Journal: PLoS ONE

Article Title: Opposing Functions of Akt Isoforms in Lung Tumor Initiation and Progression

doi: 10.1371/journal.pone.0094595

Figure Lengend Snippet: (A) Representative western blot analysis of 80 ug of total protein extracted from the lungs of 8-week-old C57BL/6 mice probed with a panel of Akt isoform-specific antibodies (Akt1, Akt2 and Akt3) as well as a pan-Akt antibody. An anti-actin antibody was used to demonstrate equal loading. Immunohistochemical staining of 8-week-old C57BL/6 mouse lungs with Akt1 (B and C), Akt2 (D and E) and Akt3 (F and G) specific antibodies.

Article Snippet: Monoclonal antibodies specific for Akt1, Akt2, Akt3, pan-Akt, phospho-Akt (Thr 308 ), phospho-Akt (Ser 473 ), phospho-GSK-3α/β (Ser 21/9 ), p27 Kip1, phospho-PRAS40 (Thr 246 ), phospho-TSC2 (Thr 1462 ), phospho-mTOR (Ser 2448 ), phospho-4E-BP1 (Ser 65 ), phospho-MDM2 (Ser 166 ), phospho-FoxO3a (Ser 318/321 ), phospho-Bad (Ser 136 ) and phospho-p44/42 MAPK (Thr 202 /Thr 204 ) were purchased from Cell Signaling Technology.

Techniques: Western Blot, Immunohistochemical staining, Staining

Journal: PLoS ONE

Article Title: Opposing Functions of Akt Isoforms in Lung Tumor Initiation and Progression

doi: 10.1371/journal.pone.0094595

Figure Lengend Snippet: Representative macroscopic images of individual lung lobes harvested from A JE JJenv infected WT (A), Akt1 −/− (B), Akt2 −/− (C), and Akt3 −/− (D) mice as well as mock infected mice (E) at 20 weeks post-infection. Representative images of hematoxylin and eosin stained sections from the lungs of WT (F to H), Akt1 −/− (I to K), Akt2 −/− (L and M) and Akt3 −/− (N to P) mice infected with A JE JJenv and harvested at 12, 20 and 32 weeks post-infection (4× magnification). Note that none of the Akt2 −/− infected mice survived past 20 weeks post-infection.

Article Snippet: Monoclonal antibodies specific for Akt1, Akt2, Akt3, pan-Akt, phospho-Akt (Thr 308 ), phospho-Akt (Ser 473 ), phospho-GSK-3α/β (Ser 21/9 ), p27 Kip1, phospho-PRAS40 (Thr 246 ), phospho-TSC2 (Thr 1462 ), phospho-mTOR (Ser 2448 ), phospho-4E-BP1 (Ser 65 ), phospho-MDM2 (Ser 166 ), phospho-FoxO3a (Ser 318/321 ), phospho-Bad (Ser 136 ) and phospho-p44/42 MAPK (Thr 202 /Thr 204 ) were purchased from Cell Signaling Technology.

Techniques: Infection, Staining

Journal: PLoS ONE

Article Title: Opposing Functions of Akt Isoforms in Lung Tumor Initiation and Progression

doi: 10.1371/journal.pone.0094595

Figure Lengend Snippet: WT, Akt1 −/− , Akt2 −/− and Akt3 −/− mice infected with A JE JJenv were euthanized at 12 (A), 20 (B) and 32 (C) weeks post infection and the number of lung tumors less than 100 μm, between 100 and 300 μm, and greater than 300 μm were quantified. Note that since all Akt2 −/− mice died around the 20-week time point there is no data for Akt2 −/− mice at the 32-week time point. Three lung lobes from three randomly selected mice per group were sectioned until the maximum surface area was exposed at which point all tumors within each lung lobe were counted. Bars on the graph labeled with an asterisk are statistically different (p<0.05).

Article Snippet: Monoclonal antibodies specific for Akt1, Akt2, Akt3, pan-Akt, phospho-Akt (Thr 308 ), phospho-Akt (Ser 473 ), phospho-GSK-3α/β (Ser 21/9 ), p27 Kip1, phospho-PRAS40 (Thr 246 ), phospho-TSC2 (Thr 1462 ), phospho-mTOR (Ser 2448 ), phospho-4E-BP1 (Ser 65 ), phospho-MDM2 (Ser 166 ), phospho-FoxO3a (Ser 318/321 ), phospho-Bad (Ser 136 ) and phospho-p44/42 MAPK (Thr 202 /Thr 204 ) were purchased from Cell Signaling Technology.

Techniques: Infection, Labeling

Journal: PLoS ONE

Article Title: Opposing Functions of Akt Isoforms in Lung Tumor Initiation and Progression

doi: 10.1371/journal.pone.0094595

Figure Lengend Snippet: Immunostaining for Jenv, SPC, and CC10 expression in lung tissue from A JE JJenv infected WT (A to C), Akt1 −/− (D to F), Akt2 −/− (G to I), and Akt3 −/− (J to L) mice at 10× magnification. Representative images of advanced neoplastic lesions show robust Jenv expression within all cells of the lung tumor (A, D, G, J). Representative images show lung tumors staining uniformly positive for SPC (B, E, H, K) and negative for CC10 (C, F, I, L).

Article Snippet: Monoclonal antibodies specific for Akt1, Akt2, Akt3, pan-Akt, phospho-Akt (Thr 308 ), phospho-Akt (Ser 473 ), phospho-GSK-3α/β (Ser 21/9 ), p27 Kip1, phospho-PRAS40 (Thr 246 ), phospho-TSC2 (Thr 1462 ), phospho-mTOR (Ser 2448 ), phospho-4E-BP1 (Ser 65 ), phospho-MDM2 (Ser 166 ), phospho-FoxO3a (Ser 318/321 ), phospho-Bad (Ser 136 ) and phospho-p44/42 MAPK (Thr 202 /Thr 204 ) were purchased from Cell Signaling Technology.

Techniques: Immunostaining, Expressing, Infection, Staining

Journal: PLoS ONE

Article Title: Opposing Functions of Akt Isoforms in Lung Tumor Initiation and Progression

doi: 10.1371/journal.pone.0094595

Figure Lengend Snippet: (A) Lung tissue sections from WT, Akt1 −/− and Akt3 −/− mice at 12 (early) and 32 (late) weeks post-infection and Akt2 −/− mice at 12 (early) and 20 (late) weeks post-infection stained with an antibody against the proliferation marker Ki67. (B) Graphical representation of the number of Ki67 positive cells. The number of Ki67-positive (proliferating) cells was measured in sections of lung derived from three randomly selected mice of each genotype and three randomly selected fields per mouse. The bars show the mean percentage of proliferating cells ± SE of the mean for each genotype. (C) Quantification of TUNEL-positive apoptotic cells in early and late (advanced) neoplastic lesions from WT, Akt1 −/− , Akt2 −/− and Akt3 −/− A JE JJenv infected mice. The number of TUNEL-positive cells was measured in sections of lung derived from three randomly selected mice of each genotype and three randomly selected fields per mouse. The bars show the mean percentage ± the SE of the mean of TUNEL-positive cells in early and late lesions. For figures B and C, 2-way ANOVA and Bonferonni's correction was used and bars on the graph with different letters are statistically different (p<0.05).

Article Snippet: Monoclonal antibodies specific for Akt1, Akt2, Akt3, pan-Akt, phospho-Akt (Thr 308 ), phospho-Akt (Ser 473 ), phospho-GSK-3α/β (Ser 21/9 ), p27 Kip1, phospho-PRAS40 (Thr 246 ), phospho-TSC2 (Thr 1462 ), phospho-mTOR (Ser 2448 ), phospho-4E-BP1 (Ser 65 ), phospho-MDM2 (Ser 166 ), phospho-FoxO3a (Ser 318/321 ), phospho-Bad (Ser 136 ) and phospho-p44/42 MAPK (Thr 202 /Thr 204 ) were purchased from Cell Signaling Technology.

Techniques: Infection, Staining, Marker, Derivative Assay, TUNEL Assay

Journal: PLoS ONE

Article Title: Opposing Functions of Akt Isoforms in Lung Tumor Initiation and Progression

doi: 10.1371/journal.pone.0094595

Figure Lengend Snippet: Representative western blot images of 80 JE JJenv infected WT (panel 1), Akt1 −/− (panel 2), Akt2 −/− (panel 3), and Akt3 −/− (panel 4) mice at 12, 20 and 32 weeks post-infection probed with Akt isoform specific antibodies (A to C), pan-Akt (D), a panel of phospho-specific (p) antibodies (E to N) and actin (O) as a loading control.

Article Snippet: Monoclonal antibodies specific for Akt1, Akt2, Akt3, pan-Akt, phospho-Akt (Thr 308 ), phospho-Akt (Ser 473 ), phospho-GSK-3α/β (Ser 21/9 ), p27 Kip1, phospho-PRAS40 (Thr 246 ), phospho-TSC2 (Thr 1462 ), phospho-mTOR (Ser 2448 ), phospho-4E-BP1 (Ser 65 ), phospho-MDM2 (Ser 166 ), phospho-FoxO3a (Ser 318/321 ), phospho-Bad (Ser 136 ) and phospho-p44/42 MAPK (Thr 202 /Thr 204 ) were purchased from Cell Signaling Technology.

Techniques: Western Blot, Infection, Control

Journal: Scientific reports

Article Title: The osteogenesis-promoting effects of alpha-lipoic acid against glucocorticoid-induced osteoporosis through the NOX4, NF-kappaB, JNK and PI3K/AKT pathways.

doi: 10.1038/s41598-017-03187-w

Figure Lengend Snippet: Figure 5. The protein expression levels of NOX4, NF-kappaB, JNK and the PI3K/AKT signalling pathway on femur. Nox4 (A), levels of phosphorylation of p65 (B), levels of phosphorylation of JNK (C) and levels of phosphorylation of AKT (D) were detected by western blotting. Data were the means ± SD (n = 6 for each group). #p < 0.05 and ##p < 0.01 compared with the SHAM group. *p < 0.05 and **p < 0.01 compared with the DEX group. GAPDH was used as loading control.

Article Snippet: Antibodies specific for Bcl-2 (Catalog No. 12789-1-AP), caspase-9 (Catalog No. 10380-1-AP), caspase-3 (Catalog No. 19677-1-AP), NOX4 (Catalog No. 14347-1-AP), JNK (Catalog No. 51151-1-AP), and AKT (Catalog No. 10176-2-AP) were purchased from Proteintech Group (Wuhan, China).

Techniques: Expressing, Phospho-proteomics, Western Blot, Control

Journal: Scientific reports

Article Title: The osteogenesis-promoting effects of alpha-lipoic acid against glucocorticoid-induced osteoporosis through the NOX4, NF-kappaB, JNK and PI3K/AKT pathways.

doi: 10.1038/s41598-017-03187-w

Figure Lengend Snippet: Figure 9. The expression levels of NOX4, NF-kappaB, JNK and the PI3K/AKT signalling pathway in MC3T3-E1 cells. NOX4 (A) and levels of phosphorylation of p65 (B) and levels of phosphorylation of JNK (C) and levels of phosphorylation of AKT (D) were detected by western blotting. GAPDH was used as loading control. Data were expressed as mean ± SD of three independent experiments. #p < 0.05 and ##p < 0.01 compared with the control group. *p < 0.05 and **p < 0.01 compared with the DEX-alone group.

Article Snippet: Antibodies specific for Bcl-2 (Catalog No. 12789-1-AP), caspase-9 (Catalog No. 10380-1-AP), caspase-3 (Catalog No. 19677-1-AP), NOX4 (Catalog No. 14347-1-AP), JNK (Catalog No. 51151-1-AP), and AKT (Catalog No. 10176-2-AP) were purchased from Proteintech Group (Wuhan, China).

Techniques: Expressing, Phospho-proteomics, Western Blot, Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: The preventive and therapeutic effects of AAV1‐KLF4‐shRNA in cigarette smoke‐induced pulmonary hypertension

doi: 10.1111/jcmm.16194

Figure Lengend Snippet: (A) Expression of KLF4 in the pulmonary vessels of rats in each group. Scale bar: 30 μm (B) Expression of PCNA in the pulmonary vascular smooth muscle of rats in each group. Scale bar: 25 μm (C) Representative bands of protein expression for KLF4 and PCNA in rat pulmonary vessels, and comparison of relative protein levels (normalized to GAPDH level). AU, arbitrary units. (D) Expression of P‐AKT in the pulmonary vessels of rats in each group. Scale bar: 20 μm. Sham (n = 8), saline (n = 6), AAV1‐control vector group (n = 6) and AAV1‐KLF4‐shRNA group (n = 11) at the end of the prevention experiment. Multiple comparisons were performed by one‐way ANOVA with SNK‐q. ** P < .01, *** P < .001

Article Snippet: Furthermore, the expression of KLF4 and P‐AKT was assessed by immunostaining with antibodies against KLF4 (ProteinTech, Wuhan, China) and P‐AKT (Gene Tex, San Antonio, USA).

Techniques: Expressing, Plasmid Preparation, shRNA

Journal: Journal of Cellular and Molecular Medicine

Article Title: The preventive and therapeutic effects of AAV1‐KLF4‐shRNA in cigarette smoke‐induced pulmonary hypertension

doi: 10.1111/jcmm.16194

Figure Lengend Snippet: (A) Expression of KLF4 in the pulmonary vessels of rats in each group. Scale bar: 30 μm (B) Expression of PCNA in the pulmonary vascular smooth muscle of rats in each group. Scale bar: 25 μm (C) Representative bands of protein expression for KLF4 and PCNA in rat pulmonary vessels, and comparison of relative protein levels (normalized to GAPDH level). AU, arbitrary units. (D) Expression of P‐AKT in the pulmonary vessels of rats in each group. Scale bar: 20 μm. Sham (n = 6), saline (n = 7), AAV1‐control vector group (n = 7) and AAV1‐KLF4‐shRNA group (n = 8) at the end of the therapeutic experiment. Multiple comparisons were performed by one‐way ANOVA with SNK‐q. * P < .05, ** P < .01, *** P < .001

Article Snippet: Furthermore, the expression of KLF4 and P‐AKT was assessed by immunostaining with antibodies against KLF4 (ProteinTech, Wuhan, China) and P‐AKT (Gene Tex, San Antonio, USA).

Techniques: Expressing, Plasmid Preparation, shRNA