MicroRNAs: Key Players in Microglia and Astrocyte Mediated Inflammation in CNS Pathologies.

The significance of microglia and astrocytes in neural development, in maintaining synaptic connections and homeostasis in the healthy brain is well established. Microglia are dynamic immune cells of the brain that elicit an immune response during brain damage and also participate in tissue repair and regeneration, while astrocytes contribute to the local inflammatory response by producing proinflammatory cytokines and resolving neuronal damage through production of anti-inflammatory cytokines and neurotrophic factors. Recent efforts have focused on elucidating the epigenetic mechanisms which regulate glial cell behavior in normal and pathologic states. An important class of epigenetic regulators is microRNAs (miRNAs) which are small non-coding RNA molecules that regulate gene expression posttranscriptionally. Certain dysregulated miRNAs contribute to chronic microglial inflammation in the brain, thereby leading to progression of neurological diseases like Alzheimer's disease, traumatic injury, amyotrophic lateral sclerosis and stroke. Further, several miRNAs are differentially expressed in astrocytes after ischemia and spinal cord injury. Despite knowledge about miRNAs in neuroinflammation, little is known about effective delivery routes and pharmacokinetic data for miRNA based therapeutics. This review summarizes the current research on the role of miRNAs in promoting and inhibiting inflammatory response of microglia and astrocytes in a disease-specific manner. In addition, miRNA delivery as a therapeutic strategy to treat neuroinflammation is discussed.

microRNA-200b modulates microglia-mediated neuroinflammation via the cJun/MAPK pathway.

Chronic activation of microglia, the macrophages of the CNS, has been shown to enhance neuronal damage because of excessive release of proinflammatory cytokines and neurotoxic molecules in a number of neurodegenerative diseases. Recent reports showed altered microRNA (miRNA) expression in immune-mediated pathologies, thus suggesting that miRNAs modulate expression of genes involving immune responses. This study demonstrates that miRNA-200b is expressed in microglia and modulates inflammatory response of microglia by regulating mitogen-activated protein kinase pathway. miRNA-200b expression was found to be down-regulated in activated microglia in vivo (traumatic brain injury rat model) and in vitro. A luciferase assay and loss- and gain-of-function studies revealed c-Jun, the transcription factor of cJun-N terminal kinase (JNK) mitogen-activated protein kinase pathway to be the target of miR-200b. Knockdown of miR-200b in microglia increased JNK activity along with an increase in pro-inflammatory cytokines, inducible nitric oxide synthase expression and nitric oxide (NO) production. Conversely, over-expression of miRNA-200b in microglia resulted in a decrease in JNK activity, inducible nitric oxide synthase expression, NO production and migratory potential of activated microglia. Furthermore, miR-200b inhibition resulted in increased neuronal apoptosis after treatment of neuronal cells with conditioned medium obtained from microglial culture. Taken together, these results indicate that miRNA-200b modulates microglial inflammatory process including cytokine secretion, NO production, migration and neuronal survival.

Analysis of epigenetic factors in mouse embryonic neural stem cells exposed to hyperglycemia.

BACKGROUND: Maternal diabetes alters gene expression leading to neural tube defects (NTDs) in the developing brain. The mechanistic pathways that deregulate the gene expression remain unknown. It is hypothesized that exposure of neural stem cells (NSCs) to high glucose/hyperglycemia results in activation of epigenetic mechanisms which alter gene expression and cell fate during brain development. METHODS AND FINDINGS: NSCs were isolated from normal pregnancy and streptozotocin induced-diabetic pregnancy and cultured in physiological glucose. In order to examine hyperglycemia induced epigenetic changes in NSCs, chromatin reorganization, global histone status at lysine 9 residue of histone H3 (acetylation and trimethylation) and global DNA methylation were examined and found to be altered by hyperglycemia. In NSCs, hyperglycemia increased the expression of Dcx (Doublecortin) and Pafah1b1 (Platelet activating factor acetyl hydrolase, isoform 1b, subunit 1) proteins concomitant with decreased expression of four microRNAs (mmu-miR-200a, mmu-miR-200b, mmu-miR-466a-3p and mmu-miR-466 d-3p) predicted to target these genes. Knockdown of specific microRNAs in NSCs resulted in increased expression of Dcx and Pafah1b1 proteins confirming target prediction and altered NSC fate by increasing the expression of neuronal and glial lineage markers. CONCLUSION/INTERPRETATION: This study revealed that hyperglycemia alters the epigenetic mechanisms in NSCs, resulting in altered expression of some development control genes which may form the basis for the NTDs. Since epigenetic changes are reversible, they may be valuable therapeutic targets in order to improve fetal outcomes in diabetic pregnancy.

CREBBP re-arrangements affect protein function and lead to aberrant neuronal differentiation.

Biallelic inactivation of the CREB-binding protein (CREBBP) a transcriptional co-activator produces an embryonic lethal phenotype in mice. In humans, re-arrangements in CREBBP are associated with the Rubinstein-Taybi Syndrome (RSTS) that is characterised by craniofacial, skeletal and neuronal symptoms. Neuronal defects in RSTS can be attributed to genetic re-arrangements in CREBBP, which has been implicated in synaptic plasticity and long-term memory. The present study was designed to investigate the role of CREBBP re-arrangements during neuronal differentiation. Towards this, deletion constructs of pCREBBP, viz. pDeltaCB-HAT and pDeltaHAT-CT were generated and transfected into NT2 cells. Expression profiling of the components of Notch, Wnt, SHH and Retinoid signaling along with screening of the neuronal markers was carried out in the NT2 cells and their mutant derivatives. ChIP-PCRs along with co-immunoprecipitations were also performed in these cells to investigate defects due to inappropriate interaction of mutated CREEBP with the corresponding transcription factor and other transcription regulatory proteins both at steady state as well as during differentiation. Mutant NT2 cells lacking the CREB, BROMO and HAT domains (CB-HAT) were highly proliferative and showed limited differentiation; while mutant NT2 cells expressing CREBBP lacking the HAT and CTAD domains (HAT-CT) are proliferation deficient and differentiate rapidly albeit generating an insufficient number of neurons. Altered CREBBP structure resulted in changes in HAT activity, cell cycle profiles and expression of basal levels of components of Notch, SHH, Wnt and retinoid pathways known to be critical in the proliferation and differentiation of neuronal progenitors. At the chromatin level, aberrant signaling correlated with altered binding affinities of the (CREBBP-transcription factor) complexes to promoter regions of components of these pathways. Thus, differentiation defects are manifested early at the genomic level leading to aberrant transcription of the genes involved in differentiation along the neuronal lineage.