Roy C. Orlando

Dilated Intercellular Spaces and Shunt Permeability in Nonerosive Acid-Damaged Esophageal Epithelium

OBJECTIVES:It has recently been established that patients with nonerosive reflux disease have on biopsy within esophageal epithelium a lesion known as dilated intercellular spaces (DIS).METHODS:To further explore the nature and implications of this lesion, in vitro models of nonerosive acid and acid-pepsin damage were created in Ussing chamber-mounted rabbit esophageal epithelium. Using these models circuit analysis and permeability studies were carried out, the latter using dextran of varying size and human epidermal growth factor (EGF).RESULTS:Luminal HCl, pH 1.1, or HCl, pH 2.0 + pepsin, 1 mg/ml, for 30 min significantly reduced transepithelial electrical resistance (RT) but produced no gross erosions or histologic evidence of cell necrosis. Transmission electron microscopy, however, documented the presence of DIS. Circuit analysis on healthy esophageal epithelium showed that shunt resistance (RS) was much lower than apical membrane, basolateral membrane and transcellular resistances (Ra, Rb, and Rcell, respectively) and approached that of RT. Further, circuit analysis on acid and acid-pepsin damaged tissues showed that the declines in RT resulted from declines in RS. Moreover, the declines in RT (and so RS) were associated with a linear increase in permeability to 4 kD dextrans as well as an increase in permeability to 6 kD EGF and dextrans as large as 20 kD.CONCLUSIONS:In nonerosive acid-damaged esophageal epithelium DIS develop in association with and as a marker of reduced transepithelial resistance and increased shunt permeability. This change in shunt permeability upon acid or acid-pepsin exposure is substantial, permitting dextran molecules as large as 20 kD (33 Å) to diffuse across the epithelium. Also, this shunt leak enables luminal EGF at 6 kD to diffuse across the acid-damaged epithelium and by so doing enables it to access its receptors on epithelial basal cells. We hypothesize that the shunt leak of EGF may in part account for the development of a reparative phenomenon on esophageal biopsy in patients with nonerosive reflux disease known as basal cell hyperplasia.

Viscous topical is more effective than nebulized steroid therapy for patients with eosinophilic esophagitis

We performed a randomized trial to compare nebulized and viscous topical corticosteroid treatments for eosinophilic esophagitis (EoE). Subjects with incident EoE (n = 25) received budesonide 1 mg twice daily, either nebulized and then swallowed (NEB) or as an oral viscous slurry (OVB), for 8 weeks. Baseline eosinophil counts for the NEB and OVB groups were 101 and 83 (P = .62). Posttreatment counts were 89 and 11 (P = .02). The mucosal medication contact time, measured by scintigraphy, was higher for the OVB group than the NEB group (P < .005) and was inversely correlated with eosinophil count (R = -0.67; P = .001). OVB was more effective than NEB in reducing numbers of esophageal eosinophils in patients with EoE. OVB provided a significantly higher level of esophageal exposure to the therapeutic agent, which correlated with lower eosinophil counts.

Clinical and manometric effects of nitroglycerin in diffuse esophageal spasm.

THE major function of the esophagus is to transport material from the pharynx to the stomach. In diffuse esophageal spasm, a neuromuscular abnormality exists that can lead to painful swallowing, im...

Pathophysiology of acute acid injury in rabbit esophageal epithelium.

To increase our understanding of the pathophysiology of reflux esophagitis, we sought the early sequence of changes in mucosal structure and function in acutely acid-damage rabbit esophagus. Using a perfused catheter technique esophageal potential difference (PD) profiles were obtained in anesthetized rabbits before, during, and after perfusion of the lower one-half of the esophagus with phosphate-buffered saline or 80 mM NaCl. When acid perfusion reduced the lower esophageal PD by 40-50% or 80-100% of the initial values, the esophagus was removed, sectioned, and the mucosa studied with light microscopy, transmission electron microscopy, and Ussing chamber technique for evaluation of sodium and mannitol transport. The earlier stage of acid damage (PD 40-50%) was associated with reduced mucosal resistance fom 2,180 +/- 199 to 673 +/- 157 ohm cm2 and increased passive transport of sodium (0.10 +/- 0.06 to 1.82 +/- 0.48 microeq/h.cm2) and mannitol (0.008 +/- 0.003 to 0.051 +/- 0.012 microM/h.cm2) (p less than 0.05). There was no significant change in shirt circuit current (0.35 +/- 0.05 to 0.35 +/- 0.04) or net sodium transport (0.32 +/- 0.06 to 0.37 +/- 0.12) at this stage, and the only morphologic finding was dilated intercellular spaces on electron microscopy. The later stage of acid damage (PD 80-100%) exhibited a further reduction in resistance to 299 +/- 65 ohm.cm2 (p less than 0.05), a finding now accompanied by a reduction in short circuit current (0.35 +/- 0.05 to 0.21 +/- 0.04 microeq/h.cm2) and complete inhibition of net sodium transport (0.32 +/- 0.06 to 0.01 +/- 0.13) (p less than 0.05). Morphologic studies at this time revealed cellular necrosis, edema, and vesicle formation in the stratum spinosum. Both gross mucosal changes and transmural necrosis were notably absent. When esophageal perfusion was performed with a combination of acid (80 mM HCl-80 mM NaCl) and pepsin (100 microgram/ml), the morphologic and physiologic findings were essentially the same as with acid alone; however, the time of perfusion to reach either the 50 or 100% reduction in PD was shortened. The findings in this model can be explained on an initial increase in cellular and/or paracellular permeability followed by inhibition of active sodium transport. The resulting loss of osmolar regulation leads to cell necrosis in the stratum spinosum.

Review article: the pathophysiology of gastro-oesophageal reflux disease - oesophageal manifestations.

The pathogenesis of gastro‐oesophageal reflux disease (GERD) is multifactorial, involving transient lower oesophageal sphincter relaxations (TLESRs) as well as other lower oesophageal sphincter (LES) pressure abnormalities. GERD is associated with a decrease in LES pressure, which can be provoked by factors such as foods (fat, chocolate, etc.), alcohol, smoking and medications. These factors have also been shown to increase TLESRs. As a result, reflux of acid, bile, pepsin and pancreatic enzymes occurs, leading to oesophageal mucosal injury, which can potentially progress to oesophageal adenocarcinoma in a minority of patients with Barretts metaplasia. In addition, duodenogastric contents can also contribute to oesophageal injury. Other factors contributing to the pathophysiology of GERD include hiatal hernia, poor oesophageal clearance, delayed gastric emptying and impaired mucosal defensive factors. Hiatal hernia has a permissive role in the pathogenesis of reflux oesophagitis by promoting LES dysfunction. Delayed gastric emptying, resulting in gastric distension, can significantly increase the rate of TLESRs, contributing to postprandial GER. The mucosal defensive factors have an important role in GERD. When excessive acid causes a breakdown in oesophageal epithelial defenses, epithelial resistance may be reduced. Nocturnal GERD is associated with prolonged acid exposure and proximal extent of acid contact, which elevates the risk for oesophageal damage and GERD‐related complications. In sum, GERD is a complex problem caused by many factors that are exacerbated when the patient is in the supine position.

Barriers to paracellular permeability in rabbit esophageal epithelium

Morphological and electrophysiological techniques were used to define the location and nature of the barriers to diffusion across the intercellular space (paracellular pathway) of rabbit esophageal epithelium. Transmission electron microscopy and light microscopy coupled with histochemistry identified a series of tight junctions and an intercellular material staining positively for neutral and acidic glycoconjugates as likely barrier candidates. Additional studies with lanthanum and horseradish peroxidase showed that the barrier to diffusion of tracers was present throughout the stratum corneum and extended to the upper three to seven layers of stratum spinosum and that these findings were most compatible with the presence of the intercellular glycoconjugate material but not the tight junctions. Further positive staining for carbohydrate moieties at the electron microscopic level with periodic acid-thiocarbohydrazide-silver proteinate suggested that the glycoconjugate material was synthesized in the cells of the barrier layers and packaged in intracellular membrane-bound vesicles before secretion into the intercellular space. Although tight junctions were present in series within stratum corneum and, less commonly, extended to two to three cell layers of upper stratum spinosum, analysis of tracer studies, freeze-fracture replicas, electrophysiological data, and mannitol fluxes, while not conclusive, provided little to support a major role for these junctions in barrier function in this tissue.

Barrett's Esophagus: Clinical, Endoscopic, Histologic, Manometric, and Electrical Potential Difference Characteristics

The clinical, endoscopic, histologic, manometric, and esophageal potential difference characteristics of 20 patients with columnar epithelia lining the lower esophagus (Barretts esophagus) are presented. Endoscopically, two distinct types were identified: a circumferential-type and an island-type Barretts esophagus. Patients with these types exhibited similarities in mean age, duration of symptoms, mean lower esophageal sphincter pressure, and frequency of gross esophagitis. Only patients with the circumferential lesion, however, had esophageal strictures or esophageal ulcers. Manometric testing revealed a range of lower esophageal sphincter pressures from 3 to 33 mmHg and qualitative motor abnormalities (i.e., aperistalsis, repetitive waves, tertiary waves) in 3 patients. Histologically, the frequency of epithelial types was junctional greater than specialized columnar greater than atrophic fundic epithelium. More importantly, dysplasia was identified in 2 patients with the circumferential lesion and in 1 patient with the island lesion. Potential difference measurements demonstrated that a high potential difference (greater than -25 mV) was highly specific (92%), but only moderately sensitive (70%) for detecting Barretts esophagus. Based on these findings, we conclude (a) that there are at least two endoscopically distinct types of Barretts esophagus involving the lower esophagus--a circumferential type and an island type, (b) that both types are associated with chronic gastroesophageal reflux, with the island type being accompanied by less severe epithelial injury than the circumferential type, and (c) that the identification of dysplasia in the two types suggests that both are unstable lesions requiring continued surveillance with endoscopy and biopsy.

Role of e-cadherin in the pathogenesis of gastroesophageal reflux disease

OBJECTIVES:An early event in the pathogenesis of gastroesophageal reflux disease (GERD) is an acid-induced increase in junctional (paracellular) permeability in esophageal epithelium (EE). The molecular events that account for this change are unknown. E-cadherin is a junctional protein important in barrier function in EE. Therefore, defects in barrier function in EE were sought in GERD as well as whether their presence correlated with abnormalities in e-cadherin.METHODS:Endoscopic biopsies of EE from GERD (n=20; male 10; female 10; mean age 50±10 years) and subjects with a healthy esophagus (controls; n=23; male 11; female 12; mean age 51±11 years) were evaluated in mini-Ussing chambers and by western blot and immunochemistry; and serum analyzed by enzyme-linked immunosorbent assay (ELISA). A role for e-cadherin was also assessed using a unique conditional knockout of e-cadherin in adult mouse esophagus.RESULTS:EE from GERD patients had lower electrical resistance and higher fluorescein flux than EE from controls; and the findings in GERD associated with cleavage of e-cadherin. Cleavage of e-cadherin in GERD was documented in EE by the presence of a 35-kDa, C-terminal fragment of the molecule on western blot and by an increase in soluble N-terminal fragments of the molecule in serum. Activation of the membrane metalloproteinase, A Disintegrin And Metalloproteinase (ADAM-10), was identified as a likely cause for cleavage of e-cadherin by western blot and immunostaining and a role for e-cadherin in the increased junctional permeability in EE from GERD supported by showing increased permeability after deletion of e-cadherin in mouse EE.CONCLUSIONS:The EE in GERD has increased junctional permeability and this is in association with proteolytic cleavage of e-cadherin. As loss of e-cadherin can, alone, account for the increase in junctional permeability, cleavage of e-cadherin likely represents a critical molecular event in the pathogenesis of GERD, and identification of cleaved fragments may, if confirmed in larger studies, be valuable as a biomarker of GERD.

Prevention of stress-induced gastric ulcers by dopamine agonists in the rat

Dopamine (DA) and DA agonists have been shown to exert a protective role against the formation of duodenal ulcers. The effect of stimulation of DA receptors on the development of stress-induced gastric ulcers is currently unknown. Accordingly, we evaluated the effect of several DA agonists on the development of gastric ulcers induced by 3 h of cold + restraint stress (CRS) in rats. Apomorphine, d-amphetamine, methylphenidate, and threo-dl-p-hydroxymethylphenidate (an hydroxylated analog of methylphenidate), significantly reduced both the incidence and severity of CRS-induced gastric ulcers. The gastric cytoprotection afforded by these agents was dose-related, and completely antagonized by pretreatment with the peripherally acting DA antagonist domperidone. Because domperidone blocks peripheral, but not central, DA receptors, and since the entry of threo-dl-p-hydroxymethylphenidate across the blood-brain barrier into the brain is restricted to a great extent, we conclude that stimulation of peripheral DA receptors is primarily involved in the gastric cytoprotection induced by dopamimetics.

Failure of clinical criteria to distinguish between primary achalasia and achalasia secondary to tumor

Three clinical criteria have been reported to distinguish patients with primary achalasia from patients with achalasia secondary to tumor invasion of the gastroesophageal junction. These criteria (age greater than 50 years, duration of symptoms less than one year, and weight loss greater than 15 pounds) are important because of their potential use for deciding between pneumatic dilation and exploratory surgery. In the present investigation we assessed the frequency of these criteria alone and in combination in 79 patients with primary and in two patients with secondary achalasia seen at our institution over a 91/2-year period. Our results indicate that while these criteria are highly sensitive and moderately specific, their predictive value for distinguishing secondary achalasia from primary achalasia is exceedingly low. For this reason, early exploratory surgery is not indicated in patients with newly diagnosed achalasia who meet these criteria unless there is prior radiologic or endoscopic evidence for tumor.