Nitric Oxide/Cyclic GMP-Dependent Calcium Signalling Mediates IL-6- and TNF-\u03b1-Induced Expression of Glial Fibrillary Acid Protein

Claudia Sticozzi,Maria Frosini,Federica Pessina,Giuseppe Belmonte

doi:10.1007/s12031-020-01708-3

Abstract

Astrocyte activation is characterized by hypertrophy with increased glial fibrillary acidic protein (GFAP), whose expression may involve pro-inflammatory cytokines. In this study, the effects of pro-inflammatory IL-6 and TNF-α and anti-inflammatory cytokines IL-4 and IL-10 on nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) signalling, intracellular calcium concentration ([Ca2+]i) and GFAP expression were investigated. In human glioblastoma astrocytoma U-373 MG cells, IL-6 and TNF-α, but not IL-4 or IL-10, increased iNOS, cGMP, [Ca2+]i and GFAP expression. The inhibitors of iNOS (1400 W), soluble guanylyl cyclase (ODQ) and IP3 receptors (ryanodine and 2-APB) reversed the increase in cGMP or [Ca2+]i, respectively, and prevented GFAP expression. In rat striatal slices, IL-6 and TNF-α, at variance with IL-4 and IL-10, promoted a concentration-dependent increase in Ca2+ efflux, an effect prevented by 1400 W, ODQ and RY/2APB. These data were confirmed by in vivo studies, where IL-6, TNF-α or the NO donor DETA/NO injected in the striatum of anaesthetised rats increased cGMP levels and increased GFAP expression. The present findings point to NO/cGMP-dependent calcium signalling as part of the mechanism mediating IL-6- and TNF-α-induced GFAP expression. As this process plays a fundamental role in driving neurotoxicity, targeting NO/cGMP-dependent calcium signalling may constitute a new approach for therapeutic interventions in neurological disorders.

Highlights

Neuroinflammation, an inflammatory response within the brain or spinal cord, involves the participation of resident immune-like glial cells such as microglia and astrocytes, endothelial cells and different peripheral immune cells including monocytes/macrophages, leukocytes and lymphocytes (DiSabato et al 2016)
In the present study, using in vitro, ex vivo and in vivo models, we investigated the effects of TNF-α and IL-6 on the expression of glial fibrillary acidic protein (GFAP), a hallmark of astrocyte activation, and the role of the nitric oxide (NO)/cyclic guanosine monophosphate (cGMP)/calcium signalling pathway in mediating this response
30 min RY (10 μM) plus 2APB (100 μM) pretreatment reversed the effects elicited by the cytokines. These results suggest that the IL-6- and TNF-α-dependent upregulation of GFAP expression is mediated by nitric oxide/cyclic guanosine monophosphate (NO/cGMP)/Ca2+ signalling and that the intracellular calcium stores are involved

Summary

Introduction

Neuroinflammation, an inflammatory response within the brain or spinal cord, involves the participation of resident immune-like glial cells such as microglia and astrocytes, endothelial cells and different peripheral immune cells including monocytes/macrophages, leukocytes and lymphocytes (DiSabato et al 2016). In astrocytes and brain striatal tissue, we previously established that nitric oxide/cyclic guanosine monophosphate (NO/cGMP)-dependent Ca2+ mobilization from intracellular pools is part of a signalling pathway subserving the pro-inflammatory/pyrogenic effect of IL-1β (Meini et al 2000; Palmi and Meini 2002) and that the same cascade, via activation of calmodulin and extracellular signal-regulated protein kinases (ERK1/2), is responsible for IL-1β upregulation of astrocyte proliferation (Meini et al 2006, 2008). In the present study, using in vitro, ex vivo and in vivo models, we investigated the effects of TNF-α and IL-6 on the expression of glial fibrillary acidic protein (GFAP), a hallmark of astrocyte activation, and the role of the NO/cGMP/calcium signalling pathway in mediating this response

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