Xinwang Lv1, Jian Liang2, Zhipu Wang3. 1. Department of Orthopaedics, Caoxian People's Hospital, Heze, China. 2. Department of Orthopaedics, Affiliated Hospital of Heze Medical College, Heze, China. 3. Department of Orthopaedics, Caoxian Panshi Hospital, Heze, China - xingll2007@163.com.
Abstract
BACKGROUND: To explore the effect of micro ribonucleic acid (miR)-21-5p on spinal cord injury (SCI) in rats and its mechanism of action. METHODS: The rat model of SCI was established, and the key miRNAs were screened using the microarray assay and miRNA-mRNA interaction network. After intrathecal injection of agomir-21 and antagomir-21, the effect of miR-21 expression on motor function recovery of rats was evaluated using the Basso-Beattie-Bresnahan (BBB) score. The expression level of miR-21 in spinal cord tissues was determined via quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and the effect of miR-21 expression on apoptosis in spinal cord tissues was determined via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay and Western blotting. Moreover, the effects of agomir-21 and antagomir-21 on SCI-induced expressions of inflammatory factors interleukin-8 (IL-8), IL-1β, IL-6 and tumor necrosis factor-α (TNF-α) in spinal cord tissues were detected through qRT-PCR. Finally, Western blotting was performed to detect the effects of agomir-21 and antagomir-21 on the phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway and its downstream molecules in each group. RESULTS: The screening results of the microarray assay revealed that the mRNA and miRNA expression profiles in spinal cord tissues had significant differences in model group from those in sham group. The BBB score was significantly higher in agomir-21 group than that in model group. Compared with that in model group, the apoptosis of spinal cord tissues was obviously weakened in agomir-21 group, while it was obviously enhanced in antagomir-21 group. Agomir-21 group had evidently lower Bax/Bcl-2, and Caspase-3 and Caspase-9 protein expressions, while antagomir-21 group had evidently higher Bax/Bcl-2 and Caspase-3 protein expression than model group. Besides, the expressions of inflammatory factors IL-8, IL-1β, IL-6 and TNF-α were remarkably lower in agomir-21 group than those in model group, while they were remarkably higher in antagomir-21 group than those in model group. Finally, it was found that the protein expressions of phosphorylated PI3K (p-PI3K)/PI3K and p-AKT/AKT rose markedly, while the protein expressions of phosphatase and tensin homolog deleted on chromosome ten (PTEN) and endothelial nitric oxide synthase (eNOS) declined markedly in agomir-21 group compared with those in model group. However, the opposite results were observed in antagomir-21 group compared with those in model group. CONCLUSIONS: MiR-21-5p may reduce the apoptosis and inflammation in spinal cord tissues of rats through the PI3K/AKT pathway.
BACKGROUND: To explore the effect of micro ribonucleic acid (miR)-21-5p on spinal cord injury (SCI) in rats and its mechanism of action. METHODS: The rat model of SCI was established, and the key miRNAs were screened using the microarray assay and miRNA-mRNA interaction network. After intrathecal injection of agomir-21 and antagomir-21, the effect of miR-21 expression on motor function recovery of rats was evaluated using the Basso-Beattie-Bresnahan (BBB) score. The expression level of miR-21 in spinal cord tissues was determined via quantitative reverse transcription-polymerase chain reaction (qRT-PCR), and the effect of miR-21 expression on apoptosis in spinal cord tissues was determined via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay and Western blotting. Moreover, the effects of agomir-21 and antagomir-21 on SCI-induced expressions of inflammatory factors interleukin-8 (IL-8), IL-1β, IL-6 and tumor necrosis factor-α (TNF-α) in spinal cord tissues were detected through qRT-PCR. Finally, Western blotting was performed to detect the effects of agomir-21 and antagomir-21 on the phosphatidylinositol 3-hydroxy kinase (PI3K)/protein kinase B (AKT) signaling pathway and its downstream molecules in each group. RESULTS: The screening results of the microarray assay revealed that the mRNA and miRNA expression profiles in spinal cord tissues had significant differences in model group from those in sham group. The BBB score was significantly higher in agomir-21 group than that in model group. Compared with that in model group, the apoptosis of spinal cord tissues was obviously weakened in agomir-21 group, while it was obviously enhanced in antagomir-21 group. Agomir-21 group had evidently lower Bax/Bcl-2, and Caspase-3 and Caspase-9 protein expressions, while antagomir-21 group had evidently higher Bax/Bcl-2 and Caspase-3 protein expression than model group. Besides, the expressions of inflammatory factors IL-8, IL-1β, IL-6 and TNF-α were remarkably lower in agomir-21 group than those in model group, while they were remarkably higher in antagomir-21 group than those in model group. Finally, it was found that the protein expressions of phosphorylated PI3K (p-PI3K)/PI3K and p-AKT/AKT rose markedly, while the protein expressions of phosphatase and tensin homolog deleted on chromosome ten (PTEN) and endothelial nitric oxide synthase (eNOS) declined markedly in agomir-21 group compared with those in model group. However, the opposite results were observed in antagomir-21 group compared with those in model group. CONCLUSIONS:MiR-21-5p may reduce the apoptosis and inflammation in spinal cord tissues of rats through the PI3K/AKT pathway.
Authors: Amir Mohammad Malvandi; Seyed Hamidreza Rastegar-Moghaddam; Saeede Ebrahimzadeh-Bideskan; Giovanni Lombardi; Alireza Ebrahimzadeh-Bideskan; Abbas Mohammadipour Journal: Mol Med Date: 2022-09-23 Impact factor: 6.376