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Biomedical subjects

M Korc

Publications and source records attributed to M Korc.

At least 181 records · Page 10Linked to original sources
PubMed1986

Processing of epidermal growth factor by suckling and adult rat intestinal cells.

Preparations of intestinal villus and crypt cells were isolated from jejunal segments of suckling (14-day-old) and adult (6- to 7-wk-old) rats. These cell preparations were incubated with 125I-labeled epidermal growth factor (EGF) at 37 degrees C to determine the extent of cellular processing of 125I-EGF in vitro. 125I-EGF bound specifically to both crypt and villus cells of suckling rats and was internalized and degraded to similar extents in both cell preparations. Analysis of the 125I radioactivity in the medium and cellular extract by gel filtration on Sephadex G-25 columns demonstrated the presence of [125I]iodotyrosine (24-31%) following 30 min of incubation. This degradation of EGF was accompanied by a loss in the capacity to bind to anti-EGF antibodies (34-52%) and A431 cells (28-48%). Binding, internalization, and processing of 125I-EGF by crypt cell preparations of adult rats was similar to that of suckling rats. In contrast, little degradation of 125I-EGF to iodotyrosine and loss of cell binding capability occurred following incubation with adult villus cells. However, a considerable loss in binding to anti-EGF antibody was detected (48%). The results indicate that isolated intestinal cells are capable of degrading 125I-EGF in vitro in a manner similar to that seen after oral feeding in vivo. They also indicate differences in the processing of 125I-EGF by isolated villus cells of adult compared with suckling rat.

Aging↗
PubMed1986

Isolated pancreatic acini from suckling and weanling rats: changes in amino acid incorporation and carbachol-stimulated amylase secretion with age.

To characterize the changes in pancreatic function during postnatal development, isolated pancreatic acini were prepared from rats aged 8-9, 12-14 and 20 days and from adult rats. Isolated acini maintained a normal microscopic appearance and viability as judged by exclusion of trypan blue and linear incorporation of tritiated leucine into total protein. The rate of incorporation in 8-day-old acini was 20% of that observed in adult rats. Significant dose-dependent increases in amylase release in response to carbachol were observed in all age groups; stimulated amylase secretion was significantly less in the 8- to 9- and 12- to 14-day-old animals than in the 20-day-old and adult rats. These data indicate that viable isolated pancreatic acini can be prepared from suckling rats and that these acini exhibit an altered in vitro responsiveness to carbachol. This preparation should therefore be a useful model for in vitro studies of the development of pancreatic function.

Amino Acids↗
PubMed1986

Effects of malnutrition and chronic reserpine treatment on pancreatic exocrine function.

The chronically reserpine-treated rat, an experimental model for cystic fibrosis, exhibits generalized exocrinopathy, impaired pancreatic secretion, and decreased pancreatic amylase. Although chronic reserpine treatment induces malnutrition by decreasing food consumption and growth, the effects of this malnutrition per se on the exocrine pancreas have not been considered. In this study, the effects of chronic reserpine treatment and malnutrition on the exocrine pancreas were determined using pair-fed controls. Male, Sprague-Dawley rats were treated daily subcutaneously for 5 to 7 days with: no injection (control), 1.0 ml/kg vehicle or sham (control-sham, pair fed-sham), or 0.5 mg/kg reserpine (chronically reserpine-treated). Both chronic reserpine-treatment and pair-feeding significantly decreased food consumption (40%), body weight (51 and 59%), total pancreatic amylase (49 and 56%) and specific amylase activity (62 and 61%), pancreatic protein (65 and 75%), and pancreatic weights (62 and 65%) compared to controls. These decreases, however, were comparable between the chronically reserpine-treated and pair fed-sham rats. In contrast, the secretory response to the biologically active cholecystokinin analog cholecystokinin octapeptide was significantly attenuated in isolated pancreatic acini prepared from reserpine-treated rats compared to that from either control or pair-fed sham rats. Malnutrition decreased pancreatic amylase activity and protein comparably to reserpine treatment, but only partially attenuated the secretory response to cholecystokinin octapeptide. Based on the results of this study, pair-fed controls should be used to distinguish between the effects of reserpine alone and the induced malnutrition on pancreatic exocrine function in studies of this experimental model of cystic fibrosis.

Amylases↗
PubMed1986

Epidermal growth factor inhibits the proliferation of a human endometrial carcinoma cell line.

Specific epidermal growth factor (EGF) receptors were measured in RL95-2 human endometrial carcinoma cells. At 37 C, binding of 125I-labeled EGF was associated with marked ligand internalization. Maximal cell surface binding occurred within 10 min. Bound [125I]EGF was partially degraded to low mol wt products, and this degradation was blocked by the lysosomotropic compound ammonium chloride. At 4 C, maximal cell surface binding occurred at 3 h. Scatchard analysis of data obtained after 3 h at 4 C revealed a single order of binding sites with a Kd of 1.8 nM and approximately 150,000 surface receptors/cell. EGF, at a concentration of 0.83 nM, inhibited the proliferation of RL95-2 cells. Inhibition was reversible and was associated with morphological changes leading to a fusiform appearance of the growth-arrested cells. These findings indicate that RL95-2 human endometrial carcinoma cells bind, internalize, and degrade EGF in a manner similar to that described for other target cells for EGF action. The ability of EGF to inhibit the growth of RL95-2 cells supports the hypothesis that this type of inhibition is dependent on postreceptor events, rather than on the presence of an overabundance of EGF receptors.

Adenocarcinoma↗
PubMed1985

Binding and processing of epidermal growth factor in Panc-I human pancreatic carcinoma cells.

The binding of 125I-labeled epidermal growth factor (EGF) was studied in Panc-I human pancreatic carcinoma cells. At 37 degrees C, binding was rapid and associated with marked endocytosis of the ligand. Bound EGF was sequentially converted to a number of more acidic species as follows: pI 4.55 to pI 4.2, to pI 4.35, to pI 4.0. EGF internalization and processing were blocked at 4 degrees C. EGF did not alter cell growth when Panc-I cells were incubated in the presence of 2 to 10% serum. In contrast, when the serum concentration was lowered to 0.1%, EGF significantly enhanced cell replication after 6 days of culture.

Autoradiography↗
PubMed1985

Residual inhibition of epidermal growth factor binding by pancreatic secretagogues and phorbol ester in rat pancreas.

Cholecystokinin-octapeptide (CCK8) inhibits 125I-labeled epidermal growth factor (EGF) cell-associated radioactivity in pancreatic acini, ostensibly as a result of its ability to mobilize cellular Ca2+. The phorbol ester tetradecanoyl phorbol acetate (TPA), a compound that activates protein kinase C, mimics the inhibitory action of CCK8. In the present study we examined the relationship between occupancy of the cholecystokinin (CCK) receptor, the subsequent inhibition of EGF binding, and the potential role of C-kinase activation in mediating this inhibition. Proglumide and dibutyryl cyclic GMP (dbGMP), two distinct competitive antagonists of CCK8, reversed the inhibitory actions of CCK8. Analysis of steady-state saturation kinetics of 125I-EGF binding indicated that CCK8 decreased the apparent affinity of the EGF receptor, mainly as a result of a marked decrease in the amount of internalized ligand. TPA also inhibited 125I-EGF internalization. Removal of CCK8 and TPA from incubation medium did not abolish their inhibitory actions. Carbachol, but not bombesin, exerted a similar residual inhibitory effect. It is suggested that in addition to acting via Ca2+, certain pancreatic secretagogues may also act through C-kinase to regulate EGF binding.

Animals↗
PubMed1985

Recycling of epidermal growth factor in a human pancreatic carcinoma cell line.

PANC-1 human pancreatic carcinoma cells readily bound and internalized 125I-labeled epidermal growth factor (EGF). Bound 125I-labeled EGF was then partially processed to a number of high molecular weight acidic species. Percoll gradient centrifugation of cell homogenates indicated that the majority of 125I activity localized to several intracellular vesicular compartments. Both intact EGF and its processed species were subsequently released into the incubation medium. A major portion of the released radioactivity was capable of rebinding to the cell. Only a small amount of bound 125I-labeled EGF was degraded to low molecular weight products, and this degradation was completely blocked by methylamine. This lysosomotropic compound did not arrest either the generation or the extrusion of the major high molecular weight species of processed EGF (pI 4.2). These findings suggest that in PANC-1 cells, bound EGF undergoes only limited processing. Both intact EGF and its major processed species bypass the cellular degradative pathways, are slowly released from the cell, and then rebind to the cell.

Biological Transport↗
PubMed1984

Epidermal growth factor binding is altered in pancreatic acini from diabetic rats.

The effects of streptozotocin-induced diabetes on 125I-labeled epidermal growth factor (EGF) binding were studied in rat pancreatic acini. 125I-EGF binding was one-half maximal at 20 min, and maximal at 90 min. Saturation data revealed a decreased binding capacity in diabetic acini when compared with normal acini. Insulin, in vivo, normalized the decreased binding capacity. 125I-EGF internalization was also decreased in diabetic rat acini. Further, the inhibitory effect of cholecystokinin-octapeptide (CCK8) on cell-associated 125I-EGF radioactivity was significantly greater in diabetic than in normal rat acini. These findings suggest that insulin deficiency may lead to defective regulation of the exocrine pancreas by EGF.

Animals↗
PubMed1984

Cytosolic calcium regulates epidermal growth factor endocytosis in rat pancreas and cultured fibroblasts.

Cholecystokinin octapeptide (CCK8), the COOH-terminal moiety of cholecystokinin (CCK), exerted a rapid inhibitory effect on total cell-associated 125I-labeled epidermal growth factor (125I-EGF) binding by decreasing the rate of EGF internalization in isolated rat pancreatic acini. Removal of CCK8 from incubation medium followed by extensive washing of acini did not abolish its inhibitory effect, indicating that its action was not readily reversible. Proglumide, a competitive antagonist of CCK8, blocked the inhibitory action of the secretagogue. Addition of CCK8 to cells previously exposed to 125I-EGF did not enhance the release of cell-associated 125I activity. CCK8 did not inhibit the binding of 125I-labeled insulin to pancreatic acini. Other pancreatic secretagogues that enhance digestive-enzyme release through Ca2+, including caerulein, bombesin, carbachol, gastrin, and the Ca2+ ionophore A23187, also inhibited cell-associated 125I-EGF radioactivity. Further, at 37 degrees C the ionophore A23187 inhibited specific 125I-EGF binding in human A-431 carcinoma cells, Swiss 3T3 cells, and Rat-1 fibroblasts, and this effect was abolished when 125I-EGF internalization was reduced by incubating cells at 4 degrees C. It is concluded that alterations in cellular Ca2+ in the pancreas and other cells lead to inhibition of EGF endocytosis.

Animals↗
PubMed1984

Manganese action on protein synthesis in diabetic rat pancreas: evidence for a possible physiological role.

The actions of manganese on [3H]phenylalanine incorporation into pancreatic acinar protein from streptozotocin-diabetic rats were compared with the actions of several divalent cations, insulin, and pancreatic secretagogues. In the presence of Ca2+, 7 X 10(-4) M manganese greatly enhanced [3H]phenylalanine incorporation. Under similar incubation conditions, magnesium exerted only a slight stimulatory effect on incorporation, whereas cobalt and nickel failed to enhance incorporation. Removal of Ca2+ from incubation medium, or replacement of Ca2+ by either barium or strontium, abolished the stimulatory effect of manganese on incorporation. Lanthanum, at a concentration that inhibits stimulated Ca2+ influx in acini (10(-4) M), also blocked the stimulatory effect of manganese but did not alter the actions of insulin on incorporation. Dibutyryl cyclic GMP, a competitive antagonist of cholecystokinin-octapeptide, and atropine, a competitive antagonist of carbachol, blocked the stimulatory effects of the respective secretagogues on incorporation. Neither antagonist altered the actions of manganese. These findings suggest that in the face of insulin deficiency manganese enhances pancreatic protein synthesis in diabetic rats via a Ca2+-dependent mechanism that is distinct from the actions of other divalent cations, insulin and pancreatic secretagogues.

Animals↗
PubMed1983

Direct modulation of epidermal growth factor binding by cholecystokinin.

The effects of cholecystokinin-octapeptide (CCK8), the biologically active C-terminal moiety of cholecystokinin (CCK), on the binding of epidermal growth factor (EGF) were studied in isolated rat pancreatic acini. CCK8 inhibited 125I-EGF binding in a dose-dependent manner. One-half maximal inhibition occurred at 5 X 10(-10)M, and maximal inhibition at 10(-8)M CCK8. This inhibitory effect was detectable within 5 minutes of addition of CCK8, and was not associated with enhanced degradation of 125I-EGF in incubation media. Unlabeled EGF exerted only a slightly greater inhibitory effect than CCK8 on 125I-EGF binding at equivalent molar concentrations. In contrast to CCK8, the gastrointestinal hormone vasoactive intestinal polypeptide (VIP) did not significantly alter EGF binding. CCK8 also inhibited EGF binding in mouse pancreatic acini, but did not alter binding in A-431 human carcinoma cells. These findings suggest that physiological levels of CCK may regulate EGF binding in the pancreas and other tissues with receptors for both hormones. They thus point to a previously unrecognized mechanism for hormonal interaction.

Animals↗
PubMed1983

Binding of epidermal growth factor in rat pancreatic acini.

Specific, saturable EGF receptors were demonstrated in isolated rat pancreatic acini. Binding of EGF to these receptors was one-half maximal at 20 min and maximal at 120 min. Scatchard analyses revealed a single order of binding sites with a Kd of 4.90 nM. Following binding, EGF was rapidly internalized and converted to two acidic species. EGF did not alter either basal amylase release or the rate of [3H]phenylalanine incorporation into TCA-precipitable protein. The finding of high affinity EGF receptors in pancreatic acinar cells supports the hypothesis that EGF participates in the long-term regulation of pancreatic exocrine function.

Animals↗
PubMed1983

Effect of lanthanum on pancreatic protein synthesis in streptozotocin-diabetic rats.

The role of extracellular Ca2+ in mediating the stimulatory effect of cholecystokinin octapeptide (CCK8) on [3H]phenylalanine incorporation into protein was studied in isolated pancreatic acini from streptozotocin-diabetic rats. The stimulatory effect of CCK8 (10(-10) M) on [3H]phenylalanine incorporation was completely abolished by preincubating acini with either 10(-4) M lanthanum or 10(-3) M manganese. At these concentrations neither compound altered the basal rate of amino acid incorporation, and both compounds inhibited CCK-mediated Ca2+ influx without affecting either basal or CCK-mediated 45Ca2+ efflux. Lanthanum (10(-4) M) also blocked the stimulatory effect of the cholinergic analogue carbachol (10(-5) M) on amino acid incorporation but did not alter the stimulatory effect of insulin (1.67 x 10(-8) M). Vasoactive intestinal polypeptide and 12-O-tetradecanoyl-phorbol-13-acetate failed to increase the incorporation of [3H]phenylalanine into acinar protein. These findings suggest that CCK and other pancreatic secretagogues that act via Ca2+ enhance protein synthesis by increasing cell membrane permeability to Ca2+ and provide additional evidence that this may be an important mechanism by which CCK regulates pancreatic exocrine function.

Animals↗
PubMed1983

Manganese action on pancreatic protein synthesis in normal and diabetic rats.

The effects of manganese on [3H]phenylalanine incorporation into protein were studied in pancreatic acini prepared from normal and streptozotocin-induced diabetic rats. In the absence of added Ca2+ manganese exerted a biphasic effect on [3H]phenylalanine incorporation in both groups of acini. Significant stimulation occurred at 3 X 10(-5) M manganese. At higher concentrations manganese inhibited incorporation. The magnitude of stimulation was similar in all acini, whereas the magnitude of inhibition was greater in acini from normal rats. Addition of Ca2+ to incubation media abolished the stimulatory effect of manganese in normal rat acini and greatly enhanced it in diabetic rat acini, significant stimulation now occurring at 10(-5) M manganese. The magnitude of inhibition was again greater in acini from normal rats. Insulin in vivo partially reversed the diabetes-induced alterations in acinar cell responsiveness to manganese. The present findings suggest that streptozotocin-induced diabetes is associated with postreceptor alterations in the pancreatic acinar cells.

Animals↗
PubMed1982

Regulation of pancreatic protein synthesis by cholecystokinin and calcium.

The effect of cholecystokinin (CCK) on [3H]phenylalanine and [14C]tyrosine incorporation into protein was studied in isolated rat pancreatic acini. CCK8, the biologically active octapeptide of CCK, had a biphasic effect on amino acid incorporation. The magnitude of the stimulatory effect (maximal at 10(-10) to 3 X 10(-10) M CCK8) was greater in diabetic rats, whereas the magnitude of the inhibitory effect (maximal at 10(-8) M CCK8) was greater in normal rats. Omission of extracellular Ca2+ from the incubation media did not diminish the stimulatory action of CCK8. Addition of 10(-4) M EGTA to media containing no added Ca2+ lowered basal incorporation, abolished CCK's stimulatory effect and enhanced its inhibitory effect. Stimulation was restored in the presence of Ca2+. The stimulatory effect of the cholinergic analogue carbachol (10(-5) M) on amino acid incorporation was also abolished by removal of extracellular Ca2+, whereas the stimulatory effect of insulin (1.67 X 10(-8) M) remained intact. These findings suggest that CCK and other pancreatic secretagogues enhance pancreatic protein synthesis via Ca2+, whereas stimulation by insulin occurs via another mechanism.

Animals↗
PubMed1981

Insulin action in pancreatic acini from streptozotocin-treated rats. I. Stimulation of protein synthesis.

Pancreatic acini were prepared from rats rendered diabetic with streptozotocin. In this tissue, insulin stimulated [3H]leucine incorporation into protein. The full effects of insulin on this function were not immediate but increased linearly with time for up to 2 h of incubation. Insulin had a detectable effect on l eucine incorporation at 50 pM, a half-maximal effect at 0.7 nM, and a maximal effect at 30 nM. Desdipeptide proinsulin was only 10% as potent as native insulin in stimulating [3H]leucine incorporation, whereas proinsulin and desoctapeptide insulin were only 1% as potent. Insulin also increased the incorporation of [3H]valine and [35S]methionine into protein but did not increase the influx of either [14C]cycloleucine or alpha-[3H]aminoisobutyric acid. These observations suggested that the increased incorporation of labeled amino acid into protein reflected stimulation of protein synthesis rather than stimulation of amino acid transport. Furthermore, insulin at 1.67 nM significantly increased the acinar cell concentration of amylase. The present findings are consistent therefore with the concept that insulin regulates pancreatic exocrine functions, including protein and enzyme synthesis.

Amino Acids↗
PubMed1981

Insulin action in pancreatic acini from streptozotocin-treated rats. II. Binding of 125I-insulin to receptors.

The binding of 125I-insulin to its receptors was investigated with isolated pancreatic acini obtained from diabetic rats under incubation conditions identical to those used to study the effects of insulin on acinar cell protein synthesis. Binding was specific, time dependent, reversible, and linearly related to the acinar protein content. Degradation of insulin after 30 min of incubation was less than 10% of the total hormone present in the incubation medium. 125I-insulin dissociated from acini with a one-half time of 9 min. Unlabeled insulin at 83.5 nM accelerated the rate of dissociation of labeled insulin. 125I-insulin binding to acini was competitively inhibited by insulin and its analogues in proportion to their biological potencies. Scatchard analysis revealed a major class of insulin-binding sites with a Kd of 1.6 nM; maximal stimulation of protein synthesis was observed when > 90% of these high-affinity receptors were occupied. These studies indicate, therefore, that insulin binding to receptors on pancreatic acini can be correlated with subsequent regulation of biological functions.

Animals↗