Enterochromaffin-like Cell Proliferation Research Articles

Proton Pump Inhibitors and Cancer Risk: A Comprehensive Review of Epidemiological and Mechanistic Evidence.

Background: Proton pump inhibitors (PPIs) are commonly prescribed long-acting drugs used to treat acid reflux, gastroesophageal reflux disease (GERD), and peptic ulcers. Recently, concerns have been raised about their safety, particularly due to the association between long-term PPI use and cancer development. Multiple comprehensive studies have consistently suggested a noteworthy link between prolonged PPI usage and an increased risk of developing gastric, esophageal, colorectal, and pancreatic cancers, yet the precise underlying mechanism remains elusive. Methods: First, we review the extensive body of research that investigates the intricate relationship between cancer and PPIs. Then, we predict PPI toxicity using the prodrug structures with the ProTox-II webserver. Finally, we predict the relative risk of cancer for each PPI, using PubMed citation counts of each drug and keywords related to cancer. Results: Our review indicates that prolonged PPI use (exceeding three months) is significantly associated with an elevated risk of cancer, while shorter-term usage (less than three months) appears to pose a comparatively lower risk. Our review encompasses various proposed mechanisms, such as pH and microbiome alterations, vitamin and mineral malabsorption, hypergastrinemia, and enterochromaffin-like cell proliferation, while ProTox-II also suggests aryl hydrocarbon receptor binding. Potentially, the PubMed citations count suggests that the PPIs omeprazole and lansoprazole are more associated with cancer than pantoprazole and esomeprazole. In comparison, the H2R blocker, famotidine, is potentially less associated with cancer than PPIs, and may serve as a safer alternative treatment for periods beyond 3 months. Conclusions: Despite the well-established cancer risk associated with PPIs, it is notable that these medications continue to be widely prescribed for periods longer than 3 months. Thus, it is of paramount importance for clinicians and patients to thoughtfully evaluate the potential risks and benefits of long-term PPI usage and explore alternative treatments before making informed decisions regarding their medical management.

Factors associated with elevated serum chromogranin A levels in patients with autoimmune gastritis.

Chromogranin A is an important tool in the diagnosis of neuroendocrine tumors. Autoimmune gastritis is an autoimmune disorder marked by hypergastrinemia, which stimulates enterochromaffin-like cell proliferation. Chromogranin A is also elevated in autoimmune gastritis patients with a different level of increase in each patient. The goal of this study is to explore constituents that influence serum chromogranin A levels in autoimmune gastritis patients. One hundred and eighty-eight autoimmune gastritis patients and 20 patients with type I gastric carcinoid tumors were analyzed retrospectively and compared to 110 functional dyspepsia patients in terms of factors that might affect serum chromogranin A levels. The mean serum chromogranin A level was 171.17±67.3 ng/mL in autoimmune gastritis patients (n=62) without enterochromaffin-like cell hyperplasia, and 303.3±102.82 ng/mL in patients (n=126) with enterochromaffin-like cell hyperplasia (p<0.001). The presence of corpus atrophy (p=0.026, OR: 5.03, CI 95%: 1.21-20.88, β=1.61) and presence of enterochromaffin-like cell hyperplasia (p=0.017, OR: 6.59, CI 95%: 1.4-31.08, β=1.88) were found as risk factors associated with serum chromogranin A level. Factors influencing raised serum chromogranin A levels in autoimmune gastritis were the presence of ECL cell hyperplasia and serum gastrin levels. Serum chromogranin A levels maybe helpful in distinguishing autoimmune gastritis patients and gastric carcinoid type I from the control group, but not useful in the differentiation of individuals with autoimmune gastritis from patients with gastric carcinoids.

The CCK 2 receptor antagonist, YF476, inhibits Mastomys ECL cell hyperplasia and gastric carcinoid tumor development

YF476 is a potent and highly selective cholecystokin 2 (CCK 2) receptor antagonist of the benzodiazepine class. It inhibits gastric neuroendocrine enterochromaffin-like (ECL) cell secretion, proliferation and spontaneous formation of gastric neuroendocrine tumors (carcinoids) in cotton rats. The Mastomys rodent species exhibits a genetic predisposition to gastric ECL neuroendocrine tumor formation which can be accelerated by acid suppression and induction of hypergastrinemia. In this respect, it mimics the human condition of atrophic gastritis, hypergastrinemia and gastric carcinoid development. We investigated whether YF476 could inhibit acid suppression-induced ECL cell hyperplasia and neoplasia in this model. In addition, we examined whether YF476 could reverse established ECL cell hyperplasia and neoplasia. Targeting the CCK 2 receptor during Loxtidine-induced hypergastrinemia resulted in a reduction in ECL cell secretion (plasma and mucosal histamine, and histidine decarboxylase ( HDC) transcripts, p < 0.05) and proliferation (numbers of HDC-positive cells, connective tissue growth factor ( CTGF) and cyclin D1 transcription). This was associated with a decrease in ECL cell hyperplasia and a 60% reduction in gastric ECL cell microcarcinoid (tumors < 0.3 mm in size) formation. YF476 inhibited ECL cell neoplasia (gastric carcinoid) in animals with hyperplasia, inhibited the formation of ECL cell tumors when co-administered with Loxtidine and reversed the growth and developement of gastric ECL cell carcinoids in long-term acid suppressed Mastomys. Variable importance analysis using a logistic multinomial regression model indicated the effects of YF476 were specific to the ECL cell and alterations in ECL cell function reflected inhibition of transcripts for HDC, Chromogranin A ( CgA), CCK 2 and the autocrine growth factor, CTGF. We conclude that specifically targeting the CCK 2 receptor inhibits gastrin-mediated ECL cell secretion and ECL cell proliferation and tumor development in vivo.

Chromogranin A in patients with acid hypersecretion and/or hypergastrinaemia

Chromogranin has been proposed as a marker for gastrin-dependent enterochromaffin-like cell proliferation. To examine this question in three populations: acid hypersecretors with gastrinoma (Zollinger-Ellison), or without gastrinoma (non-Zollinger-Ellison), and also in pernicious anaemia with achlorhydria-caused hypergastrinaemia. We measured serum chromogranin, gastrin, gastric secretion and counted and quantified hyperplasia of enterochromaffin-like cells in gastric biopsies from 38 Zollinger-Ellison and 13 non-Zollinger-Ellison patients being treated with lansoprazole, for 5 years (median) and again 2.5 years later. We also studied 12 patients with pernicious anaemia, half with gastric enterochromaffin-like cell carcinoids. Serum chromogranin was elevated in patients with gastrinoma, even without any enterochromaffin-like cell proliferation, but not in non-Zollinger-Ellison acid hypersecretors with normal gastrin (P < 0.001). In the hypersecretors chromogranin correlated well with serum gastrin (r = 0.82), but not with enterochromaffin-like cell proliferation. Moreover, chromogranin was normal or near normal (<75 ng/mL) despite very high serum gastrin in five of six patients with pernicious anaemia and enterochromaffin-like cell carcinoids. Chromogranin is not a reliable marker for enterochromaffin-like cell activity or proliferation up to and including carcinoid; chromogranin originates in the gastrinoma and, like gastrin, is a marker for gastrinoma in acid hypersecretors.

Role of CCN2/CTGF in the proliferation of Mastomys enterochromaffin-like cells and gastric carcinoid development

Mastomys enterochromaffin-like (ECL) cell proliferation is initially gastrin driven, but once neoplasia develops, cells become gastrin autonomous. We hypothesized that CCN2 (CTGF), a mitogenic growth factor, may regulate ECL cell proliferation. A Mastomys GeneChip database was examined (dCHIP) to identify CCN2 expression levels. CCN2 in normal and tumor ECL cell preparations obtained using FACS (100 nM acridine orange) was examined by real-time PCR. CCN2 protein was identified in mucosal and ECL cell preparations by immunohistochemistry. Short-term cultured cells were stimulated with either CCN2 or CCN2 + EGF, and proliferation was measured (MTT assay). The ERK1/2 inhibitor PD-98059 (0.1-100 microM) was assessed in terms of CCN2 (1 ng/ml)-mediated proliferation and ERK1/2 phosphorylation. CCN2 transcript and protein was then examined in clinical gastric carcinoids. The ccn2 transcript was upregulated in tumor samples compared with the normal mucosa (+2.36-fold, P < 0.01). PCR demonstrated that ccn2 was not expressed in FACS-prepared (>98% pure) normal ECL cells but was elevated in tumor ECL cell fractions (41.3 +/- 10.7-fold). Immunostaining of the Mastomys gastric mucosa and FACS preparations confirmed that CCN2 protein was present in ECL tumors but not in normal ECL cells. Neither CCN2 nor CCN2 + EGF stimulated normal ECL cell proliferation. CCN2 stimulated tumor proliferation (EC50 approximately 0.01 ng/ml); EGF significantly augmented (P < 0.01) CCN2-induced tumor cell proliferation (EC50 = 20 pg/ml). PD-98059 inhibited CCN2-induced proliferation (-12 +/- 3%, P < 0.05) and ERK1/2 phosphorylation (-34 +/- 5%, P < 0.05) in tumor cells. In clinical samples, both CCN2 transcript and protein were elevated in gastrin-autonomous carcinoids (P < 0.02) compared with the normal mucosa. In conclusion, CCN2 may be a proliferative regulator of Mastomys ECL neoplastic proliferation once these cells become autonomous of gastrin regulation. Identification of CCN2 in gastric carcinoid tissue may be useful both as an indicator of ECL cell transformation and may define gastrin autonomy, a criteria of gastric carcinoid malignancy.

Gastric carcinoid

Gastric carcinoid tumors are rare lesions but have been the focus of much scientific investigation. The incidence of gastric carcinoid appears to be increasing without a corresponding increase in survival, despite utilization of the latest available therapies. Therefore, there is great interest in furthering the understanding of the biologic basis of these tumors, delineating the connection between hypergastrinemia and gastric carcinoids, and most importantly, improving upon current treatment options. This review discusses the current biologic understanding of gastric carcinoid tumors, including the role of hypergastrinemia on enterochromaffin-like cell proliferation and its relation to acid-suppressive therapy. Numerous diagnostic and therapeutic modalities including endoscopic ultrasound, somatostatin receptor scintigraphy, long-acting octreotide, hepatic artery embolization, endoscopic mucosal resection, and surgical resection have also been the focus of recent investigations. Despite the many advances that have been made in both the basic science and clinical arenas, the optimal treatment of gastric carcinoid tumors is still a matter of debate. As the understanding of the biologic basis of gastric carcinoid tumors increases, the treatment will likely be a multimodal approach tailored to individual tumor biology and will incorporate a variety of diagnostic and therapeutic modalities.

Neuroendocrine tumor markers and enterochromaffin-like cell hyper/dysplasia in type 1 diabetes.

Parietal cell antibodies (PCAs) are found in 20% of type 1 diabetic patients, denoting autoimmune gastritis and pernicious anemia, which may predispose to enterochromaffin-like (ECL) cell hyper/dysplasia and gastric carcinoid tumors. We evaluated whether chromogranin A (CgA), 5-hydroxyindole acetic acid (5-HIAA), and neuron-specific enolase (NSE) contribute to screening for ECL cell hyper/dysplasia. Sera from 93 type 1 diabetic patients (53 men and 40 women, 31 PCA(+) and 62 PCA(-), aged 45 +/- 13 years) were analyzed for PCAs by indirect immunofluorescence and for CgA, NSE, and gastrin by radioimmunoassay. Urinary 5-HIAA was tested by high-performance liquid chromatography. Corpus atrophy and ECL cell proliferation were assessed in gastric biopsies. PCA(+) patients had higher gastrin (P < 0.0001) and CgA levels (P = 0.003) and were more prone to autoimmune gastritis (odds ratio [OR] 17, P < 0.0001) and ECL cell hyper/dysplasia (OR = 23, P = 0.005) than PCA(-) subjects. ECL cell hyper/dysplasia was present in seven PCA(+) patients who showed higher CgA levels (P < 0.0001) than subjects without ECL cell hyper/dysplasia, but NSE and 5-HIAA levels were similar. CgA levels correlated with gastrinemia (r = 0.50, P < 0.0001), PCA titer (r = 0.42, P = 0.001), and 5-HIAA levels (r = 0.38, P = 0.012). Logistic regression identified the CgA level (beta = 0.01, P = 0.027) as an independent risk factor for ECL cell hyper/dysplasia when PCA, CgA, 5-HIAA, NSE, gastrin, sex, and age were tested. Multivariate linear regression demonstrated that CgA level was determined by ECL cell density (r = 0.59, P < 0.0001) and gastrin level (r = 0.67, P = 0.02). One PCA(+) patient with elevated gastrin, CgA, and 5-HIAA levels had a gastric carcinoid tumor. PCA(+) patients, particularly those with high gastrin and CgA levels, risk developing ECL cell hyper/dysplasia. The determination of CgA, but not NSE and 5-HIAA, may complement histology in evaluating ECL cell mass.

Histopathology of the gastric oxyntic mucosa in two different patient groups during long-term treatment with omeprazole.

Hypochlorhydria, hypergastrinaemia, inflammation and Helicobacter pylori infection, dose and duration of omeprazole treatment may separately, or in combination, influence the proliferation of enterochromaffin-like (ECL) cells and parietal cell changes in gastric mucosa. To assess the effects of these variables comparisons were carried out in patients with the acid related Zollinger-Ellison syndrome (ZES) versus patients with progressive systemic sclerosis (PSS) and gastro-oesophageal reflux disease. Twenty-five patients with PSS and 16 patients with ZES were included and received continuous omeprazole treatment for a mean of 7.5 and 9 years. The patients were investigated every 6-12 months with endoscopy, biopsies and histology, and plasma gastrin measurements. PSS patients were titrated by 24 h pH-metry to oesophageal pH>4, and all ZES patients were titrated to a basal acid output of zero H+. Changes towards diffuse and linear ECL cell hyperplasia were observed in 41% of the PSS patients. Micronodular hyperplasia and neoplasia were not seen. In the ZES patients changes towards linear and micronodular hyperplasia were observed in all patients. Two patients developed ECL cell carcinoids; one of these had MEN-1 syndrome. Also parietal cell changes were more pronounced in the ZES group than in the PSS group. In patients without intrinsic acid hypersecretion and hypergastrinaemia significant proliferation of ECL cells is not an issue irrespective of gastric mucosal inflammation, omeprazole dose, duration of treatment and acid inhibition. The level of gastrin secretion and high plasma gastrin appear to accelerate ECL cell proliferation and parietal cell changes possibly influenced by chronic gastritis and H. pylori infection.

Histopathology of the gastric oxyntic mucosa in two different patient groups during long-term treatment with omeprazole

Objectives Hypochlorhydria, hypergastrinaemia, inflammation and Helicobacter pylori infection, dose and duration of omeprazole treatment may separately, or in combination, influence the proliferation of enterochromaffin-like (ECL) cells and parietal cell changes in gastric mucosa. To assess the effects of these variables comparisons were carried out in patients with the acid related Zollinger-Ellison syndrome (ZES) versus patients with progressive systemic sclerosis (PSS) and gastrooesophageal reflux disease. Methods Twenty-five patients with PSS and 16 patients with ZES were included and received continuous omeprazole treatment for a mean of 7.5 and 9 years. The patients were investigated every 6-12 months with endoscopy, biopsies and histology, and plasma gastrin measurements. PSS patients were titrated by 24 h pH-metry to oesophageal pH > 4, and all ZES patients were titrated to a basal acid output of zero H+. Results Changes towards diffuse and linear ECL cell hyperplasia were observed in 41% of the PSS patients. Micronodular hyperplasia and neoplasia were not seen. In the ZES patients changes towards linear and micronodular hyperplasia were observed in all patients. Two patients developed ECL cell carcinoids; one of these had MEN-1 syndrome. Also parietal cell changes were more pronounced in the ZES group than in the PSS group. Conclusions In patients without intrinsic acid hypersecretion and hypergastrinaemia significant proliferation of ECL cells is not an issue irrespective of gastric mucosal inflammation, omeprazole dose, duration of treatment and acid inhibition. The level of gastrin secretion and high plasma gastrin appear to accelerate ECL cell proliferation and parietal cell changes possibly influenced by chronic gastritis and H. pylori infection.

PACAP and gastrin regulate the histidine decarboxylase promoter via distinct mechanisms.

The enterochromaffin-like (ECL) cell controls gastric acid secretion via histamine, generated by l-histidine decarboxylase (HDC). HDC expression is regulated by gastrin. However, gastrin is not alone in controlling ECL cell function. For example, the neural peptide pituitary adenylate cyclase-activating polypeptide (PACAP) also increases ECL cell proliferation. To investigate a potential role of PACAP in regulating HDC expression, we generated a series of HDC promoter-luciferase reporter constructs and transiently transfected them into PC12 cells (stably expressing the gastrin-CCK-2 receptor). We found that PACAP regulates HDC promoter activity. This is temporally biphasic, involving both adenyl cyclase and phospholipase C-dependent pathways. Deletional analysis, block mutation, and EMSA demonstrated a PACAP-response element at -177 to -170, wholly necessary for the effects of PACAP and discrete from known gastrin-responsive elements. Discrete neural and endocrine pathways regulate ECL cells through different patterns of postreceptor signaling and promoter activation, which may be appropriate to their functions in vivo.

Progression of gastric enterochromaffin-like cells growth in Zollinger-Ellison syndrome and atrophic body gastritis patients.

Enterochromaffin-like cell hyperplasia of the gastric body mucosa occurs in hypergastrinaemic conditions such as atrophic body gastritis and Zollinger-Ellison syndrome. However, the time course of change or factors involved are not known. To compare the rate of change of enterochromaffin-like cell proliferation in patients with atrophic body gastritis and Zollinger-Ellison syndrome. From a consecutive series of atrophic body gastritis and Zollinger-Ellison syndrome patients, studied at the time of first diagnosis, 10 atrophic body gastritis (4 with pernicious anaemia) and 14 Zollinger-Ellison syndrome (4 with multiple endocrine neoplasia type 1) patients were followed-up for a median time of 48 months. At entry and during follow-up patients underwent: plasma gastrin determination, endoscopic sampling of body mucosa for qualitative assessment of enterochromaffin-like cell hyperplasia pattern and degree of glandular atrophy, qualitative and morphometric analyses of body mucosa endocrine cells. At time of diagnosis, enterochromaffin-like cell lesions were more severe in atrophic body gastritis than in Zollinger-Ellison syndrome. During follow-up, no significant variations were observed in gastrin values, enterochromaffin-like cell patterns and grade of body mucosa atrophy in atrophic body gastritis. In contrast, gastrin levels were significantly increased [median 1200 (235-2625) vs 1947 (225-5200) pg/ml; p<0.001)] as was total volume density of enterochromaffin-like cells [median 1.60 (0.53-4.06) vs 3.18 (1.35-21.13)% of mucosal epithelial component; (p<0.005)] in Zollinger-Ellison syndrome. Micronodular hyperplasia of enterochromaffin-like cells, present in only one patient at diagnosis, was observed in 8 Zollinger-Ellison syndrome patients at follow-up. These data suggest that the progression of enterochromaffin-like cell growth in human gastric mucosa requires an increase of and/or a prolonged exposure to severe hypergastrinaemia.

PACAP mediates the neural proliferative pathway of Mastomys Enterochromaffin-like cell transformation

Background and aim: Pituitary adenylate-cyclase activating peptide (PACAP) is a more potent proliferative agent than gastrin for rat enterochromaffin-like (ECL) cell proliferation in vitro. The role of this neurotransmitter during gastrin-mediated ECL cell tumor formation and gastrin-autonomous ECL cell neoplasia is unknown. Methods and results: ECL cell transformation was induced in the Mastomys using 16 wk H 2 receptor blockade of acid inhibition. Examination of the epithelial fundic mucosa demonstrated that PACAP-immunoreactivity significantly increased in the tumor mucosa compared to the naı̈ve stomach, and was associated with ECL cells. Naı̈ve and tumor ECL cells were then purified (∼95%) from Mastomys and the presence of all three PACAP/VPAC receptor subtypes was demonstrated by polymerase chain-reaction amplification. Thereafter, cells were maintained in short-term (48 h) primary cultures. PACAP significantly ( p<0.05) increased 24 h bromo-deoxyuridine uptake (∼4-fold) in both cell types with estimated EC 50 values of ∼4×10 −16 M and ∼2×10 −16 M, respectively. Specific receptor antagonists (PAC 1/VPAC 1) of PACAP competitively inhibited these proliferative effects in naı̈ve cells. Oligonucleotide antisense directed against PAC 1 significantly inhibited PACAP-stimulated DNA synthesis by ∼85% ( p<0.05) in tumor cells. Conclusion: PACAP is a potent and effective modulator of ECL cell proliferation. The expression of this neuropeptide and its receptors, particularly PAC 1, suggest the existence of a neural regulatory pathway of ECL cell proliferation and transformation.

Helicobacter pylori effects on gastritis, gastrin and enterochromaffin-like cells in Zollinger-Ellison syndrome and non-Zollinger-Ellison syndrome acid hypersecretors treated long-term with lansoprazole.

Helicobacter pylori is said to cause atrophy of the gastric corpus and enterochromaffin-like cell proliferation in gastro-oesophageal reflux disease (GERD) patients treated long-term with a proton pump inhibitor. To determine the effect of H. pylori infection on gastritis, enterochromaffin-like cell density and hyperplasia, mucosal atrophy and serum gastrin in patients with gastric hypersecretion (basal acid output gt; 15 mmol/h) with either hypergastrinemia (Zollinger-Ellison syndrome) or normal gastrin (non-Zollinger-Ellison syndrome) before and during long-term treatment with lansoprazole. Lansoprazole was individually titrated to reduce basal acid output to < 5 mmol/h (< 1 mmol/h in post-surgical Zollinger-Ellison syndrome). Gastric corpus biopsies were obtained every 6 months before treatment and up to 8 years later. H. pylori was present in corpus biopsies in approximately 50%, causing active gastritis which resolved rapidly in 15 subjects after elimination of H. pylori. Patchy mild/moderate corpus atrophy was present at entry in two and at the end in four out of 60 patients, one being H. pylori-positive. Intestinal metaplasia (< 10%) was seen in six isolated biopsies (1% of total). H. pylori did not affect serum gastrin, enterochromaffin-like cell density or hyperplasia. Enterochromaffin-like cell density was twice as high in Zollinger-Ellison syndrome as in non-Zollinger-Ellison syndrome patients (241 vs. 126 cells/mm2, P < 0.001). Enterochromaffin-like cells remained normal in the non-Zollinger-Ellison syndrome hypersecretors regardless of H. pylori status. Corpus enterochromaffin-like cell increases were related to serum gastrin elevation, but neither H. pylori nor long-term treatment with lansoprazole alone or together had any effect on enterochromaffin-like cell density or hyperplasia. Corpus acute gastritis resulted from H. pylori infection, but did not result in mucosal atrophy despite long-term proton pump inhibitor treatment and promptly resolved with loss of H. pylori.

Effect of a histamine H1 receptor antagonist on gastric endocrine cell proliferation induced by chronic acid suppression in rats.

The effect of histamine on gastrin cells and enterochromaffin-like cells has not yet been clarified. We investigated the influence of pyrilamine (a histamine H1 receptor antagonist) on serum gastrin level, gastrin cells, and enterochromaffin-like cells in rats with or without 4 weeks of famotidine treatment. The rats were divided into six groups: a control group, two pyrilamine groups (2mg/kg, or 20mg/kg, p.o.), a famotidine group (20mg/kg twice/daily i.m.), and two pyrilamine + famotidine groups. The serum gastrin concentration was determined, and gastrin cells and enterochromaffin-like cells were identified by the labeled streptavidin biotin complex method and counted. Hypergastrinemia, gastrin cell hyperplasia, and enterochromaffin-like cell hyperplasia were found after 4 weeks of famotidine treatment. Four weeks of treatment with pyrilamine alone did not affect the gastrin level, gastrin cells, or enterochromaffin-like cells in the rat stomach. When combined with famotidine, pyrilamine enhanced famotidine-induced hypergastrinemia, but it did not affect gastrin cell hyperplasia, and it significantly inhibited enterochromaffin-like cell hyperplasia. These results suggest that gastrin secretion and enterochromaffin-like cell proliferation may be regulated by histamine via the H1 receptor during acid suppression.

Pituitary adenylate cyclase-activating polypeptide modulates gastric enterochromaffin-like cell proliferation in rats

Background & Aims: Gastric carcinoids (types I and II) involve the transformation of naive enterochromaffin-like (ECL) cells to the neoplastic state and are associated primarily with hypergastrinemia. In this study, we evaluated the effects of two related neuropeptides, pituitary adenylate cyclase–activating polypeptide (PACAP) and vasoactive intestinal polypeptide (VIP), on ECL cell proliferation and characterized the receptor subtype(s) and signal transduction pathways that mediate this effect. Methods: Purified rat ECL cells were analyzed in culture for DNA synthesis as measured by 24-hour 5-bromo-2-deoxyuridine (BrdU) uptake. Reverse-transcription polymerase chain reaction (RT-PCR) with gene-specific oligonucleotide primers was performed to characterize the PACAP/VIP receptor subtype(s). Results: PACAP/VIP neuropeptide-stimulated BrdU uptake was significantly greater (3.4–3.8-fold greater than control) than that at the maximal dose of gastrin (2.2-fold greater than control). PACAP-stimulated ECL cell proliferation (EC 50, ~3 × 10 −14 mol/L) was ~100-fold more potent than VIP (EC 50, ~3× 10 −12 mol/L). The stimulated BrdU uptake by both PACAP and VIP was competitively inhibited by PACAP-receptor antagonist (IC 50, 10 −9 mol/L, 3 × 10 −9 mol/L, respectively) and VIP-receptor antagonist (IC 50, 3 × 10 −7 mol/L, 5 × 10 −7 mol/L, respectively). RT-PCR identified the presence of the PACAP-specific but not PACAP/VIP receptor subtypes. The PACAP-stimulated BrdU uptake was inhibited (70%–80%) by inhibitors of adenosine 3',5'-cyclic monophosphate, phosphatidylinositol 3 kinase, and protein tyrosine kinase as well as mitogen-activated protein kinase. Conclusions: PACAP/VIP-related peptides are more potent modulators of ECL cell proliferation than gastrin, and their effect is mediated by a PACAP-specific receptor whose activation is transduced by multiple intracellular messenger systems. GASTROENTEROLOGY 1999;116:623-635

Helicobacter pylori lipopolysaccharide stimulates histamine release and DNA synthesis in rat enterochromaffin-like cells

BACKGROUND & AIMS: Helicobacter pylori alterations in gastric acid output and mucosal proliferation may involve the enterochromaffin-like (ECL) cell. To test whether H. pylori affects ECL cell histamine secretion and proliferation, the effect of lipopolysaccharide (LPS) on ECL cell function in vitro was investigated.METHODS: Using a rat ECL cell preparation of high purity (+/-95%), basal and stimulated histamine secretion and DNA synthesis were measured by enzyme immunoassay and bromodeoxyuridine (BrdU) uptake, respectively.RESULTS: Escherichia coli LPS (10(-12) to 10(-6) mol/L) had no effect on basal and stimulated histamine secretion at concentrations of > 10(-6) mol/L. H. pylori LPS stimulated basal and gastrin-stimulated histamine secretion. These effects were completely inhibited by somatostatin (10(- 10) mol/L) but not by the gastrin receptor antagonist L365,260 at 10(- 6) mol/L. E. coli LPS had a weak stimulatory effect on ECL cell BrdU uptake at 10(-6) mol/L but had no effect on gastrin-stimulated BrdU uptake. H. pylori LPS did not stimulate basal synthesis but significantly increased (1.5-fold) gastrin-stimulated BrdU uptake.CONCLUSIONS: H. pylori influences both ECL cell proliferation and secretion in vitro. An interaction between H. pylori LPS and ECL cells may contribute to the reported abnormalities in acid secretion and gastric pathobiology noted in H. pylori infections.(Gastroenterology 1997 Oct;113(4):1110-7)

Somatostatin receptor regulation of gastric enterochromaffin-like cell transformation to gastric carcinoid

Although somatostatin is recognized as an inhibitor of neuroendocrine cell secretion, its effect on cell proliferation has not been well defined. Generation of low acid and hypergastrinemia through irreversible H2-receptor blockade (loxtidine) in the African rodent mastomys results in gastric carcinoids (ECLomas) within 4 months. This study was undertaken to evaluate and characterize the precise somatostatin receptor (SSTR) subtype on the mastomys enterochromaffin-like (ECL) cell and to define its role in the regulation of ECL cell secretion and proliferation. A pure preparation (approximately 90%) of ECL cells was derived by a combination of pronase digestion and density gradient separation. We assessed the effect of somatostatin (10(-15) to 10(-7) mol/L) on gastrin-stimulated ECL cell histamine secretion and DNA synthesis (bromodeoxyuridine uptake). SSTR2 subtype was evaluated by reverse transcription-polymerase chain reaction (RT-PCR) using gene specific primers and mRNA isolated from normal and hypergastrinemia-induced ECLoma. The polymerase chain reaction product was confirmed by Southern analysis, subcloned, and sequenced. Somatostatin inhibited both gastrin-stimulated histamine secretion (IC50, 5 x 10(-13) mol/L) and DNA synthesis (IC50, 10(-10) mol/L). SSTR2 was identified in the mastomys' brain, and both normal and tumor ECL cells and comparison of the brain and ECL cell SSTR2 nucleotide sequences revealed homology of 99%. The SSTR2 is expressed by the mastomys' ECL cell and ECLoma. Receptor activation inhibits both ECL cell secretory and proliferative functions.

Regulation of Mastomys ECL Cell Function by Transforming Growth Factor Alpha

Little progress has been made in the understanding of the pathobiology of gastric neoplasia over the past 4 decades. This reflects the paucity of information available regarding the biology of gastric mucosal cell proliferation. More recently it has become apparent that growth factor regulation of cell proliferation is of considerable relevance in initiating mucosal mitogenesis. We have recently identified the histamine secreting enterochromaffin-like (ECL) cell as a pivotal cellular regulator of gastric acid secretion. In addition to its critical role in initiating acid secretion, we have proposed that the ECL cell may produce agents responsible for the regulation of mucosal cell proliferation. We have therefore hypothesized that such a function may be subserved by production of transforming growth factor alpha (TGFα). TGFα is known to play a significant role both in normal physiology and in the transformation of naive cells into a neoplastic form. We therefore proposed that increased levels of gastrin induced by low acid states might stimulate TGFα secretion and that this agent might be capable of regulating ECL cell DNA synthesis and cell proliferation. We used the mastomys rodent to generate anin vivohy- pergastrinemia model using long-term histamine-2 receptor blockade (loxtidine 1 mg/kg/day). In order to evaluate the cell-specific effects, we developed a pure isolated ECL cell system from the mastomys stomach. This utilized pronase digestion (1.0 mg/ml) and EDTA exposure (1 mM) of the mucosa followed by particle size separation with countercurrent elutriation and density purification on a Nycodenz step gradient. ECL cells were obtained with a purity of 90–95%. Histamine secretion from ECL cells was measured by radioimmunoassay (RIA). TGFα content was measured by RIA, and TGFα expression was measured by RNAse probe protection assay. DNA synthesis was quantified by measuring bromo-deoxyuridine (BrdU) incorporation into cultured cells. TGFα levels were increased in fundic mucosa after 16 weeks of hypergastrinemia (from 4.3 ± 0.6 to 32.6 ± 2.6 fmole/mg protein,P< 0.05). TGFα message was identified in the ECL cells by RNAse probe protection assay, and was fourfold amplified in ECL cell tumors after 16 weeks of exposure to hypergastrinemia. Gastrin stimulated (10 nM) histamine secretion in isolated naive ECL cells was inhibited by TGFα (IC502 × 10−10M). DNA synthesis was stimulated by gastrin (EC502 × 10−11M) and TGFα (EC505 × 10−9M). These data are consistent with the proposal that elevated gastrin levels are associated with ECL cell TGFα production and that TGFα stimulates ECL cell DNA synthesis.

Evidence for a regulatory role for histamine in gastric enterochromaffin-like cell proliferation induced by hypergastrinemia.

Hypergastrinemia, induced by sustained suppression of gastric acid secretion, is associated with gastric enterochromaffin-like (ECL) cell hyperplasia and carcinoid tumor formation. We examined the effect of a selective H1-histamine antagonist, terfenadine, on gastric mucosal cell proliferation to determine whether histamine might modulate ECL cell generation. The rodent mastomys received the H2-antagonist loxtidine (2 g/l drinking water) alone or in combination with terfenadine (0.5 g/l or 35 mg/l drinking water) for 120 days. Controls received water or terfenadine alone. Serum gastrin levels and tissue histamine content were assayed by radioimmunoassays, and tissue chromogranin levels determined (Western blot analysis). In vivo cell proliferation was measured by bromodeoxyuridine (BrdU, 200 mg/kg/day, 3 days) incorporation. Gastric mucosal thickness was determined, ECL cell number was assessed, and the percentage of proliferating ECL cells quantitated. To evaluate the direct action on ECL cells we then studied the effect of terfenadine on histamine secretion and DNA synthesis (BrdU uptake) in an isolated preparation (approximately 90% pure) of ECL cells. Loxtidine increased serum gastrin levels, mucosal thickness, tissue chromogranin levels, tissue histamine content, BrdU incorporation, ECL cell number, and proliferating ECL cells (all parameters p < 0.05). Terfenadine alone, irrespective of dosage, had no significant effect. The high dose in combination with loxtidine significantly inhibited the increase in tissue chromogranin levels, tissue histamine content, ECL cell number and proliferating ECL cells (p < 0.05), but did not alter other parameters, compared to loxtidine alone. The low does did not alter the loxtidine-induced changes. In pure isolated ECL cells, terfenadine did not alter histamine secretion either alone or in combination with gastrin (10 nM). DNA synthesis was significantly inhibited by terfenadine (IC50 10(-10) M). Terfenadine specifically inhibited the effect of loxtidine-induced ECL cell proliferation in vivo and significantly inhibited ECL cell DNA synthesis in vitro. We postulate that histamine, through an H1 receptor, positively modulates gastric ECL cell proliferation.

Inhibition of gastrin-stimulated enterochromaffin-like cell proliferation and mucosal histamine production in the rat stomach by the somatostatin analogue octreotide

The effect of octreotide, a potent and long-acting analogue of somatostatin, on gastrin-stimulated proliferation and function of enterochromaffin-like (ECL) cells were examined in rats. Animals were divided into four groups and each group was continuously infused with saline, octreotide alone (40 μg/kg per day), gastrin alone (60 nmol/kg per day), or octreotide (40 μg/kg per day) plus gastrin (60 nmol/kg per day) respectively for 9 days via osmotic minipumps. Gastrin induced the increase of the bromodeoxyuridine labeling index and density of oxyntic mucosal ECL cells as well as oxyntic mucosal histidine decarboxylase activity. Octreotide completely abolished the gastrin-induced increases in the labeling index and density of ECL cells and oxyntic mucosal histidine decarboxylase activity. These results indicate that octreotide inhibits gastrin-stimulated proliferation of ECL cells and histamine production by these cells.