The authors have declared that no competing interests exist.
Esophageal motor abnormalities are frequently found in patients with gastroesophageal reflux disease. The effect of bile in esophageal dysmotility is probably secondary to mucosal signaling to the muscular layer and not a transmural process. This study aims to identify the mucosa-muscular signaling path by receptors blockage in an experimental study.
Fifteenguinea pig esophagi were isolated and
Contraction amplitudes after bile incubation but before antagonist incubation were 1.6±0.6 g, 1.2±0.8 g, and 1.2±0.4 g for atropine, suramin and genistein, respectively. Mean contraction amplitudes after antagonists instillation were 1.2±0.6 g, 1.4±0.5 g, 0.9±0.2 g, respectively. There was no different in contraction amplitude before and after instillation of atropine (p=0.188), suramin (p=0.488) or genistein (p=0.079).
Our results show that blockage of cholinergic (atropine), purinergic (suramin) or tyrosine kinase (genistein) paths do not change esophageal dysmotility induced by bile. Other molecular path may play the role in this scenario.
Esophageal motor abnormalities are frequently found in patients with gastroesophageal reflux disease (GERD).
It is still elusive the molecular path responsible to this signaling. Since a myriad of paths are linked to smooth muscle contraction and relaxation, this study aims to try to identify the exact nature of the mucosa-muscular signaling path by receptors blockage from 3 different paths found in the esophageal muscle in an experimental study.
Fifteen guinea pigs (3 months, males, weight 200-250g) were studied. Animals were killed by spinal cord injury and cervical incision after 24 hours fasting. Esophagi were isolated, and its contractility assessed with force transducers.
Animals were randomly assigned into 3 groups:
Group A (n=5): ursodeoxycholic acid + atropine
Group B (n=5): ursodeoxycholic acid + suramine
Group C (n=5): ursodeoxycholic acid + genistein
Contraction experiments followed previous protocol.
The esophagi were incubated in 100 µM ursodeoxycholic acid for 1 hour and 5 sequential contractions induced by 40 mM KCl spaced by 5 minutes were recorded. Wash out was carried out between contractions. Then, esophagi specimens were incubated in 3 different smooth-muscle contraction antagonists: atropine (1µM), suramin (1µM) or genistein (1µM) according to groups division. After 30 minutes, the same protocol for contractions was repeated.
Variables are expressed as mean ± standard deviation (range) and encompass the mean of 5 stimuli. Paired Student t test was used when appropriated since data followed a normal distribution according to the Kolmogorov–Smirnov test. A value of p was considered significant at the 0.05 level. A sample size calculation was performed based on the results of a previous study
The study was approved by the Institutional Review Board (#1390/11). There are no conflicts of interest. The authors are responsible for the manuscript and no professional or ghost writers were hired.
Two (13%) esophagi specimens were unresponsive after 2 hours incubation. Their data was excluded and the experiment was repeated with new specimens for a total of 15.
Esophageal contraction amplitudes before and after incubation with antagonists are depicted in
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1.609 ± 0.628 (0.708 – 2.384) | 1.232 ±0.638 (0.686 – 2.252) | 0.188 |
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1.173 ± 0.778 (0.528 – 2.464) | 1.450 ± 0.462 (0.808 – 1.786) | 0.448 |
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1.218 ± 0.406 (0.764 – 1.806) | 0.913 ± 0.191 (0.676 – 1.130) | 0.079 |
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No difference in contraction amplitudes before and after incubation with antagonists were noticed for atropine (p=0.188, paired T test), suramin (p=0.449, paired T test) or genistein (p=0.079, paired T test).
Bile duodeno-gastroesophageal reflux has been implicated in the genesis of GERD symptoms.
The objective of the current study was to evaluate changes in the amplitude of esophageal contraction of guinea pigs esophagi when exposed to 3 different contraction antagonists to try identifying mucosa-muscular signalization pathways induced by ursodeoxycholic acid exposure: (a) atropine due to its anti-cholinergic effect, (b) suramin for its purinergic blockage and (c) genistein for its anti-tyrosine kinase activity.
The study protocol employed a careful methodology with isolated organ perfusion equipment based on the expertise from previous studies from the laboratory of the Department of Biophysics.
The same methodology was previously used as a model of bile-induced dysmotility that validated the use of: (a) guinea-pigs as the animal model; (b) KCl as the contraction inductor; (c) viability of isolated esophagi specimens; and (d) ursodeoxycholic acid as one component of bile
Suramin is a molecule derived from the urea that has been used as an antimicrobial for the treatment of trypanosomiasis
Imaeda et al.
Atropine is a natural alkaloid found in some herbs, such as
In humans, atropin leads to esophageal hypocontractility measured by manometry.
Genistein, daidzein and glycitein are the main isoflavones – phytostrogens found in plants such as the soy.
Previous studies that focused on the action of genistein on the esophagus were conducted in isolated cultured cells only. Our results, similar to the other tested substances, did not support the paths inhibited by genistein as the mediators to bile-induced dysmotility.
The current experiment was limited to the study of the action of a single component of bile (ursodeoxycholic acid) in guinea-pig esophagi strips stimulated by chemical depolarization and incubated with a single dose of 3 different smooth muscle contraction antagonists. The control group to test the effects of Kcl and bile came from a previous publication