cytomegalovirus pcmv mtor his  (Sino Biological)

Journal: Nature Communications

Article Title: The tyrosine phosphatase SHP2 controls TGFβ-induced STAT3 signaling to regulate fibroblast activation and fibrosis

doi: 10.1038/s41467-018-05768-3

Figure Lengend Snippet: Knockout of Shp2 decreases JAK2/STAT3 signaling. a Knockout of Shp2 in fibroblasts ( Shp2 fl/fl x Col1a2 ; Cre ER) decreases the levels of pJAK2 and pSTAT3 and STAT3 reporter activity in cultured fibroblasts ( n = 3 different lines). Cells were stimulated with TGFβ (10 ng/ml for 6 h). b – d Conditional knockout of Shp2 reduces the levels of pJAK2 and pSTAT3 in TBRI CA (6.67 × 10 7 IFUs every 2 weeks) ( b ) and bleomycin-induced fibrosis (50 µg every other day) ( c ) and in TSK1 mice (2 mg tamoxifen over 5 days) ( d ) ( n ≥ 3). Results shown are representative of three independent experiments. All data are presented as median ± s.e.m. The p values are expressed as follows: 0.05 > p > 0.01* or # ; 0.01 > p > 0.001** or ## ; p

Article Snippet: The plasmid pCMV3-Flag-JAK2 encoding the human JAK2 was purchased from Sino Biological (Beijing, China).

Techniques: Knock-Out, Activity Assay, Cell Culture, Mouse Assay

Journal: Nature Communications

Article Title: The tyrosine phosphatase SHP2 controls TGFβ-induced STAT3 signaling to regulate fibroblast activation and fibrosis

doi: 10.1038/s41467-018-05768-3

Figure Lengend Snippet: Inhibition of SHP2 limits JAK2/STAT3 signaling and fibroblast activation. a Changes in the mRNA levels of COL1A1 and of collagen protein in human fibroblasts incubated with increasing doses of NSC-87877 (10 µM, 30 µM and 100 µM). Fibroblasts were treated with TGFβ (10 ng/ml) for 24 h. b ACTA2 mRNA. ( n ≥ 4) c Representative images of immunofluorescence stainings for α-SMA and stress fiber staining are shown at 400-fold magnification and quantification of α-SMA staining intensity as well as stress fiber staining intensity ( n ≥ 15) (TGFβ 10 ng/ml for 24 h). Horizontal scale bar, 500 μm. d Representative western blots for pJAK2 Y1007/Y1008 , pJAK2 Y570 , total JAK2, pSTAT3 Y705 and total STAT3 with β-actin as loading control and quantification of the results (TGFβ 10 ng/ml for 6 h) ( n ≥ 2). e Changes in STAT3 reporter activity ( n ≥ 6) (TGFβ 10 ng/ml for 6 h). Results shown are representative of three independent experiments All data are presented as median ± s.e.m. The p values are expressed as follows: 0.05 > p > 0.01*; 0.01 > p > 0.001**; p

Article Snippet: The plasmid pCMV3-Flag-JAK2 encoding the human JAK2 was purchased from Sino Biological (Beijing, China).

Techniques: Inhibition, Activation Assay, Incubation, Immunofluorescence, Staining, Western Blot, Activity Assay

Journal: Nature Communications

Article Title: The tyrosine phosphatase SHP2 controls TGFβ-induced STAT3 signaling to regulate fibroblast activation and fibrosis

doi: 10.1038/s41467-018-05768-3

Figure Lengend Snippet: SHP2 enhances TGFβ-induced fibroblast activation via JAK2/STAT3. a mRNA levels of SHP2 after overexpression in human dermal fibroblasts. mRNA levels of COL1A1 in human fibroblasts transfected with empty vector, SHP2 WT - and SHP2 C459S -expression vectors, with or without TGFβ1 treatment (10 ng/ml for 24 h) ( n ≥ 4). b Western blot analysis and respective quantifications for type I collagen and SHP2 in human fibroblasts transfected with empty vector, SHP2 WT - and SHP2 C459S -expression vectors, with or without TGFβ1 treatment (10 ng/ml for 24 h). Western blot for pJAK2 Y1007/Y1008 , pJAK2 Y570 , total JAK2, pSTAT3 Y705 and total STAT3 with β-actin as loading control (TGFβ 10 ng/ml for 6 h) ( n = 3). Results shown are representative of three independent experiments. c , d Representative images of immunofluorescence stainings for α-SMA and stress fiber staining are shown at 400-fold magnification ( c ) and quantification of α-SMA staining intensity as well as stress fiber staining intensity ( d ) ( n ≥ 4). Horizontal scale bar, 500 μm. All data are presented as median ± s.e.m. The p values are expressed as follows: 0.05 > p > 0.01*; 0.01 > p > 0.001**; p

Article Snippet: The plasmid pCMV3-Flag-JAK2 encoding the human JAK2 was purchased from Sino Biological (Beijing, China).

Techniques: Activation Assay, Over Expression, Transfection, Plasmid Preparation, Expressing, Western Blot, Immunofluorescence, Staining

Journal: Nature Communications

Article Title: The tyrosine phosphatase SHP2 controls TGFβ-induced STAT3 signaling to regulate fibroblast activation and fibrosis

doi: 10.1038/s41467-018-05768-3

Figure Lengend Snippet: Overexpression of JAK2 ∆Y570F prevents the inhibitory effects of SHP2 inhibitors on TGFβ-induced fibroblast activation. a mRNA levels of COL1A1 and COL1A2 (TGFβ 10 ng/ml for 24 h) ( n ≥ 5). b Release of collagen protein (TGFβ 10 ng/ml for 24 h) ( n ≥ 6). c Representative images of immunofluorescence stainings for α-SMA and stress fiber at 400-fold magnification and respective quantifications (TGFβ 10 ng/ml for 24 h) ( n ≥ 6). Horizontal scale bar, 500 μm. d STAT3 reporter Assay upon JAK2 WT and Y570F mutant overexpression. Cells were treated with TGFβ (10 ng/ml for 6 h) and NSC-87877 (100 µM) ( n ≥ 4). e Co-immunoprecipitation and respective quantifications of endogenous JAK2 with endogenous SHP2 in human fibroblasts stimulated with TGFβ (10 ng/ml for 30′) ( n = 3). Results shown are representative of ≥ 3 independent experiments. All data are presented as median ± s.e.m. The p values are expressed as follows: 0.05 > p > 0.01*; 0.01 > p > 0.001**; p

Article Snippet: The plasmid pCMV3-Flag-JAK2 encoding the human JAK2 was purchased from Sino Biological (Beijing, China).

Techniques: Over Expression, Activation Assay, Immunofluorescence, Reporter Assay, Mutagenesis, Immunoprecipitation

Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Heterochromatin protects retinal pigment epithelium cells from oxidative damage by silencing p53 target genes

doi: 10.1073/pnas.1715237115

Figure Lengend Snippet: Desumoylation of p53 is needed for heterochromatin-mediated p53 target gene repression. ( A ) FLAG-SUV39H1 was transfected into hRPE cells. The endogenous p53 protein was precipitated by p53 antibody DO-1, and a 67-kDa form of p53 was detected (labeled with *). The co-IP experiment was conducted in the presence of a desumoylation inhibitor, 20 mM NEM. ( B ) The 67-kDa p53 decreases on OS exposure. WB analysis showing the conventional p53 and a 67-kDa species. Quantification of the percentage of the 67-kDa p53 species (*p53) relative to the total p53. n = 3. ** P

Article Snippet: FLAG-SUV39H1 and HA-SUV39H1 plasmids were purchased from Sino Biological and verified by sequencing.

Techniques: Transfection, Labeling, Co-Immunoprecipitation Assay, Western Blot