Contributions
Abstract: 113
Type: Oral
Abstract Category: Pathology and pathogenesis of MS - Inflammation and tissue damage
Introduction: The progressive phase of multiple sclerosis (MS) is characterised by accumulating grey matter (GM) pathology. The presence of immune cell infiltrates in the meninges is associated with lymphoid tissue development, greater cortical demyelination, shorter disease duration and significant neuronal loss. Analysis of isolated meninges of MS cases has shown increased gene expression for the pro-inflammatory cytokines: tumour necrosis factor (TNF), lymphotoxin-a (LTa) and interferon-g (IFNg).
Aims: To test the hypothesis that chronic production of pro-inflammatory cytokines in the meningeal compartment and diffusion into underlying GM can drive MS GM pathology.
Methods: HIV-1 based VSV-G pseudotyped lentiviral vectors were stereotactically injected into the sagittal sulcus (SS) of DA rats to deliver continuous transgene expression (TNF + IFNg or LTa + IFNg) in the meninges for chronic periods. A neuropathology analysis was conducted at time points up to 3 months, together with RT-PCR to determine changes to TNF receptor-1 (TNFR1) signalling.
Results: Efficient transduction of meningeal cells resulted in cytokine expression for up to 3 months. Injection of vectors for TNF or LTa, in combination with IFNg, induced the formation of large immune cell aggregates in the meninges by 28 dpi, which remained at 3 months, containing CD4+ and CD8+ T-cells, CD79a+ B-cells and Iba1+ macrophages. These aggregates extended the length of the SS and across the surface of the cortex. Subpial demyelination was accompanied by widespread microglial activation underlying these cellular aggregates. A decrease in neurofilament expression in regions with subpial demyelination was present along with signs of neuronal stress indicated by Fluorojade-C staining of layer II-IV neurons. TNF/TNFR1 interaction can initiate cell death by activating pathways involved in necroptosis. RT-PCR on cortical RNA from TNF and IFNg vector injected animals at 28 dpi showed an increase in expression of TNFR1 and downstream necroptotic genes, RIP3 and MLKL, compared to eGFP vector control animals. RIP3+ and MLKL+ immunopositive cells with the morphology of neurons were present in TNF vector injected animals.
Conclusions: Our results suggest that TNF and LTa in the presence of IFNg are potent inducers of meningeal inflammation and can activate TNF signalling pathways in cortical cells leading to neuronal death and subpial demyelination and thus may contribute to clinical progression.
Disclosure:
Rachel James: nothing to declare.
Eleanor Browne: nothing to declare.
Nicholas Mazarakis: nothing to declare.
Richard Reynolds: nothing to declare.
The study was funded by the UK MS Society to RR and NM.
Abstract: 113
Type: Oral
Abstract Category: Pathology and pathogenesis of MS - Inflammation and tissue damage
Introduction: The progressive phase of multiple sclerosis (MS) is characterised by accumulating grey matter (GM) pathology. The presence of immune cell infiltrates in the meninges is associated with lymphoid tissue development, greater cortical demyelination, shorter disease duration and significant neuronal loss. Analysis of isolated meninges of MS cases has shown increased gene expression for the pro-inflammatory cytokines: tumour necrosis factor (TNF), lymphotoxin-a (LTa) and interferon-g (IFNg).
Aims: To test the hypothesis that chronic production of pro-inflammatory cytokines in the meningeal compartment and diffusion into underlying GM can drive MS GM pathology.
Methods: HIV-1 based VSV-G pseudotyped lentiviral vectors were stereotactically injected into the sagittal sulcus (SS) of DA rats to deliver continuous transgene expression (TNF + IFNg or LTa + IFNg) in the meninges for chronic periods. A neuropathology analysis was conducted at time points up to 3 months, together with RT-PCR to determine changes to TNF receptor-1 (TNFR1) signalling.
Results: Efficient transduction of meningeal cells resulted in cytokine expression for up to 3 months. Injection of vectors for TNF or LTa, in combination with IFNg, induced the formation of large immune cell aggregates in the meninges by 28 dpi, which remained at 3 months, containing CD4+ and CD8+ T-cells, CD79a+ B-cells and Iba1+ macrophages. These aggregates extended the length of the SS and across the surface of the cortex. Subpial demyelination was accompanied by widespread microglial activation underlying these cellular aggregates. A decrease in neurofilament expression in regions with subpial demyelination was present along with signs of neuronal stress indicated by Fluorojade-C staining of layer II-IV neurons. TNF/TNFR1 interaction can initiate cell death by activating pathways involved in necroptosis. RT-PCR on cortical RNA from TNF and IFNg vector injected animals at 28 dpi showed an increase in expression of TNFR1 and downstream necroptotic genes, RIP3 and MLKL, compared to eGFP vector control animals. RIP3+ and MLKL+ immunopositive cells with the morphology of neurons were present in TNF vector injected animals.
Conclusions: Our results suggest that TNF and LTa in the presence of IFNg are potent inducers of meningeal inflammation and can activate TNF signalling pathways in cortical cells leading to neuronal death and subpial demyelination and thus may contribute to clinical progression.
Disclosure:
Rachel James: nothing to declare.
Eleanor Browne: nothing to declare.
Nicholas Mazarakis: nothing to declare.
Richard Reynolds: nothing to declare.
The study was funded by the UK MS Society to RR and NM.