Journal: Experimental and Therapeutic Medicine

Article Title: miR-146b-5p promotes colorectal cancer progression by targeting TRAF6

doi: 10.3892/etm.2022.11155

Figure Lengend Snippet: miR-146b-5p is highly expressed in CRC. (A and B) RT-qPCR analysis of miR-146b-5p expression in CRC and normal adjacent tissue samples (n=19 pairs) and in CRC cell lines (n=3). (C and D) RT-qPCR analysis of miR-146-5p expression in HT29 and SW620 cells. (E and F) Colony formation assay in HT29 and SW620 cells. * P<0.05 and *** P<0.001. NC, negative control; miR, microRNA; RT-qPCR, reverse transcription quantitative PCR; CRC, colorectal cancer.

Article Snippet: The immortalized cell line NCM460 that was used as the normal control and the human CRC cell lines HT29, SW620, LoVo and HCT116 were purchased from the American Type Culture Collection (STR profiling authenticated).

Techniques: Quantitative RT-PCR, Expressing, Colony Assay, Negative Control, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Experimental and Therapeutic Medicine

Article Title: miR-146b-5p promotes colorectal cancer progression by targeting TRAF6

doi: 10.3892/etm.2022.11155

Figure Lengend Snippet: miR-146b-5p regulates the proliferation, migration, and invasion of HT29 and SW620 cells. HT29 and SW620 cells were transfected with the miR-146b-5p mimics or inhibitor. (A and B) Western blotting was used to determine the expression of cleaved-PARP and cleaved-caspase-3 in HT29 and SW620 cells. (C and D) Cell Counting Kit-8 assay was used to determine cell proliferation. (E and F) Transwell assay was used to evaluate the cell migration and invasion. Scale bar, 20 µm. (G and H) Quantification of Transwell assay from E and F. n=3. * P<0.05 vs. NC. NC, negative control; miR, microRNA; PARP, poly (ADP-ribose) polymerase.

Article Snippet: The immortalized cell line NCM460 that was used as the normal control and the human CRC cell lines HT29, SW620, LoVo and HCT116 were purchased from the American Type Culture Collection (STR profiling authenticated).

Techniques: Migration, Transfection, Western Blot, Expressing, Cell Counting, Transwell Assay, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: miR-146b-5p promotes colorectal cancer progression by targeting TRAF6

doi: 10.3892/etm.2022.11155

Figure Lengend Snippet: TRAF6 is the target of miR-146b-5p in CRC. (A) Correlation analysis between TRAF6 and miR-146b-5p expression in CRC tissue. (B) Schematic diagram showing the potential binding site between TRAF6 and miR-146b-5p. (C and D) Luciferase reporter assay of luciferase activities of TRAF6 3'-UTR-wt and TRAF6 3'-UTR-mut in HT29 and SW620 cells (n=3). (E and F) Western blot analysis of TRAF6 expression following transfection with miR-146b-5p mimics or inhibitor (n=3). * P<0.05 and ** P<0.01. NC, negative control; miR, microRNA; TRAF6, tumor necrosis factor receptor-associated factor 6; CRC, colorectal cancer; UTR, untranslated region; WT, wild-type; Mut, mutant.

Article Snippet: The immortalized cell line NCM460 that was used as the normal control and the human CRC cell lines HT29, SW620, LoVo and HCT116 were purchased from the American Type Culture Collection (STR profiling authenticated).

Techniques: Expressing, Binding Assay, Luciferase, Reporter Assay, Western Blot, Transfection, Negative Control, Mutagenesis

Journal: Experimental and Therapeutic Medicine

Article Title: miR-146b-5p promotes colorectal cancer progression by targeting TRAF6

doi: 10.3892/etm.2022.11155

Figure Lengend Snippet: Overexpression of TRAF6 abolishes the effects of miR-146b-5p in nude mice. (A) HT29 cells transfected with GFP-TRAF6 overexpression or GFP control vector cells subjected to western blotting analysis. (B) HT29 cells stably overexpressing GFP-TRAF6 or not were injected into nude mice. Tumor samples are presented. (C) Expression of miR-146b-5p was determined using reverse transcription-quantitative PCR. (D) Weights of xenograft tumors. (E) Volumes of xenograft tumors. n=3. * P<0.05 and ** P<0.01 vs. NC. # P<0.05 and ## P<0.01 vs. miR-146b-5p mimics. miR, microRNA; TRAF6, tumor necrosis factor receptor-associated factor 6; NC, negative control; GFP, green fluorescence protein; n.s. not significant.

Article Snippet: The immortalized cell line NCM460 that was used as the normal control and the human CRC cell lines HT29, SW620, LoVo and HCT116 were purchased from the American Type Culture Collection (STR profiling authenticated).

Techniques: Over Expression, Transfection, Control, Plasmid Preparation, Western Blot, Stable Transfection, Injection, Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Negative Control, Fluorescence

Journal: PLoS ONE

Article Title: Common Minor Histocompatibility Antigen Discovery Based upon Patient Clinical Outcomes and Genomic Data

doi: 10.1371/journal.pone.0023217

Figure Lengend Snippet: The iTopia epitope binding assay was used to measure T4A and the alternately encoded T4E peptide (GLYTYWSAGE) binding to HLA-A2 compared to the percentage of maximal binding of the iTopia control peptide (FLPSDFFPSV). T4A binding to HLA-A0201 is 95% of that compared to the positive control; however, T4E binds HLA-A0201 at 24% that of the positive control. According to the manufacturer, peptides binding of >30% that of the control peptide are candidate HLA-A0201 epitopes (4A). The iTopia assay was also used to measure the ED 50 and dissociation t 1/2 of T4A to HLA-A2 relative to the iTopia positive control (FLPSDFFPSV) labeled “POS”. T4A binds with a 50% effective dose of ED 50 = 1.3 µM and a dissociation half time of t 1/2 = 1.00 hr (4B, 4C). Western blots on whole cell lysates of normal human tissue (colon, heart, liver, skin, testis and PBMC), 4 human AML samples and Jurkat cells (positive control) are shown (4D). Western blotting of Jurkat cells reveals 2 bands of roughly 100 kDa (full length TRIM42) and 50 kDa (unknown protein product). The TRIM42 band is seen at various levels in all AML samples and faintly in PBMC. No TRIM42 protein is detected in the other human tissues. GAPDH expression (40 kDa) was used as a loading control. Subcellular fractionation was performed on AML blasts and isolated healthy donor granulocytes (4E). The fractions are labeled C, cytosolic; N, nuclear; M, membrane; Sk, cytoskeletal. No TRIM42 protein is observed in granulocytes, but the same bands seen in the Jurkat cell positive control are detected in the N and Sk fractions of the AML blasts. The allele frequencies and genotypes for rs9876490, the T4A associated cSNP, in the original ethnic populations used by the International HapMap Project (CEU, HCB, JPT, YRI) are shown (4F). The T4A recipient phenotypes (and frequencies in the CEU population) are AC (0.450) or CC (0.133), and the donor genotype is AA (0.417). The predicted gIR+ in an unrelated transplant scenario can be calculated: P(AA) × P(AC + CC) and applied over a range of allele frequencies yielding the MUD SCT curve (4G). When parental genotypes are considered in the MRD SCT setting (Supplement 1) a similar curve with generally lower gIR+ frequency at each donor allele frequency is generated (4F). In the case of rs9876490 the T4A gIR+ frequency observed in our cohort (•) fits the predicted MRD SCT curve well.

Article Snippet: Normal human tissue lysates (heart, testis, colon, skin, liver) prepared using RIPA buffer and SDS sample buffer were purchased from Prosci (Poway, CA).

Techniques: Binding Assay, Control, Positive Control, Itopia Assay, Labeling, Western Blot, Expressing, Fractionation, Isolation, Membrane, Generated