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Updated: Aug 10, 2026
Whole Cell Electrophysiology of Primary Cultured Murine Enterochromaffin Cells
Published on: September 26, 2018
The functional characterization of normal and neoplastic human enterochromaffin cells
Irvin M Modlin1, Mark Kidd, Roswitha Pfragner
1Department of Surgery, TMP202, Yale University School of Medicine, 333 Cedar Street, New Haven, Connecticut 06520, USA. imodlin@optonline.net
Context:
Neuroendocrine regulation of small intestinal (SI) function is poorly understood because pure neuroendocrine cells are unavailable, whereas the biological basis of SI carcinoid tumors is unknown because neoplastic human enterochromaffin (EC) cells are unavailable.
Objective:
The objective of this study was to define the secretory regulation and transcriptome of naive and neoplastic SI neuroendocrine cells.
Design:
EC cells from human ilea were isolated and purified, and a malignant EC cell carcinoid cell line (KRJ-I) was characterized.
Methods:
Human ilea from right hemicolectomies were pronase/collagenase digested and Nycodenz gradient centrifuged, and EC cells were fluorescence-activated cell sorting (FACS) sorted after acridine orange labeling. Enrichment was defined by immunostaining, gene expression, serotonin (5-HT) content, and real-time RT-PCR. Naive FACS-sorted EC and KRJ-I cells were cultured, and 5-HT secretion was measured after stimulation with forskolin, isoproterenol, acetylcholine, gamma-aminobutyric acid A (GABA(A)), pituitary adenylate cyclase-activating polypeptide (PACAP)-38, and gastrin. Normal and neoplastic EC cell transcriptomes were acquired by Affymetrix profiling (U133A).
Results:
FACS produced 100 +/- 0.3% (chromogranin A staining) and 99 +/- 0.7% pure EC cells by immunostaining for tryptophan hydroxylase with greater than 67-fold enrichment and a 5-HT content of 180 +/- 18 ng/mg protein (mucosa, 3.5 +/- 0.9). Forskolin- and isoproterenol-stimulated 5-HT secretion was 10-100 times more potent for naive cells (EC(50), 1.8 x 10(-9) m; 5.1 x 10(-9) m) than neoplastic cells (EC(50), 2.1 x 10(-7) m; 8.1 x 10(-8) m), but the effect of PACAP-38 was similar (EC(50), 1 x 10(-7) m). Isoproterenol stimulated cAMP levels 1.6 +/- 0.1-fold vs. basal (EC(50), 2.7 x 10(-9) m). Acetylcholine inhibited naive EC cell 5-HT secretion more potently than neoplastic (IC(50), 3.2 x 10(-9) vs. 1.6 x 10(-7) m), whereas GABA(A) was more potent in neoplastic cells (IC(50), 3.9 x 10(-10) vs. 4.4 x 10(-9) m). Octreotide inhibited naive, but not neoplastic, basal 5-HT secretion. Gastrin had no effect on 5-HT secretion. Comparison of naive and neoplastic transcriptomes revealed shared neuroendocrine and EC cell-specific marker genes. Real-time PCR confirmed that expression of adrenergic (beta1), somatostatinergic (SST(R)2), and neural (VPAC(1) and GABA(A)) receptors occurred on both cell types, but PACAP type 1 (PAC(1)) and cholecystokinin type 2 (CCK(2)) were undetectable. The putative carcinoid malignancy genes (MTA1 and MAGE-D2) were unique to the neoplastic EC cell transcriptome.
Conclusion:
These data support novel methodology to purify live human EC cells for functional characterization and transcriptome assessment, which will allow identification of new targets to control the secretion and proliferation of SI carcinoids.
Insights
Researchers developed a new method to isolate pure human enterochromaffin (EC) cells, enabling the study of small intestinal neuroendocrine regulation and carcinoid tumors. This advance identifies potential targets for controlling carcinoid tumor growth and secretion.
Area of Science:
- Gastroenterology
- Neuroendocrinology
- Oncology
Background:
- Small intestinal (SI) neuroendocrine regulation is poorly understood due to the unavailability of pure neuroendocrine cells.
- The biological basis of SI carcinoid tumors remains unknown because neoplastic human enterochromaffin (EC) cells are unavailable.
Purpose of the Study:
- To define the secretory regulation of naive and neoplastic SI neuroendocrine cells.
- To determine the transcriptome of naive and neoplastic SI neuroendocrine cells.
Main Methods:
- Isolation and purification of EC cells from human ilea using enzymatic digestion, density gradient centrifugation, and fluorescence-activated cell sorting (FACS).
- Characterization of a malignant EC cell carcinoid cell line (KRJ-I).
- Measurement of serotonin (5-HT) secretion and cAMP levels in response to various stimuli, and transcriptome profiling of normal and neoplastic EC cells.
Main Results:
- FACS yielded highly pure EC cells (>99%) with significantly enriched serotonin content.
- Differential regulation of 5-HT secretion by various agonists (e.g., isoproterenol, acetylcholine, GABA(A)) was observed between naive and neoplastic EC cells.
- Transcriptome analysis revealed shared neuroendocrine markers and identified unique genes associated with carcinoid malignancy (MTA1, MAGE-D2).
Conclusions:
- Novel methodology enables purification of live human EC cells for functional and transcriptomic assessment.
- This approach facilitates the identification of new therapeutic targets for controlling SI carcinoid secretion and proliferation.
