Journal: eLife

Article Title: A human ESC-based screen identifies a role for the translated lncRNA LINC00261 in pancreatic endocrine differentiation

doi: 10.7554/eLife.58659

Figure Lengend Snippet:

Article Snippet: Peptide, recombinant protein , KGF/FGF7 , R and D Systems , Cat# 251 KG , .

Techniques: Flow Cytometry, Control, Recombinant, Cloning, Plasmid Preparation, Expressing, Transfection, TA Cloning, Purification, cDNA Synthesis, SYBR Green Assay, Enzyme-linked Immunosorbent Assay, Bicinchoninic Acid Protein Assay, Software, Membrane, Hybridization, DNA Extraction, Modification

Journal: The Journal of investigative dermatology

Article Title: Differentiation of human embryonic stem cells into clinically amenable keratinocytes in an autogenic environment.

doi: 10.1038/jid.2012.384

Figure Lengend Snippet: Figure 1. Characterization of H9-ebF. (a, b) Qualitative analysis of gene expression profile of (a) pluripotency marker and (b) fibroblast markers. b-Actin was taken as the loading control and run on a separate gel. (c) Results of ELISA show the KGF-I secretion by P 5–P 10 of H9-ebF. Error bars indicate the mean ±SEM (n ¼ 3). *Po0.05 versus all groups. (d, e) Immunofluorescence staining shows the expression of (d) K18 and fibronectin by H9-ebF passage 8 (scale bar ¼ 100 mm), (e) K14 by HaCaT cells–H9-ebF passage 8 in coculture (scale bar ¼ 50mm). b-Act, b-actin; CRL-1486, human embryonic palatal mesenchymal cell line; DAPI, 4,6-diamidino-2-phenylindole; FAD, F12 and Dulbecco medium; H, H9-ebF; hESC, human embryonic stem cell; H9-ebF, H9-hESC-derived fibroblast; K, HaCaT cells; KGF-I, keratinocyte growth factor-I; P, passage; 4PB, prolyl 4-hydroxylase; PC, phase contrast; Vim, vimentin.

Article Snippet: ELISA was performed using the DuoSet ELISA Development System (catalog # DY251, R&D systems) as per the manufacturer’s instruc- tions.

Techniques: Gene Expression, Marker, Control, Enzyme-linked Immunosorbent Assay, Staining, Expressing, Derivative Assay

Journal: International Journal of Biological Sciences

Article Title: FGF7-induced E11 facilitates cell-cell communication through connexin43

doi: 10.7150/ijbs.65240

Figure Lengend Snippet: The high expression of Fgf7 of osteoblasts and the distribution of FGF7 in bone tissue. (A) RNA sequencing results showing the whole fibroblast growth factor family members and receptor members of primary osteoblasts. The data were presented as log (FPKM + 1) and showed the higher mRNA expression of Fgf7 among other FGF ligand members. FPKM: Fragments per Kilobase of transcript per Million fragments mapped. The results were based on three independent samples (n = 3). (B) IHC showing the distribution of FGF7 in the femur of 4 weeks wild-type C57 mice. Fast green is used to counterstain the background. Boxed areas further showing that FGF7 is partly generated by osteoblasts on the surface of trabecular bone within the secondary ossification center (boxed area 1) and subchondral bone (boxed area 2).

Article Snippet: In the treatment group, the medium was replaced with fresh 1% FBS α-MEM containing 10 ng/ml FGF7 (Recombinant Mouse KGF/FGF7, R&D Systems, Minnesota, USA, 5028-KG-025/CF).

Techniques: Expressing, RNA Sequencing, Generated

Journal: International Journal of Biological Sciences

Article Title: FGF7-induced E11 facilitates cell-cell communication through connexin43

doi: 10.7150/ijbs.65240

Figure Lengend Snippet: FGF7 promotes the expression of E11 in primary osteoblasts and MC3T3-E1 cell line. (A) RNA sequencing results showing the changes of E11 in response to FGF7 in 24 h. The data were presented as the ratio of FPKM to the inner beta-actin levels. FPKM: Fragments per Kilobase of transcript per Million fragments mapped. Cell samples from each group were obtained from three independent cell isolates (n = 3). (B) Western blotting showing the protein expression of E11 in primary osteoblasts and MC3T3-E1 cell line after treatment of FGF7 (10 ng/ml) for 24 h. beta-actin was used as the loading control. (C) Quantification of western blot analysis in (B). The results were based on three independent experiments (n = 3). * p < 0.05, ** p < 0.01. (D) Representative IF staining by CLSM showing the increased expression of E11 in primary osteoblasts and MC3T3-E1 cell line in response to FGF7 (10 ng/ml). F-actin (green) and Dapi (blue) were used to counterstain the background. The white boxed areas are showing the distribution of E11 in single cells. The yellow boxed areas are showing the distribution of E11 at cell junctions. The white arrows are revealing the details of the distribution of E11. (E) Quantification of E11 fluorescent intensity in (D) by Image J. The results were based on three independent experiments (n = 3). * p < 0.05. (F) Quantification further showing the changes in cell junction numbers induced by FGF7. The results were based on three independent experiments (n = 3). * p < 0.05. (G) Bright field showing the cell morphology at the edge of the scratch wound after FGF7 induction in a concentration-dependent manner for 36 h in MC3T3-E1 cell line and the white arrows showing the elongated osteoblasts cell processes.

Article Snippet: In the treatment group, the medium was replaced with fresh 1% FBS α-MEM containing 10 ng/ml FGF7 (Recombinant Mouse KGF/FGF7, R&D Systems, Minnesota, USA, 5028-KG-025/CF).

Techniques: Expressing, RNA Sequencing, Western Blot, Control, Staining, Concentration Assay

Journal: International Journal of Biological Sciences

Article Title: FGF7-induced E11 facilitates cell-cell communication through connexin43

doi: 10.7150/ijbs.65240

Figure Lengend Snippet: FGF7 promotes gap junction intercellular communication in primary osteoblasts and MC3T3-E1 cell line. (A) The scrape loading/dye transfer (SL/DT) assay showing functional GJIC in living primary osteoblasts and MC3T3-E1 cell line treated with FGF7 (10 ng/ml). The Lucifer yellow dye enters cells at the scratch (dotted white lines) and is transferred to cells distant from the scratch (green arrows). Intercellular gap junction transfer was calculated by measuring the distance from the scratching edge to the most distant cells with Lucifer yellow uptake. The boxed area further showing the different transmission speeds in response to FGF7. (B) Quantification showing different transmission speeds of Lucifer yellow in response to FGF7. The results were based on three independent experiments (n = 3). ** p < 0.01.

Article Snippet: In the treatment group, the medium was replaced with fresh 1% FBS α-MEM containing 10 ng/ml FGF7 (Recombinant Mouse KGF/FGF7, R&D Systems, Minnesota, USA, 5028-KG-025/CF).

Techniques: Functional Assay, Transmission Assay

Journal: International Journal of Biological Sciences

Article Title: FGF7-induced E11 facilitates cell-cell communication through connexin43

doi: 10.7150/ijbs.65240

Figure Lengend Snippet: FGF7 increases the expression of connexin 43 in primary osteoblasts and MC3T3-E1 cell line. (A) Western blotting showing the protein expression of Cx43 in primary osteoblasts and MC3T3-E1 cell line after treatment of FGF7 (10 ng/ml) for 24 h. beta-actin was used as the loading control. (B) Quantification of western blot analysis in (A). The results were based on three independent experiments (n = 3). ** p < 0.01. (C) Representative IF staining by CLSM showing the increased expression of Cx43 in primary osteoblasts and MC3T3-E1 cell line in response to FGF7 (10 ng/ml). F-actin (green) and Dapi (blue) were used to counterstain the background. The white boxed areas showing the distribution of Cx43 in single cells. The yellow boxed areas showing the distribution of Cx43 in cell junctions. The white arrows and dotted lines showing the details of the distribution of Cx43. (D) Quantification of Cx43 fluorescent intensity in (C) by Image J. The results were based on three independent experiments (n = 3). * p < 0.05. (E) Quantification further showing the changes in cell junction numbers induced by FGF7. The results were based on three independent experiments (n = 3). * p < 0.05.

Article Snippet: In the treatment group, the medium was replaced with fresh 1% FBS α-MEM containing 10 ng/ml FGF7 (Recombinant Mouse KGF/FGF7, R&D Systems, Minnesota, USA, 5028-KG-025/CF).

Techniques: Expressing, Western Blot, Control, Staining

Journal: International Journal of Biological Sciences

Article Title: FGF7-induced E11 facilitates cell-cell communication through connexin43

doi: 10.7150/ijbs.65240

Figure Lengend Snippet: E11 is involved in the FGF7-induced functional GJIC and up-regulation of Cx43. (A) The scrape loading/dye transfer (SL/DT) assay showing functional GJIC in living primary osteoblasts treated with FGF7 (10 ng/ml) in the presence or absence of E11 siRNA. The Lucifer yellow dye enters cells at the scratch (dotted white lines) and is transferred to cells distant from the scratch (green arrows). Intercellular gap junction transfer was calculated by measuring the distance from the scratching edge to the most distant cells with Lucifer yellow uptake. The boxed area further showing the different transmission speeds. (B) Quantification showing different transmission speeds of Lucifer yellow in (A). The results were based on three independent experiments (n = 3). *** p < 0.001, **** p < 0.0001. (C) Western blotting showing the protein expression of Cx43 in primary osteoblasts treated with FGF7 (10 ng/ml) in the presence or absence of E11 siRNA. (D) Quantification of western blot analysis in (C). The results were based on three independent experiments (n = 3). ***p < 0.001, **** p < 0.0001.

Article Snippet: In the treatment group, the medium was replaced with fresh 1% FBS α-MEM containing 10 ng/ml FGF7 (Recombinant Mouse KGF/FGF7, R&D Systems, Minnesota, USA, 5028-KG-025/CF).

Techniques: Functional Assay, Transmission Assay, Western Blot, Expressing

Journal: International Journal of Biological Sciences

Article Title: FGF7-induced E11 facilitates cell-cell communication through connexin43

doi: 10.7150/ijbs.65240

Figure Lengend Snippet: MAPK pathway and PI3K-AKT pathway are involved in the regulation of E11 and Cx43 in response to FGF7. (A, C) Western blotting showing ratios of P-ERK/ERK, P-JNK/JNK, P-P38/P38, and P-AKT/AKT increased in a concentration-dependent manner in primary osteoblasts and MC3T3-E1 cell line after treatment of FGF7 for 2 h. beta-actin was used as the loading control. The results were based on three independent experiments (n = 3). (B, D) Western blotting showing ratios of P-ERK/ERK, P-JNK/JNK, P-P38/P38, and P-AKT/AKT increased in a time-dependent manner in primary osteoblasts and MC3T3-E1 cell line after treatment of FGF7 (10 ng/ml) within 2 h. beta-actin was used as the loading control. The results were based on three independent experiments (n = 3). (E, F) Western blotting showing that the specific inhibitors attenuated the stimulation effect of FGF7 at E11 and Cx43 expression in primary osteoblasts and MC3T3-E1 cell line. (G-J) Quantification of western blot analysis in (E, F). The results were based on three independent experiments (n = 3). ** p < 0.01.

Article Snippet: In the treatment group, the medium was replaced with fresh 1% FBS α-MEM containing 10 ng/ml FGF7 (Recombinant Mouse KGF/FGF7, R&D Systems, Minnesota, USA, 5028-KG-025/CF).

Techniques: Western Blot, Concentration Assay, Control, Expressing