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bioRxiv · 10.1101/2020.10.16.341644

Role of TNF-α among inflammatory molecules secreted by injured astrocytes in the modulation of in vitro neuronal networks

Lantoine, J.·Proces, A.·Villers, A.·Halliez, S.·Buee, L.·Ris, L.·Gabriele, S.
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Abstract

Traumatic brain injury (TBI) remains one of the leading causes of mortality and morbidity worldwide. Despite its high prevalence and extensive efforts to develop neuroprotective therapies, effective treatments for TBI are still limited. Among important neuronal damages, TBI induces structural and functional alterations of astrocytes, the most abundant cell type in the brain. Injured astrocytes respond in diverse mechanisms that result in reactive astrogliosis and are involved in the physiopathological mechanisms of TBI in an extensive and sophisticated manner. The establishment of effective neuroprotective treatments for TBI requires to better understand the complex biochemical interactions between activated astrocytes and neurons that contribute to the secondary injury. To address this challenge, we studied in vitro the role of mechanically injured astrocytes on the growth and synaptic connections of cortical neuronal networks of controlled architectures grown on well-defined protein micropatterns. Astrocytes were cultivated on elastic membranes and mechanically activated by stretching cycles. The culture media of healthy or activated astrocytes was then introduced on neuronal networks. We analyzed the neuronal viability, the neurite growth and the synaptic density of neuronal networks to understand the role of the inflammatory molecules secreted by mechanically activated astrocytes. Furthermore, we cultivated neuronal networks during 13 days with different doses of TNF- in order to decipher its individual contribution among the other cytokines. Here we show that the ratio of tubulin to synapsin area was significantly higher in neuronal networks treated with either 4 or 2 doses of TNF-, suggesting that TNF- can promote the tubulin polymerization process. Assuming that TNF- can bind to either TNFR1 or TNFR2 receptors, which lead respectively to the cell survival or the cell apoptosis, we studied the modulation of the both TNF- receptors in response to the medium of mechanically activated astrocytes and different doses of TNF-. Our findings indicate that the amount of both receptors increases with the maturation of the network. In addition, we observed a significant modulation of the amount of TNFR1 and TNFR2 in response to the media of injured astrocytes that leads to a large imbalance between both receptors, suggesting an important role for TNF-signaling in the physiopathological mechanisms of TBI.

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Lantoine, J., Proces, A., Villers, A., Halliez, S., Buee, L., Ris, L., Gabriele, S.. 2020-10-16. Role of TNF-α among inflammatory molecules secreted by injured astrocytes in the modulation of in vitro neuronal networks. https://doi.org/10.1101/2020.10.16.341644

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