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Ablation of RIP3 protects from dopaminergic neurodegeneration in experimental Parkinson's disease.

ABSTRACT: Parkinson's disease (PD) is driven by dopaminergic neurodegeneration in the substantia nigra pars compacta (SN) and striatum. Although apoptosis is considered the main neurodegenerative mechanism, other cell death pathways may be involved. In this regard, necroptosis is a regulated form of cell death dependent on receptor interacting protein 3 (RIP3), a protein also implicated in apoptosis and inflammation independently of its pro-necroptotic activity. Here, we explored the role of RIP3 genetic deletion in in vivo and in vitro PD models. Firstly, wild-type (Wt) and RIP3 knockout (RIP3ko) mice were injected intraperitoneally with MPTP (40?mg/kg, i.p.), and sacrificed after either 6 or 30 days. RIP3ko protected from dopaminergic neurodegeneration in the SN of MPTP-injected mice, but this effect was independent of necroptosis. In keeping with this, necrostatin-1s (10?mg/kg/day, i.p.) did not afford full neuroprotection. Moreover, MPTP led to DNA fragmentation, caspase-3 activation, lipid peroxidation and BAX expression in Wt mice, in the absence of caspase-8 cleavage, suggesting intrinsic apoptosis. This was mimicked in primary cortical neuronal cultures exposed to the active MPTP metabolite. RIP3 deficiency in cultured cells and in mouse brain abrogated all phenotypes. Curiously, astrogliosis was increased in the striatum of MPTP-injected Wt mice and further exacerbated in RIP3ko mice. This was accompanied by absence of microgliosis and reposition of glial cell line-derived neurotrophic factor (GDNF) levels in the striata of MPTP-injected RIP3ko mice when compared to MPTP-injected Wt mice, which in turn showed a massive GDNF decrease. RIP3ko primary mixed glial cultures also presented decreased expression of inflammation-related genes upon inflammatory stimulation. These findings hint at possible undescribed non-necroptotic roles for RIP3 in inflammation and MPTP-driven cell death, which can contribute to PD progression.

SUBMITTER: Dionisio PA

PROVIDER: S-EPMC6831575 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Ablation of RIP3 protects from dopaminergic neurodegeneration in experimental Parkinson's disease.

Dionísio Pedro A PA Oliveira Sara R SR Gaspar Maria M MM Gama Maria J MJ Castro-Caldas Margarida M Amaral Joana D JD Rodrigues Cecilia M P CMP

Cell death & disease 20191105 11

Parkinson's disease (PD) is driven by dopaminergic neurodegeneration in the substantia nigra pars compacta (SN) and striatum. Although apoptosis is considered the main neurodegenerative mechanism, other cell death pathways may be involved. In this regard, necroptosis is a regulated form of cell death dependent on receptor interacting protein 3 (RIP3), a protein also implicated in apoptosis and inflammation independently of its pro-necroptotic activity. Here, we explored the role of RIP3 genetic ...[more]

PMID: 31690718

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Project description:Chronic microglial activation has been linked to the progressive degeneration of the nigrostriatal dopaminergic neurons evidenced in Parkinson's disease (PD) pathogenesis. The exact etiology of PD remains poorly understood. Although both oxidative stress and neuroinflammation are identified as co-contributors in PD pathogenesis, signaling mechanisms underlying neurodegenerative processes have yet to be defined. Indeed, we recently identified that protein kinase C delta (PKC?) activation is critical for induction of dopaminergic neuronal loss in response to neurotoxic stressors. However, it remains to be defined whether PKC? activation contributes to immune signaling events driving microglial neurotoxicity. In the present study, we systematically investigated whether PKC? contributes to the heightened microglial activation response following exposure to major proinflammatory stressors, including ?-synuclein, tumor necrosis factor ? (TNF?), and lipopolysaccharide (LPS). We report that exposure to the aforementioned inflammatory stressors dramatically upregulated PKC? with a concomitant increase in its kinase activity and nuclear translocation in both BV-2 microglial cells and primary microglia. Importantly, we also observed a marked upregulation of PKC? in the microglia of the ventral midbrain region of PD patients when compared to age-matched controls, suggesting a role for microglial PKC? in neurodegenerative processes. Further, shRNA-mediated knockdown and genetic ablation of PKC? in primary microglia blunted the microglial proinflammatory response elicited by the inflammogens, including ROS generation, nitric oxide production, and proinflammatory cytokine and chemokine release. Importantly, we found that PKC? activated NF?B, a key mediator of inflammatory signaling events, after challenge with inflammatory stressors, and that transactivation of NF?B led to translocation of the p65 subunit to the nucleus, I?B? degradation and phosphorylation of p65 at Ser536. Furthermore, both genetic ablation and siRNA-mediated knockdown of PKC? attenuated NF?B activation, suggesting that PKC? regulates NF?B activation subsequent to microglial exposure to inflammatory stimuli. To further investigate the pivotal role of PKC? in microglial activation in vivo, we utilized pre-clinical models of PD. We found that PKC? deficiency attenuated the proinflammatory response in the mouse substantia nigra, reduced locomotor deficits and recovered mice from sickness behavior in an LPS-induced neuroinflammation model of PD. Likewise, we found that PKC? knockout mice treated with MPTP displayed a dampened microglial inflammatory response. Moreover, PKC? knockout mice exhibited reduced susceptibility to the neurotoxin-induced dopaminergic neurodegeneration and associated motor impairments. Taken together, our studies propose a pivotal role for PKC? in PD pathology, whereby sustained PKC? activation drives sustained microglial inflammatory responses and concomitant dopaminergic neurotoxicity consequently leading to neurobehavioral deficits. We conclude that inhibiting PKC? activation may represent a novel therapeutic strategy in PD treatment.

Dataset Information

Ablation of RIP3 protects from dopaminergic neurodegeneration in experimental Parkinson's disease.

Publications

Ablation of RIP3 protects from dopaminergic neurodegeneration in experimental Parkinson's disease.

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