Project description:We have employed whole genome microarray expression profiling as a discovery platform to screen potential target genes of miR-204-5p in colorectal cancer cell HCT116. HCT116 cells were seeded in 6-cm2 tissue culture plates and transfected with the miR-204-5p mimic or negative control (NC) using Lipofectamine 2000 (Invitrogen, USA). After propagation for 48 hours, total RNA was extracted using TRIzol reagent (Invitrogen, USA). Expression profiling was performed using an Agilent human whole genome oligo microarray chip (4M-CM-^W44K) (Agilent, USA) Expression profiling in HCT116 cells was measured at 48 hours after transfection with miR-204-5p or negative control. Experiments were performed using the two samples without repeat experiment.
Project description:We have employed whole genome microarray expression profiling as a discovery platform to screen potential target genes of miR-204-5p in colorectal cancer cell HCT116. HCT116 cells were seeded in 6-cm2 tissue culture plates and transfected with the miR-204-5p mimic or negative control (NC) using Lipofectamine 2000 (Invitrogen, USA). After propagation for 48 hours, total RNA was extracted using TRIzol reagent (Invitrogen, USA). Expression profiling was performed using an Agilent human whole genome oligo microarray chip (4×44K) (Agilent, USA)
Project description:To understand molecular mechanisms underlying miR-204-5p mediated suppression of HNSCC progression, we performed RNA-seq analysis in UM-SCC1 cells transfected with miR-204-5p mimics and control mimics. The expression of JAK2, SNAI2, TGFBR2, MAP2K4, involved in both EMT and STAT3 signaling, were all decreased in cells treated with miR-204-5p mimics.
Project description:Patients with advanced colorectal cancer (CRC) are commonly treated with systemic combination therapy but suffer eventually from drug resistance. MicroRNAs (miRNAs) are suggested to play a role in treatment resistance of CRC. We studied whether restoring downregulated miR-195-5p and 497-5p sensitize CRC cells to currently used chemotherapeutics 5-fluorouracil, oxaliplatin and irinotecan. Sensitivity to 5-FU, oxaliplatin and irinotecan before and after transfection with miR-195-5p and miR-497-5p mimics was analyzed in CRC cell lines HCT116, RKO, DLD-1 and SW480. Mass spectrometry based proteomic analysis of transfected and wild-type cells was used to identify targets involved in sensitivity to chemotherapy. Proteomic analysis revealed 181 proteins with significantly altered expression after transfection with miR-195-5p mimic in HCT116 and RKO, including 118 downregulated and 63 upregulated proteins. After transfection with miR-497-5p mimic, 130 proteins were significantly downregulated and 102 were upregulated in HCT116 and RKO (P<0.05 and FC<-3 or FC>3). CHUK and LUZP1 were coinciding downregulated proteins in sensitized CRC cells after transfection with either mimic. Resistance mechanisms of these two proteins may be related to nuclear factor kappa-B signaling and G1 cell cycle arrest, respectively. Restoring miR-195-5p and miR-497-5p expression enhanced sensitivity to chemotherapy, mainly oxaliplatin, in CRC cells and could be a promising treatment strategy for patients with mCRC. Proteomics revealed potential targets of these miRNAs involved in sensitivity to chemotherapy.
Project description:To identify genes affected by miR-766-5p overexpression, we transfected with 10nmol of miR-766-5p or miR-negative control (NC) in HCT116-/-, or MIAPaCa2 cells. After 2 days, RNA was extracted, and then expression analysis was performed using agilent microarray.
Project description:Gene expression profiling of immortalized human mesenchymal stem cells with hTERT/E6/E7 transfected MSCs. hTERT may change gene expression in MSCs. Goal was to determine the gene expressions of immortalized MSCs.
Project description:Introduction: The mechanisms underlying myopia and myopia-related retinopathy remain not fully understood. We proposed to examine the function and underlying mechanisms of miR-204-5p in myopia development. Methods: The miR-204-5p expression level was assessed in the vitreous humor (VH) of a cohort consisting of 11 patients with high myopia (HM) and 16 control patients undergoing retinal surgery. The functional implications of miR-204-5p in ARPE-19 cells were assessed, encompassing cell aggressiveness. Thioredoxin-interacting protein (TXNIP) was found as a possible target of miR-204-5p through mRNA sequencing, and its interaction with miR-204-5p was confirmed employing luciferase assay and western blotting. Furthermore, the miR-204-5p function in regulating oxidative stress was examined by measuring reactive oxygen species (ROS) accumulation. Results: miR-204-5p was found to be significantly reduced in the VH of HM patients. Overexpression of miR-204-5p suppressed cell proliferation, migration, invasion, and apoptosis in ARPE-19 cells. The bioinformatics analysis demonstrated that miR-204-5p can modulate the genes associated with pathways relevant to myopia, including glycosaminoglycan (GAG) degradation, lysosome, and TGF−beta signaling pathway. The direct targeting of miR-204-5p on TXNIP has been confirmed through validation, and its downregulation mediated the miR-204-5p impacts on ARPE-19 cells. Moreover, miR-204-5p overexpression significantly reduced ROS accumulation by targeting TXNIP. Conclusion: Our findings revealed the possible contribution of the miR-204-5p/TXNIP axis in myopia development by regulating oxidative stress, which may provide new targets and novel therapeutic strategies to combat this prevalent and debilitating condition.
Project description:Gene methylation profiling of immortalized human mesenchymal stem cells comparing HPV E6/E7-transfected MSCs cells with human telomerase reverse transcriptase (hTERT)- and HPV E6/E7-transfected MSCs. hTERT may increase gene methylation in MSCs. Goal was to determine the effects of different transfected genes on global gene methylation in MSCs.
Project description:Using the highly sensitive miRNA array, we screened 220 microRNAs abundant in physiological left ventricular hypertrophy (LVH) and we explored the functions of these miRNAs in the cardiac tissue by Gene Ontology and Kyoto Encyclopedia of Genes annotation. miRNAs showed a high score in the pathway enriched in autophagy. Moreover, the expression levels of miR-26b-5p, miR-204-5p, and miR-497-3p showed an obvious increase in rat hearts. Adenovirus-mediated overexpression of miR-26b-5p, miR-204-5p, and miR-497-3p markedly attenuated IGF-1-induced hypertrophy in H9C2 cells by suppressing autophagy. Furthermore, miR-26b-5p, miR-204-5p, and miR-497-3p attenuated autophagy in H9C2 cells through targeting ULK1, LC3B and Beclin 1, respectively. Taken together, our results demonstrate that swimming exercise induced physiological LVH, at least in part, by modulating the microRNA–autophagy axis, and that miR-26b-5p, miR-204-5p, and miR-497-3p may help distinguish physiological and pathological LVH.
Project description:We established two distinct stable clones which ectopically overexpress miR-204-5p to different levels, then performed transcriptome profiling of miR-204-5p overexpressing cells compared to control 4T1 cells to understand the molecular mechanisms of the miR-204-5p’s effect on cancer cells.