miRNA expression profiles and molecular networks in resting and LPS-activated BV-2 microglia-Effect of cannabinoids.

Mammalian microRNAs (miRNAs) play a critical role in modulating the response of immune cells to stimuli. Cannabinoids are known to exert beneficial actions such as neuroprotection and immunosuppressive activities. However, the underlying mechanisms which contribute to these effects are not fully understood. We previously reported that the psychoactive cannabinoid Δ9-tetrahydrocannabinol (THC) and the non-psychoactive cannabidiol (CBD) differ in their anti-inflammatory signaling pathways. Using lipopolysaccharide (LPS) to stimulate BV-2 microglial cells, we examined the role of cannabinoids on the expression of miRNAs. Expression was analyzed by performing deep sequencing, followed by Ingenuity Pathway Analysis to describe networks and intracellular pathways. miRNA sequencing analysis revealed that 31 miRNAs were differentially modulated by LPS and by cannabinoids treatments. In addition, we found that at the concentration tested, CBD has a greater effect than THC on the expression of most of the studied miRNAs. The results clearly link the effects of both LPS and cannabinoids to inflammatory signaling pathways. LPS upregulated the expression of pro-inflammatory miRNAs associated to Toll-like receptor (TLR) and NF-κB signaling, including miR-21, miR-146a and miR-155, whereas CBD inhibited LPS-stimulated expression of miR-146a and miR-155. In addition, CBD upregulated miR-34a, known to be involved in several pathways including Rb/E2f cell cycle and Notch-Dll1 signaling. Our results show that both CBD and THC reduced the LPS-upregulated Notch ligand Dll1 expression. MiR-155 and miR-34a are considered to be redox sensitive miRNAs, which regulate Nrf2-driven gene expression. Accordingly, we found that Nrf2-mediated expression of redox-dependent genes defines a Mox-like phenotype in CBD treated BV-2 cells. In summary, we have identified a specific repertoire of miRNAs that are regulated by cannabinoids, in resting (surveillant) and in LPS-activated microglia. The modulated miRNAs and their target genes are controlled by TLR, Nrf2 and Notch cross-talk signaling and are involved in immune response, cell cycle regulation as well as cellular stress and redox homeostasis

Pathways and gene networks mediating the regulatory effects of cannabidiol, a nonpsychoactive cannabinoid, in autoimmune T cells.

BackgroundOur previous studies showed that the non-psychoactive cannabinoid, cannabidiol (CBD), ameliorates the clinical symptoms in mouse myelin oligodendrocyte glycoprotein (MOG)35-55-induced experimental autoimmune encephalomyelitis model of multiple sclerosis (MS) as well as decreases the memory MOG35-55-specific T cell (TMOG) proliferation and cytokine secretion including IL-17, a key autoimmune factor. The mechanisms of these activities are currently poorly understood.MethodsHerein, using microarray-based gene expression profiling, we describe gene networks and intracellular pathways involved in CBD-induced suppression of these activated memory TMOG cells. Encephalitogenic TMOG cells were stimulated with MOG35-55 in the presence of spleen-derived antigen presenting cells (APC) with or without CBD. mRNA of purified TMOG was then subjected to Illumina microarray analysis followed by ingenuity pathway analysis (IPA), weighted gene co-expression network analysis (WGCNA) and gene ontology (GO) elucidation of gene interactions. Results were validated using qPCR and ELISA assays.ResultsGene profiling showed that the CBD treatment suppresses the transcription of a large number of proinflammatory genes in activated TMOG. These include cytokines (Xcl1, Il3, Il12a, Il1b), cytokine receptors (Cxcr1, Ifngr1), transcription factors (Ier3, Atf3, Nr4a3, Crem), and TNF superfamily signaling molecules (Tnfsf11, Tnfsf14, Tnfrsf9, Tnfrsf18). "IL-17 differentiation" and "IL-6 and IL-10-signaling" were identified among the top processes affected by CBD. CBD increases a number of IFN-dependent transcripts (Rgs16, Mx2, Rsad2, Irf4, Ifit2, Ephx1, Ets2) known to execute anti-proliferative activities in T cells. Interestingly, certain MOG35-55 up-regulated transcripts were maintained at high levels in the presence of CBD, including transcription factors (Egr2, Egr1, Tbx21), cytokines (Csf2, Tnf, Ifng), and chemokines (Ccl3, Ccl4, Cxcl10) suggesting that CBD may promote exhaustion of memory TMOG cells. In addition, CBD enhanced the transcription of T cell co-inhibitory molecules (Btla, Lag3, Trat1, and CD69) known to interfere with T/APC interactions. Furthermore, CBD enhanced the transcription of oxidative stress modulators with potent anti-inflammatory activity that are controlled by Nfe2l2/Nrf2 (Mt1, Mt2a, Slc30a1, Hmox1).ConclusionsMicroarray-based gene expression profiling demonstrated that CBD exerts its immunoregulatory effects in activated memory TMOG cells via (a) suppressing proinflammatory Th17-related transcription, (b) by promoting T cell exhaustion/tolerance, (c) enhancing IFN-dependent anti-proliferative program, (d) hampering antigen presentation, and (d) inducing antioxidant milieu resolving inflammation. These findings put forward mechanism by which CBD exerts its anti-inflammatory effects as well as explain the beneficial role of CBD in pathological memory T cells and in autoimmune diseases

Experiments of water\u27s effect on mechanical properties of shale rocks

The multiple hydraulic fracturing is an indispensable means to improve the production mass of natural gas in development of shale gas. The fracturing water consumption of a horizontal well reaches 10x103 m3. However, the water been injected into shale layer is not reverse discharged completely. How does this part of water stays in shale layer? What\u27s the role it plays? And how does it have any effects on the development? We studied effects of water on shale rock mechanical properties experimentally to answer these questions. Please download the full abstract below

Seroprevalence of HBV Infection Among Normal Population and Healthcare Workers in Baghdad

Objective: In this study we verified the epidemiology of HBV infection among normal population and healthcare workers (HCWs) in Baghdad by analyzing the prevalence of specific viral markers (anti-HBs, anti-HBc and HBsAg). Method: A total of 797 serum specimens (588-normal population, 209-HCWs) were tested using ELISA technique and positive HBsAg specimens were confirmed by VIDAS technique. Results: In normal population group, the HBsAg, anti-HBs and anti-HBc Total were 1.02%, 10.54%, 5.44%, respectively. The HBsAg result was significantly lower (P< 0.05) than previous studies were done in Iraq. Significant correlation (P < 0.05) in prevalence of HBsAg was found between age groups and males had higher positive HBsAg marker than females (P < 0.05). The prevalence of anti-HBs was insignificant (P＞0.05) between age groups but significant importance (P < 0.05) was recorded between both sexes. Highly significant (P＜0.01) was recorded between ages groups regarding to anti-HBc Total marker but insignificantly (P＞0.05) between both sexes were noticed. In HCWs group the HBsAg, anti-HBs, anti-HBc Total were 0.96%, 26.7%, 1.44% respectively and the prevalence of HBsAg was significantly less (P < 0.05) than previous studies. The comparison between normal population and HCWs groups showed significant correlation (P < 0.05) related to HBsAg and highly significant correlation (P＜0.01) regarding anti-HBs and anti-HBc Total. The HBsAg and anti-HBs among HCWs was significantly increased (P＜0.05) with advancing ages, whereas no such variation was observed between both sexes. The HBsAg was high among those with poor health education but it was with significantly importance (P< 0.05) among different occupational types as well as among vaccinated, incomplete vaccinated and unvaccinated HCWs groups. Additionally, HCWs who had only 1st, only 2nd and those who had 3rd doses showed significant correlation between number of vaccinated individuals and prevalence of anti-HBs. Keywords: Seroprevalence of HBV, HBsAg, anti-HBs, HCWs, normal population Abbreviations: anti-HBs, antibody anti-HBs; anti-HBc, antibody anti-HBc; HBsAg, antigen HB

Co-expression networks in generation of induced pluripotent stem cells.

We developed an adenoviral vector, in which Yamanaka's four reprogramming factors (RFs) were controlled by individual CMV promoters in a single cassette (Ad-SOcMK). This permitted coordinated expression of RFs (SOX2, OCT3/4, c-MYC and KLF4) in a cell for a transient period of time, synchronizing the reprogramming process with the majority of transduced cells assuming induced pluripotent stem cell (iPSC)-like characteristics as early as three days post-transduction. These reprogrammed cells resembled human embryonic stem cells (ESCs) with regard to morphology, biomarker expression, and could be differentiated into cells of the germ layers in vitro and in vivo. These iPSC-like cells, however, failed to expand into larger iPSC colonies. The short and synchronized reprogramming process allowed us to study global transcription changes within short time intervals. Weighted gene co-expression network analysis (WGCNA) identified sixteen large gene co-expression modules, each including members of gene ontology categories involved in cell differentiation and development. In particular, the brown module contained a significant number of ESC marker genes, whereas the turquoise module contained cell-cycle-related genes that were downregulated in contrast to upregulation in human ESCs. Strong coordinated expression of all four RFs via adenoviral transduction may constrain stochastic processes and lead to silencing of genes important for cellular proliferation

Community prevalence of carbapenemase-producing Gram-negative bacteria

Purpose: To raise awareness of carbapenemase-producing organisms, identify “at-risk” patients when admitted in a medical healthcare facility, and to outline effective infection prevention and control measures in order to halt the entry and spread of these organisms. Methods: A total of 1043 un-duplicated urine specimens of healthy volunteers who had no travel history or history of hospitalization were screened. The carbapenemase genotype of each imipenem-resistant strain was determined. Molecular typing and homology analysis of the main carbapenemase-producing strains were used to reveal the mode of transmission of resistance genes. Through transfer joint experiments, the potential risk of spread of carbapenemase genes was assessed. Results: A total of 19 carbapenemase-producing strains from 1,043 non-duplicated healthy volunteers (1.82 %) were identified. The main carbapenemase-producing organism was E. coli (42.1 %, 8/19). The main carbapenemase genotype of E. coli was blaKPC-2 (7 strains). Results from multi-locus sequence typing (MLST) indicated that 7 E. coli isolates belonged to ST-10, ST-101, ST-131, ST-405, ST-410 and ST-1193 and ST-2562. Homologous cluster analysis revealed that the sequence types among the 7 E. coli were high in diversity. The blaKPC-2 gene was successfully transferred from these isolates to 10.22-14 via conjugation. All recipient cells showed marked decreases in carbapenem sensitivity to imipenem (p < 0.05)). The degrees of conjugation were 2.10±0.12 ×10-4, 1.96±0.14×10-4, 2.72±0.18 ×10-4, 3.15±0.20 × 10-4, 2.92±0.23 ×10-4, 3.50±0.20 ×10-4 and 4.12±0.24 ×10-4 in recipient cells of TC7.23-51, TC8.9-42, TC8.15-11, TC8.23-59-3, TC8.23-83, TC9.08-47 and TC10.13-15, respectively. Conclusion: The findings demonstrate the pattern and features of carbapenemase-insensitive E. coli. The blaKPC-2 was the main community-prevalent gene of carbapenem-resistant E. coli. In view of increasing incidence of resistance to multi-drug therapy, surveillance of insensitivity to antibiotics is vital, especially urinary system infection due to carbapenem-insensitive E. coli

Identification of an Efficient Gene Expression Panel for Glioblastoma Classification.

We present here a novel genetic algorithm-based random forest (GARF) modeling technique that enables a reduction in the complexity of large gene disease signatures to highly accurate, greatly simplified gene panels. When applied to 803 glioblastoma multiforme samples, this method allowed the 840-gene Verhaak et al. gene panel (the standard in the field) to be reduced to a 48-gene classifier, while retaining 90.91% classification accuracy, and outperforming the best available alternative methods. Additionally, using this approach we produced a 32-gene panel which allows for better consistency between RNA-seq and microarray-based classifications, improving cross-platform classification retention from 69.67% to 86.07%. A webpage producing these classifications is available at http://simplegbm.semel.ucla.edu