Fasl Expression On Human Nucleus Pulposus Cells Contributes To The Immune Privilege Of Intervertebral Disc By Interacting With Immunocytes

BackgroundIntervertebral disc degeneration is a major cause of many spinal disorders. Overthe past decade, the underlying mechanisms of IDD have been well documented interms of biomechanics, cell apoptosis, deregulated genes and microRNAs (miRNAs).Moreover, studies with human disc cells, especially focusing on nucleus pulposus (NP)cells, are seldom found in the literature. Despite intensive basic and clinical pieces ofstudies aiming for addressing the issues, the physiopathological mechanisms of IDDremain yet speculative with particular reference to the maintenance of immune privilege. The immune privilege of tissues is ascribed not only to the physiologicalblood-tissue barrier, but the local expression of Fas ligand (FasL).The purpose of this study was to address the role of FasL expression in humanintervertebral disc degeneration (IDD) and immune privilege in terms of the interactionbetween NP cells and immunocytes via the FasL-Fas machinery.1. FasL expresssion in human degenerative and normal NP cellsObjective: To address FasL expression in human degenerative and normal NP cells.Methods: We collected NP specimens from20patients with IDD as degenerativegroup and8normal cadaveric donors as control. FasL expression was detected byqRT-PCR, western blotting and flow cytometry (FCM). Results: We found that FasLexpression in mRNA, protein and cellular resolutions demonstrated a significantdecrease in degenerative group compared with normal control (p<0.05). Conclusions:FasL expression in normal control is significant higher than in degenerative group.2. Up-regulated FasL of NP cells in co-culture systemObjective: We aimed for addressing the role of FasL in immune privilege in terms ofthe interaction between NP cells and immunocytes via the FasL-Fas machinery.Methods: We collected macrophages and CD8+T cells from the peripheral blood ofpatients with IDD for co-cultures with NP cells. And macrophages and CD8+T cellswere harvested for apoptosis analysis by FCM after2days of co-cultures. Results: Asfor macrophages without co-cultures, the apoptosis rate was4.8%±0.007.Approximately6.5%±0.0075macrophages were apoptotic when cultured with IDD NPcells. It was noteworthy that NP cells with up-regulated FasL resulted in about17.5%±0.012apoptosis rate of macrophages (p<0.05). In the lentiviral-control groupwith scrambled sequence, the apoptosis rate of macrophages was6%±0.008. Similarfindings were observed in CD8+T cells co-cultured with NP cells. Conclusions: FCManalysis found that human NP cells with increased FasL expression resulted insignificantly increased apoptosis ratio of macrophages and CD8+T cells.