Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: Alveolar nonselective channels are ASIC1a/α-ENaC channels and contribute to AFC

doi: 10.1152/ajplung.00379.2016

Figure Lengend Snippet: Expression of α-ENaC regulates NSC channel frequency. Knockdown of α-, β-, or γ-ENaC subunits in an AT2 cell line, L2, with shRNA silencing vectors in lentivirus. A: percentage of patches with HSC channels in knockdown or sham conditions. B: percentage of patches with NSC channels in knockdown/sham conditions. Only knocking down α-ENaC reduces the surface expression of both HSC and NSC channels (n = 4 for all conditions; HSC reduction and NSC reduction are significant, P < 0.01 by z-test).

Article Snippet: This method was used to detect ENaC subunits (with in-house antibodies; 2 , 77 , 85 ) and ASIC1 (Alamone Laboratories).

Techniques: Expressing, shRNA

Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: Alveolar nonselective channels are ASIC1a/α-ENaC channels and contribute to AFC

doi: 10.1152/ajplung.00379.2016

Figure Lengend Snippet: NSC channels are sensitive to ASIC1-modifying toxins. A: NSC channels were activated by venom of the Texas coral snake (Micrurus tener tener; MitTx). It is a potent, persistent, and selective agonist for acid-sensing ion channels and is highly selective for ASIC1 at neutral pH (11). B: psalmotoxin-1 isolated from the venom of the spider, Psalmopoeus cambridgei (Trinidad chevron tarantula), is a potent and selective acid-sensing ion channel 1a (ASIC1a) blocker (IC50 = 0.9 nM) with no effect on ASIC1b, ASIC2a, ASIC3, and ENaC channels at concentrations up to 100 nM (13, 14, 19). When applied to AT2 cells in primary culture, it uniformly decreased NSC open probability. Neither toxin had any effect on HSC channels. *P < 0.05.

Article Snippet: This method was used to detect ENaC subunits (with in-house antibodies; 2 , 77 , 85 ) and ASIC1 (Alamone Laboratories).

Techniques: Isolation

Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: Alveolar nonselective channels are ASIC1a/α-ENaC channels and contribute to AFC

doi: 10.1152/ajplung.00379.2016

Figure Lengend Snippet: NSC channel frequency strongly depends on ASIC1a and α-ENaC presence. A: Western blot showing ASIC1 expression in L2 cells treated with scrambled shRNA or ASIC1 silencing vectors. Although the blot is cropped, there are no other bands in the blot. Reduction of ASIC1 protein positively correlates to amount of ASIC1a shRNA used. B, left: percentage patches with NSC channels present under ASIC1 or α-ENaC knockdown conditions. Right: percentage patches with HSC channels under ASIC1 or α-ENaC knockdown conditions. Numbers above bars indicate total number of patches.

Article Snippet: This method was used to detect ENaC subunits (with in-house antibodies; 2 , 77 , 85 ) and ASIC1 (Alamone Laboratories).

Techniques: Western Blot, Expressing, shRNA

Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: Alveolar nonselective channels are ASIC1a/α-ENaC channels and contribute to AFC

doi: 10.1152/ajplung.00379.2016

Figure Lengend Snippet: Interaction of ASIC1a and α-ENaC subunits. A, top: L2 cell protein lysate immunoprecipitated (IP) for α-ENaC and immunoblotted (IB) for ASIC1. Bottom: L2 cell protein lysate immunoprecipitated for ASIC1 and immunoblotted for α-ENaC. B: quantification of mammalian two-hybrid assay between ASIC1a and ENaC subunits. Normalized luciferase luminescence is proportional to binding affinity to ASIC1a. Negative control represents random association, and positive control represents maximum affinity. Fold increase for α-ENaC (6.6 ± 0.73) indicates a high affinity for ASIC1a. Data represent a total n of 15 (n = 3 for each condition); *P < 0.05.

Article Snippet: This method was used to detect ENaC subunits (with in-house antibodies; 2 , 77 , 85 ) and ASIC1 (Alamone Laboratories).

Techniques: Immunoprecipitation, Two Hybrid Assay, Luciferase, Binding Assay, Negative Control, Positive Control

Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

Article Title: Alveolar nonselective channels are ASIC1a/α-ENaC channels and contribute to AFC

doi: 10.1152/ajplung.00379.2016

Figure Lengend Snippet: Schematic diagram of a possible configuration of ASIC1a and ENaC subunits in NSC and HSC channels.

Article Snippet: This method was used to detect ENaC subunits (with in-house antibodies; 2 , 77 , 85 ) and ASIC1 (Alamone Laboratories).

Techniques:

Journal: PLoS ONE

Article Title: Carcinoembryonic Antigen-Related Cell Adhesion Molecules (CEACAM) 1, 5 and 6 as Biomarkers in Pancreatic Cancer

doi: 10.1371/journal.pone.0113023

Figure Lengend Snippet: (A). Basal CEACAM expression varied depending on the CEACAM subtype, highest levels were found for CEACAM 1, intermediate for CEACAM 6 and lowest levels for CEACAM 5. As seen in proliferation (B) and migration assays (C), CEACAM knock down cells showed a higher proliferative and migrative potential compared to control cells. Differences in adhesion on stimulated (TNFα) endothelial cells were not detectable between CEACAM kd and control cells. Neither the average number (D) nor the average adhesion time to the endothelial cells was different (E). Murine xenograft tumors were highly positive for CEACAM 1, 5 and 6. Immunohistochemical staining in control cells showed complete absence of CEACAM expression in immunohistochemistry in knock down cells (F).

Article Snippet: CEACAM staining was performed using a specific CEACAM monoclonal antibody (CEACAM1 clone 4D1/C2 IgG2a, in-house clone (previously described by ) at a dilution of 1∶200, CEACAM5 clone #2383 IgG1 at a dilution of 1∶50 (Cell Signaling, Beverly, USA); CEACAM6 clone IgG1 (9A6), at a dilution of 1∶40 [Sigma Aldrich, Hamburg, Germany]) overnight at 4°C.

Techniques: Expressing, Migration, Immunohistochemical staining, Staining, Immunohistochemistry

Journal: PLoS ONE

Article Title: Carcinoembryonic Antigen-Related Cell Adhesion Molecules (CEACAM) 1, 5 and 6 as Biomarkers in Pancreatic Cancer

doi: 10.1371/journal.pone.0113023

Figure Lengend Snippet: TMA immunohistochemical staining of CEACAM 1 negative (A) and positive (B), CEACAM 5 negative (C) and positive (D), CEACAM 6 negative (E) and positive (F).

Article Snippet: CEACAM staining was performed using a specific CEACAM monoclonal antibody (CEACAM1 clone 4D1/C2 IgG2a, in-house clone (previously described by ) at a dilution of 1∶200, CEACAM5 clone #2383 IgG1 at a dilution of 1∶50 (Cell Signaling, Beverly, USA); CEACAM6 clone IgG1 (9A6), at a dilution of 1∶40 [Sigma Aldrich, Hamburg, Germany]) overnight at 4°C.

Techniques: Immunohistochemical staining, Staining