MiR-7-1 potentiated estrogen receptor agonists for functional neuroprotection in VSC4.1 motoneurons
详细信息 查看全文
- 作者：M. Chakrabarti ; N.L. Banik ; S.K. Ray
- 关键词：CaMKII尾 ; Ca2+/calmodulin-dependent protein kinase II beta ; CI ; calcium ionophore ; COX-4 ; cytochrome c oxidase subunit IV ; CP伪1C ; L-type Ca2+ channel protein alpha 1C ; CREB ; cAMP response element binding protein ; DMEM ; Dulbecco&rsquo ; s Modified Eagle Medium ; DMSO ; dimethyl sulfoxide ; ECL ; enhanced chemiluminescence ; ER伪 ; estrogen receptor alpha ; ER尾 ; estrogen receptor beta ; EST ; estrogen ; FBS ; fetal bovine serum ; FITC ; fluorescein isothiocyanate ; GAPDH ; glyceraldehyde-3-phosphate dehydrogenas
- 刊名：Neuroscience
- 出版年：3 January, 2014
- 年：2014
- 卷：256
- 期：Complete
- 页码：322-333
- 全文大小：2321 K
文摘
Protection of motoneurons is an important goal in the treatment of spinal cord injury (SCI). We tested whether neuroprotective microRNAs (miRs) like miR-206, miR-17, miR-21, miR-7-1, and miR-106a could enhance efficacy of estrogen receptor (ER) agonists such as 1,3,5-tris (4-hydroxyphenyl)-4-propyl-1H-pyrazole (PPT, ER伪 agonist), Way200070 (WAY, ER尾 agonist), and estrogen (EST, ER伪 and ER尾 agonist) in preventing apoptosis in the calcium ionophore (CI)-insulted ventral spinal cord 4.1 (VSC4.1) motoneurons. We determined that 200 nM CI induced 70% cell death. Treatment with 50 nM PPT, 100 nM WAY, and 150 nM EST induced overexpression of ER伪, ER尾, and both receptors, respectively, at mRNA and protein levels. Treatment with ER agonists significantly upregulated miR-206, miR-17, and miR-7-1 in the CI-insulted VSC4.1 motoneurons. Transfection with miR-206, miR-17, or miR-7-1 mimic potentiated WAY or EST to inhibit apoptosis in the CI-insulted VSC4.1 motoneurons. Overexpression of miR-7-1 maximally increased efficacy of WAY and EST for down regulation of pro-apoptotic Bax and upregulation of anti-apoptotic Bcl-2. A search using microRNA database (miRDB) indicated that miR-7-1 could inhibit the expression of L-type Ca2+ channel protein alpha 1C (CP伪1C). miR-7-1 overexpression and WAY or EST treatment down regulated CP伪1C but upregulated p-Akt to trigger cell survival signaling. The same therapeutic strategy increased expression of the Ca2+/calmodulin-dependent protein kinase II beta (CaMKII尾) and the phosphorylated cAMP response element binding protein (p-CREB) so as to promote Bcl-2 transcription. Whole cell membrane potential and mitochondrial membrane potential studies indicated that miR-7-1 highly potentiated EST to preserve functionality in the CI-insulted VSC4.1 motoneurons. In conclusion, our data indicated that miR-7-1 most significantly potentiated efficacy of EST for functional neuroprotection and this therapeutic strategy could be used in the future to attenuate apoptosis of motoneurons in SCI.