Journal: Oncotarget

Article Title: By inhibiting snail signaling and miR-23a-3p, osthole suppresses the EMT-mediated metastatic ability in prostate cancer

doi:

Figure Lengend Snippet: A. A schematic representation of the procedure for miRNA selection. Differential expressions of miRNAs in osthole-treated cells versus vehicle-treated cells were analyzed with a OneArray ® miRNA profiling chip. B. Treatment of Du145 cells with osthole for 6 h. miR-146a, miR-22-3p, and miR-23a-3p expressions were detected by a quantitative PCR. C. Upper panel, Du145 cells were transfected with an miR-23a-3p mimic or mimic control for 24 h followed by osthole (60 μM) treatment for an additional 24 h. E-cadherin expression levels were determined by a Western blot analysis. Quantitative E-cadherin protein levels were adjusted to the β-actin protein level. Lower panel, Relative luciferase activities of DU145 cells co-transfected with an E-cadherin luciferase 3′UTR reporter vector and miR-23a-3p mimic or mimic control for 24 h. Values are presented as the mean ± SE of three independent experiments. * p < 0.05, compared to the control groups. D. DU145 cells were transfected with an miR-23a-3p mimic or mimic control for 24 h followed by osthole (60 μM) treatment for an additional 24 h. The cell-invasion ability was determined by a Matrigel invasion assay. Values are presented as the mean ± SE of three independent experiments. Data were analyzed using a one-way ANOVA with Tukey's post-hoc tests at 95% confidence intervals; different letters represent different levels of significance. E. DU145 cells were transfected with either an miR-23a-3p inhibitor or a negative control. The cell-invasion ability was determined by a Matrigel invasion assay. Values are presented as the mean ± SE of three independent experiments. * p < 0.05, compared to the control groups. F. PC3 or DU145 cells were treated with TGF-β for 6, 12, or 24 h. Expression levels of E-cadherin and miR-23a-3p were determined by Western blotting (upper panel) and a quantitative PCR (lower panel), respectively. Quantitative E-cadherin protein levels were adjusted to the β-actin protein level.

Article Snippet: To identify which miRNAs are regulated by osthole, a high-throughput and specific miRNA microarray (human miRNA OneArray ® miRNA profiling chip) using Du145 cells after osthole treatment was conducted by the Phalanx Biotech Group (Hsinchu, Taiwan).

Techniques: Selection, Real-time Polymerase Chain Reaction, Transfection, Expressing, Western Blot, Luciferase, Plasmid Preparation, Invasion Assay, Negative Control

Journal: Pathology and Oncology Research

Article Title: Identification of miRNA Signature in Breast Cancer to Predict Neoadjuvant Chemotherapy Response

doi: 10.3389/pore.2021.1609753

Figure Lengend Snippet: A workflow of the experimental design was showed (A) . Cluster analysis in chemoresistant (C) and chemosensitive (H) breast cancer tissues (B) . Hierarchical clustering of 33 miRNAs differentially expressed in 20 breast cancer patients, including miR-638, miR-23a-3p, miR-23b-3p, miR-200c-3p, miR-214-3p and miR-451a. Red: up-regulated; green: down-regulated miRNAs in the chemoresistant group. Samples are in columns; miRNAs are in rows. Expression values ranged from −3 to +3 log2. (C) 33 miRNAs are significantly expressed between chemoresistant (G1) and chemosensitive (G2) groups. Mean fold-change of chemosensitive vs. chemoresistant groups at log2 (G2/G1). qRT-PCR results showed that miR-23a-3p, miR-23b-3p, miR-200c-3p, and miR-214-3p were significantly up-regulated, and miR-638 and miR-451a were significantly downregulated in the chemoresistant group in the training set [ (D) ; p = 0.0259; 0.0379; 0.0354; 0.0367; 0.001; 0.002], internal testing set [ (E) ; p = 0.0292; 0.0486; 0.0283; 0.118; 0.0036; 0.013], in the independent set [ (F) ; p = 0.0026; 0.0195; 0.0077; 0.0162; 0.0036; 0.0209] and combined set (including training set, internal testing set and the independent set; (G) : p = 0.0015; 0.0003; 0.0037; 0.003; 0.001; 0.001). The data was represented as the mean and SE (standard error).

Article Snippet: The miRNA microarray chip covered all miRNAs in the miRBase database version 19.0 ( http://www.mirbase.org/ ; including 2019 miRNAs), and was a product of LC Sciences (Houston, TX, United States).

Techniques: Expressing, Quantitative RT-PCR

Journal: Pathology and Oncology Research

Article Title: Identification of miRNA Signature in Breast Cancer to Predict Neoadjuvant Chemotherapy Response

doi: 10.3389/pore.2021.1609753

Figure Lengend Snippet: Association between five deregulated miRNAs and endocrine features or chemotherapy drugs. miR-451a was less expressed in ER-negative cases compared to ER-positive cases [ (A) ; p = 0.0296]. miR-23a expression was greater in PR-positive cases compared to PR-negative cases [ (B) ; p = 0.0367]. miR-638 and miR-451a were down-regulated in HER2-negative cases, compared with HER2-positive cases [ (C, D) ; p = 0.0219; 0.0117]. Compared with Ki67-low cases, more expression of miR-638 and miR-200c-3p were observed in Ki67-high cases [ (E, F) ; p = 0.017; 0.0233]. miR-214-3p expression was increased in p53-positive vs. p53-negative cases [ (G) ; p = 0.0213]. miR-23a-3p expression in A and T groups were higher than in the AT group [ (H) ; p = 0.0393; 0.0430]. Upregulated miR-200c-3p was observed in A group, compared with T and AT groups [ (I) ; p = 0.0177, 0.0361]. miR-214-3p was upregulated in T group, compared with A and AT groups [ (K) ; p = 0.0254, 0.0373]. Reduced expression of miR-638 occurred in the A group, compared with T and AT groups [ (J) ; p = 0.019, 0.0162]. Significantly step-upregulated expression of miR-451a occurred in A, T and AT groups (L) . Significant difference of miR-451a expression was observed between A and T groups ( p = 0.0454), between A and AT groups ( p = 0.0177), between T and AT groups ( p = 0.0494). The data was represented as the mean and SE. ER, estrogen receptor; PR, progesterone receptor; HER2, human epidermal growth factor receptor 2; A, anthracycline; T, taxol; AT, anthracycline and taxol combination. SE, standard error.

Article Snippet: The miRNA microarray chip covered all miRNAs in the miRBase database version 19.0 ( http://www.mirbase.org/ ; including 2019 miRNAs), and was a product of LC Sciences (Houston, TX, United States).

Techniques: Expressing

Journal: Pathology and Oncology Research

Article Title: Identification of miRNA Signature in Breast Cancer to Predict Neoadjuvant Chemotherapy Response

doi: 10.3389/pore.2021.1609753

Figure Lengend Snippet: Risk scores according to five-miRNA signatures and ROC curves in training, internal testing and independent sets. ROC and AUCs of single miRNA alone and five-miRNA signature from the training (A) , internal testing (B) , and independent sets (C) . Risk scores for patients were calculated by risk scores based on a five-miRNA signature in the training (D) , internal testing. (E) and independent sets (F) .

Article Snippet: The miRNA microarray chip covered all miRNAs in the miRBase database version 19.0 ( http://www.mirbase.org/ ; including 2019 miRNAs), and was a product of LC Sciences (Houston, TX, United States).

Techniques:

Journal: Pathology and Oncology Research

Article Title: Identification of miRNA Signature in Breast Cancer to Predict Neoadjuvant Chemotherapy Response

doi: 10.3389/pore.2021.1609753

Figure Lengend Snippet: GO and KEGG analysis of predicted targets of five miRNAs. Data show that genes were enriched in biological processes of signal transduction. (A) ; eight known canonical cancer-associated pathways were predicted by KEGG. (B) , including the p53 signaling pathway, ubiquitin mediated proteolysis, pathways in cancer, the mTOR signaling pathway, the Wnt signaling pathway, regulation of actin cytoskeleton, focal adhesion, and the ErbB signaling pathway. Networks among specific deregulated miRNAs and predicted targets of the classical Wnt signaling pathway. (C) and the MAPK signaling pathway (D) .

Article Snippet: The miRNA microarray chip covered all miRNAs in the miRBase database version 19.0 ( http://www.mirbase.org/ ; including 2019 miRNAs), and was a product of LC Sciences (Houston, TX, United States).

Techniques: Transduction, Ubiquitin Proteomics

Journal: Nucleic Acids Research

Article Title: Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level

doi: 10.1093/nar/gkx1254

Figure Lengend Snippet: MiR-122 localizes in the nucleus of liver cells. ( A ) Nuclear miRNA expression profile in mouse liver cells via detecting by miRNA microarray. The most 50 enriched nuclear miRNAs were plotted in the figure. ( B ) MiR-122, miR-29b and miR-29a expression levels in mouse hepatocytes and nuclei (upper panel), human Huh-7 cells and nuclei (lower panel) by using TaqMan probe-based RT-qPCR assay. ( C ) Nuclear localization of synthetic Cy3-labeled miR-122, miR-29b and miR-29a mimics in Huh-7 cells. ( D ) 3-D re-constituted images of Z-stacks of Cy3-labeled miR-122, miR-29b and miR-29a mimics in Huh-7 cells. White arrowheads indicated the merged pink colored spots. ( E ) Distribution of miR-122 in Huh-7 cells by FISH using Dig-labeled miRCURY LNA™ microRNA Detection Probes. All images were obtained with the confocal microscope. The nuclear expression of U6 was visualized using the miRCURY LNA™ U6 probe as a positive control. DAPI was dyed with blue and probes were labeled with red by anti-Dig-rhodamine. Bars, 20 μm.

Article Snippet: Purified RNA was labeled with fluorescein, and hybridization was performed on a miRNA microarray chip (miRNA microarray V4.0, CapitalBio Corp., Beijing, China).

Techniques: Expressing, Microarray, Quantitative RT-PCR, Labeling, Microscopy, Positive Control

Journal: Nucleic Acids Research

Article Title: Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level

doi: 10.1093/nar/gkx1254

Figure Lengend Snippet: Nuclear miR-122 inhibits miR-21 biogenesis. ( A ) TaqMan Low Density Array screening for target miRNAs of miR-122. Huh-7 cells were transfected with miR-122 mimic or control oligonucleotide and then harvested 24 h after transfection. The miRNA expression profile was sorted using a hierarchical clustering method (Cluster 3.0 and Java TreeView). Twenty eight most significantly changed miRNAs were shown in the cluster. ( B ) RT-qPCR validation of decreased miRNAs screened by Low Density Array following ectopic expression of miR-122 mimic or control. ( C ) Relative pri-miR-21, pre-miR-21 and miR-21 expression levels in Huh-7 cells after miR-122 overexpression or depletion. ( D ) RT-qPCR analysis of expression levels of miR-122 in 16 paired HCC and ANCT tissue samples. ( E ) RT-qPCR analysis of miR-21 level in 16 paired HCC and ANCT samples. ( F ) Pearson's correlation scatter plot of the levels of miR-122 and miR-21 in 16 paired HCC tissues. The results are presented as the mean ± SD ( N = 3) of three independent experiments. * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: Purified RNA was labeled with fluorescein, and hybridization was performed on a miRNA microarray chip (miRNA microarray V4.0, CapitalBio Corp., Beijing, China).

Techniques: TLDA Assay, Transfection, Control, Expressing, Quantitative RT-PCR, Biomarker Discovery, Over Expression

Journal: Nucleic Acids Research

Article Title: Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level

doi: 10.1093/nar/gkx1254

Figure Lengend Snippet: Nuclear miR-122 directly binds to a cognate element on pri-miR-21 transcript. ( A ) Putative binding site for miR-122 on human pri-miR-21 as predicted using RNAhybrid. As shown in the schematic diagram, a near perfect complementary site (red rectangle) for miR-122 (black rectangle) was located 13 bp upstream of the pre-miR-21 gene loci. mfe: minimum free energy. ( B ) Schematic representations of modified GFP expression plasmid. MiR-122 binding sequences (Binding-WT) and mutant sequences (Binding-MUT) were inserted into the 3′-UTR of GFP in GFP expression plasmid. ( C ) HEK-293T cells were co-transfected with the modified vector and miR-122 mimic. After 48 h, fluorescence microscopy was used to detect GFP expression. Scale bar: 1.0 mm. ( D ) The GFP mRNA expression level was detected by RT-qPCR. NS: no significance. ( E ) Schematic illustration of pri-miR-21 pull-down strategy. ( F and G ) Levels of pri-miR-21 (F) or miR-122 and other candidate miRNAs (G) in pull-down products which were co-precipitated by anti-pri-miR-21 probe or random probe. Data are presented as mean ± SD ( N = 3) of three independent experiments. ** P < 0.01; *** P < 0.001.

Article Snippet: Purified RNA was labeled with fluorescein, and hybridization was performed on a miRNA microarray chip (miRNA microarray V4.0, CapitalBio Corp., Beijing, China).

Techniques: Binding Assay, Modification, Expressing, Plasmid Preparation, Mutagenesis, Transfection, Fluorescence, Microscopy, Quantitative RT-PCR

Journal: Nucleic Acids Research

Article Title: Nuclear miR-122 directly regulates the biogenesis of cell survival oncomiR miR-21 at the posttranscriptional level

doi: 10.1093/nar/gkx1254

Figure Lengend Snippet: MiR-122 blocks the primary miR-21 processing by in vitro pri-miRNA processing assay. ( A ) Schematic illustration of pri-miR-21 in vitro processing assay strategy. ( B ) For immunoprecipitation (IP) assays, Huh-7 cell lysates were incubated with anti-Drosha antibody and IgG control antibody, The IP-product was then detected by western blotting (WB) using anti-Drosha and anti-DGCR8 antibody. ( C ) Northern blotting analysis of the miR-122 blocking in vitro processing of the pri-miR-21-WT and pri-miR-21-MUT. The pri-miR-21-WT and pri-miR-21-MUT transcripts were incubation of synthetic mature single strand miR-122, respectively, and then cleaved by Drosha-complex in vitro . The in vitro processing products were analyzed by northern blot. ( D ) RT-qPCR validation of pre-miR-21 level in processing products. ( E ) The pri-miR-21-WT and pri-miR-21-MUT transcripts were incubation of synthetic mature single strand miR-122, respectively, and then cleaved by nuclear extracts. The in vitro processing products were analyzed by northern blot. ( F ) An unrelated pri-miR-150 was incubation of synthetic mature single strand miR-122 and then cleaved by nuclear extracts. The in vitro processing products were analyzed by northern blot. The results are presented as the mean ± SD ( N = 3) of three independent experiments. ** P < 0.01.

Article Snippet: Purified RNA was labeled with fluorescein, and hybridization was performed on a miRNA microarray chip (miRNA microarray V4.0, CapitalBio Corp., Beijing, China).

Techniques: In Vitro, Immunoprecipitation, Incubation, Control, Western Blot, Northern Blot, Blocking Assay, Quantitative RT-PCR, Biomarker Discovery

Journal: Journal of the American Society of Nephrology : JASN

Article Title: miR-335 and miR-34a Promote Renal Senescence by Suppressing Mitochondrial Antioxidative Enzymes

doi: 10.1681/ASN.2010040367

Figure Lengend Snippet: The significantly differentially expressed miRNAs in aging kidney

Article Snippet: LC Sciences (Houston, TX) provided the miRNA microarray chip (μParaflo microfluidic chip).

Techniques:

Journal: Journal of the American Society of Nephrology : JASN

Article Title: miR-335 and miR-34a Promote Renal Senescence by Suppressing Mitochondrial Antioxidative Enzymes

doi: 10.1681/ASN.2010040367

Figure Lengend Snippet: miR-184, miR-335 and miR-347 exhibited differential expressions in (A) aged rats and (B) mice. (A) Fold differences between old and young rat kidneys measured by chip and qRT-PCR. (B) Expression levels of mmu-miR-184, mmu-miR-335, and mmu-miR-347 in the aged mouse kidneys detected by qRT-PCR. n = 5 per miRNA. *P < 0.01 versus young.

Article Snippet: LC Sciences (Houston, TX) provided the miRNA microarray chip (μParaflo microfluidic chip).

Techniques: Quantitative RT-PCR, Expressing

Journal: Journal of the American Society of Nephrology : JASN

Article Title: miR-335 and miR-34a Promote Renal Senescence by Suppressing Mitochondrial Antioxidative Enzymes

doi: 10.1681/ASN.2010040367

Figure Lengend Snippet: Antioxidation-related miRNAs differentially expressed in old kidney and their target genes

Article Snippet: LC Sciences (Houston, TX) provided the miRNA microarray chip (μParaflo microfluidic chip).

Techniques:

Journal: Journal of the American Society of Nephrology : JASN

Article Title: miR-335 and miR-34a Promote Renal Senescence by Suppressing Mitochondrial Antioxidative Enzymes

doi: 10.1681/ASN.2010040367

Figure Lengend Snippet: Expressions of miR-335 and miR-34a in aging mesangial cells, endothelial cells, tubular epithelial cells and interstitial fibroblasts were significantly upregulated and expressions of SOD2 and Txnrd2 in aging mesangial cells were downregulated. (A) Expression levels of miR-335 and miR-34a in aging renal mesangial cells were detected by qRT-PCR. n = 5 per miRNA. *P < 0.01 versus young. (B) Levels of SOD2 and Txnrd2 proteins were analyzed by Western blot in aging renal mesangial cells. The graph is representative of three separate experiments. (C) Expression levels of miR-335 and miR-34a were detected by qRT-PCR in the aging glomerular epithelial cells, endothelial cells, tubular epithelial cells, and fibroblasts. n = 5 per miRNA. *P < 0.01 versus young, #P < 0.05 versus young.

Article Snippet: LC Sciences (Houston, TX) provided the miRNA microarray chip (μParaflo microfluidic chip).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Journal of the American Society of Nephrology : JASN

Article Title: miR-335 and miR-34a Promote Renal Senescence by Suppressing Mitochondrial Antioxidative Enzymes

doi: 10.1681/ASN.2010040367

Figure Lengend Snippet: (A) miR-335 and miR-34a mimics inhibit SOD2 and Txnrd2 expressions in young mesangial cells, and (B) antisense miR-335 and miR-34a increase SOD2 and Txnrd2 expressions in aging mesangial cells. Co, miRNA control; CA, antisense miRNA control; miR335 and miR34a, miR-335 and miR-34a mimics, respectively; AmiR335 and AmiR34a, antisense miR-335 and miR-34a inhibitors, respectively. (C) Analysis of expression levels of SOD2 and Txnrd2 by Western blot in aged renal tissues. The graph is representative of three separate experiments.

Article Snippet: LC Sciences (Houston, TX) provided the miRNA microarray chip (μParaflo microfluidic chip).

Techniques: Control, Expressing, Western Blot

Journal: Journal of the American Society of Nephrology : JASN

Article Title: miR-335 and miR-34a Promote Renal Senescence by Suppressing Mitochondrial Antioxidative Enzymes

doi: 10.1681/ASN.2010040367

Figure Lengend Snippet: miR-335 and miR-34a mimics induce premature senescent phenotypes in young mesangial cells, and antisense miR-335 and miR-34a relieve senescent phenotypes in old mesangial cells. (A) SA-β-gal staining in young mesangial cells transfected with miR-335 and miR-34a mimics. Blue precipitation in the cytoplasm was observed in the senescent cells. (B) Analysis of SAHF formation in young mesangial cells transfected with miR-335 and miR-34a mimics. The cells were stained with DAPI, and heterochromatin foci were observed in the senescent cells. (C) SA-β-gal staining in aging mesangial cells transfected with antisense miR-335 and miR-34a inhibitors. (D) Analysis of SAHF formation in aging mesangial cells transfected with antisense miR-335 and miR-34a inhibitors. Co, miRNA control; CA, antisense miRNA control. The above results of SA-β-gal staining and SAHF analysis are representative images of three experiments.

Article Snippet: LC Sciences (Houston, TX) provided the miRNA microarray chip (μParaflo microfluidic chip).

Techniques: Staining, Transfection, Control

Journal: Leukemia

Article Title: Regulation of PI3K signaling in T cell acute lymphoblastic leukemia: a novel PTEN/Ikaros/miR-26b mechanism reveals a critical targetable role for PIK3CD

doi: 10.1038/leu.2017.80

Figure Lengend Snippet: A: miRNA profile of Array data. KO576, KO577, KO578 and KO579 are Pten -knockout mouse T-ALL samples. WT580 and WT581 are wild-type mouse thymocytes. B: Decreased miR-26b expression level in mouse Pten deficient T-ALL cell lines (LPN248, LPN236 and LPN228) compared with mouse Ink4a/Arf knock-out T-ALL cell line (LPN211) and mouse wild type thymocytes (P<0.001). C. Decreased miR-26b in human T-ALL cell lines, CCRF-CEM, SUPT1, LOUCY, KOPT-K1, JURKAT and MOLT4 compared with postnatal normal human thymus (P<0.001). D: Decreased expression level of miR-26b in human primary T-ALL samples (P<0.001). E. Correlation of miR-26b expression level with PTEN level in human primary T-ALL samples (r=0.3987, P=0.039).

Article Snippet: Hybridization of biotin-labeled cDNA was carried out on a miRNA microarray chip (MD Anderson miRNA expression Bioarray Version 5), which contains 2300 miRNA probes, including 1400 human and 900 mouse miRNA genes, in duplicate.

Techniques: Knock-Out, Expressing