LncRNA MIAT Promotes Spinal Cord Injury Recovery in Rats by Regulating RBFOX2-Mediated Alternative Splicing of MCL-1

Abstract Background: LncRNA myocardial infarction associated transcript (MIAT) alleviates acute spinal cord injury (ASCI)-induced neuronal cell apoptosis, but the specific mechanism of it involved in regulating SCI progression needs further exploration. Methods: A SCI rat model was established, followed by administration with Adenovirus-mediated MIAT overexpression vector (Ad-MIAT) alone or together with Ad-RBFOX2 (RNA binding fox-1 homolog 2), and rat locomotor capacity was evaluated. Hematoxylin-eosin (HE), Nissl and TUNEL straining were used to analyze spinal cord tissue morphology, neuron loss and apoptosis. Then, PC-12 cells were treated with H2O2 to induce cell injury, and cell viability and apoptosis were determined. Gain- and loss-of-function experiments were carried out to explore the effect of MIAT and RBFOX2 on cell functions. The binding between MIAT and RBFOX2 was confirmed using RNA immunoprecipitation and RNA pull down assays. MCL-1(myeloid cell leukemia sequence 1)-specific steric-blocking oligonucleotides (SBOs) were used to transfer the MCL-1 pre-mRNA splicing pattern from MCL-1L to MCL-1S. Confocal laser microscopy and transmission electron microscopy were used to analyze autophagy flux and lysosome formation.Results: MIAT overexpression promoted motor function recovery, improved morphology of injured tissues, and restrained neuron loss and cell apoptosis in SCI rats, as well as hindered H2O2-caused cell apoptosis in vitro. MIAT stabilized RBFOX2 protein expression by binding to RBFOX2, thereby promoting RBFOX2-induced increase of anti-apoptotic MCL-1L and decrease of pro-apoptotic MCL-1S. Silencing RBFOX2 blocked the inhibitory effect of MIAT on cell apoptosis. And RBFOX2 protected cells from H2O2-caused damage, which was reversed by SBOs. Moreover, overexpression of MCL-1L instead of MCL-1S promoted autophagy activation in cells. Interestingly, co-overexpression of MIAT and RBFOX2 had a better promoting effect on SCI recovery.Conclusion: MIAT mitigated SCI by promoting RBFOX2-mediated alternative splicing of MCL-1. Our findings might provide a promising therapeutic target for SCI.