J. van Gool

Histophotometric estimation of volume density of collagen as an indication of fibrosis in rat liver.

SummaryThe development of fibrosis in the liver of 16 rats treated for 1, 2, 3 or 4 weeks with CCl4, has been followed with chemical hydroxyproline determination and histophotometric analysis of histological sections stained with Sirius Red F3BA in saturated aqueous picric acid. The readings were taken with a scanning and integrating microphotometer and corrected for picric acid absorbance as a measure for mean protein mass per unit area of the section. It appearts that the integrated absorbance readings of Sirius Red absorbing material in the section show a highly significant correlation with the hydroxyproline determinations. It is concluded that picrosirius photometry can be used to give a measure of the volume density of collagen in sections. An advantage of the photometric assay is that measurements are taken on the basis of the microscopic image, so that it is also possible to estimate collagen density in a selected area, e.g. a tumour formation amidst normal tissue, or to exclude necrotic areas.

Effects of monocytic products, recombinant interleukin-1, and recombinant interleukin-6 on glycosylation of alpha 1-acid glycoprotein: studies with primary human hepatocyte cultures and rats.

Changes in the carbohydrate moieties of acute-phase glycoproteins (APGPs) often accompany the increase in their secretion by the liver during inflammation. In this study, we investigated whether factors known to regulate APGP gene expression are also involved in the altered glycosylation. For this purpose, the glycosylation pattern of alpha 1-acid glycoprotein (AGP) as secreted by human hepatocytes, cultured in the presence and absence of dexamethasone and monokines, was studied by crossed affino- (concanavalin A) immunoelectrophoresis (CAIE). The monokines rIL-1 and rIL-6, in the presence of dexamethasone, both stimulated AGP secretion and caused a change in glycosylation towards an increased Con A reactivity, including the appearance of two strongly reactive forms (D and E) normally not present. Dexamethasone alone did not influence either process. When tested in vivo in rats, rIL-6 also induced an increased presence of Con A-reactive forms of AGP in serum. In conclusion, the changes in secretion and glycosylation of AGP as seen during inflammation seem to be mediated by the same factor(s).Changes in the carbohydrate moieties of acute-phase glycoproteins (APGPs) often accompany the increase in their secretion by the liver during inflammation. In this study, we investigated whether factors known to regulate APGP gene expression are also involved in the altered glycosylation. For this purpose, the glycosylation pattern ofα1,-acid glycoprotein (AGP) as secreted by human hepatocytes, cultured in the presence and absence of dexamethasone and monokines, was studied by crossed affino-(concanavalin A) immunoelectrophoresis (CAIE). The monokines rIL-1 and rIL-6, in the presence of dexamethasone, both stimulated AGP secretion and caused a change in glycosylation towards an increased Con A reactivity, including the appearance of two strongly reactive forms (D and E) normally not present. Dexamethasone alone did not influence either process. When tested in vivo in rats, rIL-6 also induced an increased presence of Con A-reactive forms of AGP in serum. In conclusion, the changes in secretion and glycosylation of AGP as seen during inflammation seem to be mediated by the same factor(s).

A method for measurement of liver iron fractions in needle biopsy specimens and some results in acute liver disease.

Methods are described for measurement of total tissue iron, ferritin iron, haem iron and ferritin protein in approx. 15 mg of tissue obtained by liver biopsy. The validity of these methods is examined by comparison with the values observed in larger samples of the same post-mortem derived liver tissue. Correlation coefficients vary between 0.80 and 0.99 (n = 11--16). It appears that in post-mortem liver tissue the haem iron concentration is higher than in biopsy specimens from patients. Analysis of liver biopsy specimens from patients with hepatitis showed a large variation in the mean iron content of the liver ferritin molecules. Also, the non-ferritin depot iron concentration and ferritin protein concentration is quite variable. It is suggested that in cases of advanced ferritin catabolism during hepatitis the mean percentage of iron in ferritin molecules often increases while at the same time the non-ferritin depot iron fraction decreases, probably because of iron release from the liver.

Inhibitory effects of rat α2 macrofetoprotein (αMFP), an acute phase globulin, on galactosamine hepatitis

Abstract Refractoriness to Gal N toxicity occurs especially in fetal rats, newborn rats, and in rats after partial hepatectomy. An injury however (laparotomy, incision on the back or ip BaSO 4 suspension), prior to Gal N administration, also inhibits Gal N toxicity. In all these circumstances high levels of rat α 2 -macrofetoprotein ( α M FP) occur. This protein is an acute phase reactant and is identical to rat α 2 -macroglobulin. α M FP isolated from the serum of injured rats and then administered to normal rats strongly inhibits Gal N toxicity. When time interval between the preceding injury, provoking α M FP production and Gal N administration shortens, the inhibiting effects are less and α M FP production remains low. During resistance to Gal N, the primary and secondary biochemical lesions of Gal N persist and the protecting effect of α M FP must be due to another mechanism, operating in later phases of cell injury. Very probably this is attributable to the stabilizing effect on membranes of hepatocytic organelles and the plasma membranes. As α M FP is an acute phase reactant the importance of these proteins to the course of hepatitis must be considered.

Pathways of enzyme transfer in sodium taurocholate-induced acute hemorrhagic pancreatitis

The pathways of enzyme transfer from the pancreas into the systemic circulation were analyzed in sodium taurocholate-induced acute hemorrhagic pancreatitis in the rat by estimating lipase concentrations in blood, lymph and ascites. During the first few hours of pancreatitis high enzyme levels were observed in thoracic duct lymph. However, cannulation of the thoracic duct did not prevent a significant increase in the lipase concentration in peripheral blood. The portal vein lipase concentration was found to exceed the peripheral values by approximately 10%. Extremely high concentrations of lipase were measured in the ascites collected during pancreatitis. When the ascites was transferred to the peritoneal space of healthy rats, a significant increase in the lipase concentration in peripheral blood was measured. This increase could not be prevented by transection of the parasternal lymphatics. It was concluded that the hematogenous rather than the lymphatic transport of lipase from the pancreas and the peritoneal surface is the most important pathway. In this respect, this study does not support thoracic duct drainage but advocates peritoneal lavage as a logical therapeutic measure to reduce the concentration of circulating toxic substances from the pancreas in acute pancreatitis.

PROGRESSIVE MULTIFOCAL LEUCOENCEPHALOPATHY ASSOCIATED WITH MULTIPLE MYELOMA.

SummaryA case of progressive multifocal leucoencephalopathy is described associated with multiple myeloma (Kahlers disease).RésuméUn cas de leucoencéphalopathie progressive multifocale associée avec de myélome multiple (maladie de Kahler) est rapporté.

Mechanisms by which acute phase proteins enhance development of liver fibrosis: Effects on collagenase and prolyl-4-hydroxylase activity in the rat liver

Previous experiments showed that the presence of high levels of acute phase reactants (APR) enhance CCl4-induced liver fibrosis in the rat. A high correlation was found between the degree of fibrosis and alpha 2-macroglobulin of the rat (alpha 2-macrofetoprotein, alpha M-FP) used for monitoring the acute phase response. This acute phase reaction was provoked by epinephrine just before CCl4 treatment was started. In the present study we analyzed the effect of APR by repeating these experiments and estimating liver neutral collagenase with a synthetic substrate and endogenous collagen as a substrate, and liver prolyl-4-hydroxylase. A strong depression of liver collagenase activity was found in rats with a preceding acute phase reaction contrary to the rats that underwent CCl4 treatment only. A high level of alpha M-FP correlated negatively with collagenase activity. Also in vitro alpha M-FP proved to inhibit collagenase activity. Prolyl-4-hydroxylase was increased in the rats during acute phase reaction and correlated highly and positively with alpha M-FP, haptoglobin, and ceruloplasmin. Thus high levels of APR promote development of CCl4-induced fibrosis, partly by anticollagenase activity and partly because of enhancement of prolyl-4-hydroxylase activity. The latter phenomenon can also be explained by the presence of APR, but this has to be proved.

Acute phase reactants enhance CCl4 induced liver cirrhosis in the rat

Abstract High levels of acute phase proteins (acute phase reactants, APR) suppress acute inflammatory reactions in the rat. As many APR have antiprotease properties, including an anticollagenase activity, the effect of APR on the development of CCl4-induced liver fibrosis was investigated in rats. APR were provoked by repeated injections of epinephrine, inducing a broad spectrum of APR. This reaction can be monitored measuring α2-macroglobulin levels in the rat (α2-macrofetoprotein, αM FP). This protein was found to inhibit both acute galactosamine hepatitis and acute CCl4-induced liver toxicity. The animals with high levels of APR at the start of CCl4 treatment developed a more severe degree of fibrosis and cirrhosis than the control group in which no acute phase reaction was induced. Epinephrine alone had no such effects. Additionally, the APR positive group showed an initially lower degree of hepatocellular damage when compared to control animals. This uncoupling of liver cell damage and subequent fibrosis may demonstrate that higher levels of APR might be important as to the development of cirrhosis, possibly based on the anticollagenase activity of these proteins.

Experimental Pancreatitis in the Rat: Role of Bile Reflux in Sodium Taurocholate-Induced Acute Haemorrhagic Pancreatitis

Mortality of sodium taurocholate-induced acute haemorrhagic pancreatitis in the rat was prevented by biliary diversion. Bile reflux into the pancreas after the induction of pancreatitis is postulated to be a major factor affecting mortality of this popular model of acute pancreatitis. The reduction of pancreatic secretory volume during pancreatitis is thought to be the cause of this phenomenon. Bile reflux augments dehydration by its stimulation of ascites production. It is suggested that sodium taurocholate-induced pancreatitis in the rat can only be extrapolated to human disease if bile reflux indeed plays a significant role in acute pancreatitis in man.

Inhibition of polymorphonuclear leukocyte chemotaxis byα-macrofetoprotein, an acute-phase reactant of the rat

Abstractα-Macrofetoprotein (αMFP) is a normal fetal plasma constituent in the rat, with very low plasma levels in the adult phase but rising sharply after injury. This fetal acute-phase protein is a strong inhibitor of inflammatory edema. Fetal inflammatory reactions show diminished exudation, but also impaired emigration of polymorph nuclear cells (PMNs). Therefore we studied the effect of (αMFP on chemotaxis of PMN in vitro and in vivo. In vitro experiments showed a strong inhibitory effect on casein-induced leukotaxis (Boyden technique) with a clear dose-effect relationship. In vivo with glycogen-induced pleurisy and peritonitis, high (αMFP levels are accompanied by diminished PMN emigration and vice versa. The significance of these findings is discussed in relation to fetal pathology and also as a model showing the modulating effects of acute-phase proteins on the inflammatory reaction induced by tissue injury.