The online version of this article (doi:10.1186/s12974-017-0805-x) contains supplementary material, which is available to authorized users.
Axon development plays a pivotal role in the formation of synapse, nodes of Ranvier, and myelin sheath. Interleukin-1β (IL-1β) produced by microglia may cause myelination disturbances through suppression of oligodendrocyte progenitor cell maturation in the septic neonatal rats. Here, we explored if a microglia-derived IL-1β would disturb axon development in the corpus callosum (CC) following lipopolysaccharide (LPS) administration, and if so, whether it is associated with disorder of synapse formation in the cerebral cortex and node of Ranvier.
Sprague-Dawley rats (1-day old) in the septic model group were intraperitoneally administrated with lipopolysaccharide (1 mg/kg) and then sacrificed for detection of IL-1β, interleukin-1 receptor (IL-1R1), neurofilament-68, neurofilament-160, and neurofilament-200, proteolipid, synaptophysin, and postsynaptic density 95 (PSD95) expression by western blotting and immunofluorescence. Electron microscopy was conducted to observe alterations of axonal myelin sheath and synapses in the cortex, and proteolipid expression was assessed using in situ hybridization. The effect of IL-1β on neurofilament and synaptophysin expression in primary neuron cultures was determined by western blotting and immunofluorescence. P38-MAPK signaling pathway was investigated to determine whether it was involved in the inhibition of IL-1β on neurofilament and synaptophysin expression.
In 1-day old septic rats, IL-1β expression was increased in microglia coupled with upregulated expression of IL-1R1 on the axons. The expression of neurofilament-68, neurofilament-160, and neurofilament-200 (NFL, NFM, NFH) and proteolipid (PLP) was markedly reduced in the CC at 7, 14, and 28 days after LPS administration. Simultaneously, cortical synapses and mature oligodendrocytes were significantly reduced. By electron microscopy, some axons showed smaller diameter and thinner myelin sheath with damaged ultrastructure of node of Ranvier compared with the control rats. In the cerebral cortex of LPS-injected rats, some axo-dendritic synapses appeared abnormal looking as manifested by the presence of swollen and clumping of synaptic vesicles near the presynaptic membrane. In primary cultured neurons incubated with IL-1β, expression of NFL, NFM, and synaptophysin was significantly downregulated. Furthermore, p38-MAPK signaling pathway was implicated in disorder of axon development and synaptic deficit caused by IL-1β treatment.
The present results suggest that microglia-derived IL-1β might suppress axon development through activation of p38-MAPK signaling pathway that would contribute to formation disorder of cortical synapses and node of Ranvier following LPS exposure.
Additional file 1: Figure S1. The CCK-8 assay (cell counting kit 8) was performed to determine the IL-1β concentration. The viability of neuronal cells was significantly reduced when neurons were treated with IL-1β at a dose exceeding 40 ng/mL. (TIF 820 kb)12974_2017_805_MOESM1_ESM.tif
Additional file 2: Figure S2. Apoptosis of neurons in the cortex. The incidence of apoptotic neurons co-labeled by caspase-3 (red) and NeuN (green) did not change markedly at 7 days (A–F), 14 days (G–L), and 28 days (M–R) in the cerebral cortex after LPS injection (D–F, J–L, P–R) when compared with controls (A–C, G–I, M–O). Scale bars: A–R 20μm. (TIF 6590 kb)12974_2017_805_MOESM2_ESM.tif
Additional file 3: Figure S3. Nissle staining shows the number of neurons in the cortex of postnatal rats at 7, 14, and 28 days after LPS injection (B, D, F) and their corresponding controls (A, C, E). (TIF 7048 kb)12974_2017_805_MOESM3_ESM.tif
Basu S, Agarwal P, Anupurba S, Shukla R, Kumar A. Elevated plasma and cerebrospinal fluid interleukin-1 beta and tumor necrosis factor-alpha concentration and combined outcome of death or abnormal neuroimaging in preterm neonates with early-onset clinical sepsis. J Perinatol. 2015;35:855–61. CrossRefPubMed
Deng YY, Lu J, Ling EA, Kaur C. Role of microglia in the process of inflammation in the hypoxic developing brain. Front Biosci (Schol Ed). 2011;3:884–900. CrossRef
Rathnasamy G, Ling EA, Kaur C. Iron and iron regulatory proteins in amoeboid microglial cells are linked to oligodendrocyte death in hypoxic neonatal rat periventricular white matter through production of proinflammatory cytokines and reactive oxygen/nitrogen species. J Neurosci. 2011;31:17982–95. CrossRefPubMed
Di Penta A, Moreno B, Reix S, Fernandez-Diez B, Villanueva M, Errea O, Escala N, Vandenbroeck K, Comella JX, Villoslada P. Oxidative stress and proinflammatory cytokines contribute to demyelination and axonal damage in a cerebellar culture model of neuroinflammation. PLoS One. 2013;8:e54722. CrossRefPubMedPubMedCentral
Shea TB, Lee S. The discontinuous nature of neurofilament transport accommodates both establishment and repair of the axonal neurofilament array. Cytoskeleton (Hoboken). 2013;70:67–73. CrossRef
Garcia ML, Lobsiger CS, Shah SB, Deerinck TJ, Crum J, Young D, Ward CM, Crawford TO, Gotow T, Uchiyama Y, Ellisman MH, Calcutt NA, Cleveland DW. NF-M is an essential target for the myelin-directed “outside-in” signaling cascade that mediates radial axonal growth. J Cell Biol. 2003;163:1011–20. CrossRefPubMedPubMedCentral
El-Husseini AE, Schnell E, Chetkovich DM, Nicoll RA, Bredt DS. PSD-95 involvement in maturation of excitatory synapses. Science. 2000;290:1364–8. PubMed
Deng YY, Lu J, Ling EA, Kaur C. Microglia-derived macrophage colony stimulating factor promotes generation of proinflammatory cytokines by astrocytes in the periventricular white matter in the hypoxic neonatal brain. Brain Pathol. 2010;20:909–25. PubMed
Larsson K, Rydevik B, Olmarker K. Disc related cytokines inhibit axonal outgrowth from dorsal root ganglion cells in vitro. Spine (Phila Pa 1976). 2005;30:621–4. CrossRef
Munoz L, Ralay Ranaivo H, Roy SM, Hu W, Craft JM, McNamara LK, Chico LW, Van Eldik LJ, Watterson DM. A novel p38 alpha MAPK inhibitor suppresses brain proinflammatory cytokine up-regulation and attenuates synaptic dysfunction and behavioral deficits in an Alzheimer’s disease mouse model. J Neuroinflammation. 2007;4:21–35. CrossRefPubMedPubMedCentral
- Microglia-derived IL-1β contributes to axon development disorders and synaptic deficit through p38-MAPK signal pathway in septic neonatal rats
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