Bystander activation of microglia by Brucella abortus-infected neurovascular cells induces neuronal death

DELPINO, M. VICTORIA; RODRÍGUEZ, JULIA; GIAMBARTOLOMEI, GUILLERMO H.; RODRÍGUEZ, ANA M.

Congreso; Reunión Anual de Sociedades de Biociencias; 2020

SAIC - SAI - SAFIS

Central nervous system invasion by bacteria of the genus Brucella results in an inflammatory disorder called neurobrucellosis. We have previously demonstrated that B. abortus is able to elicit neuronal death by activating microglia (with release of nitric oxide and proinflammatory cytokines) and inducing primary phagocytosis. The aim of this work was to investigate the role of soluble mediators released by infected cells of the neurovascular unit in the modulation of this phenomenon. For this, murine astrocytes and human brain microvasculature endothelial cells (HBMECs) were infected or not with B. abortus for 24h. After that, cell free culture supernatants were used to stimulate co-cultures of murine primary neuron/microglia during 48h. Neuronal density was evaluated by fluorescence microscopy. Stimulation of neuron/microglia co-cultures with supernatants from B. abortus-stimulated astrocytes or HBMECs induced a significant reduction in the density of neurons comparing with non-treated co-cultured or treated with supernatants from non-infected cells (p<0.005 for astrocytes supernatants and p<0.05 for HBMECs supernatants). Furthermore, supernatants obtained from infected cells increased several microglia function as proliferation (determined by microscopy, p<0.05), secretion of TNF-α (measured by ELISA; p<0.005) and phagocytic activity (evaluated by phagocytosis assay with Escherichia coli; p<0.005). In order to determine possible soluble mediators involved in these phenomenon, we use monoclonal antibodies to neutralize TNF-α and IL-6 cytokines in culture supernatants of infected astrocytes. Neutralization of IL-6 prevented neuronal loss in microglia-neurons co-cultures (p<0.05), whereas neutralization of TNF-α did not (p>0.05). Overall, these results demonstrate that soluble mediators secreted by B. abortus-infected neurovascular cells activate resting microglia and this bystander activation could be involved in worsening the neuronal death induced by B. abortus.