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Biomedical subjects

G Letko

Publications and source records attributed to G Letko.

At least 37 records · Page 2Linked to original sources
PubMed1989

Effects of trypsin, chymotrypsin, and uncoupling on survival of isolated acinar cells from rat pancreas.

To improve the knowledge of the very complex pathogenesis of acute pancreatitis, separated exocrine pancreatic cells were used as an experimental tool. Intact acinar cells were isolated from rat pancreas and the time course of their damage was studied in the presence of increasing activities of trypsin, chymotrypsin and, after, stepwise uncoupling of oxidative phosphorylation, by 2,4-dinitrophenol (DNP). With an increasing degree of uncoupling the half-life period of cells incubated was remarkably reduced. Extracellular trypsin, present in activities comparable with the endogenous trypsinogen content, was not effective in cell killing. Only when the trypsin activity was drastically enhanced was the rate of cell destruction accelerated. Compared with chymotrypsin, isolated acinar cells revealed some resistance to extracellular trypsin. But when the energy production was altered by partial uncoupling the susceptibility of these cells toward the action of trypsin was increased and their life time was remarkably reduced. The potentiation of the effects of trypsin and uncoupling on cell viability demonstrates the contribution of an intact energy metabolism to the protective potential of acinar cells against further noxae.

2,4-Dinitrophenol↗
PubMed1989

Experimental acute pancreatitis--a quantification of dynamics at enzymic and histomorphologic levels.

To study the dynamics of pathomorphologic alterations in the development of acute pancreatitis (AP) and the corresponding changes of the patterns of pancreatic enzymes in rats AP was induced by: 1) combination of a pancreatic juice edema and temporary pancreatic ischemia, ii) by intraductal instillation of trypsin, and iii) by trypsin instillation in combination with ischemia. At 4, 8 and 24 h postoperatively the histologic findings and the activities of lipase and alpha-amylase in the pancreas and the serum were analyzed. The histologic sum score of the individual rats did not correlate with their enzymic patterns in pancreas and in serum. In all three models there was a development of parenchymal necrosis independent of the existence of pancreatic fat necrosis. Therefore, it is not probable that fat necrosis represents an obligatory precondition for the initiation of autodigestion.

Acute Disease↗
PubMed1989

Effect of temporary ischemia upon development and histological patterns of acute pancreatitis in the rat.

To study the importance of a disturbed energy metabolism for the development of acute pancreatitis (AP) rats pretreated either by induction of a juice edema or by intraductal trypsin instillation were subjected to temporary pancreatic ischemia. By means of a scoring system quality and quantity of pathomorphologic parameters were quantified 24 h postoperatively. There was a clear correlation between macroscopic and histologic scores independent of the model used. While a juice edema or 40 min ischemia alone did not induce AP, a combination of both led in half of the animals treated to AP. This was mainly characterized by extrapancreatic fat necrosis. Besides less specific fat necrosis the histologic examination of the pancreas revealed acinar necroses at the periphery of the lobules as a frequent injury pattern. After trypsin injection a persistent pancreatic edema, hemorrhages, fat and parenchymal necroses were typical findings. Both focal centro-lobular and extended sublobular or lobular necroses were histologically observed. An additional temporary ischemia augmented significantly the severity of findings, however, their quality was not essentially changed. From the present results it can be concluded that an alteration of the pancreatic energy metabolism, e.g. by hypoperfusion or ischemia, may be an important pathogenic factor in precipitating experimental AP.

Acute Disease↗
PubMed1989

Pancreatic exocrine secretion--a measure of the functional capacity of the pancreatic energy metabolism.

In studies on the pathogenesis of ischemic cell injury and of other pancreatic diseases the knowledge of the actual pancreatic energy state is an important factor. Therefore, it would be advantageous to have a simple and inexpensive method to determine this parameter and its alterations in the pancreas. At uniform hormonal stimulation, the extent of exocrine pancreatic secretion showed a clear dependence on energy supply in this organ. The degree of pancreatic juice edema formed after bolus injection of cholecystokinin and secretin at ductal occlusion was found to be the most sensitive and reproducible measure of the functional capacity of the pancreatic energy metabolism. While this parameter can be applied to experimental studies, only, the juice volume secreted could be determined under clinical conditions, too. Both parameters of pancreatic secretion were clearly decreased after preceding short-term ischemia and recovered after an adequate interval of reperfusion.

Animals↗
PubMed1989

Isolated acinar cells from rat pancreas in pathogenic studies on acute pancreatitis.

In the pathogenesis of acute pancreatitis the events and mechanisms increasing the digestibility of the acinar cells are widely unknown. To study these processes at cellular level, isolated exocrine cells from rat pancreas were utilized as experimental tool. The investigations revealed that extracellularly acting noxae seem to be of less importance to triggering autodigestion. The plasma membrane of the acinar cells was relatively resistant to extracellular trypsin, while anoxia or uncoupling of oxidative phosphorylation obviously reduced the survival of these cells. Combined action of small amounts of trypsin and partial uncoupling resulted in a potentiated cell destruction. The findings presented stress the importance of a well-functioning energy metabolism for the protective potential of acinar cells to further harmful conditions.

Acute Disease↗
PubMed1988

Effects of pancreatic acinar cell surface antibodies and complement on isolated rat acinar cells in vitro.

Surface directed pancreatic acinar cell antibodies raised by immunization of rabbits with suspensions of viable isolated rat acinar cells were utilized to study immune cytolytic processes as a model of in vitro pancreatic injury. The antibodies produced were bound to rat pancreatic acinar cell surface determinants and significantly damaged freshly separated acinar cells by immune cytolytic mechanisms. Addition of complement accelerated the cytolytic effects on the target cells in a dose-dependent manner. The decline of acinar cells was dependent only on the presence of the immune cytolytic potential and not on the number of already damaged cells. Morphologic changes in the cells induced by the agents applied were revealed by both transmission and scanning electron microscopy. The presented experimental model seems a valuable tool for further investigations at the cellular level into the contribution of primarily occurring acinar cell injury in triggering the subsequent pathophysiological mechanisms initiating autodigestion of the pancreatic gland in the pathogenesis of acute pancreatitis.

Animals↗
PubMed1988

An optimized procedure for isolation of rat pancreatic acinar cells.

A simple and inexpensive procedure is described which allows reproducibly the isolation of rat pancreatic acinar cells. Using only small quantities of commercially available collagenase without addition of any further protease, a cell population consisting of about 95% of acinar cells can be obtained within about 95 min. Cell yield is 40% as calculated on a dry weight basis. Enzyme activities measured within final suspensions of isolated cells are: amylase, 1.17 +/- 0.27 amylase units.(mg d.w.)-1; lipase, 23.78 +/- 6.02 nkat.(mg d.w.)-1 and alanine aminotransferase, 0.895 +/- 0.236 nkat.(mg d.w.)-1. Isolated cells are morphologically intact as seen by electron microscopy and retain their viability for more than 3 h, even when incubated at 37 degrees C without any substrate and protease inhibitor, as revealed by their ability to exclude trypen blue. Therefore, acinar cells isolated in this manner may prove useful for investigations at cellular level into pathogenetic mechanisms underlying pancreatic diseases.

Alanine Transaminase↗
PubMed1987

Mitochondrial membrane potential in living cells: evidence from studies with rhodamine 6 G as fluorescent probe.

Rhodamine 6 G as a cationic permeant fluorophore is demonstrated to be selectively accumulated by mitochondria of living pancreatic acinar cells from guinea pigs. The accumulation of the fluorescent dye was studied by means of the application of electron transport inhibitors in the respiratory chain, ionophores and some hydrogen donors. Besides, some unspecific staining processes were excluded. Using this technique, it becomes possible to estimate the energy state of living cells under various conditions of energy supply and demand.

Animals↗
PubMed1987

Influence of pancreatic edema and short-term ischemia of rat pancreas on lipase- and alpha-amylase-activities in the serum and in the pancreas.

Acute pancreatitis was produced in rats by a combination of dyschylic edema and short-term ischemia of the pancreas. The incidence of acute pancreatitis, reflected by peri- and extra-pancreatic fat necrosis, was found to be dependent on the duration of ischemia. Under most conditions there were no significant differences in enzyme activities between the animals with macroscopic signs and those without these. The time course of alpha-amylase and lipase release was investigated within 24 hours postoperatively. Additionally the quantitative correlation was estimated of the enzyme activities released from the pancreatic tissue and the activities determined in rat serum. The data show that the serum enzymes represent only a small and variable portion of the total amount of enzymes effused from the pancreas.

Acute Disease↗
PubMed1987

Influence of antisera to pancreatic acinar cells on rat pancreas in situ.

For investigations of cell injury during pathogenesis of acute pancreatitis antisera to pancreatic acinar cells were used as experimental tool. Within one hour after intraductal injection of antiserum a strong pancreatic edema was developed. Within 24 h this edema receded to a large extent but at this time there were inflammatory cells scattered in the intra- and periductal region. As a sequel of application of antiserum pancreatic enzymes were released. At 24 h after this application serum activities of alpha-amylase and lipase were significantly increased in comparison to the control and reached that level which was found in a model of acute pancreatitis provoked by pancreatic edema plus short-term ischemia.

Acute Disease↗
PubMed1986

Morphological and biochemical characterization of pancreatic acinar cells from guinea pig.

For elucidation of pathogenesis of acute pancreatitis an evaluation of the "intactness of cellular metabolism" of the acinar cells is an important question. To this end single acinar cells were isolated from guinea pig pancreata and studied at morphological and biochemical levels. The presented procedure generally results in suspensions with more than 85 percent intact exocrine cells which maintain their typical arrangement of the zymogen granules in the apical region during isolation procedure. The cellular content of amylase, lipase, and ALAT was determined. The measurement of parameters of cellular energy metabolism was regarded as essential for determination of functional intactness of the pancreocytes. The oxygraphically measured span between unstimulated respiration and that after maximal stimulation by uncoupling was used to characterize the load capacity of cellular energy metabolism.

Amylases↗
PubMed1986

[Vital fluorochrome staining of isolated pancreatic acinar cells for the characterization of cell-structural changes].

Rhodamine 6 G as a cationic fluorophore is demonstrated to be selectively accumulated by mitochondria of living pancreatic acinar cells (cell isolation see Spormann et al. [1986]. The accumulation of rhodamine was studied under using of electron transport inhibitors, ionophores and some hydrogen donors. The application of DNP as wellknown protonophore resulted in a rapid dissipation of any fluorescent signals, whereas application of sodium succinate, hyperosmolaric exhibited a remarkable increase of fluorescence intensity. Using this technique it is possible to estimate the energy state of living cells under various conditions of energy supply and demand.

Animals↗
PubMed1986

Contribution of pancreatic edema and short-term ischemia to experimental acute pancreatitis in the rat. II. Behaviour of serum parameters.

The biochemical characterization of experimental acute pancreatitis was performed by determination of the secretory enzymes lipase and alpha-amylase, of the cytoplasmic ALAT (alanine aminotransferase), of total protein and calcium concentration in serum of rats. The moderate and protracted course of the pathological process in the small animal model presented allowed to study the initial phase from 1-24 h. In the first 4-8 h most massive enzyme release into the intravasal space was observed. The level of enzyme activities was correlating with the severity of assault. One noxa alone (ischemia or juice edema) resulted in a moderate enzyme release (lipase : 2-2.5 fold of control). The action of both noxae caused a drastical increase in enzyme activities in the initial phase lipase : 8-20 fold, ALAT: 7 fold, alpha-Amylase: 2.5 fold). 24 h postoperatively the serum enzyme activities were at distinct pathological level. At this time acute pancreatitis provoked already a decreased serum protein content. A hypocalcemia was not observed.

Acute Disease↗
PubMed1986

Production and characterization of antibodies to rat pancreatic acinar cells.

The contribution of immunologic noxae to the damage of the cells and the induction of autodigestion in the course of acute pancreatitis in rats should be regarded. The isolation of intact acinar cells from rat pancreas offers a new approach to a more specific antiserum being directed against antigens at the outside of the cell membrane. Proof of circulating antibodies was performed using the indirect immunofluorescence reaction method (IFAR). A defined membrane fluorescence was observed at the surface of the acinar cells in vitro and a fluorescence at the septa of lobules marking the borders of acini in vivo.

Acute Disease↗
PubMed1986

Effect of improved hydrogen supply on energy state, ureogenesis and gluconeogenesis in isolated hepatocytes.

The cellular energy state as well as ureogenesis and gluconeogenesis as ATP-consuming processes were studied as a function of hydrogen supply of mitochondria in isolated hepatocytes. Even though the ATP turnover was quite different during ureogenesis and gluconeogenesis and in the absence of these syntheses both the safranine signals (as a probe for the mitochondrial membrane potential) and the total ATP contents of the cells did not vary to a great extent. Partial uncoupling by FCCP destroyed the mitochondrial membrane potential and decreased the cellular ATP totally. Under these conditions the presence of ornithine plus lactate maintained a measurable membrane potential and ATP level. In order to demonstrate the relevance of hydrogen supply of mitochondria to the syntheses of glucose and urea, the load on energy metabolism was additionally increased by partial uncoupling, and the cells were incubated in a parallel mode in Krebs-Henseleit buffer and an 80 mM succinate containing medium, the latter forcing succinate into the cells. After partial uncoupling, ureogenesis and gluconeogenesis as well as the ATP level were seen to decrease markedly in Krebs-Henseleit buffer, whereas in the succinate medium the improved hydrogen supply of mitochondria counteracted the drain by FCCP, and substantially higher rates of the syntheses and elevated ATP levels were maintained.

Adenosine Diphosphate↗
PubMed1986

Contribution of pancreatic edema and short-term ischemia to experimental acute pancreatitis in the rat. I. Procedure and pathomorphological investigations.

For experimental analysis of pathogenesis of acute pancreatitis a suitable animal model is necessary. The pancreatic (juice) edema and an alteration of cellular energy metabolism are important pathogenetic factors. A partial intrapancreatic edema was induced by stimulating secretion against permanent or temporary obstruction by ductal ligation. Short-term ischemia was used as tool for alteration of cellular energy metabolism. 24 h postoperatively morphological changes were macro- and microscopically investigated. The data show a correlation between duration of ischemia (10-30 min) and the frequency of acute pancreatitis at preexisting juice edema (44-68 percent). With increasing ischemia the morphological alterations (e.g.: necroses of adipose tissue) were enhanced, too. A hemorrhagic necrotizing pancreatitis was rarely observed. In comparison to other experimental models the presented one is characterized by a mostly moderate and light course of acute pancreatitis offering a good tool for elucidation of the very first stages of the pathogenesis.

Acute Disease↗
PubMed1986

Acute pancreatitis--a quantification of dynamics at clinical and radiomorphologic levels.

To quantify the dynamics of acute pancreatitis in a prospective clinical study on 108 patients, clinical and computed tomographic classifications were performed at days 1, 3 and 6 after admission. Sixty-two percent of the patients revealed a variable clinical staging at the times considered. Progredience of pancreatic necrosis was found in 20 of the 55 patients subjected to radiomorphologic investigations. Comparison between clinical and CT findings showed an increase in the agreement of both assessments from 69 percent (day 1) to 85 percent (day 6). Despite this high-level agreement in some patients both findings diverged remarkably. Acute pancreatitis is discussed as an interplay of necrobiosis and the "compensatory capacity" of the organism, with clinical findings acting as an indicator of the overall state of this struggle.

Acute Disease↗