Temporal expression profiling of DAMPs-related genes revealed the biphasic post-ischemic inflammation in the experimental stroke model

Abstract

The neuroinflammation in the ischemic brain could occur as sterile inflammation in response to damage-associated molecular patterns (DAMPs). However, its long-term dynamic transcriptional changes remain poorly understood. It is also unknown whether this neuroinflammation contributes to the recovery or just deteriorates the outcome. The purpose of this study is to characterize the temporal transcriptional changes in the post-stroke brain focusing on DAMPs-related genes by RNA-sequencing during the period of 28 days. We conducted the RNA-sequencing on day 1, 3, 7, 14, 28 post-stroke in the mouse photothrombosis model. The gross morphological observation showed the ischemic lesion on the ipsilateral cortex turned into a scar with the clearance of cellular debris by day 28. The transcriptome analyses indicated that post-stroke period of 28 days was classified into four categories (I Baseline, II Acute, III Sub-acute-#1, IV Sub-acute-#2 phase). During this period, the well-known genes for DAMPs, receptors, downstream cascades, pro-inflammatory cytokines, and phagocytosis were transcriptionally increased. The gene ontology (GO) analysis of biological process indicated that differentially expressed genes (DEGs) are genetically programmed to achieve immune and inflammatory pathways. Interestingly, we found the biphasic induction of various genes, including DAMPs and pro-inflammatory factors, peaking at acute and sub-acute phases. At the subacute phase, we also observed the induction of genes for phagocytosis as well as regulatory and growth factors. Further, we found the activation of CREB (cAMP-response element binding protein), one of the key players for neuronal plasticity, in peri-ischemic neurons by immunohistochemistry at this phase. Taken together, these findings raise the possibility the recurrent inflammation occurs at the sub-acute phase in the post-stroke brain, which could be involved in the debris clearance as well as neural reorganization.

Sphingosine 1-phosphate, a Novel TREM2 Ligand, Promotes Phagocytosis and Reduce Ischemic Injury

“…Recent ndings reveal the importance of rapid clearance of cellular debris after ischemic stroke [31,32]. In the present study, we reveal for the rst time that S1P acts as a novel endogenous ligand of TREM2 to effectively promote microglial phagocytosis, and thereby play neuroprotective effects in stroke.…”

DAMPs and RAGE Pathophysiology at the Acute Phase of Brain Injury: An Overview

“…Necroptosis is a morphologically lytic form of cell death implicating the receptor-interacting protein kinase 1 and 3 (RIPK1-RIPK3) and the mixed-lineage kinase domain-like pseudokinase (MLKL) pathway, resulting in the release of the contents of the cell into the extracellular space [ 33 ] ( Figure 1 A). The subsequent release of DAMPs peaks around 24 h after the primary insult and decreases thereafter over several days [ 4 , 12 , 20 , 34 , 35 , 36 , 37 , 38 ]. There is a spillover of DAMPs into the core of the primary injury, with a drastic decrease of intracellular HMGB1 [ 37 ] while HMGB1 is translocated from the nucleus to the cytoplasm of neurons but not glial cells at the periphery [ 4 , 9 , 37 , 39 ].…”

“…After several days, HMGB1-positive cells are mainly phagocytic microglial cells [ 4 ] ( Figure 1 D). Some authors have also described a biphasic expression of HMGB1 and S100 proteins with a second peak 14 days later [ 34 , 35 ]. The cellular consequences of DAMPs depend on the expression of PRRs but also on their post-translational modification.…”

“…The expression of RAGE on neurons and microglial cells follows the same time course, with a biphasic pattern peaking on day 1 and 14 [ 12 , 20 , 34 , 35 , 36 , 44 , 45 ]. The amount of TLRs, which are predominantly expressed on microglial cells, increases at the subacute phase (i.e., after day 1) when activated microglia are present [ 34 ] ( Figure 1 D). The ligation of DAMPs to RAGE in turn activates the nuclear factor-kappa B (NF-κB) which in a positive feedback loop will increase the expression of RAGE.…”

“…In neurons RAGE promotes the expression of proteins involved in necroptosis (i.e., MLKL) and autophagy (i.e., Becline-1); accordingly, blocking RAGE reduces autophagy, but also increases neuronal sensitivity to injury and apoptosis [ 20 ]. Later, at the subacute phase, TLRs and RAGE activation will polarize microglial cells and infiltrating leukocytes toward a pro-inflammatory phenotype, termed M1, with the release of cytokines such as tumor necrosis factor (TNF), IL-6 or IL-1 and the up-regulation of the inducible NO synthase [ 5 , 11 , 21 , 34 , 37 , 39 ]. Experimental overactivation of RAGE or TLRs by DAMPs injection or an increase in glucose degradation products, such as during hyperglycemia, will in turn increase the cytokine levels and worsen neuronal injuries [ 5 , 11 , 37 ].…”