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W Zawalich

Publications and source records attributed to W Zawalich.

15 recordsLinked to original sources
PubMed2005

Targeted expression of the anti-apoptotic gene CrmA to NOD pancreatic islets protects from autoimmune diabetes.

The activation of apoptosis is a critical mechanism by which pancreatic beta cells are destroyed in type 1 diabetes (T1DM). Strategies aimed at interfering with the apoptotic pathways could therefore be of potential therapeutic value. To this end, we generated NOD transgenic mice with targeted expression of the anti-apoptotic gene Cytokine response modifier A (CrmA) to pancreatic beta cells using the rat insulin promoter and the reverse tetracycline transactivator to express CrmA in a temporally controlled manner. Two lines of transgenic mice were studied whose expression of CrmA occurred only after feeding doxycycline food. Islet expression of CrmA partially protected pancreatic beta cells from the cytokine-mediated cytotoxicity in vitro and reduced modestly the spontaneous development of diabetes in NOD mice in vivo. In addition, beta cells from NOD CrmA mice were significantly protected from the destruction by diabetogenic T cells after adoptive transfer. More strikingly, NODCrmA mice were significantly resistant to the diabetogenic activity of a potent insulin-specific CD8 T-cell clone. Since these adoptive transfer models mainly represent the effector phase rather than the initiation phase of autoimmune diabetes, our data suggest that the latter is more sensitive to CrmA protection. We conclude that anti-apoptotic genes such as CrmA might be potential candidates to enhance islet graft survival in T1DM.

Adoptive Transfer↗
PubMed2001

Dephosphorylation of beta2-syntrophin and Ca2+/mu-calpain-mediated cleavage of ICA512 upon stimulation of insulin secretion.

Islet cell autoantigen (ICA) 512 is a receptor-tyrosine phosphatase-like protein associated with the secretory granules of neuroendocrine cells, including pancreatic beta-cells. Binding of its cytoplasmic tail to beta2-syntrophin suggests that ICA512 connects secretory granules to the utrophin complex and the actin cytoskeleton. Here we show that stimulation of insulin secretion from INS-1 cells triggers the biosynthesis of pro-ICA512 and the degradation of its mature form. Inhibition of calpain, which is activated upon stimulation of insulin secretion, prevents the Ca2+-dependent proteolysis of ICA512. In vitro mu-calpain cleaves ICA512 between a putative PEST domain and the beta2-syntrophin binding site, whereas binding of ICA512 to beta2-syntrophin protects the former from cleavage. beta2-syntrophin and its F-actin-binding protein utrophin are enriched in subcellular fractions containing secretory granules. ICA512 preferentially binds phospho-beta2-syntrophin and stimulation of insulin secretion induces the Ca2+-dependent, okadaic acid-sensitive dephosphorylation of beta2-syntrophin. Similarly to calpeptin, okadaic acid inhibits ICA512 proteolysis and insulin secretion. Thus, stimulation of insulin secretion might promote the mobilization of secretory granules by inducing the dissociation of ICA512 from beta2-syntrophin-utrophin complexes and the cleavage of the ICA512 cytoplasmic tail by mu-calpain.

Amino Acid Sequence↗
PubMed1996

Overexpression of parathyroid hormone-related protein in the pancreatic islets of transgenic mice causes islet hyperplasia, hyperinsulinemia, and hypoglycemia.

Parathyroid hormone-related protein (PTHrP) is produced by the pancreatic islet. It also has receptors on islet cells, suggesting that it may serve a paracrine or autocrine role within the islet. We have developed transgenic mice, which overexpress PTHrP in the islet through the use of the rat insulin II promoter (RIP). Glucose homeostasis in these mice is markedly abnormal; RIP-PTHrP mice are hypoglycemic in the postprandial and fasting states and display inappropriate hyperinsulinemia. At the end of a 24-hour fast, blood glucose values are 49 mg/dl in RIP-PTHrP mice, as compared to 77 mg/dl in normal littermates; insulin concentrations at this time are 6.3 and 3.9 ng/ml, respectively. Islet perifusion studies failed to demonstrate abnormalities in insulin secretion. In contrast, quantitative islet histomorphometry demonstrates that the total islet number and total islet mass are 2-fold higher in RIP-PTHrP mice than in their normal littermates. PTHrP very likely plays a normal physiologic role within the pancreatic islet. This role is most likely paracrine or autocrine. PTHrP appears to regulate insulin secretion either directly or indirectly, through developmental or growth effects on islet mass. PTHrP may have a role as an agent that enhances islet mass and/or enhances insulin secretion.

Animals↗
PubMed1992

Immunocytochemical localization of alpha-protein kinase C in rat pancreatic beta-cells during glucose-induced insulin secretion.

To investigate the role of protein kinase C (PKC) in the regulation of insulin secretion, we visualized changes in the intracellular localization of alpha-PKC in fixed beta-cells from both isolated rat pancreatic islets and the pancreas of awake unstressed rats during glucose-induced insulin secretion. Isolated, perifused rat islets were fixed in 4% paraformaldehyde, detergent permeabilized, and labeled with a mAb specific for alpha-PKC. The labeling was visualized by confocal immunofluorescent microscopy. In isolated rat pancreatic islets perifused with 2.75 mM glucose, alpha-PKC immunostaining was primarily cytoplasmic in distribution throughout the beta-cells. In islets stimulated with 20 mM glucose, there was a significant redistribution of alpha-PKC to the cell periphery. This glucose-induced redistribution was abolished when either mannoheptulose, an inhibitor of glucose metabolism, or nitrendipine, an inhibitor of calcium influx, were added to the perifusate. We also examined changes in the intracellular distribution of alpha-PKC in the beta-cells of awake, unstressed rats that were given an intravenous infusion of glucose. Immunocytochemical analysis of pancreatic sections from these rats demonstrated a glucose-induced translocation of alpha-PKC to the cell periphery of the beta-cells. These results demonstrate that the metabolism of glucose can induce the redistribution of alpha-PKC to the cell periphery of beta-cells, both in isolated islets and in the intact animal, and suggest that alpha-PKC plays a role in mediating glucose-induced insulin secretion.

Animals↗
PubMed1992

Ca(2+)-cyclic AMP interactions in sustained cellular responses.

As early as 1970 it was apparent that the cyclic AMP (cAMP) and Ca2+ messenger systems often interact to regulate cellular responses. Work over the past 20 years has greatly expanded our knowledge of these interactions, and has shown that these signalling systems interact in complex ways to regulate sustained cellular responses such as aldosterone secretion, smooth muscle contraction and insulin secretion. The latter system is considered in detail because it illustrates several types of interactions, both positive and negative, which help to determine the normal response of beta-cells to physiological stimuli, and how abnormalities in secretory patterns can develop as a consequence of the prolonged stimulation of a messenger system.

Animals↗
PubMed1990

Multiple effects of increases in phosphoinositide hydrolysis on islets and their relationship to changing patterns of insulin secretion.

Evidence continues to accumulate which suggests that increases in phosphoinositide hydrolysis play an integral role in beta cell insulin secretory response patterns. In the present report some of the methodological approaches utilized to monitor phosphoinositide-hydrolysis in islets are reviewed. The contribution of phosphoinositide-derived second messenger molecules to acute insulin release as well as the induction of time dependent potentiation are also discussed. Finally, the role of impaired phosphoinositide hydrolysis in the induction of time dependent suppression of insulin release is also explored. It is concluded that alterations in phosphoinositide hydrolysis contribute, at least in part, to both ordered and disordered patterns of insulin release noted under a variety of experimental conditions.

Animals↗
PubMed1987

Effect of chronic hyperglycemia on in vivo insulin secretion in partially pancreatectomized rats.

We have examined the effect of chronic (4 wk) hyperglycemia on insulin secretion in vivo in an awake, unstressed rat model. Three groups of animals were examined: control, partial (90%) pancreatectomy, and partial pancreatectomy plus phlorizin, in order to normalize plasma glucose levels. Insulin secretion in response to arginine (2 mM), hyperglycemia (+100 mg/dl), and arginine plus hyperglycemia was evaluated. In diabetic compared with control animals three specific alterations were observed: (a) a deficient insulin response, in both first and second phases, to hyperglycemia; (b) an augmented insulin response to the potentiating effect of arginine under basal glycemic conditions; and (c) an inability of hyperglycemia to augment the potentiating effect of arginine above that observed under basal glycemic conditions. Normalization of the plasma glucose profile by phlorizin treatment in diabetic rats completely corrected all three beta cell abnormalities. These results indicate that chronic hyperglycemia can lead to a defect in in vivo insulin secretion which is reversible when normoglycemia is restored.

Animals↗
PubMed1987

Interactions of cholecystokinin and glucose in rat pancreatic islets.

The effects of sulfated cholecystokinin (CCK-8S) and glucose on insulin secretion and polyphosphoinositide (PPI) metabolism were studied in isolated rat islets. Both agonists stimulate PPI hydrolysis, inositol phosphate accumulation, 3H efflux from [3H]inositol-prelabeled tissue, and 45Ca efflux from prelabeled cells. However, the effects of CCK-8S on PPI metabolism are considerably greater than those of glucose. Furthermore, the effects of CCK-8S on PPI and Ca2+ metabolism are observed whether islets are incubated in either 2.75 or 7 mM glucose, but CCK-8S only stimulates insulin secretion (a biphasic response) when the higher glucose concentration is present. Addition of 1 microM forskolin to islets incubated in media containing 2.75 mM glucose does not influence basal insulin secretion but sensitizes the islets to the action of CCK-8S. In the presence of forskolin, CCK-8S induces a very marked first phase but no second phase of insulin secretion. We postulate that CCK-8S acts in this tissue via receptor-linked PPI hydrolysis, leading to an inositol trisphosphate-induced Ca2+ efflux. These receptor-mediated effects of CCK-8S are not altered either by the ambient glucose concentration or the cAMP content of the islets, but these two factors determine the responsiveness of the islets (in terms of insulin secretion) to a given CCK-8S signal.

Animals↗
PubMed1985

Effect of exogenous phospholipase A2 on insulin secretion from perifused rat islets.

Treatment of isolated, perifused rat islets with exogenous PLA2 in amounts ranging from 1 to 1000 mU/ml caused a dose-dependent increase in the rate of insulin secretion. This effect of PLA2 was rapid and seen in the absence of added exogenous fuel. It differed from glucose-induced insulin release in temporal pattern: high concentrations of PLA2 caused a single phase of secretion, and high levels of glucose caused a biphasic pattern of secretion. Like glucose-induced release, PLA2-induced release was partially dependent on extracellular calcium because D600 caused a significant inhibition of release induced by PLA2 at 5 mU/ml. Concentrations of BW755c and NDGA, inhibitors of both the cyclooxygenase and lipoxygenase or only the lipoxygenase pathways of arachidonic acid metabolism, which completely blocked the insulin secretory response to 10 mM glucose, had no effect on the secretory response to 5 mU/ml of PLA2. These inhibitors also inhibited glucose usage by the islets. Finally, although repeated brief exposure of islets to stimulatory concentrations of glucose lead to a progressive increase in the magnitude of both the first and second phases of insulin secretion, repeated brief exposures to PLA2 lead to a progressive decrease in response to each new exposure. Nonetheless, those islets that had been exposed several times to exogenous PLA2, and no longer displayed a response to a further PLA2 exposure, responded normally to the addition of 10 mM glucose. These results indicate that PLA2 is a potent insulin secretagogue, that it shares some of the characteristics of glucose as a secretagogue, but that in many significant ways differs markedly from glucose in its effects on insulin release from isolated islets.

4,5-Dihydro-1-(3-(trifluoromethyl)phenyl)-1H-pyraz↗
PubMed1984

Insulin secretion: combined tolbutamide, forskolin and TPA mimic action of glucose.

We have proposed that the two phases of glucose-induced insulin secretion are regulated by two distinct branches of the calcium messenger system: the initial phase by a calmodulin branch, and the sustained phase by a C-kinase branch. To provide further support for this concept, we examined the separate and combined effects of tolbutamide, TPA, and forskolin upon insulin secretion from rat islets perifused in the absence of added fuels. Addition of 200 microM tolbutamide to the perfusate induces only a first phase of insulin secretion, addition of 200 nM TPA only a second phase, and addition of 10 microM forskolin only a small elevation in the basal rate of secretion. The combination of tolbutamide and TPA induces a biphasic secretory response qualitatively and quantitatively similar to that evoked by an increase in glucose concentration from 2.75 to 7 mM. The combination of TPA, tolbutamide, and forskolin evokes a biphasic pattern of insulin secretion qualitatively and quantitatively similar to that evoked by an increase in glucose concentration from 2.75 to 10 mM.

Animals↗
PubMed1984

Calcium as intracellular messenger: sensitivity modulation, C-kinase pathway, and sustained cellular response.

A model of information flow in the calcium messenger system is presented. When Ca2+ serves as an intracellular messenger in cells exhibiting a sustained response to an extracellular messenger, there are two branches by which information flows from cell surface to cell interior: One operates via amplitude modulation of calmodulin-regulated reactions and the other via sensitivity modulation of the calcium-sensitive, phospholipid-dependent protein kinase, C-kinase. It is postulated that the calmodulin branch is largely responsible for initiating, and the C-kinase branch for sustaining, cellular response in cells displaying sustained responses. The role of the C-kinase branch in the calcium messenger system provides a type of gain control in the process of cell activation. It is but one element of several which provides a considerable degree of plasticity in the operation of this cellular control system. Both cAMP and the biologically active intermediates of arachidonic acid metabolism--prostaglandins, thromboxanes, prostacyclin, and leukotrienes--provide additional inputs into this messenger system and a means of extending its adaptability to specific control needs. In particular, in some cell types it is possible that the cAMP messenger system provides the means of achieving gain control in the calcium messenger system either as an alternative to the C-kinase branch of the calcium messenger system or as a supplemental pathway to it.

Animals↗
PubMed1983

Insulin secretion: combined effects of phorbol ester and A23187.

The effect of the ionophore, A23187, and/or the phorbol ester, 12-O-tetradecanoyl-phorbol-13-acetate (TPA), on insulin secretion were compared with those of glucose. Glucose induces a biphasic pattern of insulin secretion; A23187 a comparable initial spike but no second phase; and TPA a slowly progressive increase. Combined A23187 and TPA evoke a pattern similar to that induced by glucose. Forskolin enhances both phases of glucose- induced and of TPA-A23187-induced insulin secretion. These results are interpreted in terms of a model of cell activation in which two branches of the calcium messenger system, the calmodulin branch and the C-kinase branch, control, respectively, the initial and sustained phases of insulin secretion.

Animals↗
PubMed1983

Increased efficiency of weight gain and altered cellularity of brown adipose tissue in rats with impaired glucose tolerance during diet-induced overfeeding.

We examined the relationship among glucose tolerance, efficiency of weight gain, and cellularity of brown adipose tissue (BAT) in rats (initial weight: 362 +/- 1 g) made hyperphagic and obese by feeding on a highly palatable "cafeteria" (CAF) diet for 4-8 wk. As compared with chow-fed controls, CAF feeding resulted in a 45-60% increase in caloric intake (P less than 0.01), a 40-50% increase in weight gain (P less than 0.01), and hyperinsulinemia. Glucose disposal rate (K) on intravenous glucose tolerance test (IVGTT) was greater than or equal to 1.4% in all chow-fed rats, but fell to less than or equal to 1.3 in 10 of 23 CAF-fed rats. As compared with the chow-fed controls, rats with normal glucose tolerance demonstrated a 12% decline in efficiency of weight gain (g/100 kcal of food consumed) in response to CAF feeding (P less than 0.05). In marked contrast, in rats with impaired glucose tolerance, efficiency of weight gain failed to decline in response to overfeeding and was 18% higher than in the overfed group with normal glucose tolerance (P less than 0.01). Although CAF feeding increased the mass of interscapular BAT by 110-130% in rats with normal as well as impaired glucose tolerance, DNA content of BAT rose only in the normal-K CAF-fed rats (0.19 +/- 0.01 mg DNA/100 mg versus 0.12 +/- 0.02 in chow-fed controls and 0.12 +/- 0.01 in low-K rats).(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue, Brown↗
PubMed1982

Influence of physical training on insulin release and glucose utilization by islet cells and liver glucokinase activity in the rat.

The effect of physical training on insulin release and glucose utilization by perifused islets and on liver glucokinase activity was examined in rats that exercised spontaneously by running (in wheel cage) up to 4-6 mi/day for 36 +/- 4 days and in sedentary controls kept in standard cages. Perifusion of islets with 4 mM glucose resulted in comparable rates of insulin release from islets obtained from trained and sedentary control rats. In contrast, when the perifusion glucose concentration was raised to 10 mM, the biphasic increase in insulin release was 40-50% lower in the trained rats as compared with untrained rats. This decrease in glucose-stimulated insulin release occurred in the face of comparable rates of glucose utilization by islets from control and trained rats. Glucose phosphorylation by liver homogenates from trained rats was reduced at all concentrations of glucose examined (0.5-100 mM). The calculated glucokinase activity was diminished by 40%, whereas hexokinase activity was decreased by 15% in the livers from trained rats. We conclude that 1) hypoinsulinemia induced by exercise training is due to decreased sensitivity of the beta-cell to the stimulant action of glucose independent of changes in islet cell utilization of glucose, and 2) exercise training results in a diminution of liver glucokinase activity that may be a consequence of the hypoinsulinemia.

Animals↗