Journal: Digestive Diseases and Sciences

Article Title: Involvement of EP2 and EP4 Receptors in Eosinophilic Esophagitis: A Pilot Study

doi: 10.1007/s10620-019-05623-5

Figure Lengend Snippet: Bio-Rad primers used for quantitative real-time PCR

Article Snippet: For flow cytometric visualization of the EP receptors, cells were incubated with rabbit polyclonal antibodies against EP1, EP2, and EP3 (20 μg/ml; Alomone Labs, Jerusalem, Israel) and mouse polyclonal antibody against EP4 (20 μg/ml; Alomone Labs, Jerusalem, Israel), or normal rabbit or mouse IgG (20 μg/ml; Santa Cruz Biotechnology, Dallas, TX, USA) for 30 min at 4 °C.

Techniques:

Journal: Digestive Diseases and Sciences

Article Title: Involvement of EP2 and EP4 Receptors in Eosinophilic Esophagitis: A Pilot Study

doi: 10.1007/s10620-019-05623-5

Figure Lengend Snippet: Flow cytometric evaluation of EP (EP1–4) and DP (DP1, DP2) receptor expression in human blood eosinophils. Changes in receptor expression were recorded as mean fluorescence intensity and data are expressed as fold increase in fluorescence over isotype control. Expression levels of EP2 and EP4 are significantly lower in patients with eosinophilic esophagitis (EoE) compared to healthy control subjects (control). Expression levels of EP1, EP3, DP1, and DP2 do not differ between EoE and controls. Values are mean ± SEM; Student’s t test; n = 6–7. * p < 0.05, ** p < 0.01

Article Snippet: For flow cytometric visualization of the EP receptors, cells were incubated with rabbit polyclonal antibodies against EP1, EP2, and EP3 (20 μg/ml; Alomone Labs, Jerusalem, Israel) and mouse polyclonal antibody against EP4 (20 μg/ml; Alomone Labs, Jerusalem, Israel), or normal rabbit or mouse IgG (20 μg/ml; Santa Cruz Biotechnology, Dallas, TX, USA) for 30 min at 4 °C.

Techniques: Expressing, Fluorescence

Journal: Digestive Diseases and Sciences

Article Title: Involvement of EP2 and EP4 Receptors in Eosinophilic Esophagitis: A Pilot Study

doi: 10.1007/s10620-019-05623-5

Figure Lengend Snippet: Chemotaxis and adhesion assays with human blood eosinophils. Eosinophils from healthy donors were incubated with 100 nM of EP4 agonist ONO-AE1-329 or with 100 nM of the EP4 antagonist ONO-AE3-208 ( a ), with 100 and 300 nM of EP2 agonist butaprost ( b ), or with 100 nM of DP1 agonist BW245C and 1 µM of DP1 antagonist MK0524 ( c ), and chemotaxis was assessed using esophageal epithelial cell supernatant for chemoattraction. n = 6–12 (for a , and b ) and n = 3 ( c ); values are mean ± SEM; one-way ANOVA; Tukey’s post hoc test. *** p values < 0.001. Adhesion assays were performed with human blood eosinophils and esophageal epithelial cells after incubation of eosinophils with EP4 agonist ONO-AE1-329 ( d ), EP2 agonist butaprost ( e ), and with DP1 agonist BW245C ( f ) vs. untreated eosinophils (control; set at 100%). n = 5–7; values are mean ± SEM; one-way ANOVA; Tukey’s post hoc test. * p < 0.05; ** p < 0.01

Article Snippet: For flow cytometric visualization of the EP receptors, cells were incubated with rabbit polyclonal antibodies against EP1, EP2, and EP3 (20 μg/ml; Alomone Labs, Jerusalem, Israel) and mouse polyclonal antibody against EP4 (20 μg/ml; Alomone Labs, Jerusalem, Israel), or normal rabbit or mouse IgG (20 μg/ml; Santa Cruz Biotechnology, Dallas, TX, USA) for 30 min at 4 °C.

Techniques: Chemotaxis Assay, Incubation

Journal: Digestive Diseases and Sciences

Article Title: Involvement of EP2 and EP4 Receptors in Eosinophilic Esophagitis: A Pilot Study

doi: 10.1007/s10620-019-05623-5

Figure Lengend Snippet: Immunohistochemistry shows strong DP1 staining in eosinophils ( a , arrows ), while epithelial cells in esophageal mucosal biopsies are practically devoid of staining. EP4 ( b ) and EP2 ( c ) immunostaining is prominent in eosinophils ( arrows ) but also visible in epithelial cells ( arrowheads ). 3-3′-Diaminobenzidine (DAB) was used as visualization substrate for DP1 and EP4 staining (brown precipitates), while for EP2, ImmPact™ NovaRED substrate was used (red precipitate). Calibration bar: 50 µm

Article Snippet: For flow cytometric visualization of the EP receptors, cells were incubated with rabbit polyclonal antibodies against EP1, EP2, and EP3 (20 μg/ml; Alomone Labs, Jerusalem, Israel) and mouse polyclonal antibody against EP4 (20 μg/ml; Alomone Labs, Jerusalem, Israel), or normal rabbit or mouse IgG (20 μg/ml; Santa Cruz Biotechnology, Dallas, TX, USA) for 30 min at 4 °C.

Techniques: Immunohistochemistry, Staining, Immunostaining

Journal: Digestive Diseases and Sciences

Article Title: Involvement of EP2 and EP4 Receptors in Eosinophilic Esophagitis: A Pilot Study

doi: 10.1007/s10620-019-05623-5

Figure Lengend Snippet: ECIS ® cell impedance assays were performed in esophageal epithelial cells with 10 and 300 nM of EP4 agonist ONO-AE1-329 ( a ) and EP2 agonist butaprost ( b ) (or medium as control). The decrease in resistance correlates with a loss in the monolayer’s integrity indicative of cell barrier decrease. n = 5–9; values are mean ± SEM; one-way-ANOVA; Tukey’s post hoc test; * p < 0.05; *** p < 0.001

Article Snippet: For flow cytometric visualization of the EP receptors, cells were incubated with rabbit polyclonal antibodies against EP1, EP2, and EP3 (20 μg/ml; Alomone Labs, Jerusalem, Israel) and mouse polyclonal antibody against EP4 (20 μg/ml; Alomone Labs, Jerusalem, Israel), or normal rabbit or mouse IgG (20 μg/ml; Santa Cruz Biotechnology, Dallas, TX, USA) for 30 min at 4 °C.

Techniques: