cav-1 (rabbit) polyclonal antibody  (Santa Cruz Biotechnology)

Journal: eLife

Article Title: Caveolae and Bin1 form ring-shaped platforms for T-tubule initiation

doi: 10.7554/eLife.84139

Figure Lengend Snippet: List of primary antibodies.

Article Snippet: Cav-1 (Rabbit) polyclonal , Santa Cruz , sc894.

Techniques:

Journal: PLoS ONE

Article Title: SMAD-Independent Down-Regulation of Caveolin-1 by TGF-β: Effects on Proliferation and Survival of Myofibroblasts

doi: 10.1371/journal.pone.0116995

Figure Lengend Snippet: A and B , Cultured IMR-90 cells were treated with or without TGF-β1 (2 ng/ml) for 24 h and total RNA was isolated. A, cDNA Superarray analysis was performed and mRNA expression levels of Cav-1 and cyclophilin A are shown. Densitometric ratios of Cav-1:cyclophilin A are also shown. B, Histogram of real-time RT-PCR for Cav-1 expression after treated with TGF-β1 (2 ng/ml for 24 h) using triplicate samples from at least three individual experiments, normalized to 18S as mean±S.E.M. * p <0.05 compared to control. C , IMR-90 cells were treated with TGF-β1 (0–5 ng/ml) for 24 h and cell lysates extracted. Cell lysates were subjected to SDS-PAGE and immunoblotted with an antibody against Cav-1; the blot was stripped and probed for β-tubulin. Densitometric ratios of Cav-1:β-tubulin are also shown on the right, as mean±S.E.M. D , IMR-90 cells were stimulated with TGF-β1 (2 ng/ml) for the indicated times. Cell lysates were subjected to SDS-PAGE and immunoblotted with an antibody against Cav-1; the blot was then stripped and probed for β-tubulin. Densitometric ratios of Cav-1:β-tubulin are shown on the right, as mean±S.E.M. Results are averages of at least three independent experiments.

Article Snippet: Rabbit polyclonal antibody (N-20) against a peptide at the amino terminus of human Cav-1 was purchased from Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Cell Culture, Isolation, Expressing, Quantitative RT-PCR, SDS Page

Journal: PLoS ONE

Article Title: SMAD-Independent Down-Regulation of Caveolin-1 by TGF-β: Effects on Proliferation and Survival of Myofibroblasts

doi: 10.1371/journal.pone.0116995

Figure Lengend Snippet: A , IMR-90 cells were treated with inhibitors of p38 MAPK (SB203580; 6 μM) or ALK5 (SB431542; 0.5 μM) for 30 min prior to treatment with or without TGF-β1 (2 ng/ml) for a period of 48 h. Cell lysates were extracted and Western immunoblotting performed with an antibody against Cav-1; the blot was then stripped and probed for β-tubulin. B , Densitometric analyses of blots in (A) showed as % inhibition of baseline Cav-1 protein expression levels treated with TGF-β1. *indicates effect of SB203580 to completely block the inhibitory effect of TGF-β1 on Cav-1 expression. Results are averages of at least three independent experiments. Data are presented as mean±S.E.M. C , IMR-90 cells stably transfected with a kinase-deficient p38 MAPK (pcDNA-p38KM) or control vector (pcDNA) were treated with or without TGF-β1 (2 ng/ml) for 24 h. Cell lysates were obtained and subjected to SDS-PAGE and immunoblotted for Cav-1 and α-smooth muscle actin (α-SMA); blots were stripped and probed for β-tubulin. D , IMR-90 cells stably expressing SMAD2 sh RNA (pSU6H- sh SMAD2) or control vector (pSU6H) were treated with/without TGF-β1 (2 ng/ml) for 24 h. Cell lysates were immunoblotted for SMAD2, Cav-1 and α-SMA. The blots were stripped and probed for β-tubulin.

Article Snippet: Rabbit polyclonal antibody (N-20) against a peptide at the amino terminus of human Cav-1 was purchased from Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Western Blot, Inhibition, Expressing, Blocking Assay, Stable Transfection, Transfection, Plasmid Preparation, SDS Page

Journal: PLoS ONE

Article Title: SMAD-Independent Down-Regulation of Caveolin-1 by TGF-β: Effects on Proliferation and Survival of Myofibroblasts

doi: 10.1371/journal.pone.0116995

Figure Lengend Snippet: A , Human lung fibroblasts (IMR-90) were stably transfected with a plasmid encoding Cav-1 (pRC/CMV2-Cav1) or with empty vector (pRC/CMV2). Localization of Cav-1 protein was then analyzed by immunofluoresence staining with a rabbit polyclonal antibody to Cav-1. B , Stably-transfected cells described in (A) were treated with/without TGF-β1 (2 ng/ml) for 24 h. Cell lysates were obtained and subjected to immunoblotting for Cav-1, α-smooth muscle actin (α-SMA), and β-tubulin. C , Stably-transfected cells described in (A) were serum-deprived for 24 h and then treated with/without TGF-β1 (2 ng/ml) for 48 h followed by stimulation with 10% fetal bovine serum for 24 h. Cell numbers were assessed both prior to and after serum stimulation with an automated Coulter counter (n = 6 per group) as mean±S.E.M. *indicates p < 0.05 vs. control pRC/CMV2 “fibroblasts”. **indicates p < 0.05 vs. pRC/CMV2 “myofibroblasts” (TGF-β1 pre-treated). Similar results were obtained from 3 independent experiments. D , Cells in (C) were labeled with BrdU during the 24 h of serum stimulation (n = 6 per group). BrdU assays were performed as described in “Methods”. *indicates p < 0.05 vs. control pRC/CMV2 “fibroblasts”. **indicates p < 0.05 vs. pRC/CMV2 “myofibroblasts” (TGF-β1 pre-treated). Similar results were obtained from 3 independent experiments. E , Quiescent stably transfected IMR-90 cells described in (A) were treated with/without TGF-β1 (2 ng/ml) for 5 days in serum-free medium. Apoptosis assays using an ELISA for ss DNA (n = 6 for each group) were performed as described in “Methods”. *indicates p < 0.05 vs. control pRC/CMV2 “fibroblasts”. **indicates p < 0.01 vs. pRC/CMV2 “myofibroblasts” (TGF-β1 pre-treated). Similar results were obtained from 3 independent experiments.

Article Snippet: Rabbit polyclonal antibody (N-20) against a peptide at the amino terminus of human Cav-1 was purchased from Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Stable Transfection, Transfection, Plasmid Preparation, Staining, Western Blot, Labeling, Enzyme-linked Immunosorbent Assay

Journal: PLoS ONE

Article Title: SMAD-Independent Down-Regulation of Caveolin-1 by TGF-β: Effects on Proliferation and Survival of Myofibroblasts

doi: 10.1371/journal.pone.0116995

Figure Lengend Snippet: A , IMR-90 cells stably transfected with a plasmid encoding sh RNA targeted against Cav-1 (pSU6H- sh Cav1) or with control plasmid (pSU6H) were treated with/without TGF-β1 (2 ng/ml) for 24 h. Cell lysates were obtained and Western blots for Cav-1, α-smooth muscle actin (α-SMA) and β-tubulin performed. B , Stably-transfected cells described in (A) were serum-deprived for 24 h and treated with/without TGF-β1 (2 ng/ml) for 48 h followed by stimulation with 10% FBS for 24 h. Cell counts were assessed both prior to and after serum stimulation with an automated Coulter counter (n = 6 per group) shown as mean±S.E.M. *indicates p < 0.05 vs. control pSU6H “fibroblasts”. **indicates p < 0.05 vs. pSU6H “myofibroblasts” (TGF-β1 pre-treated). Similar results were obtained from 3 independent experiments. C , Stably transfected cells described in (A) were serum-deprived for 24 h and treated with/without TGF-β1 (2 ng/ml) for 48 h followed by BrdU labeling for 24 h in the presence of 10% FBS (n = 6 per group) shown as mean±S.E.M. *indicates p < 0.05 vs. control pSU6H “fibroblasts”. Similar results were obtained from 3 independent experiments. D , Quiescent stably-transfected cells described in ( A ) were treated with/without TGF-β1 (2 ng/ml) for 5 days and apoptotic assay for ss DNA performed as described in “Methods” (n = 6 per group) shown as mean±S.E.M. *indicates p < 0.05 vs. control pSU6H “fibroblasts”. **indicates p < 0.05 vs. pSU6H “myofibroblasts” (TGF-β1 pre-treated). Similar results were obtained from 3 independent experiments.

Article Snippet: Rabbit polyclonal antibody (N-20) against a peptide at the amino terminus of human Cav-1 was purchased from Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Stable Transfection, Transfection, Plasmid Preparation, Western Blot, Labeling

Journal: PLoS ONE

Article Title: SMAD-Independent Down-Regulation of Caveolin-1 by TGF-β: Effects on Proliferation and Survival of Myofibroblasts

doi: 10.1371/journal.pone.0116995

Figure Lengend Snippet: TGF-β1 activates the cell surface TGF-β receptor(s) complex that leads to rapid activation of the canonical SMAD pathway as well as the SMAD- in dependent p38 MAPK pathway. Activation of the SMAD pathway is required for the induction of a cellular program of growth-arrest and myofibroblast differentiation. In contrast, activation of the p38 MAPK pathway, independently of SMAD2/3, is required the down-regulation of Cav-1 by TGF-β1. Down-regulation of Cav-1 by TGF-β1 “primes” differentiated myofibroblasts for enhanced proliferative responses to mitogens and resistance to apoptosis. These divergent TGF-β signaling pathways may explain, in part, the contextual effects of TGF-β1 as both a growth-inhibitor and –promoter on the same target (mesenchymal) cells.

Article Snippet: Rabbit polyclonal antibody (N-20) against a peptide at the amino terminus of human Cav-1 was purchased from Santa Cruz Biotechnology, Santa Cruz, CA.

Techniques: Activation Assay