Journal: JCI insight

Article Title: DRA involvement in linaclotide-stimulated bicarbonate secretion during loss of CFTR function.

doi: 10.1172/jci.insight.172364

Figure Lengend Snippet: Figure 2. Linaclotide stimulates duodenal bicarbonate secretion independent of CFTR. (A) In vivo measurement of duodenal bicarbonate secretion in mice, similar to Figure 1A, with the exception that each perfusate also contained CFTRinh-172 (2 × 10–5 M) (n = 12). Dotted line is the mean response without CFTRinh-172 (from Figure 1A). *P < 0.05 vs. baseline by 1-way ANOVA. (B and C) In vitro duodenal mucosal bicarbonate secretion (B) and Isc (C) in wild-type (WT) and Cftr-KO mice. Data are expressed as fold increase in linaclotide-stimulated responses over baseline bicarbonate secretion or Isc in the same mouse. Each point (n = 7) is a separate piece of duodenum from 5 mice. **P < 0.01 by unpaired 2-tailed Student’s t test. (D) Transepithelial resistance measurements in WT and Cftr-KO mice. (E–G) In vitro duodenal mucosal bicarbonate secretion (E), Isc (F), and transepithelial resistance (G) in human endoscopic biopsies with or without CFTRinh-172 pretreatment (2 × 10–5 M, 40–60 minutes). Each point (n = 11–14) represents a different biopsy. All data are means ± SEM.

Article Snippet: Linaclotide was purchased from Santa Cruz Biotechnology.

Techniques: In Vivo, In Vitro

Journal: JCI insight

Article Title: DRA involvement in linaclotide-stimulated bicarbonate secretion during loss of CFTR function.

doi: 10.1172/jci.insight.172364

Figure Lengend Snippet: Figure 3. Key role of DRA in CFTR-independent linaclotide-stimulated duodenal bicarbonate secretion upon loss of CFTR. (A and B)To determine the source of linaclotide-stimulated bicarbonate transport in the absence of CFTR function, in vivo experiments were repeated, similar to Figure 2A, except in addition to CFTRinh-172 (2 × 10–5 M, n = 12), one of the following was also added to the luminal perfusate: DRAinh-A250 (10–5 M, n = 10), DIDS (2 × 10–4 M, n = 7), or S3226 (10–5 M, n = 9). (A) Linaclotide 10–7 M, (B) linaclotide 10–5 M. Each point represents a different mouse. *P < 0.05 vs. linaclotide + CFTRinh-172 (2 × 10–5 M) by 1-way ANOVA. (C) Time course with linaclotide dose response (10–9 M, 10–7 M, 10–5 M) in the presence of CFTRinh-172 (2 × 10–5 M) and DRAinh-A250 (10–5 M), as indicated by circles and whiskers (n = 12). Dotted line indicates mean response in the presence of CFTRinh-172 (2 × 10–5 M) only (from Figure 2A). *P < 0.05; **P < 0.01 vs. baseline by 1-way ANOVA. (D) Net peak linaclotide-stimulated (10–7 M, apical) mouse duodenal mucosal bicarbonate secretion (D) and Isc (E) from in vitro experiments, with or without DRA inhibition by DRAinh-A270 (10–5 M, bilateral, n = 5) or DRAinh-4a (10–5 M, bilateral, n = 14). Each point represents a separate piece of duodenum from 5–10 mice. All data are means ± SEM.

Article Snippet: Linaclotide was purchased from Santa Cruz Biotechnology.

Techniques: In Vivo, In Vitro, Inhibition

Journal: JCI insight

Article Title: DRA involvement in linaclotide-stimulated bicarbonate secretion during loss of CFTR function.

doi: 10.1172/jci.insight.172364

Figure Lengend Snippet: Figure 6. Linaclotide increases membrane expression of DRA in apical-out human duodenal enteroids. Representative images of DRA (A and E), villin (B and F), and DRA and villin and Hoechst (C and G) in apical-out healthy duodenal enteroids during control conditions (water or DMSO), CFTRinh-172 (2 × 10–5 M), or linaclotide (10–7 M) with or without CFTRinh-172 pretreatment (2 × 10–5 M) (all 40 minutes). (D, H, and I) Quantification of DRA at the apical brush border using villin to define this cell region. Apical brush border DRA MFI following linaclotide (10–7 M, 40 minutes, n = 16), CFTRinh-172 alone (2 × 10–5 M, 40 minutes, n = 31), or CFTRinh-172+linaclotide (n = 21). Treatments were normalized to vehicle controls (water for linaclotide, DMSO for CFTRinh-172, n = 12–31). Enteroids from 3 individuals were used for each condition. Significance determined by unpaired, 2-tailed Student’s t test. *P < 0.05, ***P < 0.001. Representative images of DRA (J), villin (K), and DRA and villin and Hoechst (L) in apical-out CF (F508del homozygous) duodenal enteroids during control (water, 40 minutes) or lina- clotide stimulation (10–7 M, 40 minutes). (M) Quantification of DRA present at the apical brush border using villin, similar to D. (N) Comparison of baseline DRA MFI between non-CF and CF enteroids, with data normalized to non-CF enteroids. n = 8–11 enteroids from 1 patient across 2 different passages. Significance determined by unpaired, 2-tailed Student’s t test. *P < 0.05, **P < 0.01. (O) To compare DRA membrane expression across non-CF and CF enteroids in differ- ent situations, absolute DRA membrane MFI was plotted. Data are from experiments performed in D, H, I, and M. In images, arrows assist with identifying regions of interest (scale bar, 20 µm). Columns with whiskers are mean ± SEM with each dot representing a different enteroid.

Article Snippet: Linaclotide was purchased from Santa Cruz Biotechnology.

Techniques: Membrane, Expressing, Control, Comparison

Journal: Contemporary Clinical Trials Communications

Article Title: Exploratory, multicenter, open-label study to evaluate the effects of linaclotide in patients with chronic constipation with an insufficient response to magnesium oxide: A study protocol

doi: 10.1016/j.conctc.2022.101019

Figure Lengend Snippet: Criteria for inclusion, medication initiation, and exclusion.

Article Snippet: Linaclotide provided by Astellas Pharma Inc. will be used in this study.

Techniques:

Journal: Contemporary Clinical Trials Communications

Article Title: Exploratory, multicenter, open-label study to evaluate the effects of linaclotide in patients with chronic constipation with an insufficient response to magnesium oxide: A study protocol

doi: 10.1016/j.conctc.2022.101019

Figure Lengend Snippet: Study endpoints.

Article Snippet: Linaclotide provided by Astellas Pharma Inc. will be used in this study.

Techniques: