RESUMO
We have explored whether lipopolysaccharide (LPS, endotoxin) induces pancreatic injury on pancreatic acinar cells both in vivo and in vitro. Wistar male rats were treated with four intraperitoneal injections of 10 mg/kg LPS, and AR4-2J cells were exposed to increasing doses of LPS. Expression of pancreatitis-associated-protein (PAP) mRNA was strongly induced in AR4-2J cells exposed to LPS, while amylase mRNA was reduced. LPS also induced apoptosis and expression of TNF-alpha, IL-1beta, and IL-8 mRNA in AR4-2J cells. The in vivo effect of LPS showed structural signs of cellular damage, including numerous cytoplasmic vacuoles, severe nuclear alterations, and high expression of PAP mRNA. This study demonstrated that LPS induced pancreatic damage by directly affecting the pancreatic acinar cells. The role of LPS in the pathophysiology of acute pancreatitis may be partly due to the effect LPS has on the acinar cell.
Assuntos
Antígenos de Neoplasias , Biomarcadores Tumorais , Lectinas Tipo C , Lipopolissacarídeos/toxicidade , Pâncreas/efeitos dos fármacos , Pancreatite/fisiopatologia , Síndrome de Resposta Inflamatória Sistêmica/fisiopatologia , Doença Aguda , Proteínas de Fase Aguda/genética , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Expressão Gênica/efeitos dos fármacos , Humanos , Injeções Intraperitoneais , Interleucina-1/genética , Interleucina-8/genética , Masculino , Pâncreas/patologia , Pâncreas/fisiopatologia , Pancreatite/induzido quimicamente , Pancreatite/patologia , Proteínas Associadas a Pancreatite , RNA Mensageiro/genética , Ratos , Ratos Wistar , Síndrome de Resposta Inflamatória Sistêmica/patologia , Fator de Necrose Tumoral alfa/genéticaRESUMO
To elucidate whether pancreatic acinar cell submitted to stress is able to express TNF-alpha, we studied TNF-alpha mRNA expression by Northern blot and in situ hybridization in healthy pancreas, in tissue from caerulein-induced pancreatitis and after lipopolysaccharide (LPS) treatment. In specimens from normal pancreas, TNF-alpha mRNA expression, as judged by both Northern blot and in situ hybridization, was negative, whereas a strong but transient expression was observed in acinar cells from caerulein pancreatitis and LPS treatment. TNF-alpha mRNA appeared as rapidly as 30 min after treatment, and was maximal 6 h after. At this time, there was mild infiltration consisting mostly of polymorphonuclear leukocytes (PMNL) and no signal of TNF-alpha transcript was found in their cytoplasm. Our results strongly indicate that pancreatic acinar cell is the source of TNF-alpha early in the course of acute pancreatitis and LPS treatment, and suggest that the expression of this cytokine is a part of a general response of the acinar cell to aggression.
Assuntos
Pâncreas/metabolismo , Fator de Necrose Tumoral alfa/genética , Animais , Ceruletídeo , Regulação da Expressão Gênica , Hibridização In Situ , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Pâncreas/citologia , Pancreatite/induzido quimicamente , Pancreatite/metabolismo , Pancreatite/patologiaRESUMO
Swiss mice were fed conventional lab chow and 10% ethanol or water as drinking fluid for 2 weeks. Pancreatic juice was obtained by cannulation of the bile pancreatic common duct of mice anesthetized with urethane. Isolated pancreatic lobules were also obtained. The flow rate and the amylase output were determined in pure pancreatic juice. The release of amylase was measured in pancreatic lobule preparations. The basal pancreatic juice flow rate and the amylase output were significantly increased by ethanol consumption. The magnitude of the pancreatic juice flow rate and the amylase output responses to increasing doses of bethanechol, a cholinergic agent, was significantly decreased in ethanol-fed mice. The amount of spontaneously released amylase was higher in pancreatic lobule preparations from ethanol-fed animals than that from control mice, and the difference was abolished by addition of atropine to the incubation media. The amylase release rate in response to increasing doses of bethanechol was significantly reduced in lobule preparations from the ethanol-fed group. These data indicate that ethanol intake in mice has a stimulating effect on the spontaneous pancreatic secretion and lends support to the hypothesis that ethanol consumption increases the intrapancreatic cholinergic tone.
Assuntos
Etanol/farmacologia , Suco Pancreático/metabolismo , Amilases/metabolismo , Animais , Betanecol , Compostos de Betanecol/farmacologia , Relação Dose-Resposta a Droga , Técnicas In Vitro , Masculino , Camundongos , Suco Pancreático/efeitos dos fármacosRESUMO
Expression of the c-myc and H-ras oncogenes, and of several genes specifically expressed in adult rat pancreas was investigated by monitoring changes in corresponding mRNA concentrations, by dot-blot hybridization, during the early phase of regeneration following subtotal pancreatectomy. The oncogenes c-myc and H-ras were overexpressed after 12-24 and 48 h, respectively, then returned to basal levels. The concentrations of mRNAs encoding amylase, chymotrypsinogen B, and trypsinogen I decreased during the regeneration time. By contrast, proinsulin I mRNA concentration was increased at 12-48 h after surgical resection, and actin mRNA concentration was increased at 12-48 h after surgical resection, and actin mRNA concentration was increased at 12 h after subtotal pancreatectomy and remained elevated thereafter. We concluded that regeneration after subtotal pancreatectomy is accompanied by repression of certain genes that are expressed in differentiated pancreatic tissue, and that derepression of other genes may be necessary for starting and/or maintaining the process of pancreatic regeneration.
Assuntos
Regulação da Expressão Gênica/fisiologia , Genes myc/genética , Genes ras/genética , Pâncreas/fisiologia , Regeneração/fisiologia , Animais , Colódio , Sondas de DNA , Immunoblotting , Masculino , Hibridização de Ácido Nucleico , RNA Mensageiro/metabolismo , Ratos , Ratos EndogâmicosRESUMO
To examine the effects of fasting on trophism and gene expression in pancreas, adult male rats were deprived of food from 0-6 d. Total DNA, RNA, and proteins, and specific mRNAs for rat amylase, chymotrypsinogen B, trypsinogen I, proinsulin I, and actin (assessed by employing cloned cDNAs and dot-blot hybridization) were quantitated in pancreas. Body and pancreatic wt diminished progressively to reach 65 and 75% of initial values at the 6th d of fasting. Protein/DNA and total RNA/DNA ratios decreased 2.04 and 2.31-fold, respectively, during 6 d of fasting. The concentration of amylase, chymotrypsinogen B, trypsinogen I, and actin mRNA, expressed as cpm/microgram RNA, decreased significantly throughout the study period, whereas the decrease observed in Proinsulin I mRNA concentration was not significantly different. When mRNA concentrations were refereed to the total content of DNA, however, the decrease was significant for all messengers tested. It is concluded that the prolonged absence of nutrients in the digestive tract exerts negative trophic influence on pancreas and triggers differential changes in pancreatic gene expression. These changes are gradual, asynchronic, and nonparallel.
Assuntos
Jejum/efeitos adversos , Pâncreas/metabolismo , Actinas/genética , Amilases/metabolismo , Animais , Atrofia , Quimotripsinogênio/genética , DNA/metabolismo , Jejum/metabolismo , Expressão Gênica , Masculino , Pâncreas/patologia , Proinsulina/genética , Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos , Tripsinogênio/genéticaRESUMO
Changes in pancreatic growth and in mRNA concentrations in rat pancreas were monitored by dot-blot hybridization with cloned cDNAs of rat amylase, chymotrypsinogen B, proinsulin I, and actin during the pre- and postnatal period in the rat. Wistar rats were killed at the 18th day of gestation and at the 1st, 10th, 20th, 35th, and 87th day of postnatal life. It was concluded from the ratio of pancreatic weight/body weight that pancreatic growth preceded body growth. Pancreatic protein and total RNA concentration increased 2.9 times during the period studies. All studied mRNAs increased in concentration during the postnatal development period. Messenger RNA for chymotrypsinogen B and proinsulin I exhibited a significant increase after birth, decreased by the 10th day of life, and increased thereafter. For amylase mRNA, no significant changes were observed around birth, a progressive increase occurring thereafter up to the 87th day of life. The mRNA for actin showed a progressive increase between the 18th day of gestation and the 20th postnatal day, after which it remained stable. We concluded that each mRNA showed a singular profile of increase during postnatal development.
Assuntos
Actinas/genética , Amilases/genética , Quimotripsinogênio/genética , Expressão Gênica , Pâncreas/crescimento & desenvolvimento , Proinsulina/genética , Animais , Feminino , Idade Gestacional , Masculino , Hibridização de Ácido Nucleico , Pâncreas/embriologia , Pâncreas/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos , Aumento de PesoRESUMO
The effect of repeated administration of haloperidol on the pancreatic secretion was studied in urethane-anesthetized Swiss mice. Haloperidol (2 mg/kg) injected daily i.p. for 7 days, increase the volume and protein content of the basal pancreatic juice significantly. This secretory activity was partially blocked by i.p. injection of atropine (5 mg/kg), both in control and treated animals. The volume of the secretory response to bethanechol, a cholinergic agonist, was decreased by haloperidol without any change in amylase release. From these findings it is concluded that repeated haloperidol treatment produces an increase of basal pancreatic secretion, which is probably the result of changes in the sensitivity of dopamine receptors of the gland.
Assuntos
Fibras Adrenérgicas/fisiologia , Haloperidol/farmacologia , Pâncreas/metabolismo , Suco Pancreático/metabolismo , Receptores Dopaminérgicos/fisiologia , Fibras Adrenérgicas/efeitos dos fármacos , Animais , Atropina/farmacologia , Betanecol , Compostos de Betanecol/farmacologia , Fibras Colinérgicas/efeitos dos fármacos , Fibras Colinérgicas/fisiologia , Relação Dose-Resposta a Droga , Masculino , Camundongos , Pâncreas/efeitos dos fármacos , Pâncreas/inervação , Receptores Dopaminérgicos/efeitos dos fármacosRESUMO
The effect of intraduodenal oleic acid administration on protein synthesis and enzymatic levels in rat pancreas was investigated. Sprague-Dawley rats were sacrificed at 20, 40, 60, and 80 min after intraduodenal oleic acid administration. Ten minutes before sacrifice, the rats were injected with 50 microCi 3H-Phenylalanine intraperitoneally. Amylase (Am), chymotrypsinogen (Chtg), trypsinogen (Tg) and lipase (Li) activities, and 3H-Phenylalanine incorporation to total secretory proteins were determined in pancreas homogenates. Forty minutes after oleic acid administration, the activities of Chtg, Tg and Li were significantly increased (45, 38 and 23%, respectively) above those from control rats. Amylase levels were not modified. Enzyme activities decreased below baseline levels by 60 and 80 min after oleic acid administration. The 3H-phenylalanine incorporation pattern exhibited a peak at 40 min. We conclude that intraduodenal oleic acid administration stimulates intrapancreatic enzyme content in a non-parallel fashion, before enzyme activities decreased below those from control rats. Protein synthesis was similarly affected by intraduodenal oleic acid.
Assuntos
Amilases/biossíntese , Quimotripsina/biossíntese , Lipase/biossíntese , Ácidos Oleicos/farmacologia , Pâncreas/enzimologia , Tripsinogênio/biossíntese , Animais , Duodeno , Indução Enzimática/efeitos dos fármacos , Masculino , Ácido Oleico , Ácidos Oleicos/administração & dosagem , Ratos , Ratos Endogâmicos , Estimulação QuímicaRESUMO
The effect of intraduodenal oleic acid administration on protein synthesis and enzymatic levels in rat pancreas was investigated. Sprague-Dawley rats were sacrificed at 20, 40, 60, and 80 min after intraduodenal oleic acid administration. Ten minutes before sacrifice, the rats were injected with 50 microCi 3H-Phenylalanine intraperitoneally. Amylase (Am), chymotrypsinogen (Chtg), trypsinogen (Tg) and lipase (Li) activities, and 3H-Phenylalanine incorporation to total secretory proteins were determined in pancreas homogenates. Forty minutes after oleic acid administration, the activities of Chtg, Tg and Li were significantly increased (45, 38 and 23
, respectively) above those from control rats. Amylase levels were not modified. Enzyme activities decreased below baseline levels by 60 and 80 min after oleic acid administration. The 3H-phenylalanine incorporation pattern exhibited a peak at 40 min. We conclude that intraduodenal oleic acid administration stimulates intrapancreatic enzyme content in a non-parallel fashion, before enzyme activities decreased below those from control rats. Protein synthesis was similarly affected by intraduodenal oleic acid.
RESUMO
Flow and enzymatic output from mouse pancreas were studied "in vivo" under bethanechol stimulation. A biphasic dose response curve was found in both, enzymatic output and juice flow, implicating that exocrine pancreatic juice secretion was also involved in restricted stimulation phenomenon.
Assuntos
Compostos de Betanecol/farmacologia , Pâncreas/efeitos dos fármacos , Suco Pancreático/metabolismo , Animais , Betanecol , Relação Dose-Resposta a Droga , Feminino , Masculino , Camundongos , Camundongos Endogâmicos CBA , Pâncreas/metabolismo , Suco Pancreático/enzimologia , Taxa Secretória/efeitos dos fármacos , Estimulação QuímicaRESUMO
Flow and enzymatic output from mouse pancreas were studied [quot ]in vivo[quot ] under bethanechol stimulation. A biphasic dose response curve was found in both, enzymatic output and juice flow, implicating that exocrine pancreatic juice secretion was also involved in restricted stimulation phenomenon.
RESUMO
Calcium secretion and chymotrypsin (CHT) outputs in pancreatic juice from mice were studied "in vivo". Pancreatic juice was collected after stimulation with secretin 32 mU/g. Secretin 32 mU/g plus cholecystokinin 16 mU/g (CCK) or secretin 32 mU/g plus bethanechol 0.2 ug/g. Calcium and CHT outputs were lower in mice treated with secretin alone than in those treated either with secretin plus CCK or with secretin plus bethanechol. Bethanechol or CCK addition to secretin produced a positive correlation between both parameters. Extrapolation of the regression line correlating calcium and CHT outputs indicates that at zero CHT output pancreatic juice contains calcium. These results suggest that calcium in mice pancreatic juice is produced by two different sources.
Assuntos
Compostos de Betanecol/farmacologia , Cálcio/metabolismo , Quimotripsina/metabolismo , Pâncreas/metabolismo , Secretina/farmacologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos DBA , Suco Pancreático/metabolismoRESUMO
Calcium secretion and chymotrypsin (CHT) outputs in pancreatic juice from mice were studied [quot ]in vivo[quot ]. Pancreatic juice was collected after stimulation with secretin 32 mU/g. Secretin 32 mU/g plus cholecystokinin 16 mU/g (CCK) or secretin 32 mU/g plus bethanechol 0.2 ug/g. Calcium and CHT outputs were lower in mice treated with secretin alone than in those treated either with secretin plus CCK or with secretin plus bethanechol. Bethanechol or CCK addition to secretin produced a positive correlation between both parameters. Extrapolation of the regression line correlating calcium and CHT outputs indicates that at zero CHT output pancreatic juice contains calcium. These results suggest that calcium in mice pancreatic juice is produced by two different sources.
RESUMO
An experimental model to perform dynamic studies of exocrine pancreatic secretion from mice has been developed. It consists of a microsurgical procedure in anesthetized mice using a stereoscopic microscope. The bile-pancreatic common duct was individualized, isolated and cannulated. A specially calibrated capillary tube was placed in the free end of the cannula and put on a millimetrical rule. Readings of pure pancreatic juice flow rate were performed minute by minute to obtain a flow rate curve. The procedure allowed the discrimination among increasing betanechol doses of 0.1, 0.2, and 0.4 microgram/g body weight.