RESUMEN
BACKGROUND & AIMS: Leptin is a circulating hormone that communicates the peripheral nutritional status to the hypothalamus, which controls food intake, energy expenditure, and body weight. This study characterizes leptin receptors and leptin-sensitive STAT proteins in the antrum and investigates the effects of leptin on gastric secretions. METHODS: The effects of leptin on gastrin messenger RNA (mRNA), plasma gastrin, gastric acid in vivo in the rat, and on somatostatin and gastrin secretions by isolated antral cells were determined in vitro. Leptin receptors were investigated in isolated rat antral cells by reverse transcription-polymerase chain reaction and binding of [(125)I]-leptin studies. The effects of in vivo and in vitro leptin on transduction signal STAT proteins were investigated by immunoblotting antral extracts. RESULTS: Peripheral injection of leptin inhibited in a dose-dependent manner, basal gastric secretion, gastrinemia, and mucosal gastrin mRNA in vivo. mRNAs encoding the long (Ob-Rb) and short (Ob-Ra) receptor forms were detected in rat antral mucosa, as were STAT-1, -3, and -5b immunoreactive proteins. Isolated antral cells specifically bound [(125)I]-leptin, and addition of leptin to these cells inhibited the release of somatostatin and increased the release of gastrin. These effects were associated with an increase in nuclear STAT-3 proteins in vitro and in vivo. CONCLUSIONS: This study provides the first molecular evidence for the coexpression of leptin receptors and STAT-3 in antral mucosa. It provides further evidence for the involvement of leptin in the control of gastric secretions.
Asunto(s)
Proteínas Portadoras/fisiología , Proteínas de Unión al ADN/fisiología , Mucosa Gástrica/metabolismo , Proteínas de la Leche , Receptores de Superficie Celular , Transducción de Señal/fisiología , Transactivadores/fisiología , Animales , Proteínas de Unión al ADN/metabolismo , Ácido Gástrico/metabolismo , Gastrinas/sangre , Gastrinas/genética , Gastrinas/metabolismo , Leptina/sangre , Leptina/metabolismo , Leptina/farmacología , Masculino , Ratones , Antro Pilórico , ARN Mensajero/sangre , Ratas , Ratas Wistar , Receptores de Leptina , Proteínas Recombinantes/farmacología , Factor de Transcripción STAT1 , Factor de Transcripción STAT3 , Factor de Transcripción STAT5 , Somatostatina/metabolismo , Transactivadores/metabolismoRESUMEN
Dietary proteins are mostly absorbed as di- and tripeptides by the intestinal proton-dependent transporter PepT1. We have examined the effects of leptin on PepT1 function in rat jejunum and in monolayers of the human enterocyte-like 2 cell Caco-2. Leptin is produced by the stomach and secreted in the gut lumen. We show here that PepT1 and leptin receptors are expressed in Caco-2 and rat intestinal mucosal cells. Apical (but not basolateral) leptin increased Caco-2 cell transport of cephalexin (CFX) and glycylsarcosine (Gly-Sar), an effect that was associated with increased Gly-Sar uptake, increased membrane PepT1 protein, decreased intracellular PepT1 content, and no change in PepT1 mRNA levels. The maximal velocity (Vmax) for Gly-Sar transport was significantly increased by leptin, whereas the apparent Michaelis-Menten constant (Km) did not change. Furthermore, leptin-stimulated Gly-Sar transport was completely suppressed by colchicine, which disrupts cellular translocation of proteins to plasma membranes. Intrajejunal leptin also induced a rapid twofold increase in plasma CFX after jejunal perfusion with CFX in the rat, indicating enhanced intestinal absorption of CFX. These data revealed an unexpected action of gastric leptin in controlling ingestion of dietary proteins.
Asunto(s)
Proteínas Portadoras/fisiología , Cefalexina/metabolismo , Dipéptidos/metabolismo , Intestino Delgado/fisiología , Leptina/fisiología , Receptores de Superficie Celular , Simportadores , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Transporte Biológico , Brefeldino A/farmacología , Células CACO-2 , Proteínas Portadoras/metabolismo , Colchicina/farmacología , Cartilla de ADN , Dipéptidos/química , Humanos , Intestino Delgado/metabolismo , Datos de Secuencia Molecular , Transportador de Péptidos 1 , Ratas , Receptores de LeptinaRESUMEN
Hepatocyte growth factor (HGF) and its receptor, c-Met, are involved in cell transformation. To study their role in intestinal cell differentiation, we used Caco-2 colon cancer cells, which differentiate spontaneously into enterocytes during culture. Cells grown continuously in the presence of HGF reached confluence more quickly than control cells. Markers of enterocytic differentiation, such as alkaline phosphatase and sucrase-isomaltase activities, adhesion molecules, and structural proteins such as E-cadherin, villin, and F-actin were upregulated by HGF throughout the 35 days of culture, and actin fibers were reorganized. HGF also stimulated expression and tyrosine phosphorylation of c-Met and Gab-1 as well as protein kinase C (PKC)-alpha expression. PKC-alpha has been shown to be involved in intestinal differentiation. We therefore investigated the possibility that increases in PKC-alpha protein levels were responsible for the HGF-promoted events. We did this by incubating cells with Gö-6976, an inhibitor of PKC-alpha and -beta1, concomitantly with HGF. This inhibitor abolished the HGF-induced increase in villin levels before, but not after, confluence. Thus HGF accelerates Caco-2 cell differentiation and stimulates the metabolic and structural events accompanying this process. These HGF-promoted events may be mediated partly by Gab-1, and the effects of HGF on villin before confluence seem to involve PKC.
Asunto(s)
Diferenciación Celular/efectos de los fármacos , Enterocitos/citología , Enterocitos/metabolismo , Factor de Crecimiento de Hepatocito/farmacología , Actinas/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Fosfatasa Alcalina/genética , Fosfatasa Alcalina/metabolismo , Biomarcadores , Células CACO-2 , Cadherinas/metabolismo , Proteínas Portadoras/metabolismo , Neoplasias del Colon , Enterocitos/efectos de los fármacos , Humanos , Immunoblotting , Isoenzimas/antagonistas & inhibidores , Isoenzimas/genética , Isoenzimas/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas de Microfilamentos/metabolismo , Microscopía Fluorescente , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Fosforilación , Proteína Quinasa C/antagonistas & inhibidores , Proteína Quinasa C/genética , Proteína Quinasa C/metabolismo , Proteína Quinasa C-alfa , Proteínas Proto-Oncogénicas c-met/metabolismo , Proteínas Recombinantes/farmacología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Complejo Sacarasa-Isomaltasa/genética , Complejo Sacarasa-Isomaltasa/metabolismo , Factores de Tiempo , Proteína de la Zonula Occludens-1RESUMEN
Leptin, the product of the ob gene, plays a key role in the regulation of food intake via a cross-talk between hypothalamic leptin receptors and neuropeptides that affect feeding behaviour. Recent studies have shown a synergistic interaction between leptin and cholecystokinin (CCK) leading to suppression of food intake, which involves CCK-1 receptors and capsaicin-sensitive vagal fibres. In this study, we have investigated the presence of leptin receptors in afferent and efferent neurons of the vagus nerve. By using reverse transcription-polymerase chain reaction, mRNAs encoding long (Ob-Rb) and short (Ob-Ra) leptin receptor isoforms were detected in the rat nodose ganglion, which contains the cell bodies of the vagal afferent neurons. Western blot analysis confirmed the presence of leptin receptor-immunoreactive proteins in extracts from the vagal trunk. Immunohistochemistry showed the presence of leptin receptors and the leptin-induced transcription factor STAT3 in the cytoplasm of nodose ganglion cells. In cervical vagal segments, levels of leptin receptor protein displayed physiological regulation, with decreased amounts after feeding and increased levels after food restriction. In addition, leptin receptor and STAT3 immunoreactivities were detected in neurons of the nucleus of tractus solitarius (NTS) and the dorsal motor nucleus of the vagus nerve (DMNX) by immunofluorescence histochemistry. Furthermore, direct double-labelling demonstrated colocalization of Ob-Rb and STAT3 immunoreactivities in cholinergic vagal efferent cell bodies of the DMNX. It is speculated that vagal leptin receptors, apart from being activated by adipocyte-derived leptin, may also be influenced by leptin produced by the stomach. This may explain the synergistic action of leptin and CCK on neuronal activity in the NTS and on food intake.
Asunto(s)
Vías Aferentes/metabolismo , Proteínas Portadoras/metabolismo , Ingestión de Alimentos/fisiología , Vías Eferentes/metabolismo , Leptina/metabolismo , Neuronas Aferentes/metabolismo , Receptores de Superficie Celular , Nervio Vago/metabolismo , Vías Aferentes/citología , Animales , Proteínas Portadoras/genética , Proteínas de Unión al ADN/metabolismo , Vías Eferentes/citología , Técnica del Anticuerpo Fluorescente , Masculino , Bulbo Raquídeo/citología , Bulbo Raquídeo/metabolismo , Neuronas Aferentes/citología , Ganglio Nudoso/citología , Ganglio Nudoso/metabolismo , Isoformas de Proteínas/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Receptores de Leptina , Factor de Transcripción STAT3 , Transactivadores/metabolismo , Nervio Vago/citologíaRESUMEN
We investigated the peripheral effects of an H3-receptor agonist and an H3-receptor antagonist (R)alpha-methylhistamine (Ralpha-MeHA) and thioperamide, respectively, on basal feeding and the CCK8-induced inhibition of food intake in rat. Intraperitoneal injection of thioperamide reduced food intake in a dose-dependent manner with maximal inhibition (35%, P<0.01 vs saline) at 3 mg/kg. (R)alpha-MeHA (0.3-3 mg/kg i.p.), an H3-receptor agonist alone had no effect on feeding but reversed the thioperamide-induced inhibition of food intake in a dose-dependent manner. The maximal feeding inhibitory dose of thioperamide (3 mg.kg i.p) increased by 40% and 22 % (P<0.01 vs saline) brain and stomach histamine contents, respectively. Histamine (0.3 - 6 mg/kg i.p.) and CCK-8 (3 - 30 microg/kg i.p) also inhibited food intake in a dose-dependent manner. Inhibition was 20% to 40% for histamine and 40% to 80% (P<0.01 vs saline) for CCK8. CCK-8 inhibition of feeding was increased by thioperamide and prevented by (R)alpha-MeHA in a dose-dependent way. In addition, CCK-8 did not reduce food intake if rats were pretreated with pyrilamine or ranitidine postsynaptic H1- and H2-receptor antagonists respectively. Our data suggest that the H3-receptor is involved in basal feeding. They also suggest that CCK satiety depends upon the release of histamine which acts on the H2- and H1-receptors, the final mediators of this effect.
Asunto(s)
Colecistoquinina/metabolismo , Ingestión de Alimentos/efectos de los fármacos , Agonistas de los Receptores Histamínicos/administración & dosificación , Antagonistas de los Receptores Histamínicos/administración & dosificación , Receptores Histamínicos H3/metabolismo , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Colecistoquinina/farmacología , Relación Dosis-Respuesta a Droga , Mucosa Gástrica/metabolismo , Histamina/administración & dosificación , Histamina/metabolismo , Inyecciones Intraperitoneales , Masculino , Metilhistaminas/administración & dosificación , Piperidinas/administración & dosificación , Pirilamina/administración & dosificación , Ranitidina/administración & dosificación , Ratas , Ratas Wistar , Sincalida/administración & dosificación , Estómago/efectos de los fármacosRESUMEN
Tumour progression to the metastatic phenotype is mainly dependent on tumour cell invasiveness. Cell migration is a crucial step in this process. Here we investigate the effect of hepatocyte growth factor (HGF) on the induction of in vitro invasiveness of poorly aggressive Caco-2 colonic cancer epithelial cells. Invasion assays through a Matrigel barrier were performed. Proteases were assessed by zymography, reverse transcription-polymerase chain reaction and immunoblotting. Caco-2 cells were found to express HGF receptor but not HGF and to secrete several proteases, namely matrix metalloproteinase-1 (MMP-1), MMP-2, possibly MMP-9 and urokinase plasminogen activator (uPA). Exogenous HGF promoted invasiveness of Caco-2 cells through an artificial basement membrane matrix and enhanced their production of proteases. In addition, analyses of media at the end of invasion assays indicated that anti-HGF antibody inhibited protease production in parallel with cell invasion. The involvement of proteases in the HGF-induced invasion process was further investigated using either a synthetic general MMP inhibitor or neutralizing antibodies against MMPs or uPA. All components significantly inhibited HGF-promoted cell invasion. Moreover, specific inhibitors of PKCalpha/beta1 and PI3 kinase also decreased both HGF-promoted cell invasion and protease expression in invasion assay media. Thus, our findings provide evidence that the process of HGF-activated invasiveness of Caco-2 cells involves PI3 kinase and PKC and results from close association of two events, stimulation of cell motile activity and concomitant overproduction of proteases, which permits cell migration through a degraded extracellular matrix.
Asunto(s)
Neoplasias del Colon/patología , Factor de Crecimiento de Hepatocito/fisiología , Metaloproteinasas de la Matriz/metabolismo , Invasividad Neoplásica , Metástasis de la Neoplasia , Fosfatidilinositol 3-Quinasas/metabolismo , Proteína Quinasa C/metabolismo , Células CACO-2 , Neoplasias del Colon/enzimología , Humanos , Metaloproteinasas de la Matriz/biosíntesis , Proteínas Proto-Oncogénicas c-met/metabolismoRESUMEN
The recent discovery of gastric leptin has initiated several investigations on the possible role of leptin in digestive physiology. The following clues are currently suggested: leptin might control meal size in cooperation with Cholecystokinin, help cytoprotection of the gastric mucosa, play a role in gut inflammatory processes, regulate secretion of gastric hormones such as gastrin and somatostatin, and modulate intestinal transport of small peptides. The present review is a brief survey of the most significant advances in these issues.
Asunto(s)
Regulación del Apetito/fisiología , Mucosa Gástrica/metabolismo , Leptina/fisiología , Animales , Transporte Biológico , Colecistoquinina/metabolismo , Colecistoquinina/fisiología , Citoprotección , Mucosa Gástrica/citología , Gastrinas/metabolismo , Gastrinas/fisiología , Gastritis/inmunología , Gastritis/metabolismo , Humanos , Leptina/metabolismo , Péptidos/metabolismo , Somatostatina/metabolismo , Somatostatina/fisiologíaRESUMEN
Transforming growth factor-alpha (TGF-alpha) is overexpressed in colonic carcinomas and promotes mucosal wound healing. It may be implicated in chronic inflammatory bowel disease (IBD). We analyzed the expression of TGF-alpha and its receptor, epidermal growth factor receptor (EGF-r), in the colonic mucosa of patients with Crohn's disease (CD) or ulcerative colitis (UC), in active or inactive stages, as compared with controls. Proteins and mRNA were detected in biopsies from the right and left colon and in surgical colonic specimens. Immunoblot analysis revealed TGF-alpha protein as a 29 kDa band. This band was normally expressed in uninvolved colonic mucosa of patients with CD or UC whether in active or inactive stages, but decreased or absent in involved mucosa of active IBD, even when TGF-alpha mRNA and EGF-r protein were detected. In the unaffected mucosa of CD, the intensity of TGF-alpha immunoreactivity was similar to that of controls in the right colon but stronger (P = 0.05) in the left colon. There was no TGF-alpha overexpression in dysplastic regions. In conclusion, in active IBD disease, the decreased TGF-alpha protein amount seems not only related to epithelial cell loss but reflects a down-regulation at least at the protein level. We speculate that TGF-alpha does not play a role within the active stage but may be implicated later in the repair process.
Asunto(s)
Colon/química , Receptores ErbB/aislamiento & purificación , Enfermedades Inflamatorias del Intestino/etiología , Mucosa Intestinal/química , Factor de Crecimiento Transformador alfa/aislamiento & purificación , Adulto , Anciano , Anciano de 80 o más Años , Biopsia , Western Blotting , Colitis Ulcerosa/etiología , Colonoscopía , Enfermedad de Crohn/etiología , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Distribución TisularRESUMEN
BACKGROUND AND AIM: The circulating peptide leptin produced by fat cells acts on central receptors to control food intake and body weight homeostasis. Contrary to initial reports, leptin expression has also been detected in the human placenta, muscles, and recently, in rat gastric chief cells. Here we investigate the possible presence of leptin and leptin receptor in the human stomach. METHODS: Leptin and leptin receptor expression were assessed by immunohistochemistry, reverse transcriptase-polymerase chain reaction (RT-PCR), and western blot analysis on biopsy samples from 24 normal individuals. Fourteen (10 healthy volunteers and four patients with non-ulcer dyspepsia and normal gastric mucosa histology) were analysed for gastric secretions. Plasma and fundic mucosa leptin content was determined by radioimmunoassay. RESULTS: In fundic biopsies from normal individuals, immunoreactive leptin cells were found in the lower half of the fundic glands. mRNA encoding ob protein was detected in the corpus of the human stomach. The amount of fundic leptin was 10.4 (3.7) ng leptin/g mucosa, as determined by radioimmunoassay. Intravenous infusions of pentagastrin or secretin caused an increase in circulating leptin levels and leptin release into the gastric juice. The leptin receptor was present in the basolateral membranes of fundic and antral gastric cells. mRNA encoding Ob-RL was detected in both the corpus and antrum, consistent with a protein of approximately 120 kDa detected by immunoblotting. CONCLUSION: These data provide the first evidence of the presence of leptin and leptin receptor proteins in the human stomach and suggest that gastric epithelial cells may be direct targets for leptin. Therefore, we conclude that leptin may have a physiological role in the human stomach, although much work is required to establish this.
Asunto(s)
Células Principales Gástricas/metabolismo , Leptina/biosíntesis , Receptores de Superficie Celular , Receptores de Péptidos/biosíntesis , Adulto , Biopsia , Western Blotting , Proteínas Portadoras/metabolismo , Células Principales Gástricas/patología , Femenino , Humanos , Inmunohistoquímica , Leptina/análisis , Masculino , Persona de Mediana Edad , Pentagastrina/farmacología , ARN Mensajero/análisis , Radioinmunoensayo , Receptores de Leptina , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Secretina/fisiologíaRESUMEN
In this work, we used colon carcinoma cell-line HCT116 to study the involvement of the 86-kDa subunit (Ku86) of DNA-protein kinase (DNA-PK) in human tumoural cell proliferation. We transfected these cells with a 639-bp cDNA encoding a Ku86 portion inserted into pcDNA3.1 vector in the antisense orientation. After selection by neomycin, we obtained more than 300 resistant colonies. In the Y'A5 colony that we chose as total population, we showed by PCR and RT-PCR that pcDNA3/Ku86 antisense was integrated in genomic DNA and that transcript was present. After cloning, we selected two clones, A20 and A23, which contained significatively reduced level of Ku86 protein. These two clones displayed a reduced DNA-PK activity from 44% to 71% and a slower growth than control cells. These results suggest that the HCT116 cell-line is a useful tool to investigate the role of Ku86 in the regulation of human tumoural cell growth.
Asunto(s)
Antígenos Nucleares , Neoplasias del Colon/patología , ADN Helicasas , ADN sin Sentido/genética , Proteínas de Unión al ADN/metabolismo , Proteínas Nucleares/metabolismo , Western Blotting , División Celular/efectos de los fármacos , Clonación Molecular , Neoplasias del Colon/genética , Neoplasias del Colon/metabolismo , Proteínas de Unión al ADN/genética , Resistencia a Antineoplásicos , Humanos , Autoantígeno Ku , Neomicina/farmacología , Proteínas Nucleares/genética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transfección , Células Tumorales CultivadasRESUMEN
Matrix metalloproteinases (MMP) are enzymes responsible for extracellular matrix degradation which play a role in cancer progression and metastatic spreading. We investigated the effects of the MMP inhibitor, batimastat, in vitro on the proliferation and invasiveness of the rat colon cancer cell line DHD/K12, and in vivo on the growth of an aggressive model of peritoneal carcinomatosis producing haemorrhagic ascites and metastases, obtained in the rat by i.p. injection of DHD/K12 cells. MMP production was studied in conditioned culture media, solid tumors and ascitic fluid. In vivo, after injection of tumor cells on day 0, rats received i.p. daily either batimastat (30 mg/kg) or equal volume of vehicle from day 2 until killing on day 43 (series I) or from day 13 until death (series II). The grade of peritoneal carcinomatosis, ascite volume, number and size of liver metastases were evaluated in both series, and survival in series II. MMPs-1, -2 and -9 were identified in culture media, tumors and ascites. In vitro, batimastat did not modify DHD/K12 cell proliferation and slightly reduced cell invasion. In vivo, in series I, batimastat treatment totally prevented peritoneal carcinomatosis and liver metastasis development. In series II, it significantly prolonged survival (P < 0.0002) and reduced peritoneal carcinomatosis (P < 0.001) and hepatic metastases number as compared with controls. However, batimastat-treated rats of the two series had peritoneal inflammation with marked ascites. Nevertheless, inhibition of MMP is a new therapeutic approach which may be promising in treatment of microtumors as in more advanced cancer stages.
Asunto(s)
Antineoplásicos/farmacología , Carcinoma/prevención & control , Neoplasias del Colon/enzimología , Metaloendopeptidasas/antagonistas & inhibidores , Proteínas de Neoplasias/antagonistas & inhibidores , Neoplasias Peritoneales/prevención & control , Fenilalanina/análogos & derivados , Tiofenos/farmacología , Animales , Carcinoma/mortalidad , Carcinoma/patología , Colagenasas/metabolismo , Neoplasias del Colon/mortalidad , Neoplasias del Colon/patología , Femenino , Gelatinasas/metabolismo , Masculino , Metaloproteinasa 1 de la Matriz , Metaloproteinasa 2 de la Matriz , Metaloproteinasa 9 de la Matriz , Metaloendopeptidasas/metabolismo , Invasividad Neoplásica , Neoplasias Peritoneales/mortalidad , Neoplasias Peritoneales/patología , Fenilalanina/farmacología , Ratas , Células Tumorales Cultivadas/efectos de los fármacosRESUMEN
DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylates, in the presence of double-stranded DNA, several transcription-, replication- and repair -factors. Its interaction with the DNA-binding regulatory component Ku (p86-/p70-Ku) is required for stabilization and activity. We have previously shown that p86-Ku behaves as a specific receptor for the growth inhibitory tetradecapeptide, somatostatin. In this work, we investigate a possible regulation by somatostatin analogs, of DNA-PK activity in the human gastric tumoral HGT1/clone6 cell-line. We demonstrate that a 48 h-preincubation of cells with octreotide or RC-160, stimulates DNA-PK activity by 8 and 10 fold with ED50s of 1 and 0.1 nM, respectively. These stimulations appearing only after 3 h were inhibited by cycloheximide. They were not observed in a cell clone which was transfected by a cDNA encoding p86-Ku antisense. This study demonstrates the existence of a new somatostatin signaling pathway involving the stimulation of DNA-PK activity.
Asunto(s)
Antígenos Nucleares , Antineoplásicos/farmacología , ADN Helicasas , Octreótido/farmacología , Proteínas Serina-Treonina Quinasas/metabolismo , Somatostatina/análogos & derivados , Neoplasias Gástricas/enzimología , Catálisis , Células Clonales , ADN Complementario/genética , ADN Complementario/metabolismo , Proteína Quinasa Activada por ADN , Proteínas de Unión al ADN/biosíntesis , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Relación Dosis-Respuesta a Droga , Humanos , Cinética , Autoantígeno Ku , Proteínas Nucleares/biosíntesis , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Oligodesoxirribonucleótidos Antisentido/genética , Oligodesoxirribonucleótidos Antisentido/metabolismo , Somatostatina/farmacología , Neoplasias Gástricas/patología , Transfección , Células Tumorales CultivadasRESUMEN
Hydrochloric acid (HCl) secretion into the gastric lumen is a specialized process driven primarily by H(+)/K(+)-ATPase or proton pump, a unique enzyme expressed in high quantities by the parietal cells. Physiological regulation of this secretion involves central signals conveyed by the vagus nerve and local mechanisms mediated by cholinergic and peptidergic fibers of the gastric wall, as well as amine or peptide secreting cells located in the fundic and antral epithelia. Among these cells, the enterochromaffin-like (ECL) cell plays a major role: it responds to gastrinic, cholinergic and adrenergic stimulations to secrete histamine, which in turn activates an H(2) receptor on the parietal cell. The H(2) receptor is responsible for intracellular production of cAMP, a second messenger critical for the secretory machinery to be triggered. The blockade of this receptor by specific antagonists results in a dramatic albeit surmountable inhibition of HCl output. The blockade of the proton pump is an alternative means of inhibition. This can be achieved by a series of benzimidazole derivatives which specifically accumulate in the parietal cell secretory canaliculus and covalently bind to ATPase extracellular sites. The resulting inhibition is stronger and lasts longer than with the H(2) antagonists. Furthermore, it is effective regardless of the stimulatory status. Therefore, proton pump inhibitors (PPIs) are of special interest in the treatment of acid-related diseases such as gastric and duodenal ulcer, as well as reflux esophagitis.
RESUMEN
BACKGROUND AND METHODS: Recent studies suggest that glycine-extended gastrin (G17-gly) stimulates in vitro proliferation of the pancreatic cell line AR4-2J, through selective receptors distinct from the CCK-B/G-receptor mediating the effects of amidated gastrin (G17). The aims of our study were to examine the effects of G17 and G17-gly on the growth of the colorectal cancer cell line LoVo and to determine the receptor involved by using selective receptor-antagonist. RESULTS: Both G17 and G17-gly stimulated [3H]-thymidine incorporation in a concentration-dependent fashion. Maximal stimulation (153 +/- 18% and 166 +/- 17% of control, p < 0.01) was achieved with 10 nM G17 and 100 nM G17-gly, respectively. These stimulations were fully prevented by the presence of 10 pM YM022, a G/CCK B receptor-antagonist, but unaffected by L364,718, a CCK A receptor-antagonist. Basal growth of LoVo cells was inhibited by YM022 and stimulated by L364,718. CCK A and G/CCK B receptors mRNA were detected in the cells. Gastrin immunoreactivity was detected in the cells (16 pM) and in the extracellular medium (4.5 pM). CONCLUSION: Both G17 and G17-gly stimulate LoVo cells growth through the activation of a gastrin/CCK B receptor. The evidence for secreted gastrin and CCK A and B receptors mRNA may further suggest the existence of an autocrine loop involving a stimulatory gastrin/CCK B receptor.
Asunto(s)
Neoplasias Colorrectales/patología , Gastrinas/fisiología , Receptores de Colecistoquinina/fisiología , Benzodiazepinas/farmacología , División Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Antagonistas de Hormonas/farmacología , Humanos , Receptor de Colecistoquinina A , Receptor de Colecistoquinina B , Receptores de Colecistoquinina/antagonistas & inhibidores , Células Tumorales CultivadasRESUMEN
Matrix metalloproteinases (MMPs) and growth factors such as hepatocyte growth factor (HGF) are implicated in tumoral progression of several digestive cancers. The rat DHD/K12 colonic cancer cell line is very invasive in vivo. We showed by RT-PCR and western immunoblotting the presence of HGF receptor, c-Met, in DHD/K12 cells. Then, we detected by zymography and western blots the secretion of MMP-2 and MMP-9 in the conditioned medium of these cells. After 24 or 48 h of culture in medium supplemented with HGF, transforming growth factor-alpha (TGF-alpha) or sodium butyrate, MMP production by DHD/K12 cells was stimulated by HGF and TGF-alpha and inhibited by sodium butyrate. Knowing the capacity of MMPs to degrade the extracellular matrix and thus to favour tumoral invasion, results suggest that HGF is implicated in the aggressive behaviour of DHD/K12 cells since it increased MMPs secretion by these cells.
Asunto(s)
Factor de Crecimiento de Hepatocito/farmacología , Metaloendopeptidasas/metabolismo , Animales , Butiratos/farmacología , Neoplasias del Colon/enzimología , Neoplasias del Colon/patología , Matriz Extracelular/enzimología , Factor de Crecimiento de Hepatocito/fisiología , Invasividad Neoplásica , Proteínas Proto-Oncogénicas c-met/análisis , Proteínas Proto-Oncogénicas c-met/genética , Proteínas Proto-Oncogénicas c-met/fisiología , Ratas , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factor de Crecimiento Transformador alfa/farmacología , Células Tumorales CultivadasRESUMEN
The circulating peptide leptin, which is the product of the ob gene, provides feedback information on the size of fat stores to central Ob receptors that control food intake and body-weight homeostasis. Leptin has so far been reported to be secreted only by adipocytes and the placenta. Here we show that leptin messenger RNA and leptin protein are present in rat gastric epithelium, and that cells in the glands of the gastric fundic mucosa are immunoreactive for leptin. The physiological function of this previously unsuspected source of leptin is unknown. However, both feeding and administration of CCK-8 (the biologically active carboxy-terminal end of cholecystokinin) result in a rapid and large decrease in both leptin cell immunoreactivity and the leptin content of the fundic epithelium, with a concomitant increase in the concentration of leptin in the plasma. These results indicate that gastric leptin may be involved in early CCK-mediated effects activated by food intake, possibly including satiety.
Asunto(s)
Proteínas/análisis , Estómago/química , Adipocitos/metabolismo , Animales , Mucosa Gástrica/química , Gastrinas/farmacología , Leptina , Masculino , Reacción en Cadena de la Polimerasa , Proteínas/genética , Proteínas/metabolismo , ARN Mensajero/análisis , Ratas , Ratas Wistar , Sincalida/farmacología , Estómago/efectos de los fármacosRESUMEN
We investigated the effects of the novel CCKB/gastrin antagonist YM022 on gastric acid secretion in vivo and in vitro, compared to CI-988 and L365,260 as reference antagonists. In the anaesthetized rat, pentagastrin-induced stimulation of gastric acid secretion was dose-dependently and up to 100% inhibited by i.v. administration of YM022 with an ID50 of 0.009 +/- 0.0006 mumol/kg h in comparison to 0.6 +/- 0.03 and 3.40 +/- 0.05 mumol/kg h for CI-988 and L-365,260, respectively. In the gastric fistula cat, i.v. administration of YM022 produced a similar inhibitory effect with an ID50 of 0.02 mumol/kg in comparison to 1.6 and 2.5 mumol/kg for CI-988 and L-365,260, respectively. Furthermore, bolus injection of 0.6 mumol/kg YM022 produced 100% inhibition within 30 min and 85% inhibition was still observed after 3 h. In the isolated rabbit gastric glands, CCK8-stimulated 14C-aminopyrine uptake was inhibited according to the following rank order of potency: YM022 (IC50 = 0.0012 microM) > > CI-988 (IC50 = 0.2 microM) > > L365,260 (IC50 = 2.8 microM). Unlike with L365,260, no influence of CI-988 and YM022 on histamine-stimulated acid output was shown in this study. Thus, YM022 is a highly potent and selective gastric CCKB/gastrin receptor antagonist and has a long-lasting inhibitory effect on gastric acid secretion.
Asunto(s)
Benzodiazepinas/farmacología , Ácido Gástrico/metabolismo , Mucosa Gástrica/efectos de los fármacos , Antagonistas de Hormonas/farmacología , Receptores de Colecistoquinina/antagonistas & inhibidores , Aminopirina/metabolismo , Animales , Benzodiazepinonas/farmacología , Gatos , Colecistoquinina/antagonistas & inhibidores , Relación Dosis-Respuesta a Droga , Fístula Gástrica/metabolismo , Mucosa Gástrica/metabolismo , Gastrinas/antagonistas & inhibidores , Gastrinas/farmacología , Indoles/farmacología , Masculino , Meglumina/análogos & derivados , Meglumina/farmacología , Compuestos de Fenilurea/farmacología , Conejos , Ratas , Ratas Sprague-Dawley , Receptor de Colecistoquinina A , Receptor de Colecistoquinina BRESUMEN
BACKGROUND & AIMS: Long-term survival of patients with Zollinger-Ellison syndrome (ZES) is largely determined by the presence or absence of liver metastases. However, because of the lack of precision of these criteria, development of further indicators is still required. Recent evidence showing that autoantibodies directed against the p53 protein could predict poor survival for some types of cancers prompted us to investigate the presence of such antibodies in sera from patients with ZES and their potential value as survival indicator. METHODS: Anti-p53 antibodies were detected in the sera of 44 consecutive patients with ZES using both an enzyme-linked immunosorbent assay (ELISA) and Western blotting. The mean follow-up of these patients was 92 months. RESULTS: Anti-p53 antibodies were detected in 7 of the patients with ZES (16%) by both ELISA and Western blotting. Univariate and multivariate analyses showed that the presence of anti-p53 antibodies (P = 0.0009 and P = 0.017, respectively) and liver metastases (P = 0.0009 and P = 0.012, respectively) was independently associated with shorter survival. CONCLUSIONS: These results suggest that anti-p53 antibodies are an indicator of survival and could be used in combination with staging to determine which patients with ZES have poor prognoses and therefore require reinforced therapy.
Asunto(s)
Autoanticuerpos/sangre , Proteína p53 Supresora de Tumor/inmunología , Síndrome de Zollinger-Ellison/inmunología , Síndrome de Zollinger-Ellison/mortalidad , Adulto , Autoanticuerpos/inmunología , Biomarcadores , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Análisis de Supervivencia , Síndrome de Zollinger-Ellison/sangreRESUMEN
Somatostatin and its analogues are now of current use in the management of endocrine gastroentero-pancreatic (GEP) tumours for the purpose of inhibiting hormone hypersecretion, carrying scintigraphy imaging and attempting to slow down tumour growth. Recent molecular studies have revealed the existence of up to five membrane somatostatin receptor subtypes termed SSTR1-5. However, whether or not scintigraphy imaging and tumour characteristics are correlated with specific subtype(s) remains unclear. SSTR1-5 messenger RNA (mRNA) transcripts were investigated in 38 endocrine GEP tumours (32 islet cell tumours, six carcinoid) using reverse transcriptase polymerase chain reaction (RT-PCR), and their distribution was analysed with respect to tumour characteristics and scintigraphy imaging. SSTR2, SSTR5 and SSTR4 were detected in most cases of endocrine GEP tumours (92%, 84%, and 82% respectively), but SSTR1 and SSTR3 were less frequently observed (66% and 50% respectively). No clear-cut correlation was found between tumour characteristics and subtype mRNA distribution. Moreover, no differences in mRNA subtype distribution were found between the 17 tumours detected by scintigraphy and the four tumours not detected by this method. Somatostatin receptor mRNA subtypes are widely expressed in endocrine GEP tumours, but their distribution is not correlated with tumour characteristics or scintigraphy positivity.