Intestinal Epithelial AMPK Regulates Paracellular Electrolyte Flux and Electrogenic Ion Transport Responses in Mouse Colon
Bibliographic record
Abstract
Ion transport within the colonic epithelium is crucial to maintain colonic homeostasis. During pathological states such as intestinal inflammation, alteration of ion transport processes contributes to the development of diarrhea and constipation. These pathways are tightly regulated by expression and localization to ensure fluid balance. AMP‐Activated Protein Kinase (AMPK) is a highly conserved energy‐sensing kinase that has previously been shown to regulate ion channels and transporters such as CFTR and NKCC1, in both intestinal epithelium and other epithelia. The aim of this study was to specifically test the impact of AMPK activity on ion flux across intestinal epithelia by utilizing a tissue‐specific knockout mouse model lacking both AMPK‐α catalytic isoforms. Methods Intestinal epithelial cell‐specific knockout of AMPK activity (AMPKα ΔIEC KO ) was achieved by crossing Prkaa1 and Prkaa2 floxed mice (AMPKα fl/fl ) with Villin‐Cre mice. For Ussing chamber experiments, large intestinal segments were stripped of the serosal layer and the mucosal layer was mounted in Ussing chambers containing Kreb's Ringer's buffer. Chloride secretion was stimulated using Forskolin (300μM) applied bilaterally, followed by Carbachol (10μM) applied basolaterally. For immunostaining, whole intestinal tissue was isolated and fixed in 4% paraformaldehyde overnight, washed with 1X PBS, and stored in 30% sucrose in 1X until OCT embedding. 5μm frozen sections of intestinal tissues were sliced and mounted onto glass slides. Results We previously demonstrated AMPKα ΔIEC KO mice do not have a barrier defect indicated by macromolecular intestinal permeability, histological analysis, and response to dextran sulfate sodium (DSS) compared to AMPKα fl/fl mice. We next determined if AMPK alters electrolye permeability in different regions of the large intestine. Indeed, loss of AMPK decreased transepithelial electrical resistance (TEER) in mouse proximal colon (35.21 ± 3.3 Ω·cm2, n = 6–7, p = 0.0007). Conversely, AMPKα ΔIEC KO mice exhibited an increase in basal TEER in the distal colon (62.5 ± 4.5 Ω·cm 2 , n = 2–3, p = 0.2683), although this was not statistically significant. We next tested if AMPKα ΔIEC KO mice exhibited changes in electrogenic ion transport by intestinal region. Preliminary evidence in the cecum indicated increased electrogenic ion transport responses to carbachol and Forskolin in AMPKα ΔIEC KO mice (n = 1–3) while only forskolin‐dependent ion transport was increased in the distal colon (n = 2–3). In the proximal colon, Ca 2+ ‐dependent ion transport was increased two‐fold (n = 6–7, p = 0.1131) while there was no alteration in cAMP‐dependent ion transport (n = 6–7). To determine if these region‐specific effects were associated with changes in transporter expression, colonic sections were stained for the Na + ‐K + ‐Cl − Cotransporter 1 (NKCC1) as a key contributor to electrogenic ion secretion. NKCC1 expression in AMPKα ΔIEC KO mice was similar to that of AMPKα fl/fl mice (n = 2–3). Conclusions AMP‐activated protein kinase mediates region‐specific maintenance of the intestinal barrier and regulation of electrogenic ion transport in mice. Support or Funding Information NIH 2R01DK091281 (DFM) This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.001 |
| Meta-epidemiology (broad) | 0.001 | 0.001 |
| Bibliometrics | 0.001 | 0.000 |
| Science and technology studies | 0.000 | 0.001 |
| Scholarly communication | 0.001 | 0.001 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.001 | 0.002 |
| Insufficient payload (model declined to judge) | 0.004 | 0.002 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".