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Biomedical subjects

G Letko

Publications and source records attributed to G Letko.

15 recordsLinked to original sources
PubMed1992

Influence of ethanol on survival of acinar cells isolated from rat pancreas.

The role of ethanol in precipitating acute pancreatitis has been studied in model experiments. The influence of ethanol on the survival of isolated pancreatic acinar cells (PAC) and a possible coaction with noxious agents (such as trypsin, chymotrypsin, temporary anoxia, and partial uncoupling of the oxidative phosphorylation by 2,4-dinitrophenol [DNP]) has been investigated. Isolated PACs withstood ethanol levels up to 180 mM without significant decrease of viability within 4 h incubation at 37 degrees C. A 90-min deprivation of oxygen was widely tolerated also. However, with a combination of both ethanol application and oxygen deprivation, a clearly forced cell damage was observed as was true with a combination of ethanol and chymotrypsin. The action of DNP or of extracellular trypsin on cell survival was not amplified by the addition of ethanol (180 mM). This study reveals two possible sites of action at which ethanol may contribute to the pathogenesis of acute pancreatitis.

2,4-Dinitrophenol↗
PubMed1992

Influence of trypsin-induced acute pancreatitis on survival and energy state of isolated acinar cells from rat pancreas.

To study the development of acute pancreatitis after intraductal trypsin instillation, at 4 hours, 1, 2, 4 and 6 days after this treatment and after instillation of physiologic saline viable acinar cells were isolated from rat pancreas. Gross anatomic and histologic findings were used to evaluate the time course of pathomorphologic changes. The isolated cells were incubated at 37 degrees C in Eagle's medium in a shaking water bath and the time course of their damage was studied. Additionally, by means of the active accumulation of the fluorophore rhodamine 6 G alterations of the mitochondrial membrane potential, an important parameter of the cellular energy metabolism was evaluated. The most severe histological damage was seen 1 and 2 days after trypsin instillation. At the same time yield and survivability of cells isolated, and their mitochondrial membrane potential reached a minimum. In the controls the time course of these parameters was very similar, but their decrease was less pronounced. Since a direct action of trypsin on acinar cells cannot be responsible for the findings presented a possible involvement of inflammatory cells and their products in the alteration of the cells and of their energy metabolism must be considered.

Acute Disease↗
PubMed1992

Influence of stepwise uncoupling and temporary anoxia on pancreatic enzyme secretion by isolated rat acini.

For special studies on pancreatic diseases a parameter is needed to record alterations of the cellular energy metabolism. In the in vitro model of isolated pancreatic acini, we investigated whether or not at standardized cholecystokinin stimulation the energy-consuming process of enzyme secretion can be used to monitor changes of the energy-supplying capacity. Rat pancreatic acini were isolated via collagenase digestion and characterized by basal and stimulated release of amylase and trypsin, oxygen uptake under resting and maximally uncoupled conditions and by their ability to accumulate actively rhodamine-6G, as a measure of the mitochondrial membrane potential. The stimulation of enzyme release did not find a measurable reflection in rhodamine-6G accumulation and in the respiratory rat. Stepwise uncoupling of oxidative phosphorylation by 2,4-dinitrophenol (DNP) and temporary anoxia were used to simulate disturbances of the pancreatic energy metabolism in vitro. With increasing DNP concentration the enzyme release was significantly reduced. While after 30 min anoxia the enzyme release still exceeded that of unstimulated control, after 60 min anoxia there was no further response to hormonal stimulation. At standardized stimulation and after suitable calibration the enzyme release by acini may be used to monitor alterations of the pancreatic energy metabolism in vitro.

2,4-Dinitrophenol↗
PubMed1992

Effect of phospholipase A2 and ethanol on the survival of acinar cells isolated from the rat pancreas.

The study was undertaken to investigate the effect of phospholipase A2 (PLA2) on the viability of isolated pancreatic acinar cells (PACs) and its possible synergy with ethanol. The survival of PACs may be reduced by PLA2 in a dose-dependent manner. In the presence of 0.25 microgram PLA2.(10(6) cells)-1, all cells died within less than 4 h of incubation at 37 degrees C. A quantity of 180 mM ethanol, which alone was not lethal for PACs, increased the rate of cell death induced by PLA2 at a certain concentration of this enzyme. The present results imply that (1) PLA2 is able to damage PACs in the absence of additional substrates, and (2) at an appropriate activity of PLA2, ethanol may amplify the toxic effects of this lytic enzyme and force autodigestion of the pancreas.

Acute Disease↗
PubMed1991

Involvement of free radicals in the pathophysiology of chronic pancreatitis: potential of treatment with antioxidant and scavenger substances.

The present work reviews the evidence for an involvement of free radicals in the pathophysiology of chronic pancreatitis and the potential of treatment with antioxidant and scavenger substances. Preliminary results indicate that exposure of isolated pancreatic acinar cells to a reaction mixture containing hypoxanthine, xanthine oxidase, and chelated iron causes cell damage and death probably due to generation of superoxide anion and hydrogen peroxide. It still needs to be analyzed which scavengers and antioxidants are able to ameliorate the damage due to oxidant stress in cell models. Such knowledge from cellular studies might help to plan therapeutical trials to evaluate potentially effective antioxidants and scavengers in the experimental animal and in patients with pancreatitis. As yet there are no published studies about the role of free radicals in animal models of chronic pancreatitis. This fact is probably due to the shortcomings of the animal models available. Recent studies presented evidence that activation of oxygen-derived free radicals occurs in patients with chronic pancreatitis. There is also some evidence that the dietary intake of antioxidants may be reduced in patients with chronic pancreatitis. It was suggested that such reduction of antioxidant defenses in the face of an increased demand due to heightened induction of P450 activities may facilitate lipid peroxidation. However, as yet, there is no direct evidence that a reduction of dietary antioxidants with a simultaneous increase in P450 activity is the primary mechanism which initiates chronic pancreatitis without contribution of other factors.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Influence of anoxia, reoxygenation, and uncoupling on survival, respiration, and trypsin-inhibiting capacity of isolated pancreatic acinar cells.

In the pathogenesis of acute pancreatitis, the events and mechanisms increasing the digestibility of the pancreatic acinar cells are widely unknown. Therefore, the possible contribution of a disturbed energy supply (provoked by anoxia or partial uncoupling) to the induction of autodigestion was studied in experiments on acinar cells isolated from the pancreas. During incubation viability, respiration under normal and maximally stimulated conditions, and trypsin-inhibiting capacity (TIC) of these cells were determined. With increasing duration of anoxia, the portion of surviving cells was strongly diminished, and the number of cells with blebs and vesicularly transformed endoplasmic reticulum was increased. Although the endogenous respiration was not influenced up to 1.5 h of anoxia, 30 min of anoxia substantially decreased the capacity of oxidative energy production. The survival curves were characterized by a self-accelerating course of cell destruction. The alteration of the cellular energy metabolism found its reflection in the decreased TIC of the cells.

2,4-Dinitrophenol↗
PubMed1991

Light and electron microscopic studies of lectin binding on the glycocalyx of rat pancreatic cells. I. Normal tissue and isolated cells.

Lectin binding of the glycocalyx of pancreatic tissue sections as well as of isolated pancreatic acini and acinar cells was studied of healthy wistar rats by light and electron microscopy. For light microscopy, we used FITC (WGA, RCAI, LCA) or peroxidase marked (WGA, RCAI, PNA, PHA, LCA, UEAI, LPA) as well as unmarked lectins (Con A, VAA I). Gold marked lectins were used for electron microscopy (WGA, RCAI, LCA, HPA, PNA, VAAI-B). Intact acinar cells in pancreatic tissue sections and isolated acini showed a strong binding of WGA, RACI, and HPA on the apical cell surface, whereas VAAI, UEAI, LCA, and Con A reacted strongly with the basolateral glycocalyx, but not with the apical surface. The 2 main domains of the glycocalyx of pancreatic cells showed their specific lectin binding so long as the junctional complexes between the cells are intact. The polarity of the cell surface of pancreatic acinar cells is discussed in regard to the possible function of the 2 domains.

Animals↗
PubMed1991

Influence of aprotinin and promazine on survival of isolated pancreatic acinar cells.

Aprotinin, a protease inhibitor, and promazine, an inhibitor of phospholipase A2, were tested for possible inhibition of pancreatic acinar cell (PAC) decline induced by uncoupling of oxidative phosphorylation with 2,4-dinitrophenol (DNP) or by temporary anoxia/reoxygenation. In incubates of acinar cells isolated from rat pancreas the presence of aprotinin did not influence the survival of cells treated with these noxae. This finding excludes that extracellulary acting trypsin, possibly released from damaged cells, contributes to further cell death. While promazine at concentrations of 15 to 20 nmol.(10(6) cells)-1 was well tolerated by untreated PAC, higher concentrations caused a clear reduction of cell viability. At optimum concentration promazine was without influence on DNP-treated cells, but it had a beneficial effect on survival and morphology of anoxia-treated PAC (p less than or equal to 0.05). Therefore, it can be assumed that after anoxia/reoxygenation the membrane phospholipase A2 becomes stimulated and causes phospholipid depletion with final death of the cells. It is suggested that such a mechanism may contribute to the initial cell damage in the pathogenesis of acute pancreatitis, too.

Acute Disease↗
PubMed1979

The stimulation of the mitochondrial respiration by citrulline synthesis.

1. The influence of ammonia and ornithine on the oxygen uptake and the formation of citrulline was investigated with isolated rat liver mitochondria. The experiments were performed in a cytosol-like saline medium at 38 degrees C. 2. Under these conditions an increase of the respiration rate by ammonia and ornithine was observed, but a small response to external ADP, only. The missing stimulation by ADP was due to a partial inhibition of the respiratory chain by traces of zinc (approximately 1 microM) present in the medium. This inhibition was only detected at low concentrations of mitochondria. 3. For activation of respiration by ammonia plus ornithine two different processes were responsible: (i) chelation of the inhibiting zinc by ornithine, which could be prevented by EDTA; (ii) ADP production in the matrix space during formation of carbamoyl phosphate, which could be prevented by oligomycin but not by carboxyatractyloside. 4. This stimulus of the carbamoyl phosphate formation and of the equivalent citrulline synthesis on the mitochondrial respiration ran to 12% of that increase caused by phosphorylation of external ADP. The maximum rate of citrulline formation was limited by the activity of carbamoyl phosphate synthetase. 5. Added ADP suppresses the production of citrulline probably by the exchange of extramitochondrial ADP versus intramitochondrial ATP. The data suggest a common adenine nucleotide pool delivering ATP to the adenine nucleotide translocase as well as to the carbamoyl phosphate synthetase.

Animals↗
PubMed1979

Competition between extramitochondrial and intramitochondrial ATP-consuming processes.

The relationship between intra- and extramitochondrial ATP utilization was investigated in liver mitochondria isolated from normally fed, starved and high-protein fed rats. ATP export was provoked by adding a hexokinase-glucose-trap and intramitochondrial ATP consumption by adding ammonia, bicarbonate and ornithine in order to stimulate citrulline synthesis. Both processes compete for ATP produced via oxidative phosphorylation; the rate of citrulline formation declines as the extramitochondrial [ATP]/[ADP] ratio decreases. It is concluded that ATP for adenine nucleotide translocation and that for carbamoyl phosphate synthesis are delivered from a common intramitochondrial pool of adenine nucleotides. In mitochondria from rats with a high-protein diet, citrulline synthesis greatly stimulates the rate of oxidative phosphorylation (about two thirds of state 3 respiration). Under these conditions the intramitochondrial [ATP]/[ADP] ratio is significantly reduced. The intramitochondrial [ATP]/[ADP] ratio is not in thermodynamic equilibrium with the extramitochondrial one.

Adenine Nucleotides↗
PubMed1978

Influence of Mg2+-ions on the properties of rat heart mitochondria in dependence on the preparation.

1. The preparation of rat heart mitochondria with Potter-Elvehjem homogenizer results in mitochondria showing stimualtion of respiration induced by Mg2+. This stimualtion is neither caused by adherent hexokinase nor by energy-dependent magnesium accumulation (Mg2+ content in the presence of 10 mM glutamate: 22 nmoles/mg protein; in the presence of glutamate plus antimycin A 21 nmoles/mg protein). 2. The effect of added magnesium is excluded by addition of carboxyatractyloside. This demonstrates the activity of an ATPase outside of the mitochondrial inner membrane. 3. A simple and rapid method for the preparation of Mg2+-insensitive rat heart mitochondria is presented. The minced heart is pressed through a normal syringe and then treated with trypsin. 4. A comparison of mitochondria of both preparations shows that there is no difference in magnesium content and no energy-dependent magnesium influx.

Adenylate Kinase↗
PubMed1975

Further characterization of the association of glyceraldehyde-3-phosphate dehydrogenase with reticulocyte membranes.

1. The behaviour and properties of membrane-bound GAPDH of rabbit reticulocytes were investigated. 2. The bound GAPDH is more resistant to inactivation by KCl than the soluble enzyme (allotopy). 3. The bound enzyme is released by electrolytes. This effect does not only depend on the ionic strength but additionally on the kind of ions, pH-value and protein concentration. 4. A comparison of the releasing effect of NAD analogues shows the necessity of the 5'-AMP moiety in the structure of the effector. 5. The represented results demonstrate the specifity of the GAPDH-membrane binding in rabbit reticulocytes.

Animals↗
PubMedDate not supplied

Enzymatic and histologic investigations into the course of pancreatic alterations induced by anti-acinar-cell-antiserum.

A model of acute pancreatitis (AP) was developed by application of anti-acinar-cell-antiserum in rats. Within 24 h postoperatively the binding of antibodies to the pancreas, histological findings, and the activities of lipase and a-amylase in serum and pancreas were analyzed. While after intraaortic administration morphological alterations could not be observed, after single dose intraductal injection, typical macroscopical and histological signs of AP were found. This pancreatic injury was characterized by a mild and protracted course favoring investigations of the early phase of pathogenesis. As soon as 2 h after intraductal application of antiserum, serum amylase and lipase were already increased and microscopic examination of the pancreatic tissue revealed pancreatic edema, inflammatory infiltration and parenchymal necrosis. Intra- and extrapancreatic fat necrosis occurred at 16 h post injectionem. The results suggest that parenchymal damage plays an essential role in the manifestation of this experimental AP and that fat necrosis seems to be a secondary event.

Acute Disease↗