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Biomedical subjects

H Kopelman

Publications and source records attributed to H Kopelman.

At least 19 recordsLinked to original sources
PubMed1996

Progesterone and estradiol inhibit CFTR-mediated ion transport by pancreatic epithelial cells.

The cystic fibrosis (CF), gene product, CF transmembrane conductance regulator (CFTR), is responsible for adenosine 3',5'-cyclic monophosphate (cAMP)-activated Cl- transport in epithelial cells, and mutant CFTR accounts for the pathology in the CF pancreas. We have previously shown that both isolated rabbit pancreatic acini and the human pancreatic duct cell line PANC-1 possess a cAMP-activated Cl- conductance identified as CFTR. We report here that preincubation in either of the female hormones progesterone or beta-estradiol inhibits activation by cAMP, but not by Ca2+ ionophore, of PANC-1 cell volume reduction under isotonic conditions. cAMP-activated cell volume reduction is sensitive to antisense, but not sense, CFTR oligodeoxynucleotide. Furthermore, progesterone inhibits cAMP-activated Cl- efflux from rabbit acinar cells. Moreover preincubation with progesterone, but not beta-estradiol, reduces CFTR mRNA and protein levels as measured using polymerase chain reaction amplification of reverse-transcribed acinar RNA and Western blots of protein from acinar membranes. We conclude that female hormones inhibit CFTR functional activity in pancreatic epithelial cells by different mechanisms.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1995

Rabbit pancreatic acini express CFTR as a cAMP-activated chloride efflux pathway.

Cystic fibrosis transmembrane conductance regulator (CFTR) is responsible for adenosine 3',5'-cyclic monophosphate (cAMP)-activated chloride transport in epithelial cells. Isolated rabbit pancreatic acini possess a cAMP-activated chloride efflux mechanism distinct from zymogen granule secretion. To determine whether CFTR is expressed in acini, we used polymerase chain reaction (PCR) to amplify a 480-base pair (bp) sequence from reverse-transcribed rabbit acinar RNA. The PCR product was consistent with a 480-bp band amplified in T84 cells, and its sequence was > 90% homologous to human CFTR. CFTR antibody M3A7 recognized a 180- and a 160-kDa protein from acinar membranes consistent with bands seen in Chinese hamster ovary (CHO) cells transfected with CFTR. To determine if CFTR was responsible for the cAMP-activated chloride efflux previously demonstrated in pancreatic acini, we incubated acinar cells for 20 h with 1.75 microM CFTR antisense or sense oligodeoxynucleotide. Chloride efflux, in response to 8-bromoadenosine 3',5'-cyclic monophosphate and phorbol ester but not to calcium ionophore, was selectively inhibited by CFTR antisense oligodeoxynucleotide. Antisense oligodeoxynucleotide did not inhibit acinar amylase secretion. These findings indicate that isolated pancreatic acini can be used for future studies of CFTR expression and function.

Amylases↗
PubMed1994

Developmental differences of cystic fibrosis transmembrane conductance regulator functional expression in isolated rat fetal distal airway epithelial cells.

Fluid secretion from the pulmonary epithelium may play a significant role in determining intrauterine lung development. We used suspensions of distal pulmonary epithelial cells isolated from rat fetuses to assess a shift in secretory mechanisms occurring in the lung of this species during late gestation. The impact of cAMP on distal airway epithelial cells isolated from d 18 to d 21 rat fetuses was evaluated with measurements of cell volume and 36Cl efflux rates. At d 18, 8-Br-cAMP stimulated a volume reduction measured by electronic cell sizing that was prevented by the Cl- channel blocker anthracene-9-carboxylate (A-9C) and reflected in an increased rate of A-9C sensitive 36Cl efflux. Because the cystic fibrosis transmembrane conductance regulator (CFTR) is thought to be a cAMP-regulated Cl- channel, we measured the effect of prior cell incubation with oligodeoxynucleotides antisense to the transcription site of the human CFTR gene on these events. We found that in antisense oligomer-treated cells, but not in sense oligomer-treated controls, volume and 36Cl efflux responses to 8-Br-cAMP were prevented in d 18 cells. In d 21 cells, 8-Br-cAMP did not stimulate volume reduction but the calcium ionophore A23187 did elicit cell volume reduction in cells suspended in an isotonic Ca(2+)-containing medium that was prevented by A-9C. This response to the ionophore was not found in the d 18 cells, and incubation with the antisense CFTR oligomer had no effect on the ionophore-induced responses in d 21 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1993

Antisense oligodeoxynucleotide to the cystic fibrosis transmembrane conductance regulator inhibits cyclic AMP-activated but not calcium-activated cell volume reduction in a human pancreatic duct cell line.

Cystic fibrosis (CF) is characterized by a defect in cAMP-regulated chloride channels in epithelial cells. The CF gene product CF transmembrane conductance regulator (CFTR) is expressed in the apical membrane of pancreatic duct cells, and mutant CFTR accounts for the pathology in the CF pancreas. PANC 1, a pancreatic duct cell line, has not been considered a good model for studying CFTR and pancreatic chloride transport because CFTR mRNA and protein are undetectable using standard methods. Using electronic cell sizing and cell volume reduction under isotonic conditions, PANC 1 cells were found to possess both cAMP and calcium-activated chloride conductances. Using CFTR antisense oligodeoxynucleotides, the cAMP-activated conductance could be specifically inhibited in a concentration- and time-dependent manner. These findings demonstrate that PANC 1 cells express CFTR and a CFTR-independent calcium-activated chloride channel. With electronic cell sizing and CFTR antisense oligodeoxynucleotides, PANC 1 cells can provide an ideal system for the study of pancreatic duct cell physiology and pathophysiology with respect to the role of CFTR in the pancreas. These findings also suggest that antisense oligodeoxynucleotides may provide a more sensitive yet highly specific means of detecting low levels of expression of CFTR than currently available.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1991

Cyclic AMP-sensitive chloride efflux in rabbit pancreatic acini.

We studied chloride efflux from isolated rabbit pancreatic acini in suspension, by loading with 36Cl to steady state and rapidly washing acini by filtration to determine 36Cl cpm/micrograms DNA remaining. Linear loss of acinar chloride occurred over 5 min (k = 0.038 +/- 0.008 min-1, n = 5). Forskolin (5 x 10(-5) M) increased the rate of chloride efflux (k = 0.100 +/- 0.016 min-1, n = 5, p less than 0.001) 2.6-fold. At 5 min, forskolin increased acinar cAMP levels (1065 +/- 254 versus 7 +/- 2 pmol/mL, n = 5, p less than 0.005) and percentage of chloride efflux (37.4 +/- 2.3 versus 26.0 +/- 2.2%, n = 13, p less than 0.005). The chloride channel inhibitor anthracene-9-carboxylic acid (10(-3) M) had no effect on chloride loss from acini exposed to vehicle (30.9 +/- 1.9 versus 29.9 +/- 2.3%, n = 4), but completely inhibited forskolin-stimulated efflux at 5 min (40.0 +/- 2.4 versus 29.3 +/- 2.4%, n = 5, p less than 0.005). Manipulation of extracellular calcium concentration demonstrated that chloride efflux was not coupled to zymogen granule amylase release. Secretin (10(-7) M) increased acinar cAMP levels (68 +/- 22 versus 7 +/- 2 pmol/mL, n = 5, p less than 0.05) and significantly increased the loss of chloride from acini (34.9 +/- 1.4 versus 26.1 +/- 1.7%, n = 7, p less than 0.005) without affecting amylase release. Secretagogue-stimulated amylase release by cholecystokinin octapeptide (10(-8) M) and carbamylcholine (10(-5) M), did not increase chloride efflux at 5 min.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Origins of chloride and bicarbonate secretory defects in the cystic fibrosis pancreas, as suggested by pancreatic function studies on control and CF subjects with preserved pancreatic function.

Pancreatic function studies were performed under continuous intravenous stimulation with secretin and cholecystokinin (0.125 mu/kg/min) in 40 patients with CF who had no evidence of steatorrhea and 69 control subjects without CF, who also lacked evidence of steatorrhea. A total of 41 studies in CF patients and 74 in control subjects were analysed for trypsin, electrolyte, and water secretion. 51% of the CF and 51% of the control tests showed trypsin secretion responses between 1000 mu/kg/hr (the lower limit of the normal range) and 2695 mu/kg/hr, while 16.2% of the control tests exceeded this range. Secretion of water and electrolytes, including both chloride and bicarbonate, was significantly lower in the CF subjects when compared to the appropriate controls in the normal and subnormal trypsin output range. Chloride-bicarbonate ratios, although higher in subjects with evidence of pancreatic damage, were identical in CF and control studies. A specific ductal lesion in CF which altered the balance of acinar (predominately chloride) and ductal (predominately bicarbonate) secretion, or encouraged the equilibration of ductal fluid with plasma in the distal ducts, would increase chloride-bicarbonate ratios, as in the subjects with reduced trypsin secretion. Since this did not occur in the patients with normal function, our results suggest that chloride and bicarbonate secretions are impaired to a similar extent in CF, possibly through a dependence on the same phenotypic abnormality.

Bicarbonates↗
PubMed1988

Impaired chloride secretion, as well as bicarbonate secretion, underlies the fluid secretory defect in the cystic fibrosis pancreas.

Pancreatic fluid and electrolyte secretion was assessed in 56 patients with cystic fibrosis (CF) and 56 non-CF control subjects undergoing pancreatic function testing while stimulated with cholecystokinin and secretin. Both CF patients and control subjects exhibited a wide range of pancreatic function. Fluid and trypsin outputs were positively correlated in both groups. Fluid output in CF subjects was significantly lower, however, than that of control subjects at any given level of trypsin output. Sodium, bicarbonate, and chloride secretions were all significantly decreased in CF subjects. Bicarbonate and chloride were important determinants of fluid secretion, but at any given bicarbonate or chloride output CF subjects secreted significantly less fluid than control subjects. When bicarbonate and chloride were analyzed as simultaneous predictor variables, adjusted fluid secretion was not significantly different in CF and control subjects. Diminished fluid secretion in CF subjects is therefore caused by impaired chloride, as well as bicarbonate, secretion.

Bicarbonates↗
PubMed1987

Atrial natriuretic factor and exocrine pancreas: effects on the secretory process.

Immunoreactive atrial natriuretic factor (ANF) has recently been identified in pancreatic acinar cells. The current study therefore, was, designed to ascertain whether the atrial peptide exercises any biological effects on the exocrine pancreas. When isolated rat pancreatic acinar cells were incubated with rat ANF (8-33), a concentration-dependent increase in cGMP synthesis was observed (EC50 about 5 X 10(-9) M) with the peak response occurring within 2.5 min of exposure of the cells to the peptide. ANF did not affect basal or secretagogue (carbachol +/- DbcAMP, CCK-OP, or forskolin)-induced amylase secretion from acinar cells, nor did it affect [3H]thymidine incorporation into DNA or [3H]leucine incorporation into trichloroacetic acid-precipitable protein. ANF was also infused intravenously (0.01-0.25 micrograms/min) in rabbits with cannulated pancreatic ducts, and the peptide stimulated a dose-dependent secretion of cGMP into pancreatic juice. ANF, by itself, did not affect protein or fluid secretion from rabbit pancreas; when co-infused with secretin (0.1 CU/min), which is not an acinar cell secretagogue in rabbits, ANF increased fluid secretion in one of three animals tested. The data suggest that acinar cells possess functional ANF receptors, whose activation results in both synthesis and secretion of cGMP. Intra-acinar cell cGMP is not involved in the enzyme secretory process. Secreted nucleotide is not co-released with digestive hydrolases, and does not, by itself, appear to have an intraluminal effect on fluid secretion. The mechanism by which acinar cells secrete cGMP into pancreatic juice as well as the biological significance of intracellular (acinar), and extracellular (intraluminal) cGMP remain to be elucidated.

Amylases↗
PubMed1987

Atrial natriuretic factor and exocrine pancreas: autoradiographic localization of binding sites and ultrastructural evidence for internalization of endogenous ANF.

Atrial natriuretic factor (ANF) binding sites in adult rat exocrine pancreas were studied by autoradiography using slide-mounted frozen tissue sections with mono-iodinated ANF (101-126) as the tracer. Radiolabel was displaced by unlabeled atrial peptide (IC50 = 2 X 10(-11) M). High specific labeling was found in pancreatic acini. The presence of endogenous ANF has also been demonstrated in the exocrine pancreas by immunocytochemistry on ultra-thin sections obtained by cryoultramicrotomy. ANF-like immunoreactivity was found in acinar and centro-acinar cells as well as cells of the intercalated duct. For these cells, immunostaining was observed at the plasma membrane level, in the cytoplasm and nucleus. In the cytoplasm, ANF-like immunoreactivity was observed in the cytoplasmic matrix, mitochondria, and zymogen granules. In the nucleus, ANF-like immunoreactivity was distributed in the vicinity of the heterochromatin region primarily in the euchromatin. It was also detected in the plasma membrane of microvilli of acinar and duct cells, and in the lumen of secretory ducts. In centro-acinar cells, the reaction product was also found sparsely at the nuclear envelope. No immunoreactivity was observed when anti-human ANF serum preincubated with rat ANF was used. No modifications were observed when this antiserum was preincubated with heterologous peptides (NPY, CRF, GRF, TRH, somatostatin). These data provide autoradiographic evidence of ANF binding sites, indicate the presence of this peptide in acinar and centro-acinar cells as well as cells of the intercalated duct, and immunocytochemical evidence for the internalization of endogenous ANF by exocrine pancreas.

Animals↗
PubMed1985

Pancreatic fluid secretion and protein hyperconcentration in cystic fibrosis.

To study pancreatic protein and water secretion in 28 patients with cystic fibrosis and 21 controls matched for pancreatic acinar function as defined by trypsin secretion, we used a quantitative-marker perfusion technique and continuous intravenous secretin-pancreozymin stimulation. Regardless of the level of pancreatic acinar function, secretions from the patients contained significantly higher concentrations of protein than those from the controls. Total protein output and albumin:protein ratios were not increased in secretions from the patients, but their fluid secretion was significantly decreased at any level of pancreatic function. A significant linear correlation was found between protein and volume secretion in the patients (r = 0.86, P less than 0.001), most of whom had a fluid output of less than 4.2 ml per kilogram of body weight per hour. No such relation was found in the control subjects, whose flow was always above 4.2 ml per kilogram per hour. We conclude that fluid secretion in patients with cystic fibrosis may be a rate-limiting factor in protein output and that a limited flow of hyperconcentrated protein secretions may predispose to protein precipitation and ductal obstruction in the pancreas.

Adolescent↗
PubMed1985

Bentiromide test for assessing pancreatic dysfunction using analysis of para-aminobenzoic acid in plasma and urine. Studies in cystic fibrosis and Shwachman's syndrome.

We evaluated the bentiromide test by analyzing para-aminobenzoic acid (PABA) in plasma and urine (a) for the identification of patients with complete pancreatic insufficiency and (b) as an alternative to the secretin-cholecystokinin test. Nine control subjects, 18 patients with cystic fibrosis, and 4 patients with Shwachman's syndrome were studied. Based upon the secretin-cholecystokinin test, pancreatic function was judged to be less than 0.1% of normal in 7 patients with cystic fibrosis and malabsorption and between 0.7% and 90% of control values in 11 patients with cystic fibrosis and 4 patients with Shwachman's syndrome without malabsorption. The bentiromide test was performed in two stages: first with bentiromide alone, then with equimolar free PABA. After ingestion of free PABA, the plasma profile and urinary excretion of PABA were comparable in controls, patients with cystic fibrosis, and patients with Shwachman's syndrome. Thirty minutes after oral bentiromide, plasma PABA values in patients with and without malabsorption were significantly lower than in the control group. From 60 to 180 min after ingestion, plasma PABA levels in patients without malabsorption were no different from controls; whereas levels in patients with malabsorption were significantly lower than in controls and in those without malabsorption, reaching the highest significance at 90 min. Similar results were obtained when the urinary excretion of PABA was considered. Only the 90-min plasma test reliably detected cystic fibrosis patients with steatorrhea, however. Duodenal colipase output was highly correlated with both the 90-min plasma test and the urinary excretion of PABA, with similar results for lipase and trypsin output. Reliable detection of pancreatic dysfunction, nevertheless, was not obtained even with the plasma test, in cystic fibrosis patients with greater than 5%-10% of the mean normal enzyme output. In patients with Shwachman's syndrome, none of whom had malabsorption, the plasma and urinary test failed to detect pancreatic dysfunction even with enzyme output as low as 1% of normal.

4-Aminobenzoic Acid↗
PubMed1984

Electrocardiographic changes associated with isolated right ventricular infarction.

Isolated infarction of the right ventricle is an extremely rare entity. A patient is described with diffuse interstitial lung disease who developed ST segment elevation in inferior and anterior leads on a routine electrocardiogram and at autopsy was found to have an isolated right ventricular infarct involving approximately 70% of the right ventricular circumference without involvement of the left ventricle and septum. This case illustrates that isolated right ventricular infarction in the presence of cor pulmonale and right ventricular hypertrophy can produce an injury current in the limb and precordial leads of the electrocardiogram which mimics that seen in typical transmural infarction of the left ventricle.

Electrocardiography↗
PubMed1984

Pancreatic function testing in cystic fibrosis.

None of the currently available clinical techniques for measuring pancreatic function in patients with CF can be considered "ideal". The "direct" pancreatic stimulation test provides the most information and remains the only truly accurate test of exocrine function, particularly in those individuals with pancreatic sufficiency. The invasive, complex nature of the pancreatic stimulation tests precludes its use as a routine method of assessing pancreatic function in all patients and limits its value for monitoring function in those individuals who are expected to deteriorate with time. Many of the indirect tests of pancreatic function are of value for identifying CF patients with complete pancreatic insufficiency who may require therapy with pancreatic enzyme supplements and fat soluble vitamins. However, none of the currently available "indirect" tests of exocrine function are sensitive enough to evaluate or monitor CF patients with pancreatic sufficiency. Blood tests, capable of measuring pancreatic enzymes in the circulation, such as immunoreactive trypsinogen, may be of value for monitoring pancreatic function in patients with CF, but further longitudinal studies are necessary to fully evaluate this approach. More sophisticated "indirect" tests of exocrine pancreatic function are vitally needed to facilitate adequate assessment of this important digestive organ in patients with cystic fibrosis.

Cystic Fibrosis↗
PubMed1984

Abnormal mucus: nominated but not yet elected.

Mucus secretions of the intestine may contribute to plug formation in CF, through several mechanisms acting alone or simultaneously. Excessive mucin secretion and relative dehydration of secretions would elevate mucin concentration and enhance mucus gel formation. Deficient pancreatic enzymes combined with intrinsic hyperglycosylation of mucin or an increase in covalently bound fatty acid groups on mucins, would retard normal proteolytic degradation of mucins. Excessive calcium ions in the ducts would tend to decrease mucin solubility, while excessive hydrogen ions and protein (especially albumin) together with mucins, may be responsible for the appearance of proteinaceous ductal 'plugs' characteristic of CF pathology.

Albumins↗