MicroRNAs (miRNAs) have been shown to be important regulators of TLR signaling pathway at the post-transcriptional level. In this study, the potential role of miR-149 was explored in murine alveolar macrophage RAW264.7 cells. Our results demonstrated a dynamic change of the expressions of miR-149 expression and MyD88 in macrophage RAW264.7 upon Mycobacterium bovis Bacillus Calmette-Guerlin (BCG) infection or lipopolysaccharide (LPS) stimulation. The presence of BCG or LPS dynamically reduced the miR-149 expression, along with a substantially striking increase of MyD88 expression in these cells. More importantly, overexpression of miR-149 in RAW264.7 cells was associated with a significant decrease of MyD88 protein expression, as well as a reduced production of inflammatory mediator NF-κB 1, TNF-α and IL-6 in response to BCG infection or LPS stimulation. Further studies using immunoblotting assay against anti-MyD88 antibody and microRNA targeting luciferase reporter assay revealed that miR-149 was able to directly target the 3'-UTR of MyD88 mRNA and post-transcriptionally regulated MyD88 protein expression. These data suggested that miR-149 might be a key player of immune modulator for TLR/MyD88 signaling pathway in macrophages, which may through a mechanism of negatively regulating MyD88-dependent Toll-like receptors signaling pathway.

BackgroundmicroRNA 21 (miR-21) has been demonstrated to be significantly elevated in many types of cancers, including the hepatocellular carcinoma (HCC). In the present study, we investigated the role of miR-21 in HCC by identifying its novel targets, as well as its underlying molecular mechanism.MethodsThe expression of mitogen-activated protein kinase-kinase 3 (MAP2K3) in human HCC tumor tissues and adjacent non-tumor tissues was determined by immunohistochemistry staining (IHC) analysis. The 3’-untranslated region (3’-UTR) of MAP2K3 combined with miR-21 was experimentally verified by a miRNA luciferase reporter approach. Moreover, the role of miR-21 in regulating HCC cell proliferation was analyzed by an MTT assay infected with miR-21mimics/sponge inhibitor Adenoviral viral vectors.ResultsBy immunohistochemistry staining analysis, we found that mitogen-activated protein kinase-kinase 3 (MAP2K3) was strikingly repressed in the human HCC tumor tissues, in comparison with the adjacent non-tumor tissues in clinical settings. More importantly, the repression of MAP2K3 was inversely correlated with the expression of miR-21 in HCC. Further study demonstrated that the MAP2K3 was a novel direct target of miR-21, which was experimentally validated by a miRNA luciferase reporter approach. In HepG2 cells, inhibition of miR-21 expression with an adenoviral miR-21 sponge vector profoundly suppressed cell proliferation by up-regulating MAP2K3 expression at both mRNA and protein levels.ConclusionsThese results provide a clinical evidence that MAP2K3 may be a tumor repressor gene, and it is a direct target of miR-21 in HCC, indicating an underlying mechanism by which miR-21 is able to directly target MAP2K3 and inhibit its expression during the carcinogenesis of HCC, at both transcriptional and post-translational levels. This study also suggests that targeting miR-21-MAP2K3 pathway may be a promising strategy in the prevention and treatment of HCC.

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Background The mechanisms through which Mycobacterium tuberculosis evades immune surveillance during tuberculosis (TB) infection remain complex. Previous studies have found that Mycobacteria can manipulate the miRNAs of host cells to promote their survival during host-pathogen interactions, and most of these effects occur at the cellular miRNA level. We attempted to investigate the possible related mechanisms at the exosomal miRNA level. Results High-throughput sequencing revealed that Bacillus Calmette-Guérin (BCG) infection could alter the composition of the macrophage exosome content, and the expression levels of miRNAs in exosomes derived from the cell culture media of macrophages showed significant differences between the BCG-infected and non-infected groups. Compared with the non-infected group, 20 exosomal miRNAs were up-regulated and 7 exosomal miRNAs were down-regulated in the infection group (p < 0.05), of which mmu-miR-27b-3p, mmu-miR-93-5p, mmu-miR-25-3p, mmu-miR-1198-5p, mmu-let-7c-5p and let-7a-5p were significantly up-regulated. A bioinformatic analysis indicated that these differentially expressed exosomal miRNAs were involved in multiple biological processes and pathways. The target genes of top six miRNAs in up-regulated groups were positively correlated with the regulation of apoptosis. Conclusions The expression profile of miRNA in exosomes derived from macrophage were altered after Mycobacterium Bovis Bacillus Calmette-Guérin infection, and the differentially expressed miRNAs were involved in multiple biological processes and signalling pathways. The top six up-regulated miRNAs and their targeted genes were predominantly correlated with the regulation of apoptosis.

Previous studies have linked the WNT pathway and human skeleton formation; therefore, genes related to WNT might contribute to the onset and development of osteoporosis. In this study, we investigated the potential genetic association of WLS, which encodes an important mediator in the WNT pathway, with osteoporosis and its related quantitative traits in a sample of 6,620 individuals from Han Chinese population. A two-stage approach, with a discovery stage with 859 cases and 1,690 controls and a validation stage with 1,039 cases and 3,032 controls, was applied in the study. Forty SNPs were genotyped in the discovery stage. The intronic SNP rs2566752 was identified to be significantly associated with osteoporosis (ORdiscovery = 0.78, P discovery = 3.73 × 10−5; ORvalidation = 0.80, P validation = 1.96 × 10−5). Two SNPs surrounding rs2566752 (in addition to this SNP itself) were identified to be associated with bone mineral density. In addition, we have identified a 20 kb peak region of H3K27Ac histone mark enrichment between rs2772304 and rs2566752. Our study suggested that WLS is an important locus for osteoporosis and its related quantitative phenotypes in Han Chinese population. Additional sequencing-based studies are needed to investigate the genetic architecture of this regulatory region and its relationship with osteoporosis-related phenotypes.