Upregulation of miR-9 and miR-193b over human Th17 cell differentiation.

BACKGROUND: Th17 cells are a newly discovered subset of CD4+ T cells known as key participants in various immune responses and inflammatory conditions including autoimmune diseases. Mi(cro)RNAs are a family of non-coding RNAs that regulate numerous critical immune functions. Immuno-miRNAs modulate cell biological processes in T cells, such as differentiation and function of Th17 cells. The aim of the present study is to investigate the expression of miR-9-5p, miR-193b-3p, and autoimmunity-related genes during human Th17 cells differentiation. METHODS: Human naïve CD4+ T cells were purified from peripheral blood mononuclear cells (PBMCs) by magnetic cell sorting system (MACS) and their purity was checked by flow-cytometric analysis. Naïve CD4+ T cells were cultured under Th17-polarizing condition for 6 days. IL- 17 secretion was determined by means of enzyme-linked immunosorbent assay (ELISA). Next, the expression levels of miRNAs and putative targets genes were assessed by qRT-PCR at different time points of differentiation. RESULTS: Our result showed dramatic downregulation of TCF7, MAP3K1, ENTPD1, and NT5E genes during human Th17 differentiation. Polarization also had a significant inducible effect on the expression of miR-9 and miR-193b over differentiation of human Th17 cells. According to our results, miR-9-5p and miR-193b-3p may contribute to Th17 differentiation probably by inhibiting the expression of negative regulators of Th17 differentiation. CONCLUSION: This study confirmed deregulation of TCF7, MAP3K1, ENTPD1, and NT5E genes in Th17 differentiation process and introduced miR-9 and miR-193b as Th17 cell-associated miRNAs, making them good candidates for further investigations.

Modified level of miR-376a is associated with Parkinson's disease.

Parkinson's disease (PD) is a frequent progressive neurodegenerative disorder. Impaired mitochondrial function is a major feature of sporadic PD. Some susceptibility or causative genes detected in PD are strongly associated with mitochondrial dysfunction including PGC1α, TFAM and GSK3ß. microRNAs (miRNAs) are non-coding RNAs whose altered levels are proven in disparate PD models and human brains. Therefore, the aim of this study was to detect modulations of miRs upstream of PGC1α, TFAM and GSK3ß in association with PD onset and progress. In this study, a total of 33 PD subjects and 25 healthy volunteers were recruited. Candidate miRNA (miR-376a) was selected through target prediction tools and literature survey. Chronic and acute in vitro PD models were created by MPP+ -intoxicated SHSY5Y cells. The levels of miR-376a and aforementioned genes were assessed by RT-qPCR. The expression of target genes was decreased in chronic model while there were dramatically up-regulated levels of those genes in acute model of PD. miR-376a was strongly altered in both acute and chronic PD models as well as PBMCs of PD patients. Our results also showed overexpression of PGC1α, and TFAM in PBMCs is inversely correlated with down-regulation of miR-376a, suggesting that miR-376a possibly has an impact on PD pathogenesis through regulation of these genes which are involved in mitochondrial function. miR-376a expression in PD-derived PBMCs was also correlated with disease severity and may serve as a potential biomarker for PD diagnosis. This is the first study showing altered levels of miR-376a in PD models and PBMCs, suggesting the probable role of this miRNA in PD pathogenesis. The present study also proposed TFAM and PGC1α as target genes of miR-376a for the first time, through which it possibly can exert its impact on PD pathogenesis.

Upregulation of miR-200a and miR-204 in MPP+ -treated differentiated PC12 cells as a model of Parkinson's disease.

BACKGROUND: Parkinson's disease (PD) is ranked as the second most common neurodegenerative disorder caused by loss of dopaminergic neurons in the substantia nigra. Micro(mi)RNAs are a class of small noncoding RNAs that regulate gene expression and aberrant expression of them is closely correlated with many neurodegenerative conditions including PD. Silent information regulator 1 (SIRT1) as a known deacetylase and B-cell lymphoma-2 (BCL2) as an antiapoptotic factor play vital roles in neural protection and survival. METHODS: Differentiated PC12 cells exposed to MPP+ were served here as a known PD model. Cell viability was determined by MTS assay. Apoptotic cells and ROS levels were detected using flow cytometry. Gene selection and miRNA-mRNA interaction analysis were performed through in silico methods. Relative expression of miRNAs and genes was examined by RT-qPCR. RESULTS: MPP+ exposure markedly reduced cell viability, enhanced oxidative stress, and induced apoptosis in differentiated PC12 cells. Sirt1 and BCL2were shown to be markedly declined in response to MPP+ , while miR-200a and miR-204 were significantly upregulated. CONCLUSION: The first novel finding of the current study is altered expression of miR-200a and miR-204 in differentiated PC12 cells in response to MPP+ , suggesting that deregulation of them participate in MPP+ neurotoxicity mechanisms, possibly via affecting the expression of Sirt1 and BCL2 as potential targets.

Differential expression of miR-34a, miR-141, and miR-9 in MPP+-treated differentiated PC12 cells as a model of Parkinson's disease.

Parkinson's disease (PD) is a debilitating neurodegenerative condition characterized by the loss of dopaminergic neurons in substantia nigra. MPP+ as a dopaminergic neurotoxin induces many parkinsonian-like symptoms in cell culture. MicroRNAs are noncoding RNAs which their deregulation participates in neurodegeneration by affecting most of the mechanisms responsible for neurodegenerative diseases. Differentiated PC12 cells impaired by MPP+ were served as an in vitro model of PD. The aim of present research is to evaluate expression of several miRNAs and possible target genes in MPP+-treated differentiated PC12 cells. PC12 cells were differentiated by induction of NGF. Neurite outgrowth was quantified using image analysis software. MTS assay was served to assess cell viability. DCFH-DA assay and Annexin v staining were used to detect reactive oxygen species (ROS) levels and apoptosis, respectively. An in-silico study was performed to make interaction analysis between selected mRNAs and microRNAs. The expression levels of microRNAs and target genes were examined by RT-qPCR. NGF induced differentiation led to a substantial increase in neurite lengths means and percentage of the neurite-bearing cells. NGF-differentiated PC12 cells substantially expressed TH and retained their dopaminergic characteristic after differentiation. NGF treatment enhanced TH gene expression. MPP+ exposure caused loss of cell viability and induced apoptosis and ROS overproduction. SIRT1, BCL2, and BDNF were down-regulated after MPP+-treatment. In contrast, MPP+ toxicity significantly caused up-regulation in miR-34a, miR-141, and miR-9. The present study showed altered levels of selected microRNAs in response to MPP+ for the first time, suggesting that perturbed expression of them may contribute to the PD-related pathogenic processes, probably by affecting the expression of BCL2, BDNF, and SIRT1 as potential targets.