Francisco Esteva

Risk factors associated with refractory peptic ulcers

BACKGROUND & AIMS The risk factors associated with refractory peptic ulcers are still undefined. The purpose of this study was to identify these factors in a multivariate context. METHODS Clinical and endoscopic findings as well as Helicobacter pylori status, gastric secretion analysis, serum gastrin levels, nonsteroidal anti-inflammatory drug (NSAID) use, and objective testing of aspirin use by platelet cyclooxygenase activity were studied in 60 consecutive refractory patients with peptic ulcer and 54 matched nonrefractory controls. RESULTS Refractory patients had a longer history of symptomatic ulcer, had an earlier onset, had more frequent relapses, and smoked more during the episode of refractoriness. H. pylori status was similar in both groups, but H. pylori eradication in a subset of refractory patients (23 of 26) was highly effective in healing these ulcers (14 of 23). Globally, NSAID-analgesic abuse (including > 1500 mg/day paracetamol) was present in 40% of refractory patients (P < 0.006). Objective testing showed that 43.7% of NSAID use was surreptitious. Multivariate logistic regression analysis identified only NSAID and analgesic abuse and the number of relapses as individually affecting refractoriness. CONCLUSIONS NSAID and analgesic abuse is the single most important exogenous factor associated with refractoriness. H. pylori infection emerges as an important intrinsic factor, but almost a quarter of refractory patients cannot be linked to either NSAID use or H. pylori infection.

Omeprazole Does Not Interfere with the Antiplatelet Effect of Low-Dose Aspirin in Man

Background: Experimental studies have shown that omeprazole and other anti-inflammatory agents compromise the therapeutic activity of nonsteroidal anti-inflammatory drugs and aspirin in rats. It is not known whether this effect will occur in humans. Our aim was to determine whether omeprazole affects the antiplatelet effects of low-dose aspirin in humans. Methods: Platelet lumiaggregation, skin bleeding time, plasma levels of aspirin and salicylic acid, and serum gastrin levels were determined in 14 healthy men before and after the ingestion of 125 mg aspirin with or without previous treatment with 20 mg/day of omeprazole for 4 days before testing. Results: Omeprazole increased serum gastrin levels from 64 (median; range, 52-91) pg/ml to 80.5 (median; range, 56-455) pg/ml (P < 0.05) but did not significantly affect the plasma concentration of aspirin or salicylic acid. Aspirin increased skin bleeding time in all subjects, and the increase was similar with and without previous omeprazole treatment. Aspi...Background: Experimental studies have shown that omeprazole and other anti-inflammatory agents compromise the therapeutic activity of nonsteroidal anti-inflammatory drugs and aspirin in rats. It is not known whether this effect will occur in humans. Our aim was to determine whether omeprazole affects the antiplatelet effects of low-dose aspirin in humans. Methods: Platelet lumiaggregation, skin bleeding time, plasma levels of aspirin and salicylic acid, and serum gastrin levels were determined in 14 healthy men before and after the ingestion of 125 mg aspirin with or without previous treatment with 20 mg/day of omeprazole for 4 days before testing. Results: Omeprazole increased serum gastrin levels from 64 (median; range, 52-91) pg/ml to 80.5 (median; range, 56-455) pg/ml (P < 0.05) but did not significantly affect the plasma concentration of aspirin or salicylic acid. Aspirin increased skin bleeding time in all subjects, and the increase was similar with and without previous omeprazole treatment. Aspirin inhibited platelet aggregation in response to both collagen and arachidonic acid regardless of omeprazole treatment. Conclusion: A single dose of 20 mg omeprazole daily does not interfere with the biologic activity of 125 mg aspirin on platelets in humans.

Effect of Growth Factors and Prostaglandin E2 on Restitution and Proliferation of Rabbit Esophageal Epithelial Cells

Factors and mechanisms involved in esophagealmucosal injury and repair are not well known. The aim ofthis study was to assess the effects of growth factorsand prostaglandins on esophageal mucosal cell repair activities. Rabbit esophageal cells wereisolated, cultured, and exposed to different growthfactors and prostaglandin E2. Subconfluentcell cultures were used to study proliferative responsesdetermined by [3H]thymidine incorporation intoDNA. Restitution was studied in confluent monolayerswounded by mechanical denudation. Restitution was themain mechanism involved in wound repair within the first24 hr. HGF, IGF-I, and EGF dose-dependentlystimulated cell proliferation but did not affectrestitution. TGF-β1 inhibited bothproliferation and restitution while PDGF-BB andprostaglandin E2 had no effect. Esophageal epithelial cell restitution andproliferation are affected by growth factors. HGF,IGF-I, EGF (stimulation), and TGF-β1(inhibition) are major growth factors affecting in vitroesophageal wound repair activities, which, unlike those ofother areas of the digestive tract, are not affected byprostaglandins.

Aspirin renders the oesophageal mucosa more permeable to acid and pepsin

Objective: To examine the effects of aspirin on the oesophageal mucosa and on acid- and pepsin-induced oesophagitis Design and methods: The effects both of intraluminal (18 mg/ml) and of parenteral (100 mg/kg per h) aspirin on an in-vivo rabbit model of oesophagitis induced by acidified pepsin (pH 2) were studied. Oesophageal injury was assessed by macroscopic and microscopic scoring including the cell proliferation immunohistochemical parameter mib 1. The mucosal barrier function was determined by hydrogen, potassium and haemoglobin flux rates. Results: Acidified saline alone caused no damage, but the addition of aspirin induced mucosal barrier damage (P<0.05). The exposure of the oesophageal mucosa to acidified aspirin and then acidified pepsin significantly increased mucosal injury and mucosal barrier dysfunction compared with control experiments (exposure to acidified saline and acidified pepsin). This damage was significantly (P<0.05) reduced (>40%) by prostaglandin cotherapy (prostaglandin E2) administered before acidified aspirin exposure. Mucosal damage was less severe (P<0.05) when the oesophageal mucosa was exposed to a pH 6 aspirin solution. Parenterally administered aspirin also increased the oesophageal damage induced by acidified pepsin compared with control experiments, but the damage was 23% lower than that obtained with intraluminal aspirin. Cell proliferation studies showed a significant increase in the number of positive cells in those experiments with a higher degree of damage and in those treated with prostaglandins. Conclusion: Aspirin renders the oesophageal mucosa more permeable to acid and pepsin. These effects are in part part pH-dependent and might be partially reversed by prostaglandin E2 cotherapy.

Superoxide Anion and Nitric Oxide in High-Grade Esophagitis Induced by Acid and Pepsin in Rabbits

It has been proposed that free radicals are involved in the pathogenesis of esophageal mucosal damage induced by acid and pepsin. Recent data have suggested that nitric oxide (NO) is involved in the mucosal defense of the esophagus and that superoxide anion plays a minor role in low-grade esophagitis. To study the role and potential interaction of NO and superoxide anion in an experimental model of high-grade esophagitis, acidified pepsin was perfused (45 min/12 hr) for five days in rabbits with different agents to modulate the generation of these radicals. Measurements included both macroscopic and microscopic mucosal damage, superoxide anion generation, NO synthase mucosal activity, and peroxynitrite formation. High-grade esophagitis was associated with mucosal superoxide anion generation. Treatment with exogenous superoxide dismutase completely prevented mucosal damage. The perfusion of acidified pepsin in the lumen of the esophagus was initially associated with increased NO synthase mucosal activity but decreased with the progression of damage. Generation of peroxynitrites was present in those cases with severe damage. Treatment with NO-modifying agents did not induce consistent modification of mucosal damage. It is concluded that superoxide anion is involved in the induction of high-grade esophagitis and that it interacts with nitric oxide to generate peroxynitrite radicals in this model. Superoxide dismutase but not NO-donor-modifying agents might have a therapeutic role in preventing severe esophageal mucosal damage induced by acid and pepsin.

Platelet-Derived Growth Factor and Epidermal Growth Factor Play a Major Role in Human Colonic Fibroblast Repair Activities

Background and Aims: Ulceration is a common feature of inflammatory bowel diseases, where subepithelial cell growth is frequently necessary for resolution. In order to further understand the role of colonic fibroblasts in this process, we have used an in vitro model of wound repair to study the response of human colonic fibroblasts to several growth factors expressed in colonic tissues. Methods: Proliferation was determined by [3H]thymidine incorporation into DNA in subconfluent fibroblast cultures. In vitro wound repair was determined in confluent fibroblast monolayers after mechanical denudation. The presence of growth factors secreted by fibroblasts was studied in conditioned medium by heparin affinity chromatography and immunodetection with specific antibodies. Results: Serum and platelet-derived growth factor (PDGF-BB) induced a dramatic increase in both colonic fibroblast proliferation and closure of wounded cell monolayers. Epidermal growth factor (EGF) stimulated both fibroblast activities, but the effect was less potent. However, colonic fibroblasts did not respond to transforming growth factor-β1. Conditioned medium stimulated fibroblast proliferation and wound repair activity, which was reverted by the addition of suramin. Furthermore, a PDGF-like factor was isolated from colonic fibroblast-conditioned medium. Conclusions: EGF and PDGF-BB promote human colonic fibroblast-dependent wound repair activities. Human colonic fibroblasts may exert an autocrine regulation via the production of growth factors.

Nitric oxide and superoxide anion in low-grade esophagitis induced by acid and pepsin in rabbits.

It has been suggested that free radicals are involved in esophagitis. To study the role and potential interaction of superoxide anion and nitric oxide (NO) in low-grade esophagitis, we perfused acidified pepsin (30 min every 12 hr) for seven days in rabbits treated with different agents to modulate the generation of these radicals. Measurements included macroscopic and microscopic damage, superoxide anion generation, mucosal nitric oxide synthase activity, and peroxynitrite formation. Low-grade esophagitis was associated with increased nitric oxide synthase mucosal activity and mucosal damage was dose-dependently increased by treatment with the NO synthase inhibitor NG-nitro-l-arginine. Superoxide anion was scarcely generated in the mucosa, but this was not accompanied by any change in the activity of mucosal superoxide dismutase. Treatment with superoxide dismutase did not improve mucosal damage. Generation of peroxynitrites was not detected. In conclusion, nitric oxide is involved in the mucosal defense of the esophagus against acid- and pepsin-induced damage. Superoxide anion generation seems irrelevant in the induction of low-grade esophagitis and not sufficient to interact with nitric oxide to generate measurable mucosal peroxynitrite radicals.

Selective COX-2 inhibition is associated with decreased mucosal damage induced by acid and pepsin in rabbit esophagitis.

Objective: To evaluate the effects of COX-1 and/or COX-2 inhibition in a model of chronic esophagitis in rabbits. Methods: Both high- and low-grade esophagitis were induced in rabbits by the perfusion of acidified pepsin. Rabbits were treated with either a selective COX-2 inhibitor (DFU[3-(3-Fluorophenyl)-4-(4-Methanesulfonyl)-5,5-Dimethyl-5H-Furan-2-One];30mg/Kg/day), a nonspecific COX inhibitor (indomethacin; 2 mg/Kg/day), or a COX-1 preferential inhibitor (piroxicam; 2 mg/Kg/12 h). Results: Prostaglandins are derived from COX-1 activity in the normal esophagus. Both low- and high-grade esophagitis are associated with a progressive increase of COX activity, which is partially dependent on the COX-2 isoform. DFU reduced muscosal damage in both models of esophagitis. However, indomethacin did not affect significantly mucosal damage, and piroxicam increased damage in low-grade esophagitis. Conclusions: COX-1 activity is constitutive in the rabbit esophageal mucosa, but both COX-2 and COX-1 activity are increased under the impact of acidified pepsin. Treatment with the COX-2 inhibitor DFU is associated with improvement of mucosal damage, which may have therapeutic implications.

Cyclooxygenase inhibitors decrease the growth and induce regression of human esophageal adenocarcinoma xenografts in nude mice

Cyclooxygenase (COX) inhibition has been shown to prevent the development of esophageal adenocarcinoma (EAC). However, the potential of this approach for treatment of established cancer has been poorly investigated. Our objective was to determine whether non-selective or selective inhibition of the COX pathway affects the growth of esophageal adenocarcinoma xenografts in nude mice. A human esophageal adenocarcinoma xenograft model was established by subcutaneous inoculation of OE33 cells in nude mice. Small tumor slices harvested from four OE33 xenografts were implanted in the flanks of new mice that were randomized to different treatments (6 animals per group): indomethacin (3 mg/kg/day), parecoxib (0.11 and 0.22 mg/kg/day) or a selective prostaglandin E₂ receptor antagonist (AH-23848B, 1 mg/kg/day). For each treatment, a control group of 6 animals (vehicle) carrying xenografts from the same OE33 tumor was included. Tumor growth was measured twice a week. After 8 weeks mice were euthanized. Tumors were assessed by histological analysis, mRNA expression of COX isoenzymes, PGE₂ receptors and PGE₂ content. All OE33 tumors were poorly differentiated esophageal adenocarcinomas. Tumors expressed COX-2, EP₁, EP₂ and EP₄ receptor mRNA. Treatment with parecoxib, higher dose or indomethacin significantly inhibited tumor growth. Furthermore, indomethacin induced tumor regression (74 vs 582% in control animals; p<0.01). However, AH-23848B or parecoxib low dose failed to affect tumor growth significantly. PGE₂ content in tumors was significantly decreased by high-dose parecoxib and indomethacin. Indomethacin and parecoxib inhibit the growth of human esophageal adenocarcinoma xenografts in nude mice, which suggests a potential role for NSAIDs or selective COX-2 inhibitors for EAC chemotherapy.

In vitro wound repair by human gastric fibroblasts: Implications for ulcer healing

Fibroblasts modulate epithelial biologicalactivities and play a key role in the ulcer healingprocess. There is no information regarding thebiological response of human gastric fibroblasts toregulatory compounds. The aim of this study was to assessthe effects of growth factors and prostaglandins on anin vitro model of human gastric fibroblast wound repair.Subconfluent fibroblast cultures were used to study proliferative responses, determined by[3H]thymidine incorporation into DNA. Invitro wound repair was determined in confluentfibroblast monolayers after mechanical denudation. Thepresence of putative growth factors secreted by fibroblastswas studied in conditioned medium by heparin-affinitychromatography and immunodetection with specificantibodies. Serum and platelet-derived growth factor (PDGF)-BB induced a dramatic increase in bothgastric fibroblast proliferation and closure of woundedcell monolayers, whereas these activities were inhibitedby both transforming growth factor(TGF)-β1 and prostaglandin E1. Basalactivities in unstimulated gastric fibroblasts werelower than those obtained in skin fibroblasts.Conditioned medium stimulated fibroblast proliferationand wound repair activity, which was inhibited by the addition of suramin,and was partially dependent on the presence of PDGF-likefactor. PDGF is a major, autocrine promotor of humangastric fibroblast-dependent wound repair activities, which are inhibited by prostaglandins andTGF-β. These findings might be important for futuretherapeutic ulcer healing approaches.