1. Volume 132, Issue 1, Pages 154-165 (January 2007)
Gastroesophageal Reflux Disease–Associated Esophagitis Induces Endogenous Cytokine Production Leading to Motor Abnormalities 
Florian Rieder, Ling Cheng, Karen M. Harnett, Amitabh Chak, Gregory S. Cooper, Gerard Isenberg, Monica Ray, Jeffry A. Katz, Andrew Catanzaro, Robert O’Shea, Anthony B. Post, Richard Wong, Michael V. Sivak, Thomas McCormick, Manijeh Phillips, Gail A. West, Joseph E. Willis, Piero Biancani, Claudio Fiocchi 
Gastroenterology 
Volume 132, Issue 1, Pages (January 2007)
DOI: /j.gastro
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2. Figure 1
Morphologic appearance of primary human esophageal cells in culture. (A) Tight monolayer with epithelial morphology characteristic of keratinocytes; these cells arose as outgrowths of mucosal explants and are called HEKs. (B) Monolayer with spindle-shaped morphology characteristic of fibroblasts; these cells arose as outgrowths of mucosal explants and are called HEFs. (C) Monolayer with spindle-shaped morphology characteristic of mesenchymal cells; these cells arose as outgrowths of muscularis propria explants and are called HEMCs.
Gastroenterology  , DOI: ( /j.gastro )
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3. Figure 2
Production of enhanced levels of IL-1β in mucosal biopsy specimens of the lower third of the esophagus from patients with endoscopic evidence of esophagitis. A set of 3 endoscopic biopsy specimens was obtained each from the upper, middle, and lower portions of the esophagus, organ-cultured overnight, and the concentration of IL-1β in the undernatants was measured by ELISA. The number of patients was 8. Data are expressed as the mean ± SEM.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2007 AGA Institute Terms and Conditions

4. Figure 3
Production of enhanced levels of IL-6 in mucosal biopsy specimens of the lower third of the esophagus from patients with endoscopic evidence of esophagitis. A set of 3 endoscopic biopsy specimens was obtained each from the upper, middle, and lower portions of the esophagus, organ-cultured overnight, and the concentration of IL-6 in the undernatants was measured by ELISA. Number of patients: ■, esophagitis group = 26; ☐, control group = 19. Data are expressed as the mean ± SEM. *P < .05 for IL-6 in the lower third of the esophagus in the esophagitis group compared with the control group, and for IL-6 levels in the lower compared with the middle and upper esophageal thirds of the esophagus in the esophagitis group.
Gastroenterology  , DOI: ( /j.gastro )
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5. Figure 4
Production of IL-6 in cultures and extracts of human esophageal epithelial cells. Monolayers of HET-1A cells were cultured in the presence or absence of IL-1β or IFNγ + PMA for 48 hours, and the concentration of IL-6 was measured in the supernatants by ELISA. At the end of the culture period, the cells were suspended, washed, and sonicated, and the concentration of IL-6 was measured in cell lysates. Data are expressed as the mean ± SEM of 3 separate experiments. *P < .05 for IL-6 levels in the supernatants and sonicates of stimulated compared with unstimulated cultures. ☐, Medium; , IL-1β; ■, IFNγ + PMA.
Gastroenterology  , DOI: ( /j.gastro )
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6. Figure 5
Cytokine mRNA expression by human esophageal epithelial cells. Monolayers of HET-1A cells were cultured in the presence or absence of IL-1β or IFNγ + PMA for the indicated times, and their total RNA was extracted and submitted to reverse-transcription PCR amplification using cytokine-specific primers. Results are representative of 3 separate experiments.
Gastroenterology  , DOI: ( /j.gastro )
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7. Figure 6
Production of IL-6 in primary cultures of human esophageal cells. Monolayers of HEKs (☐), HEFs (), and HEMCs (■) were cultured in the presence or absence of IL-1β or IFNγ + PMA for 48 hours, and the concentration of IL-6 was measured in the supernatants by ELISA. Data are expressed as the mean ± SEM of 2 separate experiments performed in triplicate.
Gastroenterology  , DOI: ( /j.gastro )
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8. Figure 7
Cytokine mRNA expression by primary cultures of HEFs and HEMCs. Monolayers of HEFs and HEMCs were cultured in the presence or absence of IL-1β or IFNγ + PMA for the indicated times, and their total RNA was extracted and submitted to reverse-transcription PCR amplification using cytokine-specific primers. Results are representative of 2 separate experiments.
Gastroenterology  , DOI: ( /j.gastro )
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9. Figure 8
Gastric juice–induced production of IL-6 by human esophageal epithelial cells. Monolayers of HET-1A cells were cultured in the presence or absence of (A) crude gastric juice (pH 2.5), (B) acidified culture medium (pH 2.5), (C) buffered gastric juice (pH 7.4), or (D) denatured buffered gastric juice (pH 7.4). At the indicated times, the concentration of IL-6 and LDH were measured in the supernatants. Data are expressed as the mean ± SEM of 4–6 experiments. , IL-6; - -◊- -, LDH.
Gastroenterology  , DOI: ( /j.gastro )
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10. Figure 9
Effect of human esophageal mucosal organ culture undernatants on the contraction of cat esophageal circular muscle strips in response to electrical (neural) stimulation. Contraction was induced in response to electrical stimulation consisting of 10-s trains of square wave pulses of supramaximal voltage (0.2-ms duration at 5 Hz) after incubation of the strips in undernatants of mucosal biopsy specimens taken from the upper and lower thirds of the esophagus of patients with endoscopic evidence of esophagitis, GERD symptoms, or control patients. Undernatants from cultures of mucosa from the upper or lower esophagus of the patient control group had no effect on the contraction of esophageal strips. The same results were obtained using undernatants from cultures of mucosa from the upper or lower esophageal thirds of the GERD symptoms group. Undernatants of mucosa taken from the lower third of the esophagus of patients with endoscopic evidence of esophagitis caused a significant (*P < .05, ANOVA) reduction in the amplitude of contraction. In this group, significant reduction of contraction was not observed when the mucosa derived from the esophageal upper third. Data represent the mean ± SEM of 3 separate experiments using 3 strips/cat from 3 different cats. ☐, Medium; , upper esophagus undernatants; ■, lower esophagus undernatants.
Gastroenterology  , DOI: ( /j.gastro )
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11. Figure 10
Effect of human primary esophageal cell culture supernatants on the contraction of cat esophageal circular muscle strips in response to electrical (neural) stimulation. Contraction was induced in response to electrical stimulation consisting of 10-s trains of square wave pulses of supramaximal voltage (0.2-ms duration at 5 Hz) after incubation of the strips in culture medium (☐, control), or cell culture supernatant (■, supernatant), or after washing out the supernatant with culture medium (, wash). Incubation with supernatants from HEKs significantly (P < .05, ANOVA) decreased contraction in response to electrical stimulation. Supernatants from HEMCs also caused a reduction in contractile response, which was less than that induced by HEK supernatants, and was not statistically significant. HEFs had no significant effect on contraction. Contraction of all strips returned to normal after washing out the cell culture supernatant. Data represent the mean ± SEM of 3 separate experiments using 3 strips/cat from 3 different cats.
Gastroenterology  , DOI: ( /j.gastro )
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