Electronic Poster | Session 2
072 – Expression of CD47, a “do not eat me” signal, is upregulated in neurons under pro-inflammatory conditions
Maryam Nakhaei-Nejad (1) – Dianhui Zhang (1) – David Barilla (1) – Fabrizio Giuliani (1)
University of Alberta, Department of Medicine, Edmonton, Canada (1)
Multiple Sclerosis (MS) is an autoimmune disease of the Central Nervous System characterized by immune-mediated demyelination and axonal injury. In MS patients, in addition to the loss of neurites, synaptic loss has been reported in both lesions and normal-appearing tissue. Synaptic changes have been associated with C3 complement-dependent phagocytosis in the hippocampus of MS patients. The process of phagocytosis is tightly regulated via a number of positive (“eat me”) and negative (“don’t eat me”) signals that lead to proper uptake of damaged cells or foreign molecules while preserving healthy cells. One of these “don’t eat me” signals is mediated by the interaction between CD47 on target cells and its receptor SIRPalpha on phagocytes, which has been shown to be involved in preventing excess synaptic pruning by microglia in developmental stages. Although a decrease in CD47 has been observed in MS lesions, it is not clear how CD47 is downregulated and if this decreased expression contributes to phagocyte-mediated synaptic loss and myelin stripping in MS. Thus, in our study we aimed to identify conditions that could mediate CD47 downregulation on neurons. To determine the effect of pro-inflammatory conditions on neuronal CD47 expression, primary human neurons (HN), and neurons differentiated from human induced pluripotent stem cells (iPS-Nn) were incubated with activated human lymphocyte supernatant (via CD3 and CD28 costimulation) or specific inflammatory cytokines, IFN-g and TNF-a. To our surprise, we detected an increase in CD47 mRNA levels (2.66-fold and 1.95-fold increase) in activated PBMC supernatant treated iPS-Nns and HNs, respectively, and a 4.36-fold increase in IFN-g treated iPS-Nns. TNF-alpha did not affect CD47 expression at mRNA levels. We are currently investigating the importance of CD47 upregulation in neurons using CD47 RNAi mediated knock down. Next, we tested whether CD47-SIRPalpha interaction is important in myelin uptake. Human myelin fractions were labelled with a pH sensitive dye, pHrodo, and myelin uptake and its localization to lysosomes was measured by flow cytometry. CD47 blockade significantly increased myelin uptake by phagocytes. Overall, unexpectedly, we have observed that under inflammatory conditions, neurons upregulate CD47 expression, which might have protective functions in situations of cellular stress. Also, CD47 interaction with SIRPalpha is involved in myelin debris uptake as its inhibition increases myelin uptake. The role of CD47 expression in myelin stripping needs further investigation.