A robust platform for high-throughput screening of therapeutic strategies for acute and chronic spinal cord injury

Vaibhav Patil,Enda O'Connell,Leo R Quinlan,Howard Fearnhead,Siobhan Mcmahon,Abhay Pandit

doi:10.1016/j.isci.2021.102182

Vaibhav Patil, Enda O'Connell + Show 4 more

Open Access

https://doi.org/10.1016/j.isci.2021.102182

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Journal: iScience	Publication Date: Feb 12, 2021
Citations: 7	License type: cc-by

Affiliation: Ollscoil na Gaillimhe – University of Galway

Abstract

SummaryAstrocytes and microglia are critical regulators of inflammatory cascade after spinal cord injury (SCI). Existing glial in vitro studies do not replicate inflammatory phases associated with SCI. Here, we report an in vitro model of mixed glial culture where inflammation is induced by the administration of pro-inflammatory cytokines (tumor necrosis factor-α, interleukin-1β, and interleukin-6) to promote pathologically relevant “acute” and “chronic” inflammatory phases. We observed SCI relevant differential modulation of inflammatory pathways, cytokines, chemokines, and growth factors over 21 days. Mitochondrial dysfunction was associated with a cytokine combination treatment. Highly expressed cytokine induced neutrophil chemoattractant (CINC-3) chemokine was used as a biomarker to establish an enzyme-linked immunosorbent assay-based high-throughput screening (HTS) platform. We screened a 786-compound drug library to demonstrate the efficacy of the HTS platform. The developed model is robust and will facilitate in vitro screening of anti-reactive glial therapeutics for the treatment of SCI.

Highlights

Spinal cord injury (SCI) is a catastrophic event that results in severe primary mechanical trauma followed by more complex secondary injury (Kabu et al, 2015)
Existing glial in vitro studies do not replicate inflammatory phases associated with spinal cord injury (SCI)
We report an in vitro model of mixed glial culture where inflammation is induced by the administration of pro-inflammatory cytokines to promote pathologically relevant ‘‘acute’’ and ‘‘chronic’’ inflammatory phases

Summary

Introduction

Spinal cord injury (SCI) is a catastrophic event that results in severe primary mechanical trauma followed by more complex secondary injury (Kabu et al, 2015). The dual activation (microglia and astrocytes) sparks a burst release of pro-inflammatory cytokines such as tumor necrosis factor-a (TNF-a), interleukin (IL)-1b, and interleukin (IL)-6 at the injury site (Donnelly and Popovich, 2008) These cytokines activate various pathways such as nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB), mitogen-activated protein kinase (MAPK), nitric oxide (NO) synthase, and chemokine signaling by activating their corresponding receptors which plays a vital role in upregulating the inflammatory cascade (Becher et al, 2017). Glial activation induces reactive oxygen species (ROS) production, which further elevates the inflammation and upregulates the corresponding biomarkers (Xu et al, 2005) These inflammatory stimuli can persist from acute to chronic phases after the initial trauma (Norenberg et al, 2004).

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