LaMorte, M. | Search bioRxiv

bioRxiv2026

CD40 ligand and IFN-γ induce proinflammatory activity and microglia-mediated neurodegeneration in human stem cell models

The cluster of differentiation 40 (CD40)/CD40 ligand (CD40L) co-stimulatory pathway plays a critical role in the activation and regulation of immune networks involving both adaptive and innate immune cells. Dysregulation of this pathway has been implicated in multiple autoimmune diseases, including multiple sclerosis (MS). MS is characterized by progressive neurodegeneration driven by compartmentalized neuroinflammation within the central nervous system (CNS), sustained low-grade T cell infiltration at the blood-brain barrier, and T cell reactivation at CNS border tissues. CD40L-expressing T cells can activate CD40-expressing microglia, which are a critical effector cell that contribute to neurodegeneration in MS. However, the consequence of this functional interaction in the brain remains poorly understood. Here, we demonstrate that the combination of CD40L and interferon-gamma (IFN-{gamma}), a cytokine produced by activated T cells, induces a robust inflammatory response and cell death in human induced pluripotent stem cells (hiPSC)-derived microglia. Stimulation with CD40L elicited microglial clustering, morphological transformation, and robust release of proinflammatory cytokines including interleukin-12 (IL-12), tumor necrosis factor-alpha (TNF-), and IL-6. Anti-CD40L antibody treatment reversed both cell death and inflammatory response and partially normalized the associated transcriptional changes. Furthermore, using an hiPSC-derived tri-culture system comprising neurons, astrocytes, and microglia, we demonstrated that CD40L and IFN-{gamma} trigger microglia-dependent neuronal dysregulation and death. Anti-CD40L treatment substantially attenuated neuronal injury. Finally, transcriptomic enrichment analysis identified CD40L-mediated microglial gene expression signatures that correlated with CD40 expression in primary progressive MS brain tissue but not in the normal-appearing white matter of healthy controls. Collectively, these findings support a role for the microglial CD40/T cell CD40L axis in MS pathology and suggest CD40L blockade as a potential therapeutic strategy warranting further investigation.

neuroscience↗

bioRxiv2024

Glial state changes and neuroinflammatory RIPK1 signaling are a key feature of ALS pathogenesis

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that causes motor neuron loss in the brain and spinal cord. Neuroinflammation driven by activated microglia and astrocytes is prominent in ALS, but an understanding of cell state dynamics and which pathways contribute to the disease remains unclear. Single nucleus RNA sequencing of ALS spinal cords demonstrated striking changes in glial cell states, including increased expression of inflammatory and glial activation markers. Many of these signals converged on RIPK1 and the necroptotic cell death pathway. Activation of the necroptosis pathway in ALS spinal cords was confirmed in a large bulk RNA sequencing dataset and at the protein level. Blocking RIPK1 kinase activity delayed symptom onset and motor impairment and modulated glial responses in SOD1G93A mice. We used a human iPSC-derived motor neuron, astrocyte, and microglia tri-culture system to identify potential biomarkers secreted upon RIPK1 activation, inhibited pharmacologically in vitro, and modulated in the CSF of people with ALS treated with a RIPK1 inhibitor. These data reveal ALS-enriched glial populations associated with inflammation and suggest a deleterious role for neuroinflammatory signaling in ALS pathogenesis.

neuroscience↗