Microglia-derived IL-1β contributes to axon development disorders and synaptic deficit through p38-MAPK signal pathway in septic neonatal rats
© The Author(s). 2017
Received: 11 August 2016
Accepted: 26 January 2017
Published: 14 March 2017
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.
KeywordsMicroglia LPS IL-1β PWMD Axon Neurofilament Synapse
Neonatal sepsis may cause a systemic inflammatory response which is an important risk factor for periventricular white matter (PWM) damage (PWMD) in the developing brain [1–3]. A large number of immune effector cells are mobilized into the neonatal circulation [1–3]. Concomitantly, these immune cells release proinflammatory cytokines such as tumor necrosis factor (TNF-α) and interleukin-1β (IL-1β) which readily cross the blood-brain barrier into the brain parenchyma . In the latter, the serum-derived proinflammatory cytokines can activate microglia, the resident immune cells in the central nervous system (CNS), which initiates complex inflammatory cascades including excessive release of proinflammatory cytokines, reactive oxygen species (ROS), and glutamate excitotoxicity [5, 6]. It has been reported that this may induce the injury of immature oligodendrocytes and axons resulting in hypomyelination, a hallmark feature of PWMD [7–9].
Various inflammatory mediators play different roles in the pathogenesis of PWMD [10–16]. Among them, the most widely studied mediators are the proinflammatory cytokines including TNF-α and IL-1β [17, 18]. TNF-α produced by activated microglia may elicit the apoptosis of oligodendrocytes via its TNFR1 which activate the signal pathway of apoptosis in the oligodendrocytes [19, 20]. There is also mounting evidence suggesting that IL-1β is a crucial contributor to various acute and chronic neurodegenerative diseases [21–23]. Unlike TNF-α, IL-1β was documented as being nontoxic to oligodendrocyte lineage cells in that it could not induce oligodendrocyte apoptosis through its receptors . However, some studies have demonstrated that IL-1β can suppress oligodendrocyte proliferation at the late developmental stage of oligodendrocyte progenitor cell (OPC) . Our previous studies have found that microglia-derived IL-1β could affect OPC maturation and induce hypomyelination in the PWM of septic neonatal brain . In primary cultured neurons administrated with recombinant IL-1β, a significant increase in the phosphorylation of neuronal tau was accompanied by a decline in synaptophysin levels . IL-1 receptor antagonist (IL-1ra) and anti-IL-1β antibody attenuated the effects of IL-1β on neuronal tau and synaptophysin . Systemic inflammation activated innate immune response in the CNS and induced the release of IL-1β from activated microglia, which increased axon injury and synaptic deficit [26–29]. However, the underlying molecular mechanisms whereby IL-1β is involved in PWMD in septic neonatal rats have not been fully addressed. Here, we provide evidences that IL-1β produced by activated microglia could induce disorder of axon development and synaptic deficit in septic neonatal brain. Expression of IL-1β in microglia and its receptor 1 on developing axons was first observed by double immunofluorescence. The axon development, node of Ranvier, and myelin sheath in the PWM and synapse formation in the cerebral cortex were examined in septic rats in comparison with the controls. Furthermore, the signaling pathway via which IL-1β could suppress axon development and synapse formation was investigated. It is suggested that microglia-derived IL-1β may have a negative impact on axon development and synapse formation through activation of p38-MAPK signaling pathway after LPS administration.
Number of rats killed at various time points after the LPS exposure (in brackets) and their age-matched controls for various methods (outside the brackets)
In situ hybrization
Primary cultures of cortical neurons
Primary cortical neuron culture was performed using neonatal SD rats (1-day old), as described previously  with some modifications. The cerebral cortices were dissected from the neonatal brain, minced into small tissue pieces of size 1 mm3 excluding the hippocampus and meninges, trypsinized for 15 min with 0.125% trypsin (Gibco) at 37 °C, and then neutralized with fetal bovine serum (FBS) (Life Technology). Cells were dissociated by passage through a pasteur pipette. The suspension containing neural cells was centrifuged at 1100 rpm for 5 min; thereafter, the cells were resuspended in Dulbecco’s modified eagle medium (DMEM) containing 10% fetal bovine serum (Life Technology) and plated in 6-well plates (Corning) for various experiments, or plated in 96-well plates (Corning) specifically for the CCK-8 test. All the plates were pre-coated with poly-l-lysine (Sigma). At 6 h after plating, DMEM/10% FBS was replaced with neurobasal medium containing 2% B27 and 1% glutamine; half of the medium was replaced with neurobasal containing 2% B27 without glutamine 3 days later. For immunocytochemistry, primary cultured neurons were detached from 75-cm2 flask, then plated at a density of 2.5 × 105/well in a 24-multiwell culture dish. For western blotting, primary cultured neurons were plated at 1 × 106 cells per flask; thereafter, they received different treatments according to experimental protocols on the following day. The purity of cortical neurons was examined through immunocytochemical staining using MAP-2 (a marker of neurons) and 4′6-diamidino-2-phenylindole (DAPI). The purity of primary neuron cultures in this study was above 95%.
Treatment of primary cultured neurons
To determine the IL-1β concentration, the CCK-8 assay (cell counting kit 8) was performed following the manufacturer’s instructions (DOJINDO). Primary cultured neurons (3000 cells/well in 100-ul neurobasal medium) were incubated with different concentrations (0, 5, 10, 20, 40, 80, 100, and 200 ng/ml) of IL-1β for 24 h in 96-well plates (Corning) at 37 °C. Ten-microliter CCK-8 solution was then added to each well, and the plates were incubated at 37 °C for 6 h. The optical density (OD) in each well was measured at 450 nm using an enzyme-linked immunosorbent assay (ELISA) reader (BioTek, Winooski, VT, USA) according to the manufacturer’s instructions. The neuronal cell activity remained relatively changed when neurons were treated with IL-1β at a dose less than 40 ng/mL (Additional file 1: Figure S1).
To examine the effects of IL-1β on expression of NFL, NFM, and synaptophysin in primary cultured neurons by western blotting analysis and immunocytochemical staining, the cells were cultured in neurobasal medium containing 2% B27 and 1% glutamine in a humidified atmosphere of 95% air and 5% CO2 for 24 h. The neurons were plated in 6-well plates at the density of 2 × 106/well for western blotting analysis and in 6-well plates at the density of 1.2 × 106/well for immunofluorescence staining. The neurons in group I were randomized into four groups including the control group (0.01 M PBS), IL-1β (40 ng/mL) group, IL-1β (40 ng/mL) + IL-1Ra (40 ng/mL) group, and IL-1Ra (40 ng/mL) group.
The primary cultured cortical neurons in group II were used to investigate the effects of IL-1β on the phosphorylation of p38-MAPK pathway. For this, the primary neurons were administrated with 40-ng/ml IL-1β for 0.5, 1, 2, 4, and 6 h before harvest.
To determine whether p38-MAPK signaling pathway is implicated in the effects of IL-1β on expression of NFL, NFM, and synaptophysin in primary cultured cortical neurons, the cells (2 × 106/well) were cultured in 6-well plates in a humidified atmosphere of 95% air and 5% CO2 at 37 °C. Subsequently, the cells were randomized into four subgroups including the control group (0.01 M PBS), IL-1β (40 ng/ml) group, IL-1β (40 ng/ml) + SB203580 (10 μmol/L) group, and SB203580 (10 μmol) group. After incubation with above-mentioned reagents for 24 h, the cells were harvested.
Antibodies used in experiments
Cell Sigaling Technology, Danvers, MA, USA
Cell Sigaling Technology, Danvers, MA, USA
Cell Sigaling Technology, Danvers, MA, USA
Chemicon International, Temecula, CA, USA
Santa Cruz Biotechnology, Santa Cruz, CA, USA
Sigma-Aldrich, Saint Louis, MO, USA
Sigma-Aldrich, Saint Louis, MO, USA
Sigma-Aldrich, Saint Louis, MO, USA
Abcam, Cambridge, MA, USA
Abcam, Cambridge, MA, USA
Abcam, Cambridge, MA, USA
Abcam, Cambridge, MA, USA
Abcam, Cambridge, MA, USA
Invitrogen Life Technologies Corporation
Invitrogen Life Technologies Corporation
Invitrogen Life Technologies Corporation
Sigma-Aldrich, St. Louis, MO
Coronal frozen brain sections of 10-μm thickness were incubated with 0.3% hydrogen peroxide in methanol to deactivate endogenous peroxidase for 20 min. After washing three times with PBS, the sections were blocked with a mixed solution composed of 5% BSA and 0.3% Triton X-100 in PBS for 30 min at room temperature. Subsequently, the brain sections from rats at different time points (n = 3 at each time point) after PBS or LPS injection were randomized into five groups. The sections in group I from rats at 7, 14, and 28 days after PBS or LPS injection were incubated with antibody against NFL (Table 2). The sections in group II from the control and septic rats at 14 and 28 days were incubated with PLP antibody (Table 2). The sections in group III from the control and septic rats at 14 and 28 days were incubated with antibodies against PSD95 (Table 2) and synaptophysin (Table 2). The sections in group IV from rats at 6 h and 2, 4, and 6 days after PBS and LPS injection were incubated with IL-1β antibody (Table 2) and Lectin (Table 2). The brain sections in group V from the control and septic rats at 2 and 4 days were incubated with antibody against IL-1R1 (Table 2) and NFL (Table 2). On the next day, after washing three times with PBS, the sections were incubated with secondary antibodies: Alexa Fluor555 goat anti-mouse IgG (H + L) (Table 2) for NFL in group I, Alexa Fluor555 donkey anti-rabbit IgG (H + L) (Table 2) for PLP in group II, Alexa Fluor555 donkey anti-rabbit IgG (H + L) (Table 2) and Alexa Fluor488 donkey anti-mouse IgG (H + L) (Table 2) for PSD95/synaptophysin in group III and for IL-1R1/NFL in group V, and Alexa Fluor555 donkey anti-rabbit IgG (H + L) (Table 2) for IL-1β in group IV for 1 h. After three rinses in PBS, the sections in group IV were incubated with Lectin (Table 2) for 1 h. Incubation of sections for all groups was carried out at room temperature. Finally, all sections were counterstained with DAPI (Sigma-Aldrich, St. Louis, MO, USA, Cat. No. D9542) and then observed using a fluorescence microscope (Olympus System Microscope Model BX53, Olympus Company Pte, Tokyo, Japan).
For primary cultured cortical neurons, the cells were treated with protein IL-1β, IL-1β + IL-1Ra, IL-1Ra, and the equal volume of PBS for 1 day. After three rinses in PBS, the cells were fixed in 4% paraformaldehyde for 30 min and then blocked in 1% BSA for 1 h. After this, the cells were incubated with NFM antibody (Table 2) overnight at 4 °C. On the next day, after three rinses in PBS (10 min each time), the cells were incubated with Alexa Fluor488 donkey anti-mouse IgG (H + L) (Table 2) for 1 h. Following three washes in PBS, the cells were incubated with DAPI for 5 min and observed under a fluorescence microscope (Olympus System Microscope Model BX53, Olympus Company Pte, Tokyo, Japan).
LPS-injected rats (n = 3 at 28 days) and littermate controls (n = 3 at 28 days) were transcardially perfused with a mixed aldehyde fixative composed of 2% paraformaldehyde and 3% glutaraldehyde. Coronal sections of the brain at about 1-mm thick were prepared. Blocks of the corpus callosum (CC) and cerebral cortex were trimmed from the brain slices. These blocks were cut into vibratome sections of 80–100-μm thickness by a vibratome (Model 3000™, The Vibratome™ Company, St Louis, MO, USA). The vibratome sections were then washed overnight in 0.1-M phosphate buffer, postfixed for 2 h in 1% osmium tetroxide, dehydrated, and embedded in Araldite mixture. Ultrathin sections, doubly stained with uranyl acetate and lead citrate, were observed under a Philips CM 120 electron microscope (FEI™ Company, Hillsboro, OR, USA). Four different areas of the CC or cerebral cortex from each of the brain were scrutinized and photographed at three different magnifications. Image J software (SummaSketch III Summagraphics, Seattle, WA) was used to measure the diameter of each axon magnified at 6800 times by a blind researcher.
In situ hybridization
We performed in situ hybridization on 10-μm-thick coronal frozen brain sections as previously described [32, 33]. Briefly, brain sections were incubated with proteinase K (S3004, Dako, Carpinteria, CA, USA) for 10 min and then rinsed in distilled water in 96% ethanol and in isopropanol for 5 min each. Following this, the brain sections were incubated with 125 μL of hybridization mixtures composed of 15 μL of distilled water, 25 μL of 20× saline-sodium citrate (SSC) buffer, 62.5 μL of 50% formamide, 12.5 μL of 50% dextran sulfate, 2.5 μL of Denhardt’s solution (D2532, Sigma-Aldrich, Saint Louis, MO,USA), 6.25 μL of herring sperm DNA (D7290, Sigma-Aldrich), and 1.25 μL of 3′-digoxigenin-conjugated probe (presented from prof fu hui’ lab). The probe, 5′-CAAGGGAAGGGAGGAAGAGACAG-3′, in final concentration of 100 ng/mL, detects a segment of the 5.8S ribosomal RNA of PLP. The sections were incubated at 95 °C for 6 min, immediately chilled in ice, and then incubated at 40 °C for 14–16 h in a humidified chamber. After this, the sections were washed in 2× SSC, 1× SSC, and 0.1× SSC buffer for 5 min each, followed by incubation with the anti-digoxigenin antibody conjugated to alkaline phosphatase, diluted in tris-buffered saline (TBS) (1:200, 11093274910, Roche Diagnostics, Indianapolis, IN, USA) for 1 h, and then washed in TBS. Visualization was achieved using NBT/BCIP (nitro blue tetrazolium/5-bromo-4-chloro-3-indolyl-phosphate) (11681451001, Roche Diagnostics) for 1 h in the dark. The reaction was stopped with TE buffer (pH 8.0) for 10 min and then washed in distilled water. The sections were counterstained with Mayer’s hematoxylin and mounted in aqueous medium (Faramount, S3025, Dako). PLP-positive cells were examined under a microscope (Olympus System Microscope Model BX53, Olympus Company Pte, Tokyo, Japan). Four nonoverlapping regions of the corpus callosum from each animal were photographed at two different magnifications. PLP-positive cells were enumerated under ×10 magnifications.
The present data were analyzed by the SPSS 20.0 statistical software (IBM, Armonk, New York, USA). The results were presented as mean ± SD. Statistical significance was examined by Student’s t test. The statistical significance of the results was considered at P < 0.05.
Neurofilament protein expression in the CC
Number of neurons in the cerebral cortex
To ascertain if there was a significant loss of neurons in the whole cortex in the frontal lobe after LPS treatment, the sections from the control and septic rats at 7, 14, and 28 days were incubated with antibodies against NeuN and Caspase-3, an apoptotic marker. After LPS treatment, the number of mature and apoptotic neurons at 7, 14, and 28 days in the cerebral cortex (Additional file 2: Figure S2D–F, J–L, and P–R) was comparable to that in the matching controls (Additional file 2: Figure S2A–C, G–I, and M–O). To confirm this, Nissl’s staining was carried out in sections from the control and septic rats at 7, 14, and 28 days. The results showed that the number of neurons was not significantly changed in the cerebral cortex in the frontal lobe at 7, 14, and 28 days after LPS injection (Additional file 3: Figure S3B, D, and F) in comparison with the corresponding control (Additional file 3: Figure S3A, C, and E).
PLP protein expression in the CC
Synaptophysin and PSD95 protein expression in the cortex
IL-1β and IL-1R1 protein expression in the CC
NFL, NFM, NFH, and synaptophysin immunoreactivity in primary cortical neuron cultures following administration with IL-1β
Effect of IL-1β on p38-MAPK signaling pathway in primary cortical neurons
IL-1β inhibited the protein expression of NFL, NFM, and synaptophysin through promoting the phosphorylation of p38-MAPK pathway in primary cortical neurons
To verify that IL-1β could suppress NFL, NFM, and synaptophysin expression in primary cortical neurons by activating the p38-MAPK pathways, NFL, NFM, and synaptophysin protein levels were detected by western blotting in primary cortical neurons treated with IL-1β + SB203580, a selective inhibitor of p38. Incubation of primary cortical neurons with SB203580 30 min prior to IL-1β administration for 24 h reversed the inhibition of NFL, NFM, and synaptophysin protein expression induced by IL-1β (Fig. 10c–f).
There are five principal subunit proteins of neuron-specific intermediate filaments (IFs) including the light, medium, and heavy molecular mass neurofilament (NF) triplet proteins (NFL, NFM, and NFH, respectively), α-internexin, and peripherin [34–36]. Mature filaments are made up of different combinations of these five subunits . NFL, NFM, and NFH are the main cytoskeletal elements in mature neurons, although NFH expression is usually delayed relative to NFL and NFM [38, 39]. Their main role is to increase the axonal caliber of myelinated axons and consequently axonal conduction velocity [38, 39]. NFM and NFL are required for axonal radial growth [40–42]. Loss of NFM and NFL results in small caliber axons [40–42]. The axons are wrapped by many processes of oligodendrocytes to form myelin sheath in the CNS . The axolemma is relatively uncovered at regularly spaced nodes of Ranvier . These structures allow rapid and efficient saltatory propagation of action potentials along Ranvier nodes, which enhance information transmission on axons . The formation of synapses occurs between axons and dendrites of different neurons in the CNS . Synaptic proteins including synaptophysin and post-synaptic density-95 (PSD-95) are involved in synaptic plasticity . Synaptophysin is localized in presynaptic vesicle membranes, which play important roles in docking, fusion, endocytosis, and membrane trafficking . PSD-95 is a post-synaptic protein, which is associated with regulating the number and size of dendritic spines and developing glutamatergic synapses . Changes in these synaptic proteins have been used to evaluate synaptic deficit . The present results have shown that NFL, NFM, and NFH protein expression level was significantly reduced in the CC at different times after LPS injection. By electron microscopy, a lesser number and smaller diameter of axons were observed. A striking structural feature or alteration in the LPS-injected rats was the occurrence of some “darkened neurons” which were not present in the cerebral cortex of control rats. Interestingly, some of the “darkened neurons” were seen in juxtaposition to normal looking neurons which have also been found in ischemic monkey brain [50, 51]. Therefore, it can be confidently argued that they were not fixation artifacts rather they represent degenerating neurons which have also been reported in traumatic brain injuries in the primates . In light of this, it is suggested that LPS injection had caused damage or structural alterations to some neurons in the developing cerebral cortex as well as their associated synapses. As a result, it is possible that the neuronal proteins including those that constitute the synapses in the cortex would be altered. The close association of activated microglia with the “darkened neuron” may not be fortuitous. LPS is known to stimulate microglia in the brain and their release of proinflammatory cytokines such as IL-1β that might have induced the neuronal changes affecting the soma, axons, dendrites, and synapses as demonstrated in this study. It is suggested that these are associated with decreased level of synaptophysin, Ranvier node structural damage, and reduction of mature oligodendrocytes. In other words, axon development in the CC was inhibited in the septic brain that ultimately would lead to presynaptic deficit, axonal hypomyelination, along with Ranvier node structural damage.
It is well documented that complex crosstalks occur between developing axons and oligodendrocyte progenitor cell processes during myelin sheath formation . Normal axon development is indispensable for oligodendrocyte proliferation, maturation, and subsequent myelination . The present results have shown that PLP protein level was drastically downregulated in the CC in septic brain coupled with reduction in the number of PLP-positive oligodendrocytes. It stands to reason therefore that inhibition of axon development can cause disorder of oligodendrocyte maturation and hypomyelination in the CC in LPS-injected rats.
We show here that microglia, especially those closely associated with the callosal axons in the CC, were activated and generated excess amounts of IL-1β after LPS injection. Microglia activation may be elicited by serum-derived proinflammatory mediators which have gained access to the brain tissue by passing through the disrupted blood-brain barrier . Interestingly, microglial activation was sustained up to nearly a week, suggesting that it is a persistent and intense inflammatory response in the CC in septic rats. Furthermore, at a late stage of the inflammatory response, the reactive astrocytes would be an additional cellular source for IL-1β . Besides IL-1β, activated microglia release other proinflammatory mediators in adverse conditions including TNF-α, inducible nitric oxide synthase (iNOS), and NO; all these have been reported to cause the loss of oligodendrocyte and myelination deficit through the corresponding signaling pathways [19, 28, 55]. The exhibition of IL-1R1 expression on NFL immunoreactive axons was augmented and remained to be so for about a week after LPS challenge. It has been documented that IL-1β exerts direct inhibitory effect on axonal growth of developing superior cervical ganglion sympathetic neurons via activating NF-κB signaling pathway . In agreement with this, IL-1β exerted an inhibitory effect on the outgrowth of axons from cultured dorsal root ganglion cells in vitro . In light of this, it was surmised that IL-1β derived from the microglia might suppress the development of axons via IL-1R1 in the PWM of septic neonatal rats. We show here reduction of NFM, NFL in the CC, and synaptophysin expression in the cerebral cortex in vivo. However, the number of neurons was not decreased significantly in the cortex, albeit the identification by electron microscopy of some “darkened neurons” indicative of neuronal degeneration or death. Additionally, IL-1β administration in vitro was found to decrease expression of NFM, NFL, and synaptophysin in primary neurons. Remarkably, IL-1 receptor antagonist neutralized the inhibitory effect of IL-1β on expression of NFM, NFL, and synaptophysin. Taken together, these results suggest that IL-1β treatment could inhibit axonal development and synapse formation in primary culture neurons. More importantly, the in vitro results corroborated with in vivo findings.
This study has shown reduction of axonal neurofilament protein expression coupled with disorder of axonal myelin sheath formation and synaptic deficit in the PWM and cerebral cortex, respectively, of septic developing brain. Concomitantly, AMCs associated with the axons were activated and produced a large amount of IL-1β. The possible crosstalk between AMCs and axons through IL-1β and its receptor 1 localized on axons would perturb the development of axons, which would contribute to disorder of myelin sheath formation and synaptic deficit. In vitro, IL-1β inhibited the expression of NFL, NFM, NFH, and synaptophysin in primary neurons via p38-MAPK signaling pathway. Therefore, inhibition of the biochemical and/or molecular processes mentioned above may represent one potential therapeutic strategy in mitigating PWMD induced by LPS in the developing brain.
Amoeboid microglial cells
Bovine serum albumin
Cell counting kit 8
Central nervous system
Dulbecco’s modified eagle medium
Enzyme-linked immunosorbent assay
Fetal bovine serum
- IL-1R1 :
IL-1 receptor antagonist
Nitro blue tetrazolium/5-bromo-4-chloro-3-indolyl-phosphate
Oligodendrocyte progenitor cell
p38-mitogen-activated protein kinase
Postsynaptic density 95
Periventricular white matter
Periventricular white matter damage
Reactive oxygen species
Tris-buffered saline Tween-20
- TNFR1 :
Tumor necrosis factor receptor
Tumor necrosis factor-α
This study was supported by National Natural Science Foundation of China (Grant No. 81271329 and 81471237), National Clinical Key Subject Construction Project (2012-649), Guangzhou Clinical Medicine Research and Translational Centre Construction Project (201508020005), and Natural Science Foundation of Guangdong Province; Grant number: 2015A030313538.
Availability of data and materials
The datasets during and/or analyzed during the current study are available from the corresponding author on reasonable request.
YD and HZ conceived and designed the experiments. QH performed the experiments. QH analyzed the data. YD contributed reagents/materials/analysis tools: YD. Wrote the paper: QH, YD. All authors read and approved the final manuscript.
The authors declare that they have no competing interests.
Consent for publication
All the authors have consent to publish the data in the manuscript.
All animals were handled according to the protocols of the Institutional Animal Care and Use Committee, Guangdong Province, China (Animal Certificate No.: SYXK2012-0081).
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