title: Deregulated miR-155 promotes Fas-mediated apoptosis in human intervertebral disc degeneration by targeting FADD and caspase-3.
abstract: The role of apoptosis in the pathogenesis of intervertebral disc degeneration (IDD) remains enigmatic. Accumulating evidence has shown that the apoptotic machinery is regulated by miRNAs. We hypothesized that miRNAs might contribute to apoptosis in IDD. We have found that 29 miRNAs were differentially expressed and miR-155 was down-regulated in degenerative nucleus pulposus (NP). The deregulation of miR-155 was further verified using real-time PCR (0.56 fold, p < 0.05). Bioinformatics target prediction identified FADD and caspase-3 as putative targets of miR-155. Furthermore, miR-155 inhibited FADD and caspase-3 expression by directly targeting their 3'-UTRs, which was abolished by mutation of the miR-155 binding sites. In vitro up-regulation of miR-155 in human NP cells by transfection with lentiviral pre-miR-155 resulted in repression of FADD and caspase-3; whereas knockdown of miR-155 with lentiviral antigomiR-155 led to over-expression of FADD and caspase-3. Also, Fas-mediated apoptosis was increased when antagonizing miR-155 and decreased when using pre-miR-155 in human NP cells. In addition, we presented direct evidence of NP cells undergoing apoptosis in IDD tissues using transmission electron microscopy analysis. Moreover, a combination of in situ hybridization (ISH) and immunohistochemistry (IHC) revealed that miR-155 expressed in the cytoplasm of human NP cells with reverse correlation with FADD and caspase-3. In summary, this is the first study addressing the underlying mechanisms of IDD in terms of apoptosis and miRNAs. Furthermore, caspase-3 is identified as a novel target of miR-155. Our results suggest that deregulated miR-155 promotes Fas-mediated apoptosis in human IDD by targeting FADD and caspase-3, implicating an aetiological and therapeutic role of miR-155 in IDD.
authors.affiliation: Institute of Orthopaedics, Xijing Hospital, Fourth Military Medical University, 15 Changle Western Road, Xi'an, People's Republic of China.
authors.forename: Hai-Qiang
authors.initials: HQ
authors.lastname: Wang
authors.forename: Xiao-Dong
authors.initials: XD
authors.lastname: Yu
authors.forename: Zhi-Heng
authors.initials: ZH
authors.lastname: Liu
authors.forename: Xin
authors.initials: X
authors.lastname: Cheng
authors.forename: Dino
authors.initials: D
authors.lastname: Samartzis
authors.forename: Lin-Tao
authors.initials: LT
authors.lastname: Jia
authors.forename: Sheng-Xi
authors.initials: SX
authors.lastname: Wu
authors.forename: Jing
authors.initials: J
authors.lastname: Huang
authors.forename: Jing
authors.initials: J
authors.lastname: Chen
authors.forename: Zhuo-Jing
authors.initials: ZJ
authors.lastname: Luo
mesh_terms: D000328:Adult; D017209:Apoptosis / Q000235:genetics*; D015153:Blotting, Western; D053148:Caspase 3 / Q000235:genetics* / Q000378:metabolism; D002469:Cell Separation; D053200:Fas-Associated Death Domain Protein / Q000235:genetics* / Q000378:metabolism; D005434:Flow Cytometry; D015870:Gene Expression; D005786:Gene Expression Regulation / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry; D055959:Intervertebral Disc Degeneration / Q000235:genetics* / Q000378:metabolism; D035683:MicroRNAs / Q000235:genetics*; D046529:Microscopy, Electron, Transmission; D020411:Oligonucleotide Array Sequence Analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019014:fas Receptor / Q000235:genetics* / Q000378:metabolism
chemical_list: C505533:FADD protein, human; D053200:Fas-Associated Death Domain Protein; C524942:MIRN155 microRNA, human; D035683:MicroRNAs; D019014:fas Receptor; C505407:CASP3 protein, human; D053148:Caspase 3