Journal: American Journal of Physiology - Heart and Circulatory Physiology

Article Title: Enhanced large conductance K + channel activity contributes to the impaired myogenic response in the cerebral vasculature of Fawn Hooded Hypertensive rats

doi: 10.1152/ajpheart.00636.2013

Figure Lengend Snippet: Comparison of BK-α- and BK-β-subunit expression in cerebral vessel homogenates isolated from FHH and FHH.1BN rats. BK-α- and BK-β-subunits were probed on different blots. The same blots were then reprobed for β-actin. A: representative bands corresponding to the molecular weight of the BK-α-subunit (∼100 kDa), BK-β-subunit (∼25 kDa), and β-actin (∼42 kDa) in cerebral vessels obtained from FHH and FHH.1BN rats (n = 8 animals each; 3 sets of experiments). B: a comparison of the expression of α- and β-subunits. Numbers in parentheses indicate the number of samples studied.

Article Snippet: After transfer, the membrane was blocked with TBS-T buffer containing 20 mM Tris pH 7.5, 150 mM NaCl, 0.05% Tween, and a 5% blocking powder (Bio-Rad) at 4°C for 1 h. The blot was probed with primary antibody against BK α- and β-subunit [1:500 and 1:200, respectively; polyclonal rabbit Anti-K Ca 1.1, to amino acids 1184–1200 and Anti-sloβ1 (KCNMB1); Alomone Labs, Jerusalem, Israel] overnight at 4°C.

Techniques: Expressing, Isolation, Molecular Weight

Journal: Diabetes

Article Title: Gene Silencing of Phogrin Unveils Its Essential Role in Glucose-Responsive Pancreatic β-Cell Growth

doi: 10.2337/db08-0970

Figure Lengend Snippet: Effects of shRNA-expressing adenoviruses on GSIS and the proliferation of MIN6 cells. A : MIN6 cells were infected with adenoviruses integrating shVector, shPh1, shPh2, shPh3, or shIA2 for 72 h. The expression levels of endogenous phogrin and IA-2 proteins were determined by immunoblotting with specific antibodies for phogrin and IA-2. α-Tubulin is a nontargeting control. B : MIN6 cells infected with Ad-shVec, shPh1, shPh2, or shPh3 were immunostained with anti-phogrin antibody. C : MIN6 cells expressing shPh3 were fixed at time points 24, 48, and 72 h. Cells were then immunostained with anti-phogrin and anti-insulin antibodies. D : The infected cells were incubated for 2 h in modified KRB (2 mmol/l glucose) before high-glucose stimulation. After stimulation for 30 min with 2 or 25 mmol/l glucose, culture media ( left ; secretion) and cells ( right ; content) were collected and subjected to RIA for insulin. Data are given as means ± SE of five independent experiments. E : Time course of MIN6 cell proliferation 24–96 h after infection with Ad-shVec, shPh1, shPh2, shPh3, or shIA2. Cell number was measured as indicated in research design and methods . Phogrin-KD or IA2-KD cells grew more slowly than the shVec-infected cells (* P < 0.001, ** P < 0.005). F : Cells infected by Ad-shVec or shPh3 for 72 h were assayed for BrdU incorporation by immunofluorescence analysis. Experiments were performed four times ( n = 12). (Please see http://dx.doi.org/10.2337/db08-0970 for a high-quality digital representation of this figure.)

Article Snippet: Anti-adaptin, anti–carboxypeptidase E (anti-CPE), anti–calcium-dependent activator protein (anti-CAPS), anti-munc18, anti-dynamin, anti–clathrin heavy chain, anti–phosphatidylinositol 3-kinase (anti-PI3K), anti-Grb2, and anti–extracellular signal–related kinase (anti-ERK) mouse monoclonal antibodies and anti–insulin receptor β-subunit (anti-IRβ) rabbit polyclonal antibodies were from BD Biosciences.

Techniques: shRNA, Expressing, Infection, Western Blot, Incubation, Modification, BrdU Incorporation Assay, Immunofluorescence

Journal: American Journal of Physiology - Renal Physiology

Article Title: NBCe1-A is required for the renal ammonia and K + response to hypokalemia

doi: 10.1152/ajprenal.00481.2019

Figure Lengend Snippet: Antibodies used (alphabetical order)

Article Snippet: ENaCβ , StressMarq Biosciences (SPC-404, Victoria, BC, Canada) , WB, 1:5,000.

Techniques: Generated