Journal: Stem Cells and Development

Article Title: MicroRNAs Involved in Asthma After Mesenchymal Stem Cells Treatment

doi: 10.1089/scd.2015.0339

Figure Lengend Snippet: miRNAs differently expressed during asthma and after BM-MSCs treatment. (A) Hierarchical clustering analysis of all the aberrant miRNAs that were differently expressed either between the model ( n = 3) and control groups ( n = 3) or between the model+BM-MSCs ( n = 3) and model groups (total n = 9, greater than twofold; P < 0.05). (B) The miRNA development patterns after asthma induction and then with the BM-MSCs. (a) The first lines in the boxes represent increased or decreased miRNA expression after asthma induction compared with the control groups. The second lines in the boxes represent increased or decreased miRNA expression after the BM-MSCs treatment compared with the model group. The numbers in the boxes represent the number of miRNAs that obey the development pattern of that box. Two patterns with a total 14 aberrant miRNAs were selected and marked with orange for next experiments. (b) A detailed pattern of 10 miRNAs in the upper orange box and 4 miRNAs in the lower orange box in (B, a ) for the three groups. (C) Hierarchical clustering analysis of the said 14 miRNAs marked with orange in (B, a ) . Red , expression above the median; green , expression below the median. miRNAs, microRNAs.

Article Snippet: For the miRNA array, the Exiqon miRCURY LNA TM Universal RT miRNA PCR profiling kit with two 384-well plates (Ready-to-Use Mouse & Rat Panel I+II V3, Cat. No. 20379; Exiqon; www.exiqon.com ), which covered 752 murine miRNAs, was used with quantitative real-time PCR (qRT-PCR) technology.

Techniques: Control, Expressing

Journal: Stem Cells and Development

Article Title: MicroRNAs Involved in Asthma After Mesenchymal Stem Cells Treatment

doi: 10.1089/scd.2015.0339

Figure Lengend Snippet: Mmu-miR-21a-3p , mmu-miR-449c-5p , and mmu-miR-496a-3p were significantly differentially expressed in asthma and with BM-MSCs treatment. (A) The fold changes of the 14 selected miRNAs with the log ratio of the mean value are shown. The arrows indicate the miRNAs that were selected for the next experiments. The mmu-miR-21a-3p (B) , mmu-miR-449c-5p (C), and m mu-miR-496a-3p (D) expression levels were confirmed with real time PCR in subsequent lung tissue samples from the control ( n = 14), model ( n = 12), and model+BM-MSC ( n = 14) groups. ** P < 0.01; *** P < 0.001. PCR, polymerase chain reaction.

Article Snippet: For the miRNA array, the Exiqon miRCURY LNA TM Universal RT miRNA PCR profiling kit with two 384-well plates (Ready-to-Use Mouse & Rat Panel I+II V3, Cat. No. 20379; Exiqon; www.exiqon.com ), which covered 752 murine miRNAs, was used with quantitative real-time PCR (qRT-PCR) technology.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Control, Polymerase Chain Reaction

Journal: Stem Cells and Development

Article Title: MicroRNAs Involved in Asthma After Mesenchymal Stem Cells Treatment

doi: 10.1089/scd.2015.0339

Figure Lengend Snippet: The mRNA profile in asthma with the treatment of BM-MSCs and the relationship between mmu-miR-21a-3p and Acvr2a . (A) A hierarchical clustering analysis of all the aberrant protein-coding genes in the PCR array that were differentially expressed either between the model and control groups or between the model+BM-MSCs and model groups (≥1.5-fold; P < 0.05). (B) A hierarchical clustering analysis of 19 aberrant protein-coding genes in the PCR array. Red , expression above the median; green , expression below the median in (A) . (C) Schematic diagrams showing the interaction sites between the mmu-miR-21a-3p , mmu-miR-449c-5p , and mmu-miR-496a-3p and the 3′ UTR of their corresponding mRNAs. (D) Mmu-miR-21a-3p was negatively correlated with the Acvr2a gene in miRNAs array and mRNA PCR array. y , the mean Acvr2a Ct value in the three groups. x , the mean mmu-miR-21a-3p Ct value in the three groups. (E) qRT-PCR analysis of Acvr2a in lung tissue samples from the control, model, and model+BM-MSC groups. n = 6 in each group. (F) Dual luciferase assay of 293T cells cotransfected with firefly luciferase vector containing the Acvr2a 3′ UTRs and the mmu-miR-21a-3p mimics or scrambled oligonucleotides as the miRNA negative control. * P < 0.05; ** P < 0.01. Acvr2a, activin A receptor, Type IIA; mRNA, messenger RNA; UTR, untranslated region; qRT-PCR, quantitative real-time PCR.

Article Snippet: For the miRNA array, the Exiqon miRCURY LNA TM Universal RT miRNA PCR profiling kit with two 384-well plates (Ready-to-Use Mouse & Rat Panel I+II V3, Cat. No. 20379; Exiqon; www.exiqon.com ), which covered 752 murine miRNAs, was used with quantitative real-time PCR (qRT-PCR) technology.

Techniques: Control, Expressing, Quantitative RT-PCR, Luciferase, Plasmid Preparation, Negative Control, Real-time Polymerase Chain Reaction

Journal: Biological Research

Article Title: MicroRNA-148b secreted by bovine oviductal extracellular vesicles enhance embryo quality through BPM/TGF-beta pathway

doi: 10.1186/s40659-024-00488-z

Figure Lengend Snippet: Mature miRNA forward primers (used with Qiagen miScript Universal Primer as reverse primer)

Article Snippet: The resulting aqueous layer was collected, and cDNA synthesis using the miRCURY LNA miRNA PCR Starter Kit (Qiagen #3,390,320) was performed following the manufacturer’s instructions.

Techniques: Sequencing, Control

Journal: Biological Research

Article Title: MicroRNA-148b secreted by bovine oviductal extracellular vesicles enhance embryo quality through BPM/TGF-beta pathway

doi: 10.1186/s40659-024-00488-z

Figure Lengend Snippet: MiR-148b mimics is taken up by bovine embryos and influences on embryo development. ( A ) Representative fluorescence images that illustrate the passage of miRNA mimics through the zona pellucida of 16-cell stage bovine embryos and blastocyst. Presumptive zygotes were cultured from 21 h post-insemination (hpi) until day 8 pi (CMimic) along with commercial triple-RNA-strand mimics prelabeled with 5′ FAM (5-carboxyfluorescein, green) or PBS (Control). Zona-intact bovine embryos at 16-cell stage and blastocyst (5′FAM-labeled or PBS) were washed, fixed, and stained with Hoechst (blue) to visualize the nuclei. The merged images demonstrate uptake of green, fluorescent–labeled mimics by 16-cell stage embryos and blastocyst. ( B ) Relative expression level of miR-148b of bovine embryo cultured from Day 1 until Day 7 pi with or without miR-148b mimics or control mimics. ( C ) Developmental rates of in vitro produced bovine embryos cultured from Day 1 until Day 7 p.i. with or without miR-148b mimics or control mimics. Results are expressed as mean ± s.e.m. Significant differences ( P < 0.001) are indicated with different letters

Article Snippet: The resulting aqueous layer was collected, and cDNA synthesis using the miRCURY LNA miRNA PCR Starter Kit (Qiagen #3,390,320) was performed following the manufacturer’s instructions.

Techniques: Fluorescence, Cell Culture, Control, Labeling, Staining, Expressing, In Vitro, Produced

Journal: Biological Research

Article Title: MicroRNA-148b secreted by bovine oviductal extracellular vesicles enhance embryo quality through BPM/TGF-beta pathway

doi: 10.1186/s40659-024-00488-z

Figure Lengend Snippet: Embryo quality assessment of bovine embryos after IVC with or without miRNA-148b mimic or control mimics

Article Snippet: The resulting aqueous layer was collected, and cDNA synthesis using the miRCURY LNA miRNA PCR Starter Kit (Qiagen #3,390,320) was performed following the manufacturer’s instructions.

Techniques: Control

Journal: Biological Research

Article Title: MicroRNA-148b secreted by bovine oviductal extracellular vesicles enhance embryo quality through BPM/TGF-beta pathway

doi: 10.1186/s40659-024-00488-z

Figure Lengend Snippet: pSMAD1/5 phosphorylation level in blastocysts after IVC with or without miRNA-148b mimic. ( A ) Blastocyst analyzed for localization and abundance of phosphorylated (active) form of SMAD1/5 (pSMAD1/5) transcription factor. pSMAD1/5 Immunostaining (green) counterstained with Hoechst 33,342 for nuclear staining (blue). Embryos cultured from presumptive zygotes to blastocyst in SOF + 0.3% BSA (Control), supplemented with 1 µM control mimics (CMimic), or supplemented with 1 µM of miR-148b (miR148b) or embryos cultured from presumptive zygotes to 16- cell stage in SOF + 0.3% BSA supplemented with 1 µM of miR-148b (miR148b-OV), or embryos cultured from 16-cell stage to blastocyst in SOF + 0.3% BSA supplemented with 1 µM of miR-148b (miR148b-UT). ( B and C ) Quantification of phosphorylation levels and representative WB images of pSMAD1/5 signal from each group. WB data were normalized relative to the abundance of β-actin as a loading control and expressed as mean ± SEM. Values with different superscripts indicate statistically significant differences at ( P ≤ 0.05)

Article Snippet: The resulting aqueous layer was collected, and cDNA synthesis using the miRCURY LNA miRNA PCR Starter Kit (Qiagen #3,390,320) was performed following the manufacturer’s instructions.

Techniques: Phospho-proteomics, Immunostaining, Staining, Cell Culture, Control