Journal: Gene therapy

Article Title: The use of polyethylenimine-DNA to topically deliver hTERT to promote hair growth.

doi: 10.1038/gt.2011.62

Figure Lengend Snippet: Figure 1 Flow cytometric analysis and fluorescence detection of sorted cells double stained for integrin b1 and CD 71. The integrin b1CD71 (a) and integrin b1+CD71 (b) cells were collected by FACS and examined by flow cytometry, respectively. Integrin b1CD71 cells are shown in the lower left-hand quadrant of each contour plot, whereas integrin b1+CD71 cells are shown in the lower right-hand quadrant, with percentage of each sorted cell population. Photo images of integrin b1CD71 (c) and integrin b1+CD71 cells (d). Integrin b1 was obvious by green fluorescence and CD71 was obvious by red fluorescence. Nuclei were counterlabeled with 4,6-diamidino-2-phenylindole (DAPI) (scale bar¼10 mm). The color reproduction of this figure is available on the html full text version of the manuscript.

Article Snippet: Stem cells, which expressed integrin a6, were detected by staining with anti-integrin a6 mouse monoclonal antibody (sc-59920; Santa Cruz Biotechnology, Inc.) followed by anti-mouse secondary antibody conjugated with Alexa Fluor 647 (A21235; Invitrogen) and with anti-hTERT with anti-rabbit secondary antibody conjugated with fluorescein isothiocyanate.

Techniques: Staining, Cytometry

Journal: Gene therapy

Article Title: The use of polyethylenimine-DNA to topically deliver hTERT to promote hair growth.

doi: 10.1038/gt.2011.62

Figure Lengend Snippet: Figure 3 hTERT expression in DNA–PEI complex transfected b1+CD71 keratinocytes increased expression of cell proliferation marker. (a–d) Detection of hTERT expression was detected by immunofluorescent staining (Allophycocyanin, APC) in integrin b1+CD71 keratinocytes after (a) treatment of water (Neg), (b) naked pLC–hTERT plasmid (DNA), (c) PEI vector only (PEI) and (d) PEI–pLC–hTERT complex at N/P¼7 (N/P). Panels (e–h) showed that immunofluorescent staining of PCNA (Allophycocyanin, APC) after treatment of water (e), DNA (f), PEI (g) and hTERT–DNA–PEI complex (h). Sections stained with antibodies that were color-coded according to the fluorescent tags of the secondary antibodies. Western blot analyses of hTERT (i) and PCNA (j) expression in cells transfected with the different treatments indicated on the top (Neg, water; DNA, pLC–hTERT plasmid; P7, PEI at molar ratio¼7; P15, PEI at molar ratio¼15; N/P7, hTERT–DNA–PEI complex at N/P ratio¼7; N/P15, hTERT–DNA–PEI complex at N/P ratio¼15). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as a control for normalization of the total protein amounts. The color reproduction of this figure is available on the html full text version of the manuscript.

Article Snippet: Stem cells, which expressed integrin a6, were detected by staining with anti-integrin a6 mouse monoclonal antibody (sc-59920; Santa Cruz Biotechnology, Inc.) followed by anti-mouse secondary antibody conjugated with Alexa Fluor 647 (A21235; Invitrogen) and with anti-hTERT with anti-rabbit secondary antibody conjugated with fluorescein isothiocyanate.

Techniques: Expressing, Transfection, Marker, Staining, Plasmid Preparation, Western Blot, Control

Journal: European Journal of Medical Research

Article Title: RNA interference targeting human integrin α6 suppresses the metastasis potential of hepatocellular carcinoma cells

doi: 10.1186/2047-783X-18-52

Figure Lengend Snippet: HepG2 and Bel-7402 cells were infected with integrin α6 short hairpin RNA (shRNA) plasmids and control shRNA plasmids respectively. Stable transfectant clones were picked up by puromycin. Integrin α6 expression levels remarkably reduced. (A) Western blot assays yielded the expression level of integrin α6 on protein level in HepG2 and Bel-7402 cell lines. (B) The qRT-PCR assays showed the expression level of integrin α6 on mRNA level in HepG2 and Bel-7402 cell lines. * P < 0.05; *** P < 0.001 compared with control group.

Article Snippet: Monoclonal rabbit anti-human CD49f (1:100, Abgent, San Diego, CA, USA), rabbit anti-human AKT (1:200); rabbit anti-human p-AKT (1:200); rabbit anti-human ERK (1:200); and rabbit anti-human p-ERK (1:200); antibody (these last four supplied by Santa Cruz Biotechnology, Inc., CA, USA), were used to detect the expression of integrin α6, AKT, p-AKT, ERK, and p-ERK respectively.

Techniques: Infection, shRNA, Control, Transfection, Clone Assay, Expressing, Western Blot, Quantitative RT-PCR

Journal: European Journal of Medical Research

Article Title: RNA interference targeting human integrin α6 suppresses the metastasis potential of hepatocellular carcinoma cells

doi: 10.1186/2047-783X-18-52

Figure Lengend Snippet: Cells were incubated in 96-well plates coated with laminin (LN) or fibronectin (FN) respectively for 48 hours. The MTT assays showed that the lower-level expression of integrin α6 caused decreased proliferation of HCC cells with significant statistical difference ( P < 0.05) in the cells incubated on LN, while the cells incubated on FN did not show any statistically significant effect ( P > 0.05). * P < 0.05, **** P < 0.0001 compared with control group.

Article Snippet: Monoclonal rabbit anti-human CD49f (1:100, Abgent, San Diego, CA, USA), rabbit anti-human AKT (1:200); rabbit anti-human p-AKT (1:200); rabbit anti-human ERK (1:200); and rabbit anti-human p-ERK (1:200); antibody (these last four supplied by Santa Cruz Biotechnology, Inc., CA, USA), were used to detect the expression of integrin α6, AKT, p-AKT, ERK, and p-ERK respectively.

Techniques: Incubation, Expressing, Control

Journal: European Journal of Medical Research

Article Title: RNA interference targeting human integrin α6 suppresses the metastasis potential of hepatocellular carcinoma cells

doi: 10.1186/2047-783X-18-52

Figure Lengend Snippet: Cells were cultured in Transwell chambers with the upper part coated with laminin (LN) or fibronectin (FN) respectively for 48 hours. The invaded cells were numerated under a light microscope (×400). (A) HepG2 infected by control shRNA plasmids invaded LN. (B) HepG2 cells infected by control shRNA plasmids invaded FN. (C) HepG2 cells infected by integrin α6 shRNA plasmids invaded LN. HepG2 had a decreased invasion potential after the knockdown of integrin α6 with significant statistical difference ( P < 0.0001). It also proved that LN can increase invasion potential of HepG2 ( P < 0.05).

Article Snippet: Monoclonal rabbit anti-human CD49f (1:100, Abgent, San Diego, CA, USA), rabbit anti-human AKT (1:200); rabbit anti-human p-AKT (1:200); rabbit anti-human ERK (1:200); and rabbit anti-human p-ERK (1:200); antibody (these last four supplied by Santa Cruz Biotechnology, Inc., CA, USA), were used to detect the expression of integrin α6, AKT, p-AKT, ERK, and p-ERK respectively.

Techniques: Cell Culture, Light Microscopy, Infection, Control, shRNA, Knockdown

Journal: European Journal of Medical Research

Article Title: RNA interference targeting human integrin α6 suppresses the metastasis potential of hepatocellular carcinoma cells

doi: 10.1186/2047-783X-18-52

Figure Lengend Snippet: Cells were cultured in 96-well plates coated with laminin (LN) or fibronectin (FN) respectively for 30 minutes and then cultured with acid phosphatase substrate at 37°C. The reaction was stopped two hours later. (A) The adhesion assays showed that lower-level integrin α6 reduced the adhesion of HCC cells with significantly statistical difference ( P < 0.0001) in the cells incubated with LN, while the cells incubated with FN did not show any significant effect ( P > 0.05). (B) Cells cultured in 96-well plates were coated with LN for 150 minutes. The spreading cells were numerated under a light microscope (×200). HepG2 cells with lower level of integrin α6 had a lower spreading rate. (C) The qRT-PCR assay showed that MMP-2 expression reduced significantly at mRNA level ( P < 0.01). (D) The qRT-PCR assay showed that MMP-9 expression reduced significantly at RNA level ( P < 0.01). * P < 0.05, ** P < 0.01, **** P < 0.0001 compared with control.

Article Snippet: Monoclonal rabbit anti-human CD49f (1:100, Abgent, San Diego, CA, USA), rabbit anti-human AKT (1:200); rabbit anti-human p-AKT (1:200); rabbit anti-human ERK (1:200); and rabbit anti-human p-ERK (1:200); antibody (these last four supplied by Santa Cruz Biotechnology, Inc., CA, USA), were used to detect the expression of integrin α6, AKT, p-AKT, ERK, and p-ERK respectively.

Techniques: Cell Culture, Incubation, Light Microscopy, Quantitative RT-PCR, Expressing, Control

Journal: European Journal of Medical Research

Article Title: RNA interference targeting human integrin α6 suppresses the metastasis potential of hepatocellular carcinoma cells

doi: 10.1186/2047-783X-18-52

Figure Lengend Snippet: Expression levels determination of PI3K/AKT and MAPK/ERK. (A) Western blot assay revealed the expression level of p-ERK was lower in HepG2 and Bel-7402 cells transfected with integrin α6 short hairpin RNA (shRNA) plasmids, while the expression of ERK remained unchanged. Cells transfected with control shRNA plasmids show no difference on p-ERK and ERK expression. (B) Western blot assay revealed the expression of p-AKT was lower in HepG2 and Bel-7402 cells transfected with integrin α6 shRNA plasmids, while the expression level of AKT remained unchanged. Cells transfected with control shRNA plasmids show no difference in p-AKT and AKT expression. (C) Western blot assay showed that PI3K inhibitor LY294002 reduced the expression of both p-AKT and p-ERK. (D) MTT assay showed that the proliferation of HepG2 cells cultured with LY294002 was dramatically reduced ( P < 0.05). (E) Transwell assay showed that the invasion potential of HepG2 cells cultured with LY294002 was dramatically reduced ( P < 0.05). * P < 0.05 compared with control.

Article Snippet: Monoclonal rabbit anti-human CD49f (1:100, Abgent, San Diego, CA, USA), rabbit anti-human AKT (1:200); rabbit anti-human p-AKT (1:200); rabbit anti-human ERK (1:200); and rabbit anti-human p-ERK (1:200); antibody (these last four supplied by Santa Cruz Biotechnology, Inc., CA, USA), were used to detect the expression of integrin α6, AKT, p-AKT, ERK, and p-ERK respectively.

Techniques: Expressing, Western Blot, Transfection, shRNA, Control, MTT Assay, Cell Culture, Transwell Assay