Journal: Cancer Research

Article Title: A lncRNA TCL6-miR-155 Interaction Regulates the Src-Akt-EMT Network to Mediate Kidney Cancer Progression and Metastasis

doi: 10.1158/0008-5472.can-20-0832

Figure Lengend Snippet: Figure 2. lncTCL6 is tumor suppressor with an antimetastatic role in RCC. To evaluate the functional significance of TCL6 in renal cancer, TCL6 gene was overexpressed in Caki-1/786-O cell lines by transfection, followed by selection of cells stably overexpressing TCL6 stably (TCL6-OE). For comparison, cells transfected with control plasmid (OE-control) were used. Later, these TCL6-OE cells were used to perform rescue experiments. A, Relative expression levels of TCL6 in stable TCL6-OE cells as compared with control cells. B, Relative expression levels of TCL6 in cells transfected with siTCL6 as compared with control cells. C, Proliferation of TCL6-OE stable Caki-1 (left) and 786-O (right) cells by MTS assay. D, Proliferation of TCL6-suppressed Caki-1 (left) and 786-O (right) cells by MTS assay. E, Colony formation assay (graphical representation) in TCL6-OE-stable cells (left) and cells transfected with siTCL6 compared with control cells (right). F, Apoptosis assay represented as a graph to show total apoptosis (early þ late) in stable TCL6-OE cells. G, Immunoblots showing apoptotic proteins in TCL6-OE Caki-1 and 786-O cells compared with control cells with GAPDH as endogenous control. H, Cell-cycle analysis showing significant increase in G2–M phase in TCL6-OE Caki-1 and 786-O cells compared with control cells. I, Cell-cycle analysis showing significant increase in S-phase in Caki-1 and 786-O cells with suppressed TCL6 expression compared with control cells. J and K, Transwell migration assays (J) and invasion assays (K) in TCL6-OE– and si-TCL6– treated Caki-1 and 786-O cells as compared with their respective controls. L, Immunoblot assay showing EMT-related proteins in Caki-1 and 786-O cells stably expressing TCL6 and transient transfected siTCL6 as compared with control cells. b-Actin was used as endogenous control. The numbers below the blot represent relative protein level, which was determined from band intensity using Image studio software of LI-COR and normalized relative to cells stably expressing control plasmid or treated with si-control. Graphs are mean of three independent experiments (n ¼ 3). P value was calculated by Student t test. Bar, mean SEM.

Article Snippet: Establishing stable anti-miR-155 and lncTCL6-overexpressing cell lines For lncTCL6 overexpression, cells were transfected with 10 mg of plasmid DNA, (TCL6 (NM_020550) Human Tagged ORF Clone), purchased from OriGene using JetPRIME (Polyplus Transfection).

Techniques: Functional Assay, Transfection, Selection, Stable Transfection, Comparison, Control, Plasmid Preparation, Expressing, MTS Assay, Colony Assay, Apoptosis Assay, Western Blot, Cell Cycle Assay, Migration, Software

Journal: Cancer Research

Article Title: A lncRNA TCL6-miR-155 Interaction Regulates the Src-Akt-EMT Network to Mediate Kidney Cancer Progression and Metastasis

doi: 10.1158/0008-5472.can-20-0832

Figure Lengend Snippet: Figure 4. miR-155 overexpression and TCL6 suppression increases invasiveness and proliferation of normal immortalized renal epithelial cell line, HK2. HK2 was treated with miRVANA miR-155 mimic/control mimic (miR-CON) as well as si-TCL6 and corresponding control, followed by functional assays (performed 72 hours posttransfection). A, Relative miR-155 expression as assessed by real-time PCR. B, Relative expression of TCL6 cells treated with siTCL6. C, Cell viability in HK2 cells after miR-CON/miR-155 mimic transfections and after si-control/si-TCL6 transfections as assessed by MTS assay at 72-hour timepoint. D, Colony formation assay shows increase in clonogenicity of HK2 cells post miR-155 overexpression and post TCL6 suppression as compared with control. E, Cell-cycle analysis showing significant increase in S-phase for the HK2 cells transfected with miR-155 mimic as well as the HK2 cells transfected with si-TCL6 as compared with cells transfected with control. F, Transwell migration assays and invasion assays in miR-CON/miR-155 mimic–transfected cells. G, Transwell migration assays and invasion assays in si- Control/si-TCL6 –transfected cells. H, Relative expression of EMT markers (fibronectin, Claudin, Src) in miR-155–overexpressing and TCL6-suppressed HK2 cells as compared with control cells. I, TCL6 expression in miR-155–overexpressing HK2 cells as compared with control cells. The bar graph (mean SEM) represents the data from three technical replicates, n ¼ 3. P value was calculated by Student t test.

Article Snippet: Establishing stable anti-miR-155 and lncTCL6-overexpressing cell lines For lncTCL6 overexpression, cells were transfected with 10 mg of plasmid DNA, (TCL6 (NM_020550) Human Tagged ORF Clone), purchased from OriGene using JetPRIME (Polyplus Transfection).

Techniques: Over Expression, Control, Functional Assay, Expressing, Real-time Polymerase Chain Reaction, Transfection, MTS Assay, Colony Assay, Cell Cycle Assay, Migration

Journal: Cancer Research

Article Title: A lncRNA TCL6-miR-155 Interaction Regulates the Src-Akt-EMT Network to Mediate Kidney Cancer Progression and Metastasis

doi: 10.1158/0008-5472.can-20-0832

Figure Lengend Snippet: Figure 5. LncTCL6 degrades Src mRNA by STAU1-mediated RNA degradation and thereby attenuates Src-Akt mediated EMT transition in renal carcinoma. A, Negative correlation between TCL6 and Src expression in KIRC-TCGA dataset. Spearman test was used to analyze the correlation. B–D, Relative expression of Src (mRNA) in cells with TCL6 overexpression (B), in TCL6-OE cells transfected with si-TCL6 (C), and in cells with stable miR-155 inhibition (D) as compared with their respective control cells. E–G, Immunoblot of Src in Caki-1 and 786-O cells stably overexpressing TCL6 (E), cells expressing anti-miR-155 (F), and cells transfected with si-TCL6 (G) as compared with respective control cells. H, RNA-immunoprecipitation using STAU1 antibody shows enrichment of TCL6 and Src as compared with control antibody (IgG) in TCL6-overexpressing cells. I, STAU1 expression in TCL6-OE cells transfected with siSTAU. J, Src mRNA expression in TCL6-OE cells transfected with siSTAU. K, Immunoblot showing expression of Src and EMT markers (fibronectin, vimentin, Claudin) in TCL6-OE cells transfected with siSTAU. L, Immunoblot showing suppression of phosphorylated Akt in cells stably expressing TCL6 and anti-miR-155 as compared with respective controls. M, Immunostaining of p--catenin (red) counterstained with DAPI (blue) in Caki-1 and 786-O cells expressing anti-miR-155 stably and overexpressing TCL6 as compared with respective control cells. Scale bar, 2 mm (bottom right). N, Immunoblot showing suppression of metastatic marker CD44 in cells stably expressing TCL6 as compared with respective controls. The numbers below the blot represent relative protein level, which was determined from band intensity using Image studio software of LI-COR and normalized relative to cells stably expressing control plasmid or treated with si-control. The bar graphs represent mean SEM from three sets of experiments (N ¼ 3). P value was calculated by Student t test. Immunoblot and immunofluorescence staining was performed for three and two biological replicates, respectively.

Article Snippet: Establishing stable anti-miR-155 and lncTCL6-overexpressing cell lines For lncTCL6 overexpression, cells were transfected with 10 mg of plasmid DNA, (TCL6 (NM_020550) Human Tagged ORF Clone), purchased from OriGene using JetPRIME (Polyplus Transfection).

Techniques: Expressing, Over Expression, Transfection, Inhibition, Control, Western Blot, Stable Transfection, RNA Immunoprecipitation, Immunostaining, Marker, Software, Plasmid Preparation, Staining

Journal: Journal of Medical Virology

Article Title: SARS‐CoV‐2 pseudovirus infectivity and expression of viral entry‐related factors ACE2, TMPRSS2, Kim‐1, and NRP‐1 in human cells from the respiratory, urinary, digestive, reproductive, and immune systems

doi: 10.1002/jmv.27244

Figure Lengend Snippet: Cell lines used in the present study

Article Snippet: BC‐3 , Primary effusion lymphoma , ATCC CRL‐2277 , RPMI‐1640 complete medium.

Techniques: Cell Culture

Journal: European journal of haematology

Article Title: Etodolac inhibits EBER expression and induces Bcl-2-regulated apoptosis in Burkitt's lymphoma cells.

doi: 10.1111/j.1600-0609.2005.00498.x

Figure Lengend Snippet: Fig. 7. RT-PCR analysis of EBER-1 and -2 mRNA in Burkitt’s lymphoma cells treated with etodolac. Daudi and Raji cells were treated with etodolac at the indicated con- centration for 8 h. The PCR products are 166 bp in size for EBER-1 (upper panel), 172 bp for EBER-2 (middle panel), and 245 bp for G3PDH (bottom panel). (1): Cells were cultured without agents, (2): cells were cultured with 50 lm etodolac, and (3): cells were cultured with 100 lm etodolac.

Article Snippet: Cell lines and cell culture Human Burkitt’s lymphoma cells, Daudi and Raji, were obtained from American Type Culture Collection (ATCC) (Rockville, MD, USA).

Techniques: Reverse Transcription Polymerase Chain Reaction, Cell Culture