Necrostatin-1 Suppresses Autophagy and Apoptosis in Mice Traumatic Brain Injury Model

Abstract

Traumatic brain injury (TBI) results in neuronal apoptosis, autophagic cell death and necroptosis. Necroptosis is a newly discovered caspases-independent programmed necrosis pathway which can be triggered by activation of death receptor. Previous works identified that necrostatin-1 (NEC-1), a specific necroptosis inhibitor, could reduce tissue damage and functional impairment through inhibiting of necroptosis process following TBI. However, the role of NEC-1 on apoptosis and autophagy after TBI is still not very clear. In this study, the amount of TBI-induced neural cell deaths were counted by PI labeling method as previously described. The expression of autophagic pathway associated proteins (Beclin-1, LC3-II, and P62) and apoptotic pathway associated proteins (Bcl-2 and caspase-3) were also respectively assessed by immunoblotting. The data showed that mice pretreated with NEC-1 reduced the amount of PI-positive cells from 12 to 48 h after TBI. Immunoblotting results showed that NEC-1 suppressed TBI-induced Beclin-1 and LC3-II activation which maintained p62 at high level. NEC-1 pretreatment also reversed TBI-induced Bcl-2 expression and caspase-3 activation, as well as the ratio of Beclin-1/Bcl-2. Both 3-MA and NEC-1 suppressed TBI-induced caspase-3 activation and LC3-II formation, Z-VAD only inhibited caspase-3 activation but increased LC3-II expression at 24 h post-TBI. All these results revealed that multiple cell death pathways participated in the development of TBI, and NEC-1 inhibited apoptosis and autophagy simultaneously. These coactions may further explain how can NEC-1 reduce TBI-induced tissue damage and functional deficits and reflect the interrelationship among necrosis, apoptosis and autophagy.

Impaired autophagy flux is associated with neuronal cell death after traumatic brain injury

“…21,23,[26][27][28] Our data indicate a transient block of autophagic clearance at the early time points after TBI, which is than relieved later after injury. Therefore, the contradictory observations of both beneficial and detrimental effects of autophagy after TBI described by previous studies may reflect this time-dependent alteration in autophagic flux in the injured brain.…”

“…Moreover, the function of autophagy following TBI is controversial, with both beneficial and detrimental roles suggested. [25][26][27][28] Here we examined levels of autophagy and autophagic flux following TBI induced by controlled cortical impact in wild-type and transgenic GFP-Lc3 autophagy reporter mice. Our data demonstrate that LC3 and autophagosomes accumulate in ipsilateral cortex and hippocampus within hours after injury, and remain elevated for at least 1 wk.…”

Molecular and Cellular Mechanisms Responsible for Cellular Stress and Low-grade Inflammation Induced by a Super-low Dose of Endotoxin

“…Unlike programmed apoptotic cell death, which resolves inflammation, necroptosis is associated with non-resolving inflammation and persistent activation of stress kinases such as JNK (41,42). As a consequence, cell necroptosis is associated with a multitude of inflammatory complications such as atherosclerosis, reduced wound repair, inflammatory bowel diseases, and ischemic injury (13,(42)(43)(44)(45)(46). Our finding that a super-low dose of endotoxin elicits cell necroptosis potentially explains the detrimental effect of superlow-grade endotoxemia in humans.…”

Traumatic Brain Injury-Induced Neuronal Apoptosis is Reduced Through Modulation of PI3K and Autophagy Pathways in Mouse by FTY720

“…Inhibition of autophagy can protect injured brain from undergoing apoptosis . However, in other cases, autophagy also leads to brain injury by promoting apoptosis (Lin et al 2014;Wang et al 2012). The discrepancies may be due to the complex and diverse interplays between autophagy and apoptosis, and further studies are needed to clarify their relationship.…”