our results showed that miR-125b could directly target p53. As observed, overexpression of miR-125b could obviously reduce the neurological score, infarct size, and brain water content after CIR in rats, which also improved the morphology of cortical neurons, increased the number of neurons, reduced neuronal apoptosis, and inhibited the expressions of Bax/Cytochrome C/Caspase-3 pathway. Moreover,the similar results were observed in rats with CIR after injected with PFT-α. But no significant differences in each index were found in CIR group and CIR + anti-miR-125b + PFT-α group. CONCLUSION: MiR-125b exerts protective effects on CIR injury through inhibition of Bax/Cytochrome C/Caspase-3signaling pathway via targeting p53, which is likely to be a promising treatment for CIR. ABBREVIATIONS: 3'-UTR: 3-untranslated region; CIR: cerebral ischemia-reperfusion; CIS: cerebral ischemic stroke; PFT-α: Pifithrin-α; PVDF: polyvinylidene fluoride; SD: Sprague-Dawley; TBST: tris buffered saline with tween. TTC staining: Triphenyl tetrazolinm chloride staining; TUNEL: Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling.
OBJECTIVE: To explore the potential effect of miR-125b on p53-mediated regulation of Bax/Cytochrome C/Caspase-3 apoptotic signaling pathway in rats with cerebral ischemia-reperfusion (CIR) injury. METHODS: Sprague-Dawley (SD) rats were used to conduct CIR injury and injected with miR-125b mimic/inhibitor or p53 inhibitor (Pifithrin-α, PFT-α). Dual-luciferase reporter gene assay was used to analyze the targeting relationship between miR-125b and p53. Longa scoring and Triphenyl tetrazolinm chloride (TTC) staining were used to test the neurologic function and determine infarct size, respectively. Hematoxylin-eosin (HE) and Nissl's stainings were conducted to observe the morphology of cortical neurons. Neuronal nuclei (NeuN) expression was detected by immunohistochemical staining. QRT-PCR was performed to detect the expressions of miR-125b and p53. TUNEL staining and Western blotting was used to determine neuronal apoptosis and expressions of Bax/Cytochrome C/Caspase-3 signaling pathway-related proteins, respectively. RESULTS: Our results showed that miR-125b could directly target p53. As observed, overexpression of miR-125b could obviously reduce the neurological score, infarct size, and brain water content after CIR in rats, which also improved the morphology of cortical neurons, increased the number of neurons, reduced neuronal apoptosis, and inhibited the expressions of Bax/Cytochrome C/Caspase-3 pathway. Moreover,the similar results were observed in rats with CIR after injected with PFT-α. But no significant differences in each index were found in CIR group and CIR + anti-miR-125b + PFT-α group. CONCLUSION:MiR-125b exerts protective effects on CIR injury through inhibition of Bax/Cytochrome C/Caspase-3signaling pathway via targeting p53, which is likely to be a promising treatment for CIR. ABBREVIATIONS: 3'-UTR: 3-untranslated region; CIR: cerebral ischemia-reperfusion; CIS: cerebral ischemic stroke; PFT-α: Pifithrin-α; PVDF: polyvinylidene fluoride; SD: Sprague-Dawley; TBST: tris buffered saline with tween. TTC staining: Triphenyl tetrazolinm chloride staining; TUNEL: Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling.