Roles and regulation of microglia activity in multiple sclerosis: insights from animal models.

As resident macrophages of the CNS, microglia are critical immune effectors of inflammatory lesions and associated neural dysfunctions. In multiple sclerosis (MS) and its animal models, chronic microglial inflammatory activity damages myelin and disrupts axonal and synaptic activity. In contrast to these detrimental effects, the potent phagocytic and tissue-remodelling capabilities of microglia support critical endogenous repair mechanisms. Although these opposing capabilities have long been appreciated, a precise understanding of their underlying molecular effectors is only beginning to emerge. Here, we review recent advances in our understanding of the roles of microglia in animal models of MS and demyelinating lesions and the mechanisms that underlie their damaging and repairing activities. We also discuss how the structured organization and regulation of the genome enables complex transcriptional heterogeneity within the microglial cell population at demyelinating lesions.

Cellular Proteomic Profiling Using Proximity Labeling by TurboID-NES in Microglial and Neuronal Cell Lines.

Different brain cell types play distinct roles in brain development and disease. Molecular characterization of cell-specific mechanisms using cell type-specific approaches at the protein (proteomic) level can provide biological and therapeutic insights. To overcome the barriers of conventional isolation-based methods for cell type-specific proteomics, in vivo proteomic labeling with proximity-dependent biotinylation of cytosolic proteins using biotin ligase TurboID, coupled with mass spectrometry (MS) of labeled proteins, emerged as a powerful strategy for cell type-specific proteomics in the native state of cells without the need for cellular isolation. To complement in vivo proximity labeling approaches, in vitro studies are needed to ensure that cellular proteomes using the TurboID approach are representative of the whole-cell proteome and capture cellular responses to stimuli without disruption of cellular processes. To address this, we generated murine neuroblastoma (N2A) and microglial (BV2) lines stably expressing cytosolic TurboID to biotinylate the cellular proteome for downstream purification and analysis using MS. TurboID-mediated biotinylation captured 59% of BV2 and 65% of N2A proteomes under homeostatic conditions. TurboID labeled endolysosome, translation, vesicle, and signaling proteins in BV2 microglia and synaptic, neuron projection, and microtubule proteins in N2A neurons. TurboID expression and biotinylation minimally impacted homeostatic cellular proteomes of BV2 and N2A cells and did not affect lipopolysaccharide-mediated cytokine production or resting cellular respiration in BV2 cells. MS analysis of the microglial biotin-labeled proteins captured the impact of lipopolysaccharide treatment (>500 differentially abundant proteins) including increased canonical proinflammatory proteins (Il1a, Irg1, and Oasl1) and decreased anti-inflammatory proteins (Arg1 and Mgl2).

Profiling the neuroimmune cascade in 3xTg-AD mice exposed to successive mild traumatic brain injuries.

Repetitive mild traumatic brain injuries (rmTBI) sustained within a window of vulnerability can result in long term cognitive deficits, depression, and eventual neurodegeneration associated with tau pathology, amyloid beta (Aß) plaques, gliosis, and neuronal and functional loss. However, a comprehensive study relating acute changes in immune signaling and glial reactivity to neuronal changes and pathological markers after single and repetitive mTBIs is currently lacking. In the current study, we addressed the question of how repeated injuries affect the brain neuroimmune response in the acute phase of injury (< 24 h) by exposing the 3xTg-AD mouse model of tau and Aß pathology to successive (1x-5x) once-daily weight drop closed-head injuries and quantifying immune markers, pathological markers, and transcriptional profiles at 30 min, 4 h, and 24 h after each injury. We used young adult 2-4 month old 3xTg-AD mice to model the effects of rmTBI in the absence of significant tau and Aß pathology. We identified pronounced sexual dimorphism in this model, with females eliciting more diverse changes after injury compared to males. Specifically, females showed: (1) a single injury caused a decrease in neuron-enriched genes inversely correlated with inflammatory protein expression and an increase in AD-related genes within 24 h, (2) each injury significantly increased a group of cortical cytokines (IL-1α, IL-1ß, IL-2, IL-9, IL-13, IL-17, KC) and MAPK phospho-proteins (phospho-Atf2, phospho-Mek1), several of which co-labeled with neurons and correlated with phospho-tau, and (3) repetitive injury caused increased expression of genes associated with astrocyte reactivity and macrophage-associated immune function. Collectively our data suggest that neurons respond to a single injury within 24 h, while other cell types, including astrocytes, transition to inflammatory phenotypes within days of repetitive injury.

Unique molecular characteristics and microglial origin of Kv1.3 channel-positive brain myeloid cells in Alzheimer's disease.

Kv1.3 potassium channels, expressed by proinflammatory central nervous system mononuclear phagocytes (CNS-MPs), are promising therapeutic targets for modulating neuroinflammation in Alzheimer's disease (AD). The molecular characteristics of Kv1.3-high CNS-MPs and their cellular origin from microglia or CNS-infiltrating monocytes are unclear. While Kv1.3 blockade reduces amyloid beta (Aß) burden in mouse models, the downstream immune effects on molecular profiles of CNS-MPs remain unknown. We show that functional Kv1.3 channels are selectively expressed by a subset of CD11b+CD45+ CNS-MPs acutely isolated from an Aß mouse model (5xFAD) as well as fresh postmortem human AD brain. Transcriptomic profiling of purified CD11b+Kv1.3+ CNS-MPs, CD11b+CD45int Kv1.3neg microglia, and peripheral monocytes from 5xFAD mice revealed that Kv1.3-high CNS-MPs highly express canonical microglial markers (Tmem119, P2ry12) and are distinct from peripheral Ly6chigh/Ly6clow monocytes. Unlike homeostatic microglia, Kv1.3-high CNS-MPs express relatively lower levels of homeostatic genes, higher levels of CD11c, and increased levels of glutamatergic transcripts, potentially representing phagocytic uptake of neuronal elements. Using irradiation bone marrow CD45.1/CD45.2 chimerism in 5xFAD mice, we show that Kv1.3+ CNS-MPs originate from microglia and not blood-derived monocytes. We show that Kv1.3 channels regulate membrane potential and early signaling events in microglia. Finally, in vivo blockade of Kv1.3 channels in 5xFAD mice by ShK-223 reduced Aß burden, increased CD11c+ CNS-MPs, and expression of phagocytic genes while suppressing proinflammatory genes (IL1b). Our results confirm the microglial origin and identify unique molecular features of Kv1.3-expressing CNS-MPs. In addition, we provide evidence for CNS immunomodulation by Kv1.3 blockers in AD mouse models resulting in a prophagocytic phenotype.

Osteoinductive effect of soluble transforming growth factor beta receptor 3 on human osteoblast lineage.

The development of bone requires carefully choregraphed signaling to bone progenitors to form bone. Our group recently described the requirement of transforming growth factor beta receptor 3 (TGFßR3), a receptor involved in TGFß pathway signaling, during osteoblast lineage commitment in mice. The TGFß pathway is known to play multiple osteo-inductive and osteo-inhibitory roles during osteoblast development and TGFßR3 human mutations are associated with reduced bone mineral density, making TGFßR3 a unique target for bone inductive therapy. In this article, we demonstrated increased mineralization of human pediatric bone-derived osteoblast-like cells (HBO) when treated with soluble TGFßR3 (sR3) using Alizarin Red staining. Osteogenic commitment of HBO cells was demonstrated by induction of osteogenic genes RUNX2, osteocalcin, osteopontin, and osterix. Evaluation of the canonical TGFß pathway signaling demonstrated that sR3 was able to induce bone formation in HBO cells, mainly through activation of noncanonical targets of TGFß pathway signaling including AKT, ERK, and p38 MAP kinases. Inhibition of these osteogenic noncanonical pathways in the HBO cells also inhibited mineralization, suggesting they are each required. Although no induction of SMAD1, 5, and 9 was observed, there was the activation of SMAD2 and 3 suggesting that sR3 is primarily signaling via the noncanonical pathways during osteogenic induction of the HBO. Our results highlight the important role of TGFßR3 in osteoblast induction of mineralization in human bone cells through noncanonical targets of TGFß signaling. Future studies will focus on the ability of sR3 to induce bone regeneration in vivo using animal models.

Frequency and duration of sensory flicker controls astrocyte and neuron specific transcriptional profiles in 5xFAD mice.

Background: Current clinical trials are investigating gamma frequency sensory stimulation as a potential therapeutic strategy for Alzheimer's disease, yet we lack a comprehensive picture of the effects of this stimulation on multiple aspects of brain function. While most prior research has focused on gamma frequency sensory stimulation, we previously showed that exposing mice to visual flickering stimulation increased MAPK and NFκB signaling in the visual cortex in a manner dependent on duration and frequency of sensory stimulation exposure. Because these pathways control multiple neuronal and glial functions and are differentially activated based on the duration and frequency of flicker stimulation, we aimed to define the transcriptional effects of different frequencies and durations of flicker stimulation on multiple brain functions. Methods: We exposed 5xFAD mice to different frequencies of audio/visual flicker stimulation (constant light, 10Hz, 20Hz, 40Hz) for durations of 0.5hr, 1hr, or 4hr, then used bulk RNAseq to profile transcriptional changes within the visual cortex and hippocampus tissues. Using weighted gene co-expression network analysis, we identified modules of co-expressed genes controlled by frequency and/or duration of stimulation. Results: Within the visual cortex, we found that all stimulation frequencies caused fast activation of a module of immune genes within 1hr and slower suppression of synaptic genes after 4hrs of stimulation. Interestingly, all frequencies of stimulation led to slow suppression of astrocyte specific gene sets, while activation of neuronal gene sets was frequency and duration specific. In contrast, in the hippocampus, immune and synaptic modules were suppressed based on the frequency of stimulation. Specifically,10Hz activated a module of genes associated with mitochondrial function, metabolism, and synaptic translation while 10Hz rapidly suppressed a module of genes linked to neurotransmitter activity. Conclusion: Collectively, our data indicate that the frequency and duration of flicker stimulation controls immune, neuronal, and metabolic genes in multiple regions of the brain affected by Alzheimer's disease. Flicker stimulation may thus represent a potential therapeutic strategy that can be tuned based on the brain region and the specific cellular process to be modulated.

Microglia morphological response to mesenchymal stromal cell extracellular vesicles demonstrates EV therapeutic potential for modulating neuroinflammation.

BACKGROUND: Mesenchymal stromal cell derived extracellular vesicles (MSC-EVs) are a promising therapeutic for neuroinflammation. MSC-EVs can interact with microglia, the resident immune cells of the brain, to exert their immunomodulatory effects. In response to inflammatory cues, such as cytokines, microglia undergo phenotypic changes indicative of their function e.g. morphology and secretion. However, these changes in response to MSC-EVs are not well understood. Additionally, no disease-relevant screening tools to assess MSC-EV bioactivity exist, which has further impeded clinical translation. Here, we developed a quantitative, high throughput morphological profiling approach to assess the response of microglia to neuroinflammation- relevant signals and whether this morphological response can be used to indicate the bioactivity of MSC-EVs. RESULTS: Using an immortalized human microglia cell-line, we observed increased size (perimeter, major axis length) and complexity (form factor) upon stimulation with interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α). Upon treatment with MSC-EVs, the overall morphological score (determined using principal component analysis) shifted towards the unstimulated morphology, indicating that MSC-EVs are bioactive and modulate microglia. The morphological effects of MSC-EVs in TNF-α /IFN-γ stimulated cells were concomitant with reduced secretion of 14 chemokines/cytokines (e.g. CXCL6, CXCL9) and increased secretion of 12 chemokines/cytokines (e.g. CXCL8, CXCL10). Proteomic analysis of cell lysates revealed significant increases in 192 proteins (e.g. HIBADH, MEAK7, LAMC1) and decreases in 257 proteins (e.g. PTEN, TOM1, MFF) with MSC-EV treatment. Of note, many of these proteins are involved in regulation of cell morphology and migration. Gene Set Variation Analysis revealed upregulation of pathways associated with immune response, such as regulation of cytokine production, immune cell infiltration (e.g. T cells, NK cells) and morphological changes (e.g. Semaphorin, RHO/Rac signaling). Additionally, changes in microglia mitochondrial morphology were measured suggesting that MSC-EV modulate mitochondrial metabolism. CONCLUSION: This study comprehensively demonstrates the effects of MSC-EVs on human microglial morphology, cytokine secretion, cellular proteome, and mitochondrial content. Our high-throughput, rapid, low-cost morphometric approach enables screening of MSC-EV batches and manufacturing conditions to enhance EV function and mitigate EV functional heterogeneity in a disease relevant manner. This approach is highly generalizable and can be further adapted and refined based on selection of the disease-relevant signal, target cell, and therapeutic product.

Microglia Morphological Response to Mesenchymal Stromal Cell Extracellular Vesicles Demonstrates EV Therapeutic Potential for Modulating Neuroinflammation.

Background: Mesenchymal stromal cell derived extracellular vesicles (MSC-EVs) are a promising therapeutic for neuroinflammation. MSC-EVs can interact with microglia, the resident immune cells of the brain, to exert their immunomodulatory effects. In response to inflammatory cues, such as cytokines, microglia undergo phenotypic changes indicative of their function e.g. morphology and secretion. However, these changes in response to MSC-EVs are not well understood. Additionally, no disease-relevant screening tools to assess MSC-EV bioactivity exist, which has further impeded clinical translation. Here, we developed a quantitative, high throughput morphological profiling approach to assess the response of microglia to neuroinflammation-relevant signals and whether this morphological response can be used to indicate the bioactivity of MSC-EVs. Results: Using an immortalized human microglia cell-line, we observed increased size (perimeter, major axis length) and complexity (form factor) upon stimulation with interferon-gamma (IFN-γ) and tumor necrosis factor-alpha (TNF-α). Upon treatment with MSC-EVs, the overall morphological score (determined using principal component analysis) shifted towards the unstimulated morphology, indicating that MSC-EVs are bioactive and modulate microglia. The morphological effects of MSC-EVs in TNF-γ/IFN-α stimulated cells were concomitant with reduced secretion of 14 chemokines/cytokines (e.g. CXCL6, CXCL9) and increased secretion of 12 chemokines/cytokines (e.g. CXCL8, CXCL10). Proteomic analysis of cell lysates revealed significant increases in 192 proteins (e.g. HIBADH, MEAK7, LAMC1) and decreases in 257 proteins (e.g. PTEN, TOM1, MFF) with MSC-EV treatment. Of note, many of these proteins are involved in regulation of cell morphology and migration. Gene Set Variation Analysis revealed upregulation of pathways associated with immune response, such as regulation of cytokine production, immune cell infiltration (e.g. T cells, NK cells) and morphological changes (e.g. Semaphorin, RHO/Rac signaling). Additionally, changes in microglia mitochondrial morphology were measured suggesting that MSC-EV modulate mitochondrial metabolism. Conclusion: This study comprehensively demonstrates the effects of MSC-EVs on human microglial morphology, cytokine secretion, cellular proteome, and mitochondrial content. Our high-throughput, rapid, low-cost morphological approach enables screening of MSC-EV batches and manufacturing conditions to enhance EV function and mitigate EV functional heterogeneity in a disease relevant manner. This approach is highly generalizable and can be further adapted and refined based on selection of the disease-relevant signal, target cell, and therapeutic product.

Pathogenic Th17 cells in autoimmunity with regard to rheumatoid arthritis.

Th17 cells contribute the pathobiology of autoimmune diseases, including rheumatoid arthritis (RA). However, it was shown that differentiated Th17 cells display a high degree of plasticity under the influence of inflammatory conditions. In some autoimmune diseases, the majority of Th17 cells, especially at sites of inflammation, have a phenotype that is intermediate between Th17 and Th1. These cells, which are described as Th17.1 or exTh17 cells, are hypothesized to be more pathogenic than classical Th17 cells. In this review, the involvement of Th17.1 lymphocytes in RA, and potential features that might render these cells to be more pathogenic are discussed.

Brain rhythms control microglial response and cytokine expression via NF-κB signaling.

Microglia transform in response to changes in sensory or neural activity, such as sensory deprivation. However, little is known about how specific frequencies of neural activity, or brain rhythms, affect microglia and cytokine signaling. Using visual noninvasive flickering sensory stimulation (flicker) to induce electrical neural activity at 40 hertz, within the gamma band, and 20 hertz, within the beta band, we found that these brain rhythms differentially affect microglial morphology and cytokine expression in healthy animals. Flicker induced expression of certain cytokines independently of microglia, including interleukin-10 and macrophage colony-stimulating factor. We hypothesized that nuclear factor κB (NF-κB) plays a causal role in frequency-specific cytokine and microglial responses because this pathway is activated by synaptic activity and regulates cytokines. After flicker, phospho-NF-κB colabeled with neurons more than microglia. Inhibition of NF-κB signaling down-regulated flicker-induced cytokine expression and attenuated flicker-induced changes in microglial morphology. These results reveal a mechanism through which brain rhythms affect brain function by altering microglial morphology and cytokines via NF-κB.

Profiling the neuroimmune cascade in 3xTg mice exposed to successive mild traumatic brain injuries.

Repetitive mild traumatic brain injuries (rmTBI) sustained within a window of vulnerability can result in long term cognitive deficits, depression, and eventual neurodegeneration associated with tau pathology, amyloid beta (Aß) plaques, gliosis, and neuronal and functional loss. However, we have limited understanding of how successive injuries acutely affect the brain to result in these devastating long-term consequences. In the current study, we addressed the question of how repeated injuries affect the brain in the acute phase of injury (<24hr) by exposing the 3xTg-AD mouse model of tau and Aß pathology to successive (1x, 3x, 5x) once-daily weight drop closed-head injuries and quantifying immune markers, pathological markers, and transcriptional profiles at 30min, 4hr, and 24hr after each injury. We used young adult mice (2-4 months old) to model the effects of rmTBI relevant to young adult athletes, and in the absence of significant tau and Aß pathology. Importantly, we identified pronounced sexual dimorphism, with females eliciting more differentially expressed proteins after injury compared to males. Specifically, females showed: 1) a single injury caused a decrease in neuron-enriched genes inversely correlated with inflammatory protein expression as well as an increase in AD-related genes within 24hr, 2) each injury significantly increased expression of a group of cortical cytokines (IL-1α, IL-1ß, IL-2, IL-9, IL-13, IL-17, KC) and MAPK phospho-proteins (phospho-Atf2, phospho-Mek1), several of which were co-labeled with neurons and correlated with phospho-tau, and 3) repetitive injury caused increased expression of genes associated with astrocyte reactivity and immune function. Collectively our data suggest that neurons respond to a single injury within 24h, while other cell types including astrocytes transition to inflammatory phenotypes within days of repetitive injury.

Systems Analysis of the Neuroinflammatory and Hemodynamic Response to Traumatic Brain Injury.

Mild traumatic brain injuries (mTBIs) are a significant public health problem. Repeated exposure to mTBI can lead to cumulative, long-lasting functional deficits. Numerous studies by our group and others have shown that mTBI stimulates cytokine expression and activates microglia, decreases cerebral blood flow and metabolism, and impairs cerebrovascular reactivity. Moreover, several works have reported an association between derangements in these neuroinflammatory and hemodynamic markers and cognitive impairments. Herein we detail methods to characterize the neuroinflammatory and hemodynamic tissue response to mTBI in mice. Specifically, we describe how to perform a weight-drop model of mTBI, how to longitudinally measure cerebral blood flow using a non-invasive optical technique called diffuse correlation spectroscopy, and how to perform a Luminex multiplexed immunoassay on brain tissue samples to quantify cytokines and immunomodulatory phospho-proteins (e.g., within the MAPK and NFκB pathways) that respond to and regulate activity of microglia and other neural immune cells. Finally, we detail how to integrate these data using a multivariate systems analysis approach to understand the relationships between all of these variables. Understanding the relationships between these physiologic and molecular variables will ultimately enable us to identify mechanisms responsible for mTBI.

BIN1 is a key regulator of proinflammatory and neurodegeneration-related activation in microglia.

BACKGROUND: The BIN1 locus contains the second-most significant genetic risk factor for late-onset Alzheimer's disease. BIN1 undergoes alternate splicing to generate tissue- and cell-type-specific BIN1 isoforms, which regulate membrane dynamics in a range of crucial cellular processes. Whilst the expression of BIN1 in the brain has been characterized in neurons and oligodendrocytes in detail, information regarding microglial BIN1 expression is mainly limited to large-scale transcriptomic and proteomic data. Notably, BIN1 protein expression and its functional roles in microglia, a cell type most relevant to Alzheimer's disease, have not been examined in depth. METHODS: Microglial BIN1 expression was analyzed by immunostaining mouse and human brain, as well as by immunoblot and RT-PCR assays of isolated microglia or human iPSC-derived microglial cells. Bin1 expression was ablated by siRNA knockdown in primary microglial cultures in vitro and Cre-lox mediated conditional deletion in adult mouse brain microglia in vivo. Regulation of neuroinflammatory microglial signatures by BIN1 in vitro and in vivo was characterized using NanoString gene panels and flow cytometry methods. The transcriptome data was explored by in silico pathway analysis and validated by complementary molecular approaches. RESULTS: Here, we characterized microglial BIN1 expression in vitro and in vivo and ascertained microglia expressed BIN1 isoforms. By silencing Bin1 expression in primary microglial cultures, we demonstrate that BIN1 regulates the activation of proinflammatory and disease-associated responses in microglia as measured by gene expression and cytokine production. Our transcriptomic profiling revealed key homeostatic and lipopolysaccharide (LPS)-induced inflammatory response pathways, as well as transcription factors PU.1 and IRF1 that are regulated by BIN1. Microglia-specific Bin1 conditional knockout in vivo revealed novel roles of BIN1 in regulating the expression of disease-associated genes while counteracting CX3CR1 signaling. The consensus from in vitro and in vivo findings showed that loss of Bin1 impaired the ability of microglia to mount type 1 interferon responses to proinflammatory challenge, particularly the upregulation of a critical type 1 immune response gene, Ifitm3. CONCLUSIONS: Our convergent findings provide novel insights into microglial BIN1 function and demonstrate an essential role of microglial BIN1 in regulating brain inflammatory response and microglial phenotypic changes. Moreover, for the first time, our study shows a regulatory relationship between Bin1 and Ifitm3, two Alzheimer's disease-related genes in microglia. The requirement for BIN1 to regulate Ifitm3 upregulation during inflammation has important implications for inflammatory responses during the pathogenesis and progression of many neurodegenerative diseases.

Cell type-specific biotin labeling in vivo resolves regional neuronal and astrocyte proteomic differences in mouse brain.

Proteomic profiling of brain cell types using isolation-based strategies pose limitations in resolving cellular phenotypes representative of their native state. We describe a mouse line for cell type-specific expression of biotin ligase TurboID, for in vivo biotinylation of proteins. Using adenoviral and transgenic approaches to label neurons, we show robust protein biotinylation in neuronal soma and axons throughout the brain, allowing quantitation of over 2000 neuron-derived proteins spanning synaptic proteins, transporters, ion channels and disease-relevant druggable targets. Next, we contrast Camk2a-neuron and Aldh1l1-astrocyte proteomes and identify brain region-specific proteomic differences within both cell types, some of which might potentially underlie the selective vulnerability to neurological diseases. Leveraging the cellular specificity of proteomic labeling, we apply an antibody-based approach to uncover differences in neuron and astrocyte-derived signaling phospho-proteins and cytokines. This approach will facilitate the characterization of cell-type specific proteomes in a diverse number of tissues under both physiological and pathological states.

Bifidobacteria: A probable missing puzzle piece in the pathogenesis of multiple sclerosis.

Multiple sclerosis (MS) is an autoimmune disorder in which the immunopathogenesis is not fully understood. In the recent years, the role of gut microbiome in the pathogenesis of this disorder has been highlighted. Bifidobacteria as a component of gut microbiome might also be involved in MS pathogenesis. Being emerged in early days after birth, bifidobacteria have a prominent role in immune system maturation and function. Some factors like mode of delivery, breast feeding, mother's blood group and her secretory state and also environmental factors could influence its level in the early infancy, which may remain throughout lifetime. In this review, we discussed possible immunopathogenic link between the bifidobacteria and MS.

Association of Increased Levels of lncRNA H19 in PBMCs with Risk of Coronary Artery Disease.

OBJECTIVE: Considerable research shows that long non-coding RNAs, those longer than 200 nucleotides, are involved in several human diseases such as various cancers and cardiovascular diseases. Their significant role in regulating the function of endothelial cells, smooth muscle cells, macrophages, vascular inflammation, and metabolism indicates the possible effects of lncRNAs on the progression of atherosclerosis which is the most common underlying pathological process responsible for coronary artery disease (CAD). The aim of present study was to assess whether the expression of the lnc RNA H19 was associated with a susceptibility to CAD by evaluating the expression level of H19 in the peripheral blood. MATERIALS AND METHODS: A case-control study of 50 CAD patients and 50 age and sex-matched healthy controls was undertaken to investigate whether the H19 lncRNA expression level is associated with a CAD using Taqman Real-Time polymerase chain reaction (PCR). RESULTS: The subsequent result indicated that the H19 lncRNA was over-expressed in CAD patients in comparison with the controls. However, it was not statistically significant. This overexpression may be involved in coronary artery disease progression. CONCLUSION: We report here, the up-regulation of H19 lncRNA in the whole blood of CAD patients and suggest a possible role for H19 in the atherosclerosis process and its consideration as novel biomarker for CAD.

Association between Long Noncoding RNA ANRIL Expression Variants and Susceptibility to Coronary Artery Disease.

Animal cells possess thousands of long non-coding (lnc) RNAs, such as antisense noncoding RNA in the INK4 locus (ANRIL), which have regulatory roles in the cells' molecular mechanisms, including X-chromosome inactivation, and developmental processes. These lnc RNAs are known to influence the extensive spectrum of age-related disorders. Accordingly, there is evidence for the role of these lnc RNAs in cardiovascular diseases, particularly coronary artery diseases (CAD). The aim of this study was to assess whether the expression of the lnc RNA ANRIL was associated with a susceptibility to CAD by evaluating the expression level of the two transcripts of ANRIL. Peripheral blood was taken from fifty patients affected by CAD and relative expression of ANRIL was determined by Real-Time PCR assay. The obtained data indicated that the EU741058 transcript expression level significantly decreased in CAD patients in comparison with the healthy individuals (P= 0.001). Furthermore, there was no significant association between the NR_003529 transcript expression, and CAD risk in Iranian patients (P=0.751). Our results suggest that the expression level of the EU741058 transcript of ANRIL may be implicated in CAD development, creating a predictive biomarker for CAD patients in future.