Alteration of protein glycosylation in liver diseases.

Chronic liver diseases are a serious health problem worldwide. The current gold standard to assess structural liver damage is through a liver biopsy which has several disadvantages. A non-invasive, simple and non-expensive test to diagnose liver pathology would be highly desirable. Protein glycosylation has drawn the attention of many researchers in the search for an objective feature to achieve this goal. Glycosylation is a posttranslational modification of many secreted proteins and it has been known for decades that structural changes in the glycan structures of serum proteins are an indication for liver damage. The aim of this paper is to give an overview of this altered protein glycosylation in different etiologies of liver fibrosis / cirrhosis and hepatocellular carcinoma. Although individual liver diseases have their own specific markers, the same modifications seem to continuously reappear in all liver diseases: hyperfucosylation, increased branching and a bisecting N-acetylglucosamine. Analysis at mRNA and protein level of the corresponding glycosyltransferases confirm their altered status in liver pathology. The last part of this review deals with some recently developed glycomic techniques that could potentially be used in the diagnosis of liver pathology.

Occult pancreaticobiliary reflux in gallbladder cancer and benign gallbladder diseases.

BACKGROUND AND OBJECTIVES: It was proposed that occult pancreaticobiliary reflux (OPBR) was associated with precancerous mucosal changes in the gallbladder, hence the importance of this disorder. There are no published reports investigating the incidence of OPBR in patients operated on for the entire spectrum of benign gallbladder diseases and gallbladder cancer. Our aim was to determine the incidence of OPBR and measure the levels of active pancreatic enzymes (amylase and lipase) in gallbladder bile of patients undergoing cholecystectomy for benign and malignant gallbladder diseases. METHODS: One hundred eight patients with normal pancreaticobiliary junction evidenced by operative cholangiography were included in the study. RESULTS: According to gallbladder bile amylase and lipase levels, 84.2% and 89% patients respectively had OPBR. OPBR was present in all gallbladder cancer patients; in these patients the biliary levels of amylase and lipase were significantly higher than the levels found in patients with benign gallbladder pathology (P < 0.0001). CONCLUSIONS: OPBR could lead to inflammatory changes of the biliary epithelium and progress towards the development of precancerous mucosal changes and gallbladder cancer. The reason why such high levels of pancreatic enzymes are regurgitated into the biliary tree of patients with gallbladder cancer should be clarified.

Liver enzyme alterations after laparoscopic cholecystectomy.

AIMS: We aimed to investigate the alterations in serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl-transferase (GGT), alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) in patients who had undergone laparoscopic cholecystectomy (LC) and compare these changes with those occurring after open cholecystectomy (OC). METHODS: Of 267 patients who underwent LC between January 2003 and December 2005, 86 patients without complications were eligible for study. Twenty-six patients who underwent OC during the same interval were also enrolled in the study as a control group. Blood samples were taken preoperatively and 24 hours after operation for biochemical tests. RESULTS: Statistical analyses revealed significant increases in AST, ALT, GGT and LDH levels in the LC group postoperatively. Compared with the OC group, the differences between elevations of enzyme levels were also significant for LC group. CONCLUSION: We conclude that these enzyme elevations could mostly be attributed to the negative effects of the pneumoperitoneum on the hepatic blood flow. Although these changes do not seem to be clinically important, care should be taken before deciding to perform LC in patients with hepatic insufficiency.

Studies on the mechanism of accumulation of cholesterol in the gallbladder mucosa. Evidence that sterol 27-hydroxylase is not a pathogenetic factor.

BACKGROUND/AIMS: Cholesterolosis is characterized by accumulation of esterified cholesterol in human gallbladder mucosa. The present study aimed at investigating possible pathogenetic factors for cholesterolosis. The hypothesis was tested that a reduced sterol 27-hydroxylase or an increased amount of ACAT-1 enzyme may be of importance. METHODS: Gall bladder mucosa and bile were obtained from patients with cholesterol gallstones undergoing cholecystectomy (30 with and 43 without cholesterolosis). RESULTS: In cholesterolosis, the gall bladder mucosa was characterized by a several-fold increase in esterified cholesterol and normal content of free cholesterol. The amount of ACAT-1 protein, measured by immunoblotting, was similar in patients with and without cholesterolosis. The level of 27-hydroxycholesterol in gallbladder mucosa was elevated sevenfold as compared with cholesterol in patients with cholesterolosis. Most (87%) of this oxysterol was esterified and the accumulation is most probably secondary to the higher total amount of cholesterol in the cells. Patients with cholesterolosis had normal levels of both sterol 27-hydroxylase mRNA (real time polymerase chain reaction) and protein (immunoblotting). The enzymatic activity of the sterol 27-hydroxylase in gallbladder mucosa was normal or increased in cholesterolosis. CONCLUSIONS: The pathogenesis of cholesterolosis may be multifactorial, but is not caused by reduced efflux of cholesterol due to a defect sterol 27-hydroxylase mechanism.

Increased acylCoA-cholesterol ester acyltransferase activity in gallbladder mucosa in patients with gallbladder cholesterolosis.

OBJECTIVE: Although cholesterolosis of the human gallbladder is a relatively common disease, its etiology has not been fully understood. The aim of this study was to determine this etiology. METHODS: The lipid composition of the gallbladder mucosa and gallbladder bile and the enzyme activities (acylCoA-cholesterol ester acyltransferase [ACAT] and cholesterol ester hydrolase [CEH]) of the gallbladder mucosa were measured in control subjects, patients with cholesterolosis, and patients with cholesterol gallstone disease. RESULTS: Levels of cholesterol ester in gallbladder mucosa in patients with cholesterolosis (n = 12) were higher than those in control subjects (n = 8). With regard to the lipid content in gallbladder bile, no differences were found in concentrations of cholesterol, phospholipids, and bile acids among control subjects (n = 11), patients with cholesterolosis (n = 13), and those with cholesterol gallstone disease (n = 15). In gallbladder mucosa, ACAT activity was significantly higher in patients with cholesterolosis (n = 10) than in control subjects (n = 8), whereas CEH activity did not differ between the two groups. As a result, the ACAT/CEH activity ratio was higher in patients with cholesterolosis than in control subjects. CONCLUSIONS: It would be suggested that cholesterol ester synthesis of gallbladder mucosa might play an etiological role in the development of cholesterolosis.

Identification of an alkaline sphingomyelinase activity in human bile.

The hydrolysis of sphingomyelin has been found to generate important signals regulating cell proliferation, differentiation and apoptosis. However, the enzymes responsible for digestion of dietary sphingomyelin have not been well documented. This study demonstrates the occurrence of a sphingomyelinase (SMase) in both human hepatic bile and gallbladder bile. The enzyme was equally found in both bacteria negative and positive bile samples and in samples obtained from patients with or without gallbladder diseases. A bacteria-free gallbladder bile was used for characterization. It was found that bile SMase hydrolyzed sphingomyelin to phosphorylcholine and ceramide with negligible activity against either phosphatidylcholine or p-nitrophenyl phosphate. The enzyme preferred an alkaline condition and the optimal pH was 9. The activity of this alkaline SMase was bile salt dependent and was fully activated by 4-6 mM bile salts. Triton X-100, the non-ionic detergent did not activate bile SMase. Ca2+ and Mg2+ ions had no significant effect at optimal bile salt concentration. The molecular mass of this enzyme was about 85 kDa as measured by Sephadex G200 gel chromatography. In conclusion, we demonstrated a SMase in bile which differs markedly from the known acid and neutral SMase. Its potential important roles in sphingomyelin digestion and gallbladder diseases require further investigation.

Clinical utility of a wheat-germ precipitation assay for determination of bone alkaline phosphatase concentrations in patients with different metabolic bone diseases.

Bone alkaline phosphatase was evaluated by wheat-germ lectin precipitation in several clinical conditions. The study included 33 premenopausal healthy women, 46 postmenopausal apparently healthy women, 19 growing children, 24 patients with Paget's disease, 31 patients with primary hyperparathyroidism and 66 patients with hepatobiliary diseases. In postmenopausal women the mean T score (i.e.: the number of SD below or above the mean for premenopausal women) was 2.6 +/- 1.3 (SD) for bone alkaline phosphatase and 1.61 +/- 1.21 for total alkaline phosphatase (p < 0.001). The T score for bone alkaline phosphatase provided a better discrimination from normals for both Paget's disease (22.1 +/- 27.8 versus 12.8 +/- 16 p < 0.001) and primary hyperparathyroidism (8.2 +/- 4.3 versus 4.6 +/- 3.7 p < 0.005 for bone alkaline phosphatase and total alkaline phosphatase respectively). After treatment with intravenous bisphosphonate the percent decrease of bone alkaline phosphatase was larger than that of total alkaline phosphatase both in patients with Paget's disease (-46% versus -72% p < 0.01) and in patients with primary hyperparathyroidism (-21% versus -47% p < 0.02) and an estimate of the precision (delta mean/SD of the delta mean) for bone alkaline phosphatase was 1.9-3.7 times higher than that of total alkaline phosphatase. In twelve osteoporotic patients treated for six months with oral alendronate the decrease in bone turnover was detected with significantly higher precision with bone alkaline phosphatase than with total alkaline phosphatase (p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

[Immunohistochemical demonstration of pepsinogens I and II in the gallbladder]. / Demostración inmunohistoquímica de pepsinógeno I y II en la vesícula biliar.

The immunohistochemical expression of pepsinogen I (PI) and pepsinogen II (PII) was studied in 103 gallbladder carcinomas, 25 non tumoral gallbladder lesions and 23 gallbladder cancer metastases. PI was positive in 6% of carcinomas and only in 2% of non tumoral contiguous mucosa. PII was positive in 46% of carcinomas and 43% of controls. Metastases were negative for both enzymes. Pyloric gland metaplasia of non tumoral mucosa adjacent to the tumor expressed PII in 68.9% and PI in 3.4% of cases. The same lesion in non tumoral gallbladders was positive for PII in 66.6% of cases, and no case expressed PI. There was no relationship between PI expression and the histological degree of differentiation. Only tumors with serosal or subserosal involvement were positive for PI. PII had a higher expression in early or well differentiated tumors. Tumors with pyloric gland metaplasia in the contiguous non tumoral mucosa expressed PII more frequently. Our results, based on the expression of PI and PII, suggest a relationship between the presence of pyloric gland metaplasia and some types of gallbladder cancer.

Intestinal metaplasia and altered enzyme expression in propylnitrosamine-induced Syrian hamster cholangiocellular and gallbladder lesions.

Intrahepatic bile duct and gallbladder preneoplastic and neoplastic lesions induced in Syrian golden hamsters by propylnitrosamine treatment were investigated for the presence of polysaccharides and assayed immunohistochemically for expression of the enzymes glucose-6-phosphate dehydrogenase (G6PD) and glutathione-S-transferase (GST) molecular forms. On the basis of an increase in G6PD and the GST-placental form, a sequence of altered cell populations ranging from simple ductular proliferation through dysplasia and cholangiofibroma to cholangiocellular carcinoma could be established, the latter three lesions being characterized by marked increase in polysaccharide production. While similar goblet cell (intestinal) metaplasia and increased polysaccharide storage were also evident in carcinogen-induced gallbladder lesions G6PD and GST-P expression was decreased when compared with control epithelium.

Cholecystectomy with and without intraperitoneal drain.

383 patients undergoing cholecystectomy for non-acute gallbladder pathology were randomized with regard to the use of an intraperitoneal drain. 187 patients were drained, 196 patients were not drained. The postoperative course of temperature and liver laboratory tests, the duration of hospital stay and the postoperative morbidity were studied. There were no obviously significant differences between drained and undrained patients. Thus drainage of the subhepatic space after cholecystectomy as performed in this study, could not be associated with any disadvantages for the patient. The advantage of the intraperitoneal drain, could, because of a failing randomization in 13% of the patients not be completely evaluated. The preventive effect of the drain with regard to the incidence of postoperative complications however appeared to be very limited. The use of a prophylactic intraperitoneal drain after cholecystectomy could therefore safely be limited to appropriate patients as judged by the operating surgeon.

[Diagnostic-prognostic significance of long-term (5-11 years) variations in serum GOT-GPT levels in the blood]. / Sul significato diagnostico-prognostico delle variazioni a lungo termine (5-11 anni) dei valori sierici di AST e ALT.

A group of 48 patients (42 suffering from hepato-biliary diseases and 6 without hepatic diseases) was followed by the authors for a period lasting from 5 to 8 years, 13 out of them for longer. The hepatic disease was assessed on the basis of physical examination, current liver chemistry and proper and specific instrumental procedures. Initial and final diagnosis and the aminotransferases (AST, ALT) trend in years were carefully considered. First of all it was concluded that no advantage is obtained in monitoring the two aminotransferases instead of one alone. Moreover it is stressed the opportunity of referring aminotransferases activities in a simple way such as per cent of variation as referred to considered upper normal value differing from one to the other laboratory. The aminotransferase increase maintains an important and diagnostic significance in acute liver damage such as in acute hepatitis. An inappreciable prognostic value may be drawn from the follow up of these enzymatic parameters: for example the development of posthepatic fibrosis, or cirrhosis or hepatoma cannot be foreseen on the basis of the aminotransferases trend. A greater variability and sharp increases in AST-ALT values are recorded in patients with biliary gallstones.

Serum chemistry templates of disease in liver, pancreas, and gallbladder.

On routine hospital admission, 23,714 patients received a 28-test serum metabolic profile. The 33 most common diseases (4,132 patients) of liver, pancreas, and gallbladder (LPG) had unique chemical templates averaging 15 significant serum deviations. Each LPG disease differed from all others by elevations of both leucine-aminopeptidase (LAP) and alkaline phosphatase (AP) levels. LAP level was low or normal and serum glutamic oxaloacetic transaminase (SGOT) and AP levels were elevated in 43 non-LPG diseases. Patients with acute and chronic pancreatitis had elevated amylase levels. The four nonmalignant diseases of the gallbladder were associated with normal levels of amylase and lactic dehydrogenase (LDH); except for silent cholelithiasis, each showed elevated total bilirubin (BIL) levels. Patients with solitary or scattered lesions of the liver had normal bilirubin levels (2,115 patients), and those with diffuse interstitial or parencymal disease had elevated BIL levels. Cancer patients had elevated LDH and alpha1 globulin (A1G) levels, but low albumin levels. The importance of comprehensive liver profiles in the treatment of psychoses is emphasized by significant liver damage in a number of these patients. A1G was normal and LDH was elevated in patients having mononucleosis, hepatitis, lupus erythematosus, alcoholism, and alcoholic cirrhosis.

Effectof dialysis on tissue and serum alkaline phosphatase heat stability.

Tissue extracts and serum samples were dialyzed against TRIS buffer and the heat stability of the alkaline phosphatase (AP) was examined before and after dialysis for various periods of time. The effect of adding a mixture of ions or heat inactivated polled serum (HIS) to the dialyzed samples was also investigated. Dialysis of tissue extracts or of serum resulted in an increase in the mean AP heat stability. Addition of the ion mixture to dialyzed bone or liver extracts decreased the AP heat stability. Addition of HIS to these extracts decreased the AP heat stability of dialyzed bone extract but had little effect on dialyzed liver extract. These results are discussed.

[Serum lipase activity and endoscopic retrograde pancreatography in chronic pancreatitis and pancreatic neoplasm (author's transl)]. / Serum-Lipase-Aktivitat und endoskopische retrograde Pankreatikographie bei chronischer Pankreatitis un Pankreasneoplasma, Vergleichende Untersuchungen

Serum lipase activity was measured in 360 patients with the clinical suspicion of chronic pancreatic disease, 60 of them also having the lipase evocation test (serum lipase activity before and after pancreatic stimulation with secretin and pancreozymin). Of 48 with chronic pancreatitis (40 confirmed at operation) the diagnosis was made by endoscopic retrograde pancretography in all but one. Serum lipase activity was abnormal in 38. Without those cases associated with pancreatic insufficency, serum lipase activity-spontaneously and after the evocation test-was abnormal in 46 patients. Nine of 10 patients with papillary stenosis had the diagnosis confirmed at surgery, the pancretographic findings co-inciding with the surgical ones in all instances. All the five patients with abnormally high serum lipase activity also had chronic pancreatitis on pancreatography. In all of the 18 patients with pancreatic neoplasm pancreatography gave the same results as operation or post-mortem findings. In eight of these serum lipase activity was spontaneously elevated. The lipase evocation test was shown to be most effective if 2 C.H.R.- U/KG-h each of pancreozymin and secretin were administered. Serum lipase results were falsely positive in 17 of 300 patients with clinical suspicion of pancreatic disease but normal pancreatographic findings.