Immunoglobulins G from Sera of Amyotrophic Lateral Sclerosis Patients Induce Oxidative Stress and Upregulation of Antioxidative System in BV-2 Microglial Cell Line

Abstract

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder with a very fast progression, no diagnostic tool for the presymptomatic phase, and still no effective treatment of the disease. Although ALS affects motor neurons, the overall pathophysiological condition points out to the non-cell autonomous mechanisms, where astrocytes and microglia play crucial roles in the disease progression. We have already shown that IgG from sera of ALS patients (ALS IgG) induce calcium transients and an increase in the mobility of acidic vesicles in cultured rat astrocytes. Having in mind the role of microglia in neurodegeneration, and a well-documented fact that oxidative stress is one of the many components contributing to the disease, we decided to examine the effect of ALS IgG on activation, oxidative stress and antioxidative system of BV-2 microglia, and to evaluate their acute effect on cytosolic peroxide, pH, and on reactive oxygen species (ROS) generation. All tested ALS IgGs (compared to control IgG) induced oxidative stress (rise in nitric oxide and the index of lipid peroxidation) followed by release of TNF-α and higher antioxidative defense (elevation of Mn- and CuZn-superoxide dismutase, catalase, and glutathione reductase with a decrease of glutathione peroxidase and glutathione) after 24 h treatment. Both ALS IgG and control IgG showed same localization on the membrane of BV-2 cells following 24 h treatment. Cytosolic peroxide and pH alteration were evaluated with fluorescent probes HyPer and SypHer, respectively, having in mind that HyPer also reacts to pH changes. Out of 11 tested IgGs from ALS patients, 4 induced slow exponential rise of HyPer signal, with maximal normalized fluorescence in the range 0.2–0.5, also inducing similar increase of SypHer intensity, but of a lower amplitude. None of the control IgGs induced changes with neither of the indicators. Acute ROS generation was detected in one out of three tested ALS samples with carboxy-H2DCFDA. The observed phenomena demonstrate the potential role of inflammatory humoral factors, IgGs, as potential triggers of the activation in microglia, known to occur in later stages of ALS. Therefore, revealing the ALS IgG signaling cascade in microglial cells could offer a valuable molecular biomarker and/or a potential therapeutic target.

Immunity in amyotrophic lateral sclerosis: blurred lines between excessive inflammation and inefficient immune responses

“…IgGs accumulate in motoneurons; IgG from ALS patients enter and stimulate motoneurons, astrocytes and microglia ( Engelhardt and Appel, 1990 ; Appel et al , 1991 ; Pagani et al , 2011 ; Bataveljic et al , 2014 ; Miloševic et al , 2017 ).…”

Serum IgG-induced microglial activation enhances neuronal cytolysis via the NO/sGC/PKG pathway in children with opsoclonus-myoclonus syndrome and neuroblastoma

“…Notably, increased microglial activation is not specific to only OMS, since previous literatures have documented that serum IgG from patients with PD [12] or ALS [13] enhances the activation of microglia and the production of NO, and serum IgG from patients with SLE induced behavioral changes are mediated by microglial activation [14][15][16]. Moreover, several autoantibodies found in patients with OMS [45][46][47][48] also exist in other diseases, such as autoantibody against glycine receptor in progressive encephalomyelitis with rigidity and myoclonus [47,49] or autoantibody against glutamic acid decarboxylase in stiff-person syndrome [49].…”

“…Moreover, the expression of microglial marker soluble CD14 and proinflammatory cytokines is enhanced in CSF from patients with pediatric OMS [10,11]. Additionally, serum IgG from patients or autoantibody existed in patients directly enhances microglial activation in PD [12], ALS [13] and SLE [14][15][16], or initially binds with astrocytes or neurons and further indirectly affects microglial activation [43,44]. Autoantibodies are also detectable in sera and CSF of children with OMS [25,26] and may be contained in serum IgG from children with OMS and NB in our study, although we did not identify these autoantibodies and autoantibodies include IgM besides IgG.…”

“…Notably, patients with pediatric OMS exhibit increased expression of a microglial marker and proinflammatory cytokines in cerebrospinal fluid (CSF) and some children with OMS are post-infectious [10,11]. Furthermore, serum IgG or autoantibody existed in patients enhances microglial activation in Parkinson disease (PD) [12], amyotrophic lateral sclerosis (ALS) [13] and systemic lupus erythematosus (SLE) [14][15][16]. Thus, it is reasonable to hypothesize that serum IgG from children with OMS and NB may impact the activation of microglia.…”

Defining novel functions for cerebrospinal fluid in ALS pathophysiology

“…Out of these, IgG, also present in post-mortem ALS samples, has been found to favour neurodegeneration, both in vitro and in vivo [49,155,156]. IgG further possesses pro-inflammatory properties, promoting microglia recruitment and upregulation of inflammatory cytokines [124,141,142]. Some of the other chemokines found to be elevated in ALS-CSF include IL-6, TNF-α and TGF-β [81,130,172], with the finding of raised TNF-α levels being particularly interesting, given its associations with glutamate excitotoxicity [43,178,218].…”