Michael C. Geokas

Methemalbumin in the Diagnosis of Acute Hemorrhagic Pancreatitis

Abstract The early differentiation between acute edematous and hemorrhagic pancreatitis is crucial for prognosis and treatment. By testing serum and ascites or pleural effusion for methemalbumin th...

Ethanol, the Liver, and the Gastrointestinal Tract

Ethanol is easily absorbed from the intestine and diffuses quickly throughout body water. The bulk of ethanol is metabolized in the liver, where alcohol dehydrogenase, a complex mixture of isoenzymes, oxidizes ethanol to acetaldehyde. Ethanol abuse produces functional and structural changes in the gastrointestinal tract, such as in the stomach, small intestine, liver, and pancreas. Accumulating evidence suggests direct toxicity of ethanol and possibly of acetaldehyde. Fatty liver, alcoholic hepatitis, liver cirrhosis, acute and chronic gastritis, deranged structure and function of the small intestine, acute and chronic pancreatitis, and pancreatic lithiasis are some of the sequelae of ethanol abuse. Recent investigations have enhanced our understanding of the functional and structural changes of the gastrointestinal tract produced by the abuse of ethanol.

Effect of ethanol on cholecystokinin-induced enzyme secretion from isolated rat pancreatic acini ☆

Cholecystokinin octapeptide (CCK8)-stimulated amylase release in isolated rat pancreatic acini was inhibited over 30% by 600 mM ethanol. The configuration of the dose-response curve for CCK8, however, in the presence of ethanol was similar to that of the control. Amylase release elicited by maximal concentrations of CCK8 (300 pM) was inhibited by increasing concentrations of ethanol (0.3 to 1.3 M), and this inhibition was concentration dependent. In addition, the binding of [125I]CCK33 to specific membrane receptors on acini was inhibited by ethanol in a dose-dependent manner. A positive correlation between the inhibitory effects of ethanol on CCK binding and CCK-induced amylase release was observed. Furthermore, these inhibitory effects of ethanol were reversible. Basal amylase release, however, was increased 20-50% by ethanol between the concentrations of 0.3 and 1.3 M; higher concentrations caused a leakage of amylase from the acini both in the absence and presence of 300 pM CCK8. This is confirmed by 51Cr release from prelabeled acini which revealed no significant damage to acinar cell membrane between 0.3 and 1.6 M ethanol, but significant damage to acini at higher concentrations. These data suggest that the 600 mM ethanol-induced inhibition of CCK action in acini is due to reversible perturbation of the acinar cell membrane.

Proinsulin conversion to desalanyl insulin by alpha2-macroglobin-bound trypsin.

HARPEL and Mosesson1 observed that α2-macroglobulin-bound trypsin hydrolysed low molecular weight ester and amide substrates at rates comparable with those of free protease, but degradation of high molecular weight proteins was markedly inhibited. To account for this finding, Barrett and Starkey2 introduced the novel concept of steric entrapment of endopeptidases by α2-macroglobulin. Neurath and Walsh3 have recently reviewed the role of proteolytic enzymes in the conversion of a variety of enzymes, hormones, and other physiologically active proteins from inactive precursors to active forms by limited proteolysis. Many of these physiologically active proteins exist in the bloodstream in both precursor and active forms. We report here that proinsulin, a large polypeptide of 9,000 molecular weight, is rapidly hydrolysed by α2-macroglobulin-bound trypsin. This finding suggests that pro-teases bound to α2-macroglobulin may be involved in limited proteolytic cleavage associated with activation of precursor molecules as well as the degradation of biologically active polypeptides in the circulation.

Altered exocrine pancreatic function in rats treated with nicotine

The present study investigates the effects of nicotine treatment on exocrine pancreatic function. Adult male, Sprague-Dawley rats received nicotine via a time-release pellet, at a rate of 1.65 micrograms/min for 3 weeks. At the end of the experimental period, it was observed that although nicotine did not affect final body or pancreatic weight, the activities of amylase, trypsin, and chymotrypsin in pancreatic homogenates from nicotine-treated rats were 51, 29, and 35% higher, respectively, than in controls. Levels of immunoreactive cationic trypsin(ogen) were significantly higher in pancreatic homogenates and serum from nicotine-treated rats as compared with controls. In addition, concentrations of mRNA, encoding for pancreatic amylase, were higher in pancreatic homogenates from the nicotine-treated rats than in controls. In dispersed pancreatic acini isolated from nicotine-treated rats, basal secretion of amylase, trypsinogen, and chymotrypsinogen was 50% higher than controls and enzyme release following CCK-8 (100 pM), secretin (1 microM), and carbachol (7.5 microM) stimulation was also significantly higher. These data indicate that nicotine treatment, at levels comparable to those expected in moderate cigarette smokers, increases the content of digestive enzymes in rat pancreas, as well as their basal and secretagogue-induced release.

[17] Radioimmunoassay determination of circulating pancreatic endopeptidases

Publisher Summary This chapter describes the radioimmunoassay determination of circulating pancreatic endopeptidases. The radioimmunoassay technique has been applied to the detection of pancreatic endopeptidases in plasma and serum. These studies have demonstrated that, while the bulk of the digestive enzymes enter the pancreatic ductal system, a small fraction enters the bloodstream. The application of the radioimmunoassay methodology has permitted the characterization of the circulating molecular forms of human cationic trypsin, anionic trypsin, elastase 2, and chymotrypsin. The ability to quantitate the levels of total immunoreactive trypsin in biological fluids as well as to identify the molecular forms of trypsin present in the sample has resulted in several reports of clinical applications for the radioimmunoassay of cationic trypsin. The lack of cross-reactivity in the radioimmunoassay of a given pancreatic endopeptidase, with the same protease, from other mammalian species, demonstrates that a separate radioimmunoassay must be developed for each enzyme to be investigated. The lack of cross-reactivity between the various pancreatic endopeptidases of the same species permits the specific identification of each protease in a biological sample.

Ethanol and the Pancreas

The acute and chronic effects of ethanol on pancreatic structure and function are discussed. Acute necrotizing, acute edematous, acute relapsing, chronic relapsing, and painless pancreatitis have an established association with ethanol abuse. The management of these disorders is outlined.

Plasma immunoreactive pancreatic cationic trypsinogen in cystic fibrosis: a sensitive indicator of exocrine pancreatic dysfunction.

Summary: Plasma immunoreactive cationic trypsin(ogen) levels were determined in 32 control subjects and 43 patients with varying degrees of pancreatic insufficiency including 35 with cystic fibrosis (CF) and eight with Shwachmans syndrome. In six CF infants less than 2 years of age, plasma trypsin(ogen) levels were significantly elevated (97.3 ± 62.2 ng/ml) above the normal range for nine controls (7.0 ± 5.9 ng/ml; P < 0.025). Four of these infants had steatorrhea, three of whom had undetectable duodenal trypsin activity after stimulation with secretin-cholecystokinin. In two CF infants, molecular size fractionation by gel filtration of plasma followed by radioimmunoassay of the column fractions demonstrated that trypsinogen was the only immunoreactive species in the circulation.In contrast, in older CF patients with steatorrhea (mean age, 15.3 ± 4.6 years), plasma cationic trypsin(ogen) levels were undetectable or low (1.1 ± 1.7 ng/ml). This finding clearly distinguished them from older CF patients without steatorrhea (mean age, 14.3 ± 3.9 years) in whom cationic trypsin(ogen) levels were significantly higher (23.3 ± 17.6 ng/ml; P < 0.01). The mean trypsin(ogen) concentration in the older CF patients without steatorrhea did not differ from the mean value for 23 normal subjects of similar age. Plasma cationic trypsin(ogen) levels in two Shwachmans patients with steatorrhea (0.19 and 0.86 ng/ml) were significantly lower than the values found in six Shwachmans patients without steatorrhea (5.9 ± 2.3 ng/ml; P < 0.025). Furthermore, in nine older CF patients and eight Shwachmans patients, circulating trypsin(ogen) levels were highly correlated with duodenal trypsin output after secretin-cholecystokinin stimulation (r = 0.946, P < 0.01; r = 0.899, P < 0.01, respectively). These results suggest that in CF infants high levels of circulating trypsin(ogen) persist even in those with complete pancreatic insufficiency. In older CF patients and those with Shwachmans syndrome, however, circulating trypsin(ogen) accurately reflects residual pancreatic function.Speculation: Cystic fibrosis (CF) infants often possess viable but ductally obstructed pancreatic tissue, which may be destroyed with disease progression. The correlation between pancreatic exocrine function and circulating trypsin(ogen) in older CF patients and Shwachmans patients, however, indicates that ductal obstruction is not a prominent feature in these patients.

Dynamic changes of pancreatic structure and function in rats treated chronically with nicotine

Adult, male Sprague-Dawley rats weighing initially 185-225 g, were treated with 5, 15, or 50 mg nicotine or placebo 3-week-release pellets by sc implantation, for 1.5, 3, 6 and 12 weeks. These doses of nicotine correspond to infusion rates of 9.9, 29.8, and 99.2 micrograms/h, respectively. At the highest nicotine dose trypsin and chymotrypsin activities were markedly higher in pancreas from 12-week nicotine-treated rats compared with controls. This was associated with a fourfold increase in steady-state amylase mRNA levels in comparison to placebo controls. In addition, secretagogue-stimulated enzyme release from pancreatic acini isolated from rats treated with 50 mg nicotine pellets was significantly higher than controls at 1.5 and 3 weeks and declined below control levels after 12 weeks of treatment. In rats treated with 15-mg nicotine pellets, maximal secretagogue-stimulated enzyme release from isolated acini occurred at 1.5 weeks, declining thereafter to control levels. Electron microscopy of pancreas from rats treated with the 50 mg nicotine dose revealed intracytoplasmic vaculoes appearing after 3 weeks of treatment, and persisting throughout the remaining experimental period. It is concluded that 12-week nicotine treatment results in increased pancreatic enzyme biosynthesis and accumulation of digestive enzymes within the pancreas. This is associated with altered responsiveness to secretagogues and evidence of morphological damage.

Proteolysis of parathyroid hormone in vitro by sera from acute pancreatitis patients.

Abstract The degradation of 125I-labeled parathyroid hormone in vitro has been investigated in order to test the hypothesis that proteolytic degradation of PTH may be related to the hypocalcemia associated with acute pancreatitis. A polyacrylamide gel electrophoresis technique has been employed to separate bovine PTH (1-84) from lower molecular weight peptides produced by proteolytic cleavage. It has been demonstrated that trypsin bound to the plasma protease inhibitor α2-macroglobulin rapidly degrades PTH and destroys its biological activity in vitro in a kidney cortex adenyl cyclase assay system. Substantial PTH degrading activity was also detected in serum samples obtained from nine patients with severe acute pancreatitis. In contrast, significant PTH degradation could not be detected in normal serum or plasma. Furthermore, no degradation was observed with sera from two patients with mild pancreatitis due to biliary tract stones or four patients with hyperamylasemia but no evidence of pancreatic disease. The PTH degrading activity in patient sera could be blocked by the combination of two peptide chloromethylketones that specifically inhibit the pancreatic endopeptidases trypsin, chymotrypsin, and elastase 2. These results suggest that the FTH degradation assay represents a sensitive test for biologically relevant proteolytic activity in acute pancreatitis, and that it can specifically detect molecular forms of the pancreatic proteases in blood reflecting the intrapancreatic activation of zymogens in the course of pancreatic inflammation.