Abstract
Toll-like receptor 4 (TLR4) activation is pivotal to innate immunity and has been shown to regulate proliferation and differentiation of human neural stem cells (hNSCs) in vivo. Here we study the role of TLR4 in regulating hNSC derived from the human telencephalic-diencephalic area of the fetal brain and cultured in vitro as neurospheres in compliance with Good Manifacture Procedures (GMP) guidelines. Similar batches have been used in recent clinical trials in ALS patients. We found that TLR2 and 4 are expressed in hNSCs as well as CD14 and MD-2 co-receptors, and TLR4 expression is downregulated upon differentiation. Activation of TLR4 signaling by lipopolysaccharide (LPS) has a positive effect on proliferation and/or survival while the inverse is observed with TLR4 inhibition by a synthetic antagonist. TLR4 activation promotes neuronal and oligodendrocyte differentiation and/or survival while TLR4 inhibition leads to increased apoptosis. Consistently, endogenous expression of TLR4 is retained by hNSC surviving after transplantation in ALS rats or immunocompromised mice, thus irrespectively of the neuroinflammatory environment. The characterization of downstream signaling of TLR4 in hNSCs has suggested some activation of the inflammasome pathway. This study suggests TLR4 signaling as essential for hNSC self-renewal and as a novel target for the study of neurogenetic mechanisms.
Highlights
Stem cells in brain niches are responsible for neurogenesis and integration of new neurons into functional circuits, with inflammatory and immune system mediators playing critical roles in neurogenesis and in several diseases of the nervous system[1,2]
We observed by immunofluorescence that both TLR2 and Toll-like receptor 4 (TLR4) are expressed by undifferentiated human neural stem cells (hNSCs) and undergo downregulation with differentiation (Fig. S1)
The expression of TLR4, CD14, and myeloid differentiation factor 2 (MD-2) was detected in undifferentiated hNSC, but less than in peripheral blood mononuclear cells (PBMC) (Fig. 1a). hNSC were treated with LPS 10 and 100 nM and with the synthetic antagonist FP7 used at two concentrations, 1 and 10 μM11,12
Summary
Stem cells in brain niches are responsible for neurogenesis and integration of new neurons into functional circuits, with inflammatory and immune system mediators playing critical roles in neurogenesis and in several diseases of the nervous system[1,2]. A family of innate immune receptors, the Toll-like receptors (TLRs), respond to the presence of minute amounts of circulating pathogen-associated molecular patterns (PAMPs) and. These receptors are expressed in the central nervous system by microglia, perivascular dendritic cells, and neural progenitor cells (NPCs) and can respond to endogenous DAMPs5. TLR2 activation, in infectious, ischemic, and inflammatory diseases, could negatively impinge on brain development by inhibition of NPC proliferation[8]. Previous studies in murine cells or animal models have shown a multifaceted role
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