Journal: Bioscience Reports

Article Title: Bovine tongue epithelium-derived cells: A new source of bovine mesenchymal stem cells

doi: 10.1042/BSR20181829

Figure Lengend Snippet: List of primers used for reverse transcription-polymerase chain reaction

Article Snippet: To identify stem cell surface markers, cells (5 × 10 5 ) were stained with mouse anti-bovine CD29 (Kingfisher Biotech, WS0577B, U.S.A.), phycoerythrin (PE)–conjugated mouse anti-bovine CD44 (AbD serotec, MCA2433PE, U.K.), Fluorescein isothiocyanate (FITC)–conjugated mouse anti-bovine CD45 (AbD serotec, MCA832F, U.K.), FITC–conjugated mouse anti-human CD90 (Novusbio, NBP2-47755F, U.S.A.), mouse anti-bovine major histocompatibility complex (MHC) class I (MHC-I; Kingfisher Biotech, WS0558B, U.S.A.), PE–conjugated mouse anti-bovine MHC-II (Mybiosource, MBS224588, U.S.A.), FITC–conjugated mouse anti-bovine CD11b (Bio-Rad, MCA1425F, U.K.), FITC–conjugated mouse anti-bovine CD80 (Bio-Rad, MCA2436F, U.K.), and FITC–conjugated mouse anti-bovine CD86 (Bio-rad, MCA2437F, UK) for 30 min at 4°C.

Techniques: Sequencing

Journal: Bioscience Reports

Article Title: Bovine tongue epithelium-derived cells: A new source of bovine mesenchymal stem cells

doi: 10.1042/BSR20181829

Figure Lengend Snippet: ( A ) Cell surface markers (positive: CD29, CD44, CD90, and MHC-I; negative: CD45, CD11b, CD80, CD86, and MHC-II) of boT-MSCs were observed (P3) by flow cytometry analysis. Data are expressed as means ± standard error of values, and were obtained by three determinations. ( B ) Cell surface markers (CD13, CD14, CD29, CD31, CD34, CD44, CD45, CD73, CD90, CD105, CD106, CD116, and MHC-II), and ( C ) pluripotency markers (Sox2 and Oct3/4) of boT-MSCs were observed (P3) by qRT-PCR. GAPDH was used as a housekeeping control gene. Results are shown as means ± standard error ( n =3) (* P <0.05, ** P <0.001 and *** P <0.0001).

Article Snippet: To identify stem cell surface markers, cells (5 × 10 5 ) were stained with mouse anti-bovine CD29 (Kingfisher Biotech, WS0577B, U.S.A.), phycoerythrin (PE)–conjugated mouse anti-bovine CD44 (AbD serotec, MCA2433PE, U.K.), Fluorescein isothiocyanate (FITC)–conjugated mouse anti-bovine CD45 (AbD serotec, MCA832F, U.K.), FITC–conjugated mouse anti-human CD90 (Novusbio, NBP2-47755F, U.S.A.), mouse anti-bovine major histocompatibility complex (MHC) class I (MHC-I; Kingfisher Biotech, WS0558B, U.S.A.), PE–conjugated mouse anti-bovine MHC-II (Mybiosource, MBS224588, U.S.A.), FITC–conjugated mouse anti-bovine CD11b (Bio-Rad, MCA1425F, U.K.), FITC–conjugated mouse anti-bovine CD80 (Bio-Rad, MCA2436F, U.K.), and FITC–conjugated mouse anti-bovine CD86 (Bio-rad, MCA2437F, UK) for 30 min at 4°C.

Techniques: Flow Cytometry, Quantitative RT-PCR, Control

Journal: Theranostics

Article Title: Carcinoma-associated fibroblasts promote the stemness and chemoresistance of colorectal cancer by transferring exosomal lncRNA H19

doi: 10.7150/thno.25541

Figure Lengend Snippet: H19 activates the β-catenin pathway by acting as a competing endogenous RNA sponge for miR-141 in colorectal cancer. (A) H19 was overexpressed by transfection with the pcDNA3.1-H19 plasmid, and luciferase experiments with Top-Luc Flash reporter and pRL-TK (as an internal control) were performed to analyze the Wnt/β-catenin pathway. (B) Western blotting assessed the level of β-catenin in SW480 and HCT116 cells after indicated treatments. (C) The expression of the downstream genes of the Wnt/β-catenin pathway (c-myc, CCND1 and CD44) was analyzed qRT-PCR. (D) SW480 cells were treated with exosomes (10 µg/mL) derived from CAFs or NFs for 48 h, and Western blotting analyzed the protein level of β-catenin. (E) Immunohistochemistry analysis of β-catenin levels in two xenograft tumor tissues in Figure and Figure . Scale bar, 50 μm. (F) Immunoprecipitation using anti-AGO2 antibody (lane 3) or IgG (lane 2) followed by Western blot analysis. Immunoprecipitated RNA was isolated by TRIzol reagent, and the level of H19 was analyzed by qRT-PCR. (G) The putative binding sites between H19 and miR-141 were predicted. (H) HEK293T cells were co-transfected with luciferase plasmids (psicheck2 vector, psicheck2-H19-wt, psicheck2-H19-mut) and miR-141 mimics or mimic controls, and dual-luciferase reporter gene assays were performed. (I) The mRNA levels of β-catenin were analyzed by qRT-PCR. The SW480 cells were transfected with empty vector (pcDNA3.1), H19-overexpressing plasmids (pcDNA3.1-H19), miR-NC (mimic control) or miR-141 (mimic), and mRNA levels of β-catenin were analyzed by qRT-PCR. (J) The protein levels of β-catenin were analyzed by Western blotting in SW480 cells transfected with plasmids overexpressing H19 (pcDNA3.1-H19), an empty vector (pcDNA3.1), miR-NC (mimic control) or miR-141 (mimic). (K) Representative immunostaining graphs of β-catenin expression in H19-high and H19-low CRC tissues. Scale bar, 200 μm. (L) Correlation between relative H19 level and β-catenin immunostaining scores in CRC tissues with linear regression lines. Data are shown as the means ± SD from three independent experiments. Error bars, SD. *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: The protein concentrations were tested by BCA Protein Assay Kit (Pierce, USA). β-catenin, vimentin, α-SMA, HSP70, CD63, GM130, Sox2, Oct4, Nanog, CD44, CD133 and GAPDH antibodies were used in the study. β-catenin, α-SMA, HSP70, CD63, and GM130 antibodies (1:1000) were purchased from Cell Signaling Technology (MA, USA).

Techniques: Transfection, Plasmid Preparation, Luciferase, Control, Western Blot, Expressing, Quantitative RT-PCR, Derivative Assay, Immunohistochemistry, Immunoprecipitation, Isolation, Binding Assay, Immunostaining

Journal: Cancers

Article Title: CD44 Promotes Lung Cancer Cell Metastasis through ERK–ZEB1 Signaling

doi: 10.3390/cancers13164057

Figure Lengend Snippet: CD44 is overexpressed in primary tumors and lymph nodes, and is correlated with poor survival. ( A ) Kaplan–Meier analysis of OS, according to the expression of CD44 mRNA by using lung cancer online microarray datasets software. ( B ) Representative images of IHC for CD44 expression in primary tumors and lymph node metastases ( left ) and bar graph showing the expression of CD44 in primary tumors and metastatic lymph nodes of lung cancer patients ( right ). ( C ) Flow cytometry results showing the percentage of CD44 positive cells in primary tumors and metastatic lymph nodes analyzed by flow cytometer ( left ) and bar figure showing the average of CD44 positive cells in primary lung tumors and metastatic lymph nodes ( right ). * p < 0.05.

Article Snippet: Antibodies for CD44, α-tubulin, p-ERK 1/2, ERK, ZEB1, and Claudin-1 were purchased from GeneTex (US).

Techniques: Expressing, Microarray, Software, Flow Cytometry

Journal: Cancers

Article Title: CD44 Promotes Lung Cancer Cell Metastasis through ERK–ZEB1 Signaling

doi: 10.3390/cancers13164057

Figure Lengend Snippet: CD44 knockdown reduced migration and invasion abilities of lung cancer cells in vitro. ( A ) Expression of CD44 in different lung cancer cell lines and quantification of western data. ( B ) CD44 knockdown efficiency of different lentivirus clones and quantification of western data. ( C ) CD44 knockdown efficiency in A549 and H1299 cells determined by Western blot ( left ), and effects of CD44 knockdown on Transwell migration ( middle ) and invasion ( right ) abilities of A549 and H1299 cells. ( D ) CD44 overexpression efficiency in H441 and CL1-0 cells ( left ), and effects of CD44 overexpression on Transwell migration ( middle ) and invasion ( right ) abilities of H441 and CL1-0 cells. * p < 0.05, ** p < 0.01.

Article Snippet: Antibodies for CD44, α-tubulin, p-ERK 1/2, ERK, ZEB1, and Claudin-1 were purchased from GeneTex (US).

Techniques: Knockdown, Migration, In Vitro, Expressing, Western Blot, Clone Assay, Over Expression

Journal: Cancers

Article Title: CD44 Promotes Lung Cancer Cell Metastasis through ERK–ZEB1 Signaling

doi: 10.3390/cancers13164057

Figure Lengend Snippet: CD44 expression has no significant effects on cell proliferation and cell cycle. ( A ) Cell viability analysis in A549 and H1299 cells with CD44 knockdown. ( B ) Cell viability analysis in H441 and CL1-0 cells with CD44 overexpression. ( C ) Cell cycle analysis in A549 and H1299 cells with CD44 knockdown.

Article Snippet: Antibodies for CD44, α-tubulin, p-ERK 1/2, ERK, ZEB1, and Claudin-1 were purchased from GeneTex (US).

Techniques: Expressing, Knockdown, Over Expression, Cell Cycle Assay

Journal: Cancers

Article Title: CD44 Promotes Lung Cancer Cell Metastasis through ERK–ZEB1 Signaling

doi: 10.3390/cancers13164057

Figure Lengend Snippet: CD44 promoted migration and invasion abilities of lung cancer cells through ERK–ZEB1 signaling. ( A ) Western blot showing the expression pattern of p-ERK, ERK, Claudin-1, and ZEB1 in CD44-knockdowned A549 and H1299 cells. ( B ) Western blot showing the expression pattern of p-ERK, ERK, Claudin-1, and ZEB1 in CD44-overexpressing H441 and CL1-0 cells. ( C ) Effect of ERK inhibitor (PD98059, 25 uM) treatment for 48 h on migration and invasion abilities of CD44-overexpressing H441 and CL1-0 cells. ( D ) Western blot showing the expression pattern of p-ERK, ERK, Claudin-1, and ZEB1 in CD44-overexpressing H441 and CL1-0 cells after ERK inhibitor PD98059 (25 uM for 48 h) treatment. * p < 0.05, ** p < 0.01.

Article Snippet: Antibodies for CD44, α-tubulin, p-ERK 1/2, ERK, ZEB1, and Claudin-1 were purchased from GeneTex (US).

Techniques: Migration, Western Blot, Expressing

Journal: Cancers

Article Title: CD44 Promotes Lung Cancer Cell Metastasis through ERK–ZEB1 Signaling

doi: 10.3390/cancers13164057

Figure Lengend Snippet: CD44 knockdown decreased lung cancer cell metastasis in tail-vein injection mouse model. ( A ) H & E staining showing representative photos for positive and negative metastatic nodules in the lung. ( B ) Bar graph showing the rate of lung metastasis in control and CD44 knockdown groups.

Article Snippet: Antibodies for CD44, α-tubulin, p-ERK 1/2, ERK, ZEB1, and Claudin-1 were purchased from GeneTex (US).

Techniques: Knockdown, Injection, Staining, Control

Journal: Oncoimmunology

Article Title: Loss of p27Kip1 leads to expansion of CD4+ effector memory T cells and accelerates colitis-associated colon cancer in mice with a T cell lineage restricted deletion of Smad4

doi: 10.1080/2162402X.2020.1847832

Figure Lengend Snippet: The population of effector memory CD4 + cells in each genotype . A) Analysis of total spleen cells and CD4 + T cells of each genotype at 4 months of age. Cell suspensions were prepared from spleens by filtering through nylon mesh. The cell numbers were counted by trypan blue exclusion assay. The population and number of CD4 T cells in each genotype. Splenocytes were stained with antibodies for CD4 and CD8. Events are gated on the live lymphocyte gate, based on forward and side light scatter (FSC X SSC) by FACS analysis. Bar graphs show the number of spleen cells and the proportion and absolute number of CD4 T cells, respectively. B) Analysis of memory markers (CD44 High and CD62L Low ) in CD4 + T cell compartment of each genotype. The bar graphs show the proportion and absolute number of CD4 T EM cells from the same experiment depicted in Supplementary Figure 2B. C) Analysis of effector CD4 + T cells expressing pro-inflammatory cytokines of each genotype. Splenocytes per genotype were collected, stained on cell surface with antibody for CD4 and intracellularly with IFN-γ or TNF-γ antibody, and analyzed on CD4 + T cells by FACS. The bar graphs are from the same experiment depicted in Supplementary Figure 2 C. D) IHC staining for CD3 + T cells in colon of each genotype. E) Colon lamina propria per each genotype at 4 months of age were collected, stained with antibodies for CD4 and CD8, and analyzed for effector memory markers (CD44 High and CD62L Low ) in CD4 + T cell compartment. Bar graphs show the proportion and absolute number of CD4 T EM cells and effector CD4 T cells producing IFN-γ, respectively. ***P < .001, **P < .01, *P < .05. Scale bar = 200 µm

Article Snippet: Anti-CD3, anti-CD28, anti-CD44, anti-CD62L, anti-IFN-γ, and anti-TNF-α were purchased from BD Biosciences.

Techniques: Trypan Blue Exclusion Assay, Staining, Expressing, Immunohistochemistry