Journal: The FASEB Journal

Article Title: Extracellular vesicles extracted from young donor serum attenuate inflammaging via partially rejuvenating aged T-cell immunotolerance

doi: 10.1096/fj.201800059R

Figure Lengend Snippet: Characteristics of serum-extracted EVs. A) Comparison of the sizes of EVs extracted from young murine serum by the ExoQuick reagent pretreated with and without using 0.2-μm filters. B) Morphology of EVs from young murine serum used in this project for rejuvenation of inflammaging, photographed by atomic force microscopy (AFM). C, D) Different miRNA expression profiles in heatmap (C) and quantified summary (D) of EVs from young vs. old murine serum, analyzed by murine miRNA microarray with Mus musculus miRBase version-21 arrays that contained 1900 unique mature miRNA probes (miRNA microarray service via LC Sciences).

Article Snippet: Then, 1–3 μg of total RNAs for each sample were used for customized miRNA microarray service from LC Sciences (Houston, TX, USA).

Techniques: Comparison, Microscopy, Expressing, Microarray

Journal: Gene

Article Title: Comparative profiling of miRNA expression of lung adenocarcinoma cells in two-dimensional and three-dimensional cultures

doi: 10.1016/j.gene.2012.09.093

Figure Lengend Snippet: Distinct miRNA profiles of mK-ras-LE cells in 2-D and rBM 3-D cultures. A) mK-ras-LE cells were grown in 2-D and rBM 3-D culture. Morphology of the cells was visualized using fluorescent staining for F-actin. The images of rBM 3-D culture were captured at the central planes of cell clusters using a confocal fluorescent microscope on day 12 post-seeding. The images were captured at 600x magnification. Representative images of each culture condition were displayed. B) Total cell RNA was extracted from mK-ras-LE cells in 2-D and rBM 3-D cultures. miRNA expression profiles were compared between the two culture conditions using miRNA microarrays. A heatmap of the significantly differentially expressed miRNAs was generated using geWorkbench suite (n = 3, P < 0.01, FDR < 0.05). C) The expression of miR-17, miR-92b, and miR-21 was measured using qRT-PCR in total cell RNA collected from 2-D and rBM 3-D cultures of mK-ras-LE cells. A fold change of each miRNA of interest was obtained by setting the values of each miRNA in 2-D culture to one and normalization to the values of U6. D) Similar to part C except that the expression of miR-200a was compared between 2-D and rBM 3-D cultures. E) The culture conditions were similar to part A. The cell lysates were collected from the cell colonies extracted from the culture. The protein levels of ZEB2, PTEN, and -actin were determined using immunoblots. The data were presented in averages and standard deviations obtained from three independent experiments. and indicated a P value < 0.05 and 0.01, respectively.

Article Snippet: The miRNA expression profiling service was provided by LC Sciences (Houston, TX) using μParaflo® technology and proprietary probe hybridization (Cy3 and Cy5 dendrimer dyes) that enables highly sensitive and specific direct detection of mouse miRNAs.

Techniques: Staining, Microscopy, Expressing, Generated, Quantitative RT-PCR, Western Blot

Journal: Gene

Article Title: Comparative profiling of miRNA expression of lung adenocarcinoma cells in two-dimensional and three-dimensional cultures

doi: 10.1016/j.gene.2012.09.093

Figure Lengend Snippet: miRNA expression associated with aincar morphogenesis. A) A549 and A549LC cells were cultured in 2-D (A1 & A2) and rBM 3-D (A3 & A4) cultures. Morphology was recorded using an inverse phase contrast microscope. Polarized monolayer of the cells and the central lumen were pointed at by white arrows. B) The expression of miR-17, miR-92, and miR-21 was measured using qRT-PCR in total cell RNA collected from 2-D and rBM 3-D of A549 and A549LC cells. A fold change of each miRNA was obtained by setting the values of each miRNA from A549 cells in 2-D culture to one and normalization to the values of U6. C) Similar to part A except that the expression of miR-200a was compared across the groups. D) The culture conditions were similar to part A. The RNA levels of pri-miR-21 and VMP1 were measured in A549 cells using qRT-PCR. A fold change of each RNA was obtained by setting the values of RNA from A549 cells in 2-D culture to one and normalization to the values of 36B4. The data were presented in averages and standard deviations obtained from three independent experiments. , , and indicated a P value < 0.05, 0.01, and 0.001, respectively.

Article Snippet: The miRNA expression profiling service was provided by LC Sciences (Houston, TX) using μParaflo® technology and proprietary probe hybridization (Cy3 and Cy5 dendrimer dyes) that enables highly sensitive and specific direct detection of mouse miRNAs.

Techniques: Expressing, Cell Culture, Microscopy, Quantitative RT-PCR

Journal: International Journal of Molecular Sciences

Article Title: Downregulation of miR-140-3p Contributes to Upregulation of CD38 Protein in Bronchial Smooth Muscle Cells

doi: 10.3390/ijms21217982

Figure Lengend Snippet: List of differentially expressed microRNAs (miRNAs) in bronchial smooth muscle of antigen-induced airway hyperresponsive (AHR) mice.

Article Snippet: miRNA microarray services were contracted out to Cosmo Bio Co., Ltd. (Tokyo, Japan).

Techniques: Sequencing, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Downregulation of miR-140-3p Contributes to Upregulation of CD38 Protein in Bronchial Smooth Muscle Cells

doi: 10.3390/ijms21217982

Figure Lengend Snippet: Downregulation of miR-140-3p in bronchial smooth muscles of antigen-induced airway hyperresponsive (AHR) mice. The ovalbumin (OA)-immunized animals were repeatedly challenged with aerosolized OA solution, and total RNAs including miRNAs were extracted from the main bronchi 24 h after the last challenge. The miR-140-3p levels were determined by quantitative real-time reverse transcriptase-polymerase chain reaction. The relative expression of miR-140-3p to U6 snRNA was calculated by the 2 − ΔΔCT methods as described in the methods section. Results are presented as mean ± S.E. from six animals, respectively. ** p < 0.01 versus control animal (Cont) by unpaired Student’s t -test.

Article Snippet: miRNA microarray services were contracted out to Cosmo Bio Co., Ltd. (Tokyo, Japan).

Techniques: Muscles, Reverse Transcription, Polymerase Chain Reaction, Expressing, Control

Journal: International Journal of Molecular Sciences

Article Title: Downregulation of miR-140-3p Contributes to Upregulation of CD38 Protein in Bronchial Smooth Muscle Cells

doi: 10.3390/ijms21217982

Figure Lengend Snippet: Upregulation of CD38 protein ( A ) but not mRNA ( B ) in bronchial smooth muscles of antigen-induced airway hyperresponsive (AHR) mice. The ovalbumin (OA)-immunized animals were repeatedly challenged with aerosolized OA solution, and proteins and total RNAs including miRNAs were extracted from the main bronchi 24 h after the last challenge. The protein ( A ) and mRNA ( B ) levels of CD38 were determined by immunoblotting and quantitative real-time reverse transcriptase-polymerase chain reaction, respectively. The relative expressions of CD38 to glyceraldehyde 3-phosphate dehydrogenase (GAPDH; ( A , B )) were calculated as described in the methods section. Results are presented as mean ± S.E. from six animals, respectively. ** p < 0.01 versus control animal (Cont) by unpaired Student’s t -test.

Article Snippet: miRNA microarray services were contracted out to Cosmo Bio Co., Ltd. (Tokyo, Japan).

Techniques: Muscles, Western Blot, Reverse Transcription, Polymerase Chain Reaction, Control

Journal: International Journal of Molecular Sciences

Article Title: Downregulation of miR-140-3p Contributes to Upregulation of CD38 Protein in Bronchial Smooth Muscle Cells

doi: 10.3390/ijms21217982

Figure Lengend Snippet: Effects of interleukin-13 (IL-13) on the expressions of miR-140-3p ( A ) and CD38 mRNA ( B ) in cultured human bronchial smooth muscle cells. Cells were treated with IL-13 (100 ng/mL) or its vehicle (Veh) for 24 h and total RNAs including miRNAs were extracted. The gene expressions were determined by quantitative real-time reverse transcriptase-polymerase chain reaction. The relative gene expressions of miR-140-3p to U6 snRNA ( A ) and CD38 to GAPDH mRNAs ( B ) were calculated by the 2 − ΔΔCT methods as described in the methods section. Results are presented as mean ± S.E. from six independent experiments. ** p < 0.01 versus Veh by unpaired Student’s t -test.

Article Snippet: miRNA microarray services were contracted out to Cosmo Bio Co., Ltd. (Tokyo, Japan).

Techniques: Cell Culture, Reverse Transcription, Polymerase Chain Reaction