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The effect of gastric metaplasia (GM) of the duodenal bulb on proximal duodenal mucosal bicarbonate secretion (DMBS)
April 1995
•
HIGH SYSTEMIC HCO3 AND TOPICAL 16,16-DIMETHYL PGE2 PROTECT THE GASTRIC MUCOSA AGAINST LUMINAL ACID BY ENHANCING ITS PRE-EPITHELIAL BUFFER CAPACITY T.Ranta-Knuuttila, H.Mustonen, E.Kivilaakso. II Dept. of Surgery, Helsinki University Central Hospital, Helsinki, Finland Systemic metabolic (high-HCO3) alkalosis (Gastro 81:921,1981) and topical prostaglandins (DDS 34:1028,1989) protect the gastric mucosa against luminal acid. This study investigates whether this protection is mediated by increased epithelial HC03 secretion with resultant alkalisatinn of the pre-epithelial mucus HCO3 buffer layer. Methods: Surface pH (pHs) of chambered ex rive rat gastric epithelium was measured with blunt-tipped liquid sensor pH-microelectrodas positioned under microscopic control in light contact with the epithelial surface. The lumen was perfused with increasing luminal acidities (0, 10, 30, 50 and 100 mM HC1). The experimental groups were; I : Control (N=9)(arterial pH 7.3020.04, aHC03 19.16~-1A9 mM), II: topical 16,16-dimethyl-PGE2 (100~g/100ml) treatment (N=7) (a-pH 7.30±0.01, a- HC03 16.7320.78 r a M ) , III: high HC03 metabolic alkalosis (N=7) ( 3.6 mmol/kg/h of NaHCO3 i.v., a-pH 7.37±0:02, a-HCO3 33A4-3.5mM), III: low -HC03 respiratory aikalosis by mandatory hyperventilation (N=9) (a- pH 7.4920.02, a-HC03 g.3± 0.9 raM). Results: pHs:s during exposure to increasing luminal acidities are given in the Table. PGE2 treated and high-HCO3 alkalotic rats tolerated significantly better luminal acid than controls, but in low-HCO3 alkaiosis tolerance to acid was significantly impaired. In alkalotie rats there was a significant correlation between arterial HCO3 concentration and pHs at luminal 30 mM HC1 (r=0.81, p<0.01), whereas no correlation was found between arterial pH and pHs (i"=0.27, p>0.05), suggesting that high HC03 concentration rather than high systemic pH provides protection. pHs (N) pH6 HCI 10 HCI 30 HCI 50 HCI 100 Control (9) 7.704-0.08 7.4620.10 7.204-0.11 6.93±0.14 6.34±0.25 PGE2 (7) 7A920.10 7.4020.11 7.214-0.17 7.034-0.18 7.0120.16" High-HCO 8.034-0.04 * 7.904-0.07* 7.724-0.11" 7.474-0.20* 7.344-0.22* Low-HCO 7.804-0.07 7.434-0.13 6.54±0.28* 6.124-0.09" • =p<0.05 vs. control Conclusions: The protective actions of systemic metabolic alkalosis and topical PGE2 treatment in gastric mueosa against luminal acid are, at least in part, mediated by enhanced buffer capacity of the pro-epithelial mueus-HC03 layer.
• INCREASED T R A N S L O C A T I O N OF pp600"~ FROM PLASMA MEMBRANE TO CYTOSOL BY TGF-a DURING THE E A R L Y R E P A R A T I V E PHASE O F GASTRIC MUCOSA AND EFFECTS O F AGING. N.K. Relan, R.R. Matta and A.P.N. Majumdar, VA Medical Center, Allen Park, MI 48101 and Dept. Med. Wayne State Univ. Detroit, MI. Although the gastric mucosa of healthy adult animals possesses a remarkable capacity to regenerate promptly after an acute injury, we have shown that aging diminishes its regenerative ability. Little is known about the regulatory mechanisms for the reparative processes. We suggested that decreased ligand-induced activation of EGF-reeeptor (EGF-R) tyrosine kinase (Tyr-k) in aged rats during the early reparative phase (restitution) may contribute to diminished mucosal repair in these animals (Gastroenterology 106: A629, 1994). To further elucidate the EGF-R-induced signal transduction processes during restitution, we examined the changes in Tyr-k activity of pp60 ~ and its translocation from membrane t o cytosol in the gastrie mucosa of young (4-me) and aged (24-m0) rats during the first 60 rain after i.g. administration of either 2 M NaCI (1.3 mill00 g) or water (controls). Data from control animals revealed that Tyr-k activity of pp60" ~ was about 50% higher in the gastric mucosa of aged than in young rats. Injury caused a prompt stimulation of mucosal pp60 ~ Tyr-k (measured in detergent-soinbillzed mucosa) in both age groups which attained its maximal stimulation (70-80% increase) at 30 rain after injury, where,after it declined sharply. However, when the relative concentration of pp60"I~ (assessed by Western-blot) was determined in the cytosolie fraction of the gastric mucosa (measure of pp60 ~ release) at 30 rain after injury, we observed a siguificanfly greater release (2-3-fold) in young than in aged rats, when compared with the corresponding controls. These changes were accompanied by parallel alterations in Tyr-k activity of pp6ff "~ in the cytosolic fraction. To further determine the rate of release of pp60~ from membrane to cytosol, the 30,000g membrane fraction (0.5 mg protein) from 30 rain post-injured rats was incubated in the absence (basal) or presence of lxl0 -9 M TGF-~t for up to 20 rain. The incubation medium (containing released pp60~ ) and the membrane fraction, recovered by centrifugation, were assayed for pp60 ~ levels (Western-blot) and its Tyr-k activity. TGF-c~ caused a significant 80-100% increase in pp60 ~ release from membranes of 30 rain post-injured young but not from aged rats. The membrane fraction from young injured rats showed a corresponding reduction in pp60 ~ concentration. These changes were also accompanied by parallel alterations in Tyr-k activity of pp60~ in the cytosol. Our observation suggests that ligand-induced activation of EGF-R Tyr-k in the gastric mueosa during the early reparative phase results in pp6ff "~ release from the membrane, and that pp60 ~ may play a role in signal transduction process.
Esophageal, Gastric, and Duodenal Disorders
A201
• THE EFFECT OF GASTRIC METAPLASIA (GM) OF THE DUODENAL BULB ON PROXIMAL DUODENAL MUCOSAL BICARBONATE SECRETION (DMBS). R.C. Rapier, M.A. Koss, A.D. Dreilinger, L.M. Nyberg, V.S. Pratha, D.L. Hogan, J.l. Isenberg. Depts of Medicine and Pathology, University of California at San Diego, San Diego, CA. Gastric metaplasia (GM) of the duodenal bulb is reportedly more common in patients with DU vs normal (NL} subjects and is required for adherence of Helicobacterpylori (HP;Gut 1993;34:1510). Our aim was to determine the frequency and effect of GM on human proximal DMSS in both NL and DU subjects. To date, 13 NL subjects (mean age 49 yr, 12 males) and 8 DU patients with healed ulcers (mean age 5t yr, 5 males) were evaluated prospectively. Each subject underwent upper gastrointestinal endoscopy. Antral biopsies were obtained to determine HP status (CLOtest ® and histology, observer blinded). Also, 2 "jumbo" duodenal bulbar biopsies, directed at areas of gastric metaplasia (mucosa "washed" with acetylcysteine, then stained with methylene blue; (Gastrointest Endosc 1992;38:696) were obtained for histological examination (observer blinded). DMBS was measured by isolating the proximal 4 cm of duodenum between occluding balloons using previously validated methods. Basal and acid-stimulated (20 ml, 100 mM HCI, 37°C, over 5 min) DMBS were measured. Ten of 13 NL (77%) and 5 of 8 DU patients (63%) had evidence of GM, an insignificant difference (P > 0.5). In the NL subjects, basal, peak and total acid-stimulated DMBS were comparable in those with GM (Table; means -+ SEMs). Also, in the DU patients (all HP+) basal and acid-stimulated DMBS was similar in those with, versus those without GM (Table). Moreover, in the 10 NL subjects with GM, DMBS was similar in those HP- (n = 8) versus those HP+ (n = 2). DUODENAL MUCOSAL BICARBONATESECRETION BASAL (pmol/cm-h)
PEAK (pmol/cm-h)
TOTAL (/.'mol/h)
NL GM+(IO)
217 ± 24
338 2 37
987 2 97
NLGM- (3)
139 2 5
405 2 109
1059+_ 136
DU GM+(5)
118 + 23
233 + 45
633 2 105
164 2 63 226 ± 81 619 2 164 DU GM- (3) Thus, these findings suggest that the presence of gastric metaplasia of the duodenum alters neither basal nor acid-stimulated proximal duodenal mucosal bicarbonate secretion in normal and duodenal ulcer patients.
STRESS ULCERATION:ROLE OF CAPSAJCIN-SENS1TIVE NERVES IN RAT MODEL. J Ren, H Ojeas, M Kids, K Brewer, J Gao, T Hoag and R Harry. Department of Medicine, University of Oklahoma and Oklahoma City DVAMC, Oklahoma City, OK. Acute stimulation of capsaicin-sensitlve nerves has been shown to protect gastric mucosa from various noxious insults. In contrast, chronic capsaicin treatment which causes sensory deafferentation enhances mucosal injury to luminal agents. The objectives of this study were to determine the effect of acute and chronic capsaiein administration in a rodent model of stress ulceration and to establish correlations between gastric calcitonin gone-related peptide (CGR15) content and the degree of gastric mucosal injury. Methods: The following groups of adult S-D rats were subjected to orogastric intubation without or with subsequent stress by restraint and water immersion for 4 hours: The control group (I) consisted of rats not subjected to stress; II-stress: orogastric administration of vehicle (1 ml solution: ethanol 10%, Tween 80 10%, saline 80% v/v/v) at the beginning of the experiment CI'o~and after 2 hours (T2) of stress; Ill-acute capsaicin: orogastric dosing (1.3 mg/kg) at TO and T2; and IV-chronic capsaicin treatment: 150 mg/kg subcutaneously on 3 consecutive days, 2 weeks prior to orognstric vehicle administration. The number of ulcers and the area of ulceration (ram 2) were quantitated. Data shown for each group represent the results of 8 experiments. CGRP antral content was determined by radioimmunoassay. Results: In group II (stress) there were 7.6 -. 0.4 ulcers encompassing 40.9 ~ m 2. Group III (acute capsaicin) the ulcer incidence and area of injury were reduced significantly to 3.2 ± 0.4 and 5.1 +- 0.2 mm2 , respectively (p < 0.01). In contrast, group IV animals (chronic eapsaicin) demonstrated significant increase in lesion number and ulcer area: 14.4 ± 2.6 and 69.5 ± 5.4 mm2 . CGRP antrat content of group I non-stressed controls was 62.4 ± 3.5 pmol/gram tissue. CGRP antral contents in group II (stress) was 32.4 ± 1.2 pmol. This represented a 48% decrease in CGRP antral content compared to control (p < 0.01). Group III (acute capsaicin) contained 59.4 ± 7.8 pmol/g tissue and group IV (chronic capsaic'm) 8.1 ± 0.5 pmol/g tissue; both values were significantly different from values recorded for stress animals (group I1) (p<0.01). Linear regression analysis of group data for area of stress ulceration and CGRP content revealed a significant negative correlation with R value of-0.8379 (p < 0.001 for linearity). Conclusions: 1) Acute capsaicin administration intragastrically Protects against stress ulceration. 2) Chronic capsaicin treatment enhances the degree of gastric injury in stress ulceration. 3) Stress causes a significant decrease in antral CGRP content. 4) There is an inverse relation between CGRP antral content and area of gastric ulceration. In summary, these data indicate that gastric capsaicin-senstive primary sensory afferent nerves and CGRP may be involved in the pathogenesis of stress ulceration in rats.
•
HIGH SYSTEMIC HCO3 AND TOPICAL 16,16-DIMETHYL PGE2 PROTECT THE GASTRIC MUCOSA AGAINST LUMINAL ACID BY ENHANCING ITS PRE-EPITHELIAL BUFFER CAPACITY T.Ranta-Knuuttila, H.Mustonen, E.Kivilaakso. II Dept. of Surgery, Helsinki University Central Hospital, Helsinki, Finland Systemic metabolic (high-HCO3) alkalosis (Gastro 81:921,1981) and topical prostaglandins (DDS 34:1028,1989) protect the gastric mucosa against luminal acid. This study investigates whether this protection is mediated by increased epithelial HC03 secretion with resultant alkalisatinn of the pre-epithelial mucus HCO3 buffer layer. Methods: Surface pH (pHs) of chambered ex rive rat gastric epithelium was measured with blunt-tipped liquid sensor pH-microelectrodas positioned under microscopic control in light contact with the epithelial surface. The lumen was perfused with increasing luminal acidities (0, 10, 30, 50 and 100 mM HC1). The experimental groups were; I : Control (N=9)(arterial pH 7.3020.04, aHC03 19.16~-1A9 mM), II: topical 16,16-dimethyl-PGE2 (100~g/100ml) treatment (N=7) (a-pH 7.30±0.01, a- HC03 16.7320.78 r a M ) , III: high HC03 metabolic alkalosis (N=7) ( 3.6 mmol/kg/h of NaHCO3 i.v., a-pH 7.37±0:02, a-HCO3 33A4-3.5mM), III: low -HC03 respiratory aikalosis by mandatory hyperventilation (N=9) (a- pH 7.4920.02, a-HC03 g.3± 0.9 raM). Results: pHs:s during exposure to increasing luminal acidities are given in the Table. PGE2 treated and high-HCO3 alkalotic rats tolerated significantly better luminal acid than controls, but in low-HCO3 alkaiosis tolerance to acid was significantly impaired. In alkalotie rats there was a significant correlation between arterial HCO3 concentration and pHs at luminal 30 mM HC1 (r=0.81, p<0.01), whereas no correlation was found between arterial pH and pHs (i"=0.27, p>0.05), suggesting that high HC03 concentration rather than high systemic pH provides protection. pHs (N) pH6 HCI 10 HCI 30 HCI 50 HCI 100 Control (9) 7.704-0.08 7.4620.10 7.204-0.11 6.93±0.14 6.34±0.25 PGE2 (7) 7A920.10 7.4020.11 7.214-0.17 7.034-0.18 7.0120.16" High-HCO 8.034-0.04 * 7.904-0.07* 7.724-0.11" 7.474-0.20* 7.344-0.22* Low-HCO 7.804-0.07 7.434-0.13 6.54±0.28* 6.124-0.09" • =p<0.05 vs. control Conclusions: The protective actions of systemic metabolic alkalosis and topical PGE2 treatment in gastric mueosa against luminal acid are, at least in part, mediated by enhanced buffer capacity of the pro-epithelial mueus-HC03 layer.
• INCREASED T R A N S L O C A T I O N OF pp600"~ FROM PLASMA MEMBRANE TO CYTOSOL BY TGF-a DURING THE E A R L Y R E P A R A T I V E PHASE O F GASTRIC MUCOSA AND EFFECTS O F AGING. N.K. Relan, R.R. Matta and A.P.N. Majumdar, VA Medical Center, Allen Park, MI 48101 and Dept. Med. Wayne State Univ. Detroit, MI. Although the gastric mucosa of healthy adult animals possesses a remarkable capacity to regenerate promptly after an acute injury, we have shown that aging diminishes its regenerative ability. Little is known about the regulatory mechanisms for the reparative processes. We suggested that decreased ligand-induced activation of EGF-reeeptor (EGF-R) tyrosine kinase (Tyr-k) in aged rats during the early reparative phase (restitution) may contribute to diminished mucosal repair in these animals (Gastroenterology 106: A629, 1994). To further elucidate the EGF-R-induced signal transduction processes during restitution, we examined the changes in Tyr-k activity of pp60 ~ and its translocation from membrane t o cytosol in the gastrie mucosa of young (4-me) and aged (24-m0) rats during the first 60 rain after i.g. administration of either 2 M NaCI (1.3 mill00 g) or water (controls). Data from control animals revealed that Tyr-k activity of pp60" ~ was about 50% higher in the gastric mucosa of aged than in young rats. Injury caused a prompt stimulation of mucosal pp60 ~ Tyr-k (measured in detergent-soinbillzed mucosa) in both age groups which attained its maximal stimulation (70-80% increase) at 30 rain after injury, where,after it declined sharply. However, when the relative concentration of pp60"I~ (assessed by Western-blot) was determined in the cytosolie fraction of the gastric mucosa (measure of pp60 ~ release) at 30 rain after injury, we observed a siguificanfly greater release (2-3-fold) in young than in aged rats, when compared with the corresponding controls. These changes were accompanied by parallel alterations in Tyr-k activity of pp6ff "~ in the cytosolic fraction. To further determine the rate of release of pp60~ from membrane to cytosol, the 30,000g membrane fraction (0.5 mg protein) from 30 rain post-injured rats was incubated in the absence (basal) or presence of lxl0 -9 M TGF-~t for up to 20 rain. The incubation medium (containing released pp60~ ) and the membrane fraction, recovered by centrifugation, were assayed for pp60 ~ levels (Western-blot) and its Tyr-k activity. TGF-c~ caused a significant 80-100% increase in pp60 ~ release from membranes of 30 rain post-injured young but not from aged rats. The membrane fraction from young injured rats showed a corresponding reduction in pp60 ~ concentration. These changes were also accompanied by parallel alterations in Tyr-k activity of pp60~ in the cytosol. Our observation suggests that ligand-induced activation of EGF-R Tyr-k in the gastric mueosa during the early reparative phase results in pp6ff "~ release from the membrane, and that pp60 ~ may play a role in signal transduction process.
Esophageal, Gastric, and Duodenal Disorders
A201
• THE EFFECT OF GASTRIC METAPLASIA (GM) OF THE DUODENAL BULB ON PROXIMAL DUODENAL MUCOSAL BICARBONATE SECRETION (DMBS). R.C. Rapier, M.A. Koss, A.D. Dreilinger, L.M. Nyberg, V.S. Pratha, D.L. Hogan, J.l. Isenberg. Depts of Medicine and Pathology, University of California at San Diego, San Diego, CA. Gastric metaplasia (GM) of the duodenal bulb is reportedly more common in patients with DU vs normal (NL} subjects and is required for adherence of Helicobacterpylori (HP;Gut 1993;34:1510). Our aim was to determine the frequency and effect of GM on human proximal DMSS in both NL and DU subjects. To date, 13 NL subjects (mean age 49 yr, 12 males) and 8 DU patients with healed ulcers (mean age 5t yr, 5 males) were evaluated prospectively. Each subject underwent upper gastrointestinal endoscopy. Antral biopsies were obtained to determine HP status (CLOtest ® and histology, observer blinded). Also, 2 "jumbo" duodenal bulbar biopsies, directed at areas of gastric metaplasia (mucosa "washed" with acetylcysteine, then stained with methylene blue; (Gastrointest Endosc 1992;38:696) were obtained for histological examination (observer blinded). DMBS was measured by isolating the proximal 4 cm of duodenum between occluding balloons using previously validated methods. Basal and acid-stimulated (20 ml, 100 mM HCI, 37°C, over 5 min) DMBS were measured. Ten of 13 NL (77%) and 5 of 8 DU patients (63%) had evidence of GM, an insignificant difference (P > 0.5). In the NL subjects, basal, peak and total acid-stimulated DMBS were comparable in those with GM (Table; means -+ SEMs). Also, in the DU patients (all HP+) basal and acid-stimulated DMBS was similar in those with, versus those without GM (Table). Moreover, in the 10 NL subjects with GM, DMBS was similar in those HP- (n = 8) versus those HP+ (n = 2). DUODENAL MUCOSAL BICARBONATESECRETION BASAL (pmol/cm-h)
PEAK (pmol/cm-h)
TOTAL (/.'mol/h)
NL GM+(IO)
217 ± 24
338 2 37
987 2 97
NLGM- (3)
139 2 5
405 2 109
1059+_ 136
DU GM+(5)
118 + 23
233 + 45
633 2 105
164 2 63 226 ± 81 619 2 164 DU GM- (3) Thus, these findings suggest that the presence of gastric metaplasia of the duodenum alters neither basal nor acid-stimulated proximal duodenal mucosal bicarbonate secretion in normal and duodenal ulcer patients.
STRESS ULCERATION:ROLE OF CAPSAJCIN-SENS1TIVE NERVES IN RAT MODEL. J Ren, H Ojeas, M Kids, K Brewer, J Gao, T Hoag and R Harry. Department of Medicine, University of Oklahoma and Oklahoma City DVAMC, Oklahoma City, OK. Acute stimulation of capsaicin-sensitlve nerves has been shown to protect gastric mucosa from various noxious insults. In contrast, chronic capsaicin treatment which causes sensory deafferentation enhances mucosal injury to luminal agents. The objectives of this study were to determine the effect of acute and chronic capsaiein administration in a rodent model of stress ulceration and to establish correlations between gastric calcitonin gone-related peptide (CGR15) content and the degree of gastric mucosal injury. Methods: The following groups of adult S-D rats were subjected to orogastric intubation without or with subsequent stress by restraint and water immersion for 4 hours: The control group (I) consisted of rats not subjected to stress; II-stress: orogastric administration of vehicle (1 ml solution: ethanol 10%, Tween 80 10%, saline 80% v/v/v) at the beginning of the experiment CI'o~and after 2 hours (T2) of stress; Ill-acute capsaicin: orogastric dosing (1.3 mg/kg) at TO and T2; and IV-chronic capsaicin treatment: 150 mg/kg subcutaneously on 3 consecutive days, 2 weeks prior to orognstric vehicle administration. The number of ulcers and the area of ulceration (ram 2) were quantitated. Data shown for each group represent the results of 8 experiments. CGRP antral content was determined by radioimmunoassay. Results: In group II (stress) there were 7.6 -. 0.4 ulcers encompassing 40.9 ~ m 2. Group III (acute capsaicin) the ulcer incidence and area of injury were reduced significantly to 3.2 ± 0.4 and 5.1 +- 0.2 mm2 , respectively (p < 0.01). In contrast, group IV animals (chronic eapsaicin) demonstrated significant increase in lesion number and ulcer area: 14.4 ± 2.6 and 69.5 ± 5.4 mm2 . CGRP antrat content of group I non-stressed controls was 62.4 ± 3.5 pmol/gram tissue. CGRP antral contents in group II (stress) was 32.4 ± 1.2 pmol. This represented a 48% decrease in CGRP antral content compared to control (p < 0.01). Group III (acute capsaicin) contained 59.4 ± 7.8 pmol/g tissue and group IV (chronic capsaic'm) 8.1 ± 0.5 pmol/g tissue; both values were significantly different from values recorded for stress animals (group I1) (p<0.01). Linear regression analysis of group data for area of stress ulceration and CGRP content revealed a significant negative correlation with R value of-0.8379 (p < 0.001 for linearity). Conclusions: 1) Acute capsaicin administration intragastrically Protects against stress ulceration. 2) Chronic capsaicin treatment enhances the degree of gastric injury in stress ulceration. 3) Stress causes a significant decrease in antral CGRP content. 4) There is an inverse relation between CGRP antral content and area of gastric ulceration. In summary, these data indicate that gastric capsaicin-senstive primary sensory afferent nerves and CGRP may be involved in the pathogenesis of stress ulceration in rats.