Characterization of m6A RNA methylation mediated immune heterogeneity and functional validation in hepatocellular carcinoma.

BACKGROUND: N6 -methyladenosine (m6 A) mediates RNA modification in various biological processes. It plays a key role in hepatocellular carcinoma (HCC) through regulating methyltransferase. The present study aims to analyze the correlation between the m6 A and the immune status of HCC, and to construct an m6 A-related prognostic signature for HCC. METHODS: HCC subtypes with different m6 A modification activities were identified based on the m6 A-related genes. Lasso Cox regression was applied to construct an m6 A-related prognostic model for HCC. Then, the prognostic potential of the constructed signature was evaluated and validated in the external validation dataset. Small interfering RNAs were designed to knockdown FBXO5. CCK-8 assay, Edu staining, wound healing assay, and Transwell cell invasion assay were used to detect cell proliferation, migration, and invasion ability. RESULTS: Two m6 A-related HCC subtypes were identified. The m6 A modification active group showed an immune suppressive microenvironment compared to the m6 A modification inactive group. The differentially expressed genes (DEGs) between the HCC subtypes were screened. Enrichment analysis was performed using the DEGs. Subsequently, an m6 A-related prognostic model was established. The prognostic model performed well in both training and validation datasets. Moreover, knockdown of FBXO5, one of the genes in the prognostic model, inhibited the proliferation, migration, and invasion of HepG2 cells. CONCLUSIONS: The heterogeneity of m6 A RNA methylation is associated with immune status in HCC. The constructed m6 A-related gene-based signature can predict the prognosis of HCC patients. The genes in the prognostic model also have therapeutic potential for HCC.

MicroRNA-182 exacerbates blood-brain barrier (BBB) disruption by downregulating the mTOR/FOXO1 pathway in cerebral ischemia.

Cerebral ischemia causes damage to the structure and function of the blood-brain barrier (BBB) and alleviating BBB destruction will be of great significance for the treatment and prognosis of ischemic stroke. Recently, microRNAs have been shown to play a critical role in BBB integrity. However, the potential mechanism by which microRNA-182 (miR-182) affects the BBB in ischemic stroke remains unclear. We demonstrated for the first time that cerebral ischemia leads to a significant progressive increase in miR-182 after pMCAO, and bEnd.3 cells are the primary target cells of miR-182. In miR-182 KD transgenic mice, infarct volume, and BBB permeability were attenuated, and tight junction (TJ) proteins increased. Inhibition of miR-182 with an antagomir reduced OGD-induced apoptosis of bEnd.3 cells and the loss of ZO-1 and Occludin. To further explore the mechanism by which miR-182 regulates BBB integrity, we detected the apoptotic proteins Bcl-2/Bax and demonstrated that mTOR and FOXO1 were the targets of miR-182. Inhibition of mTOR/FOXO1 by rapamycin/AS1842856 decreased the ratio of Bcl-2/Bax and exacerbated TJ protein loss. Taken together, inhibition of miR-182 protects BBB integrity by reducing endothelial cell apoptosis through the mTOR/FOXO1 pathway. Thus, miR-182 may be a potential target for the treatment of BBB disruption during cerebral ischemia.

MicroRNA 182 promotes T helper 1 cell by repressing hypoxia induced factor 1 alpha in experimental autoimmune encephalomyelitis.

MicroRNA 182 is important for the clonal expansion of CD4+ T cells (Th) following IL-2 stimulation and is a potential therapeutic target for autoimmune diseases. In the present study, we investigated the role of microRNA 182 in the differentiation of pro-inflammatory CD4+ T helper cell by overexpressing or silencing microRNA 182 expression both in in vivo and in vitro settings. We report that in the studied Chinese cohort, microRNA 182 is upregulated in patients with relapse and remitting multiple sclerosis (RRMS) and this upregulation is associated with increased IFN-γ producing CD4+ Th1 cells in the circulation. In the murine experimental autoimmune encephalomyelitis (EAE) model, global microRNA 182 overexpression exacerbates clinical symptoms and results in augmented CD4+ IFN-γ+ Th1 and CD4+ IL-17+ Th17 differentiation in vivo. Addition of microRNA 182 mimics in vitro represses both the protein expression and transcriptional activity of hypoxia induced factor 1α (HIF-1α) but increases the level of IFN-γ transcripts in sorted murine CD4+ T cells. Together, our results provide evidence that microRNA 182 may be one of the transitional hubs contribution to regulate Th cells expansion in response to self-antigens and differentiation of antigen specific Th cells during the progression of autoimmune inflammations.

Hypobaric hypoxia regulates iron metabolism in rats.

Intermittent hypobaric hypoxia can produce a protective effect on both the nervous system and non-nervous system tissues. Intermittent hypobaric hypoxia preconditioning has been shown to protect rats from cardiac ischemia-reperfusion injury by decreasing cardiac iron levels and reactive oxygen species (ROS) production, thereby decreasing oxidative stress to achieve protection. However, the specific mechanism underlying the protective effect of hypobaric hypoxia is unclear. To shed light on this phenomenon, we subjected Sprague-Dawley rats to hypobaric hypoxic preconditioning (8 hours/day). The treatment was continued for 4 weeks. We then measured the iron content in the heart, liver, spleen, and kidney. The iron levels in all of the assessed tissues decreased significantly after hypobaric hypoxia treatment, corroborating previous results that hypobaric hypoxia may produce its protective effect by decreasing ROS production by limiting the levels of catalytic iron in the tissue. We next assessed the expression levels of several proteins involved in iron metabolism (transferrin receptor, L-ferritin, and ferroportin1 [FPN1]). The increased transferrin receptor and decreased L-ferritin levels after hypobaric hypoxia were indicative of a low-iron phenotype, while FPN1 levels remained unchanged. We also examined hepcidin, transmembrane serine proteases 6 (TMPRSS6), erythroferrone (ERFE), and erythropoietin (EPO) levels, all of which play a role in the regulation of systemic iron metabolism. The expression of hepcidin decreased significantly after hypobaric hypoxia treatment, whereas the expression of TMPRSS6 and ERFE and EPO increased sharply. Finally, we measured serum iron and total iron binding capacity in the serum, as well as red blood cell count, mean corpuscular volume, hematocrit, red blood cell distribution width SD, and red blood cell distribution width CV. As expected, all of these values increased after the hypobaric hypoxia treatment. Taken together, our results show that hypobaric hypoxia can stimulate erythropoiesis, which systemically draws iron away from nonhematopoietic tissue through decreased hepcidin levels.

Functional network analysis reveals biological roles of lncRNAs and mRNAs in MOG35-55 specific CD4+T helper cells.

Long non-coding RNAs have the potential to regulate immune responses. Their impact on multiple sclerosis has remained elusive. For illustrating their roles in experimental autoimmune encephalomyelitis (EAE) pathogenesis, we investigated the differential expression of lncRNAs and mRNAs in CD4+Th cells obtained from myelin oligodendrocytic glycoprotein35-55(MOG35-55)-induced EAE and complete Freund's adjuvant (CFA) controls. We observed differential expression of 1112 lncRNAs and 519 mRNAs in CD4+Th cells. The functional network showed lncRNAs had the capacity to modulate EAE pathogenesis via regulating many known EAE regulators such as Ptpn6. Predicting the function of lncRNAs demonstrated that dysregulated lncRNAs were closely associated with the development of EAE. These dysregulated lncRNAs may have function in EAE and they could be novel biomarkers and therapeutic targets of EAE. However, the precise mechanisms and biological functions of these specific lncRNAs in EAE pathogenesis require further study.

Regulator of G protein signaling 5 (RGS5) inhibits sonic hedgehog function in mouse cortical neurons.

Regulator of G protein signaling 5 (RGS5) acts as a GTPase-activating protein (GAP) for the Gαi subunit and negatively regulates G protein-coupled receptor signaling. However, its presence and function in postmitotic differentiated primary neurons remains largely uncharacterized. During neural development, sonic hedgehog (Shh) signaling is involved in cell signaling pathways via Gαi activity. In particular, Shh signaling is essential for embryonic neural tube patterning, which has been implicated in neuronal polarization involving neurite outgrowth. Here, we examined whether RGS5 regulates Shh signaling in neurons. RGS5 transcripts were found to be expressed in cortical neurons and their expression gradually declined in a time-dependent manner in culture system. When an adenovirus expressing RGS5 was introduced into an in vitro cell culture model of cortical neurons, RGS5 overexpression significantly reduced neurite outgrowth and FM4-64 uptake, while cAMP-PKA signaling was also affected. These findings suggest that RGS5 inhibits Shh function during neurite outgrowth and the presynaptic terminals of primary cortical neurons mature via modulation of cAMP.

MicroRNA 182 inhibits CD4+CD25+Foxp3+ Treg differentiation in experimental autoimmune encephalomyelitis.

MicroRNA 182 has been found to have a distinct contribution in the clonal expansion of activated- and functioning of specialized-helper T cells. In this study we knocked down microRNA 182 in vivo and induced experimental autoimmune encephalomyelitis (EAE) to determine the influences of microRNA 182 in the Treg cells functional specialization through Foxo1 dependent pathway in the peripheral lymphoid organs. Down-regulation of microRNA 182 significantly increased the proportions of Foxp3+ T cells in the peripheral lymph nodes and spleen. In vivo study verified a positive correlation between microRNA 182 levels and symptom severity of EAE, and a negative correlation between microRNA 182 and the transcriptional factor Foxp3. In vitro polarization study also confirmed the contribution of Foxo1 in microRNA 182 mediated down-regulation of Foxp3+ T cells. Together, our results provide evidence that during the development of EAE, microRNA 182 repressed Treg cells differentiation through the Foxo1 dependent pathway.

Bone Marrow Stromal Cells Inhibit the Activation of Liver Cirrhotic Fat-Storing Cells via Adrenomedullin Secretion.

BACKGROUND: Cirrhosis, or liver fibrosis, which is mainly triggered by cirrhosis fat-storing cells (CFSCs) activation, has traditionally been considered an irreversible disease. However, recent observations indicate that even advanced fibrosis is still reversible by removing the causative agents. Anti-fibrotic effects of bone marrow-derived stromal cells (BMSCs) have been demonstrated by inhibiting CFSCs via cytokines secretion; however, the mechanisms are still unclear. AIMS: The purpose of this study was to explore the underlying mechanisms by which BMSCs modulate the function of activated CFSCs. METHODS: After the co-culture of CFSCs with BMSCs supernatants with or without the addition of recombinant rat adrenomedullin (AM)/AM-specific siRNA, western blot analysis was mainly used to detect the differences of relative protein expression on CFSCs. RESULTS: BMSC-secreted adrenomedullin (AM) effectively inhibited the proliferation and activation of CFSCs by suppressing the expression of Ang II and its binding receptor, AT1, which resulted in a reduction of p47-phox formation. CONCLUSIONS: Our data suggested that BMSCs inhibited CFSC activation in vitro via the AM-Ang II-p47-phox signaling pathway, and since CFSC activation is an essential part of hepatic fibrosis process, this inhibition by BMSCs implies us new insights into the potential treatment of hepatic fibrosis via BMSCs.

Neutralization of interleukin-9 ameliorates symptoms of experimental autoimmune myasthenia gravis in rats by decreasing effector T cells and altering humoral responses.

Interleukin-9 (IL-9) was initially thought to be a type 2 T helper (Th2)-associated cytokine involved in the regulation of autoimmune responses by affecting multiple cell types. However, it was recently shown that IL-9-producing CD4+ T cells represent a discrete subset of Th cells, designated Th9 cells. Although Th9 cells have been shown to be important in many diseases, their roles in myasthenia gravis (MG) are unclear. The aim of this study was to determine whether IL-9 and Th9 cells promote the progression of experimental autoimmune myasthenia gravis (EAMG). The results showed that the percentage of Th9 cells changed during the progression of EAMG, accompanied by an up-regulation of IL-9. Blocking IL-9 activity with antibodies against IL-9 inhibited EAMG-associated pathology in rats and reduced serum anti-acetylcholine receptor IgG levels. Neutralization of IL-9 altered the Th subset distribution in EAMG, reducing the number of Th1 cells and increasing the number of regulatory T cells. Administration of an anti-IL-9 antibody may represent an effective therapeutic strategy for MG-associated pathologies or other T-cell- or B-cell-mediated autoimmune diseases.

t-PA acts as a cytokine to regulate lymphocyte-endothelium adhesion in experimental autoimmune encephalomyelitis.

In this study, the capacity for t-PA to affect T cell-brain microvascular endothelial cell adhesion by acting as a cytokine was investigated. Following the treatment of a brain-derived endothelial cell line, bEnd.3, with various concentrations of t-PA, adhesion and transwell migration assays were performed. In the presence of t-PA, enhanced adhesion of T cells to bEnd.3 cells was observed. Using western blot analysis, an increase in ICAM-1 expression was detected for both t-PA-treated bEnd.3 cells and bEnd.3 cells treated with a non-enzymatic form of t-PA. In contrast, when LRP1 was blocked using a specific antibody, upregulation of ICAM-1 was inhibited and cAMP-PKA signaling was affected. Furthermore, using an EAE mouse model, administration of t-PA was associated with an increase in ICAM-1 expression by brain endothelial cells. Taken together, these findings suggest that t-PA can induce ICAM-1 expression in brain microvascular endothelial cells, and this may promote the development of EAE.

Accumulation of natural killer cells in ischemic brain tissues and the chemotactic effect of IP-10.

BACKGROUND: Stroke is accompanied by a distinguished inflammatory reaction that is initiated by the infiltration of immunocytes, expression of cytokines, and other inflammatory mediators. As natural killer cells (NK cells) are a type of cytotoxic lymphocyte critical to the innate immune system, we investigated the mechanism of NK cells-induced brain injuries after cerebral ischemia and the chemotactic effect of IP-10 simultaneously. METHODS: NK cells infiltration, interferon-gamma (IFN-γ) and IP-10 expression were detected by immunohistochemistry, immunofluorescence, PCR and flow cytometry in human and C57/BL6 wild type mouse ischemic brain tissues. The ischemia area was detected via 2,3,5-triphenyltetrazolium chloride staining. CXCR3 mean fluorescence intensity of isolated NK cells was measured by flow cytometry. The neuronal injury made by NK cells was examined via apoptosis experiment. The chemotactic of IP-10 was detected by migration and permeability assays. RESULTS: In human ischemic brain tissue, infiltrations of NK cells were observed and reached a peak at 2 to 5 days. In a permanent middle cerebral artery occlusion (pMCAO) model, infiltration of NK cells into the ischemic infarct region reached their highest levels 12 hours after ischemia. IFN-γ-positive NK cells and levels of the chemokine IP-10 were also detected within the ischemic region, from 6 hours up to 4 days after pMCAO was performed, and IFN-γ levels decreased after NK cells depletion in vivo. Co-culture experiments of neural cells with NK cells also showed that neural necrosis was induced via IFN-γ. In parallel experiments with IP-10, the presence of CXCR3 indicates that NK cells were affected by IP-10 via CXCR3, and the effect was dose-dependent. After IP-10 depletion in vivo, NK cells decreased. In migration assays and permeability experiments, disintegration of the blood-brain barrier (BBB) was observed following the addition of NK cells. Moreover, in the presence of IP-10 this injury was aggravated. CONCLUSIONS: All findings support the hypothesis that NK cells participate in cerebral ischemia and promote neural cells necrosis via IFN-γ. Moreover, IP-10 intensifies injury to the BBB by NK cells via CXCR3.

Adenosine receptor expression in a rat model of experimental autoimmune myasthenia gravis.

Extracellular adenosine is an essential negative regulator of immune reactions that acts by signaling via 4 distinct adenosine receptors. We evaluated adenosine receptor expression in Lewis rats presenting with experimental autoimmune myasthenia gravis (EAMG) to determine whether the expression of adenosine receptors are changed in the development and progression of EAMG. Lymphocyte A1AR and A2AAR mRNA and protein levels from lymphocytes harvested from the lymph nodes, spleen, and peripheral blood mononuclear cells (PBMCs) of EAMG rats were decreased. A modest but not significant increase in A2BAR levels was observed in EAMG lymphocytes harvested from lymph nodes and PBMCs. No changes in A3AR expression were observed in lymphocytes harvested from lymph nodes, spleen, or PBMCs following EAMG induction. Results presented in this report showed that the expression levels and the distribution pattern of adenosine receptors were altered in EAMG lymphocytes.

DHA plays a protective role in cerebral ischemia-reperfusion injury by affecting macrophage/microglia type polarization.

A close correlation exists between the macrophage/microglia(MΦ/MG) polarization states and the development of cerebral ischemia and reperfusion (I/R). Therefore it is of great significance to research on how to modulate the MΦ/MG states for improved patient outcomes. In particular, regulatory mechanisms involved in this process remain to be identified. Hereby, we aim to shed light on how docosahexaenoic acid (DHA) actively modulates the switch between M1 and M2 macrophage states by restraining the NACHT-LRR-PYD-containing protein three inflammasome (NALP3). We found that NALP3-positive cells were detected in clinical human cerebral infarction tissue samples and the mouse tMCAO model. In mice after DHA treatment, the number of NALP3-positive cells was significantly reduced, significantly decreasing infarct volume and improving the postoperative physical status of mice. NALP3-positive cells were found to be MΦ/MG after co-staining with CD11b. By extracting peritoneal macrophages, it was verified that DHA inhibited the activation of NALP3 and regulated the transformation of M1 and M2 cells, thereby reducing I/R injury.

Astrocyte-secreted C3 signaling impairs neuronal development and cognition in autoimmune diseases.

Neuromyelitis optica (NMO) arises from primary astrocytopathy induced by autoantibodies targeting the astroglial protein aquaporin 4 (AQP4), leading to severe neurological sequelae such as vision loss, motor deficits, and cognitive decline. Mounting evidence has shown that dysregulated activation of complement components contributes to NMO pathogenesis. Complement C3 deficiency has been shown to protect against hippocampal neurodegeneration and cognitive decline in neurodegenerative disorders (e.g., Alzheimer's disease, AD) and autoimmune diseases (e.g., multiple sclerosis, MS). However, whether inhibiting the C3 signaling can ameliorate cognitive dysfunctions in NMO remains unclear. In this study, we found that the levels of C3a, a split product of C3, significantly correlate with cognitive impairment in our patient cohort. In response to the stimulation of AQP4 autoantibodies, astrocytes were activated to secrete complement C3, which inhibited the development of cultured neuronal dendritic arborization. NMO mouse models exhibited reduced adult hippocampal newborn neuronal dendritic and spine development, as well as impaired learning and memory functions, which could be rescued by decreasing C3 levels in astrocytes. Mechanistically, we found that C3a engaged with C3aR to impair neuronal development by dampening ß-catenin signalling. Additionally, inhibition of the C3-C3aR-GSK3ß/ß-catenin cascade restored neuronal development and ameliorated cognitive impairments. Collectively, our results suggest a pivotal role of the activation of the C3-C3aR network in neuronal development and cognition through mediating astrocyte and adult-born neuron communication, which represents a potential therapeutic target for autoimmune-related cognitive impairment diseases.

All-trans-retinoic acid ameliorates experimental allergic encephalomyelitis by affecting dendritic cell and monocyte development.

Experimental allergic encephalomyelitis (EAE) can be induced in animal models by injecting the MOG35-55 peptide subcutaneously. Dendritic cells (DCs) that are located at the immunization site phagocytose the MOG35-55 peptide. These DCs mature and migrate into the nearest draining lymph nodes (dLNs), then present antigen, resulting in the activation of naive T cells. T helper type 1 (Th1) and Th17 cells are the primary cells involved in EAE progression. All-trans-retinoic acid (AT-RA) has been shown to have beneficial effects on EAE progression; however, whether AT-RA influences DC maturation or mediates other functions is unclear. In the present study, we showed that AT-RA led to the down-regulation of MHC class II, CD80 (B7-1) and CD86 (B7-2) expressed on the surface of DCs that were isolated from dLNs or spleen 3 days post-immunization in an EAE model. Changes to DC function influenced Th1/Th17 subset polarization. Furthermore, the number of CD44(+) monocytes (which might trigger EAE progression) was also significantly decreased in dLNs, spleen, subarachnoid space and the spinal cord parenchyma after AT-RA treatment. These findings are the first to demonstrate that AT-RA impairs the antigen-presenting capacity of DCs, leading to down-regulation of pathogenic Th1 and Th17 inflammatory cell responses and reducing EAE severity.

ATRA alters humoral responses associated with amelioration of EAMG symptoms by balancing Tfh/Tfr helper cell profiles.

All-trans retinoic acid (ATRA) is a vitamin A metabolite with diverse immunomodulatory actions used therapeutically in the treatment of some autoimmune diseases. However, the effects that ATRA may have on diminishing myasthenia gravis (MG) symptoms remain undefined. This study investigated the effect of ATRA on experimental autoimmune myasthenia gravis (EAMG) in vivo and in vitro. Data presented in this study demonstrated that intraperitoneal injection of ATRA ameliorated EAMG pathology in rats associated with reduced total anti-acetylcholine receptor (AChR) serum IgG levels. We observed that EAMG development was accompanied by an increase in follicular helper T cells (Tfh, defined as CD4(+)CXCR5(+)ICOS(high)) and a decrease of follicular regulatory T cells (Tfr, defined as CD4(+)Foxp3(+)CXCR5(+)ICOS(median)) and that the Tfh:Tfr ratio was altered following ATRA administration. In addition, ATRA treatment restored the Th1/Th2/Th17/Treg balance. In vitro, ATRA inhibited AChR-specific cell proliferation and eliciting apoptosis in these cells without affecting the cell cycle. ATRA also altered the Th distribution in animals presenting with EAMG resulting in a reduction in Th1/Th17/Tfh cells and increasing the number of Th2/Treg/Tfr cell types. These results suggested that ATRA reduced EAMG severity by regulating Th cell profiles thereby providing new insights into the development of novel MG (or related) therapies.

Activation of the adenosine A2A receptor attenuates experimental autoimmune myasthenia gravis severity.

The adenosine A2A receptor (A2AR) is the major cellular adenosine receptor commonly associated with immunosuppression. Here, we investigated whether A2AR activation holds the potential for impacting the severity of experimental autoimmune myasthenia gravis (EAMG) induced following immunization of Lewis rats with the acetylcholine receptor (AChR) R97-116 peptide. This report demonstrates reduced A2AR expression by both T cells and B cells residing in spleen and lymph nodes following EAMG induction. A2AR stimulation inhibited anti-AChR antibody production and proliferation of AChR-specific lymphocytes in vitro. Inhibition was blocked with the A2AR antagonists or protein kinase A inhibitor. We also determined that the development of EAMG was accompanied by a T-helper cell imbalance that could be restored following A2AR stimulation that resulted in increased Treg cell levels and a reduction in Th1-, Th2-, and Th17-cell subtypes. An EAMG-preventive treatment regimen was established that consisted of (2-(p-(2-carbonylethyl)phenylethylamino)-5-N-ethylcarboxamidoadenosine) (CGS21680; A2AR agonist) administration 1 day prior to EAMG induction. Administration of CGS21680 29 days post EAMG induction (therapeutic treatment) also ameliorated disease severity. We conclude that A2AR agonists may represent a new class of compounds that can be developed for use in the treatment of myasthenia gravis or other T-cell- and B-cell-mediated autoimmune diseases.

The Aldose Reductase Inhibitor Epalrestat Maintains Blood-Brain Barrier Integrity by Enhancing Endothelial Cell Function during Cerebral Ischemia.

Excessive activation of aldose reductase (AR) in the brain is a risk factor for aggravating cerebral ischemia injury. Epalrestat is the only AR inhibitor with proven safety and efficacy, which is used in the clinical treatment of diabetic neuropathy. However, the molecular mechanisms underlying the neuroprotection of epalrestat remain unknown in the ischemic brain. Recent studies have found that blood-brain barrier (BBB) damage was mainly caused by increased apoptosis and autophagy of brain microvascular endothelial cells (BMVECs) and decreased expression of tight junction proteins. Thus, we hypothesized that the protective effect of epalrestat is mainly related to regulating the survival of BMVECs and tight junction protein levels after cerebral ischemia. To test this hypothesis, a mouse model of cerebral ischemia was established by permanent middle cerebral artery ligation (pMCAL), and the mice were treated with epalrestat or saline as a control. Epalrestat reduced the ischemic volume, enhanced BBB function, and improved the neurobehavior after cerebral ischemia. In vitro studies revealed that epalrestat increased the expression of tight junction proteins, and reduced the levels of cleaved-caspase3 and LC3 proteins in mouse BMVECs (bEnd.3 cells) exposed to oxygen-glucose deprivation (OGD). In addition, bicalutamide (an AKT inhibitor) and rapamycin (an mTOR inhibitor) increased the epalrestat-induced reduction in apoptosis and autophagy related protein levels in bEnd.3 cells with OGD treatment. Our findings suggest that epalrestat improves BBB function, which may be accomplished by reducing AR activation, promoting tight junction proteins expression, and upregulating AKT/mTOR signaling pathway to inhibit apoptosis and autophagy in BMVECs.

miR-3934 regulates the apoptosis and secretion of inflammatory cytokines of basophils via targeting RAGE in asthma.

BACKGROUND: Several miRNAs are now known to have clear connections to the pathogenesis of asthma. The present study focused on the potential role of miR-3934 during asthma development. METHODS: miR-3934 was detected as a down-regulated miRNA in basophils by sequencing analysis. Next, the expression levels of miR-3934 in peripheral blood mononuclear cells of 50 asthma patients and 50 healthy volunteers were examined by RT-qPCR methods. The basophils were then treated with AGEs and transfected with miR-3934 mimics. The apoptosis levels were examined by flow cytometry assay; and the expression levels of cytokines were detected using the ELISA kits. Finally, the Western blot was performed to examined the expression of key molecules in the TGF-ß/Smad signaling pathway. RESULTS: miR-3934 was down-regulated in the basophils of asthmatic patients. The expression of the pro-inflammatory cytokines IL-6, IL-8 and IL-33 was enhanced in basophils from asthmatic patients, and this effect was partially reversed by transfection of miR-3934 mimics. Furthermore, receiver operating characteristics analysis showed that miR-3934 levels can be used to distinguish asthma patients from healthy individuals. miR-3934 partially inhibited advanced glycation end products-induced increases in basophil apoptosis by suppressing expression of RAGE. CONCLUSION: Our results indicate that miR-3934 acts to mitigate the pathogenesis of asthma by targeting RAGE and suppressing TGF-ß/Smad signaling.

Activation of the receptor for advanced glycation end products (RAGE) exacerbates experimental autoimmune myasthenia gravis symptoms.

RAGE belongs to immunoglobulin superfamily and serves as a ligand for various immunoregulatory molecules including S100B that has been demonstrated important to T cell mediated autoimmune diseases. In this context, we hypothesized that RAGE could also impact B cell mediated, T cell-dependent autoimmune diseases. This was tested using myasthenia gravis (MG) animal model, EAMG. We show that expression of both RAGE and S100B are increased during EAMG and the interaction between RAGE and S100B affected the Th1/Th2/Th17/Treg cell equilibrium, up-regulate AChR-specific T cell proliferation. Furthermore, addition of S100B in vitro stimulated splenocyte activity linked to COX-2 up-regulation. NS-398, a selective COX-2 inhibitor, effectively diminished S100B mediated activity of AChR-specific antibody secreting splenocytes. These findings suggested that a reciprocal relationship between RAGE and S100B promoted the development of EAMG, highlighting the importance of understanding the mechanisms of EAMG disease as a means of developing new therapies for the treatment of MG.