G392E neuroserpin causing the dementia FENIB is secreted from cells but is not synaptotoxic

Abstract

Familial encephalopathy with neuroserpin inclusion bodies (FENIB) is a progressive neurodegenerative disease caused by point mutations in the gene for neuroserpin, a serine protease inhibitor of the nervous system. Different mutations are known that are responsible for mutant neuroserpin polymerization and accumulation as inclusion bodies in many cortical and subcortical neurons, thereby leading to cell death, dementia and epilepsy. Many efforts have been undertaken to elucidate the molecular pathways responsible for neuronal death. Most investigations have concentrated on analysis of intracellular mechanisms such as endoplasmic reticulum (ER) stress, ER-associated protein degradation (ERAD) and oxidative stress. We have generated a HEK-293 cell model of FENIB by overexpressing G392E-mutant neuroserpin and in this study we examine trafficking and toxicity of this polymerogenic variant. We observed that a small fraction of mutant neuroserpin is secreted via the ER-to-Golgi pathway, and that this release can be pharmacologically regulated. Overexpression of the mutant form of neuroserpin did not stimulate cell death in the HEK-293 cell model. Finally, when treating primary hippocampal neurons with G392E neuroserpin polymers, we did not detect cytotoxicity or synaptotoxicity. Altogether, we report here that a polymerogenic mutant form of neuroserpin is secreted from cells but is not toxic in the extracellular milieu.

Polymerogenic neuroserpin causes mitochondrial alterations and activates NFκB but not the UPR in a neuronal model of neurodegeneration FENIB

“…The expression of G392E NS caused an increase in the percentage of neurons with mitochondrial localisation to the perinuclear area, indicative of alterations in mitochondrial dynamics that can have multiple origins, as seen in other neurodegenerative conditions [35]. We have previously described the presence of G392E NS polymers in the culture medium in this and other cellular systems [7,12,13,17,36] and have shown that NS polymers are secreted and not released by cell death [32], similarly to polymers of the Z variant of A1AT [37]. Our present results support the idea that mitochondrial alterations are caused by intracellular G392E NS, since the culture medium conditioned by G392E NS cells did not cause the same phenotype in control GFP or WT NS neurons.…”

“…Since polymers of NS have been consistently found in the culture medium of G392E NS cells in this (Fig. 1B, nondenaturing PAGE media panel) and other cellular models of FENIB [7,17,32], we investigated their potential effects in mitochondrial distribution. We first quantified the amount of G392E NS polymers in the culture media by sandwich ELISA using a monoclonal antibody with high affinity for polymeric NS (7C6, [7]), and found an average concentration of 1.25 ng/ml.…”

Neuroserpin, a crucial regulator for axogenesis, synaptic modelling and cell–cell interactions in the pathophysiology of neurological disease

“…Furthermore, it is suggested that neuroserpin polymer formation at the neuronal synapse may give rise to chronic localised inflammatory changes, thus participating in the loss of synaptic plasticity in FENIB-associated neurodegeneration [14] FENIB pathophysiology is characterised by the accumulation of mutant neuroserpin in the neuronal ER, triggering an ER overload response, with the protein overload potentially activating an altogether different set of stress signalling pathways in diseased conditions, such as NFκB activation, as was observed in the cells in culture [27]. The proteinase inhibitory activity of the serpin and its ability to be secreted is also affected under such conditions [9,11,73]. The expression of polymerogenic mutant forms of neuroserpin has been shown to induce the upregulation of antioxidant defence mechanisms in neural cell cultures derived from mice brain, and the disruption of these antioxidant mechanisms was shown to promote pro-apoptotic pathways [52].…”

Neuroserpin: structure, function, physiology and pathology

“…Mutant mice were more susceptible to kainite-induced seizures Takasawa et al [ 113 ] Overexpression of wild type and S49P human NS Correlation between mutant NS accumulation and neurodegeneration. Transient induction of the UPR in young mice Schipanski et al [ 69 ] Overexpression of wild type and S49P human NS Transient inflammatory responses and UPR activation at middle stage of the disease, sequestration of UPR activators GRP78 and GRP94 in NS-positive inclusions Lopez-Gonzalez et al [ 118 ] Overexpression of wild type and S49P human NS Increased expression of the postsynaptic protein PSD-95 in the hippocampus of S49P NS mice Ingwersen et al [ 159 ] NS deficiency (knock-out) Unaltered tPA activity and behavioural abnormalities: reduced locomotor activity in novel environments, anxiety-like response on the O-maze, neophobic phenotype in the novel object test Madani et al [ 84 ] NS deficiency (knock-out) crossed with human APP-J20 transgenic mice Rapid clearance of Abeta 1–42 injected into the frontal cortex in the absence of NS. Following crossing with human APP-J20 transgenic mice, decrease in amyloid-beta peptides, reduction in number and size of plaques, increased activity of tPA associated with plaques, rescue of spatial memory defects compared to J20 mice Fabbro et al [ 135 ] NS deficiency (knock-out) Following induction of focal ischemic stroke, aggravated infarct size and neurological outcome and increased activation of proinflammatory microglia Gelderblom et al, 2013 [ 123 ] NS deficiency (knock-out) and NS/tPA double deficient mice Following kainic acid injection into the amygdala, reduced latency to seizure onset and generalisation, shorter mean time of survival and increase in blood–brain barrier permeability compared to wild type mice.…”