Central laboratory quality control in the National Cooperative Gallstone Study

This article presents the rationale, selection, operation, and quality control of the Central Serum and Central Bile Laboratories utilized by the National Cooperative Gallstone Study. The external quality control protocols were designed to monitor long-term stability of the analytical procedures and to measure the precision of the measurements as affected by the collection, labelling, storage, shipping, and laboratory methods. For both laboratories, the assessment of long-term stability by pool standards failed to produce the data necessary to come to relevant conclusions. Several of the problems involved, however, did lead to protocol changes that increased the reliability of the laboratory data. The use of duplicate measurements to monitor precision was more successful and demonstrated acceptable performance of these systems. This article describes the external quality control surveillance procedures employed in the NCGS, their strengths and weaknesses, statistical methods for the analysis of such quality control programs, and the implications for the final statistical analyses of the clinical trial patient data.     

External monitoring of a data coordinating center: experience of the National Cooperative Gallstone Study

A Biostatistical Monitoring Committee was established to review periodically the procedures and performance of the data coordinating center of the National Cooperative Gallstone Study. The functions of this committee, the types of data coordinating center activities reviewed, the manner in which monitoring of these activities was carried out, and an assessment of the value of this committee to the study are discussed in this article.     

Mass spectrometry identification of biliary bile acids in bile from patients with gallstones before and during treatment with chenodeoxycholic acid. An ancillary study of the National Cooperative Gallstone Study

The chemical structure of individual bile acids in 255 duodenal bile samples obtained from patients with radiolucent gallstones before and during treatment with chenodeoxycholic acid (375 or 750 mg/day) was determined by coupled gas chromatography/mass spectrometry. The two primary bile acids, cholic acid and chenodeoxycholic acid, and their metabolic products, deoxycholic acid, lithocholic acid, and ursodeoxycholic acid, were present in all bile samples and constituted greater than 97% of all bile acids. In pretreatment samples, the 12-oxo derivative of deoxycholic acid (3 alpha-hydroxy-12-oxo-cholanoic acid) was the next most abundant bile acid, being present in 62% of the samples; the average concentration was 1%, but three individuals had 6% to 8% of this bile acid. The 7-oxo derivative of chenodeoxycholic acid was also present in the majority of samples, but at a lower proportion (0.3%); five individuals had 2% to 3%. The 7-oxo derivative of cholic acid was present in a minority of samples (37%) in trace concentrations; isodeoxycholic acid and the 3-oxo derivatives of chenodeoxycholic acid and deoxycholic acid were also present in trace amounts. Four patients had 1% to 11% ursocholic acid in bile. During treatment with chenodeoxycholic acid, bile became enriched in it in direct relation to dosage; the concentration of its bacterial metabolites increased, and the proportion of cholic acid and its bacterial metabolites showed a reciprocal decrease. No unusual bile acids appeared, indicating that treatment with these doses of chenodeoxycholic acid does not result in the occurrence of additional uncommon bile acids in bile. It is suggested that the paucity of uncommon bile acids in bile, which contrasts strikingly with the great variety of uncommon bile acids known to be present in urine and feces, is the result of two factors: (1) the conversion of uncommon bile acids to common bile acids by reduction, hydroxylation, or epimerization during hepatic passage; and (2) efficient hepatic transport of common but not uncommon bile acids into bile.     

Persistent effects of chenodeoxycholic acid on biliary lipids in the hamster.

The effects of feeding chenodeoxycholic acid (CDC) on biliary lipid composition, on the rate-limiting enzymes of hepatic cholesterol and bile acid synthesis, and on hepatic cholesterol and bile acids were determined in hamsters. The goals were to study the mechanism and duration of the cholesterol desaturation action of CDC. Administration of CDC for 30 days significantly increased the biliary bile acid and lecithin to cholesterol ratio and the percentage of CDC in bile (p less than 0.01). These effects persisted for 20 days after discontinuing CDC (p less than 0.01) and were no longer evident at 30 days. HMG CoA reductase and 7 alpha-hydroxylase activities were significantly reduced by CDC (p less than 0.01). After discontinuing CDC, these effects persisted for 10 days at which time HMG CoA reductase was still decreased by 50 per cent (p less than 0.01) and 7 alpha-hydroxylase by only 12 per cent (p less than 0.01) and were no longer evident by 20 days. Hepatic cholesterol did not change, while hepatic CDC was significantly elevated throughout the experiment. Conclusions: (1) CDC has a salutory effect on biliary lipid composition while causing an increase of exogenous CDC in bile and a decrease of endogenous cholesterol synthesis. (2) The persistence of decreased cholesterol synthesis and of improved biliary lipid composistion after discontinuing CDC provides a rationale for studying this in man and then testing intermittent CDC regimes for gallstone dissolution.     

Acute effects of HMG-CoA reductase inhibitors on biliary lipids in patients with interrupted enterohepatic circulation

HMG-CoA reductase inhibitors decrease serum cholesterol by inhibiting hepatic cholesterol synthesis, but their influence on biliary lipids is not well characterized. In the present study Pravastatin (80 mg) was administered as a single oral dose to 10 patients with external bile fistula, after 1 week of interruption of the enterohepatic circulation, in order to assess the effect of inhibition of hepatic cholesterol synthesis on biliary lipids in conditions of stimulated bile acid synthesis. Bile was collected every hour for 12 h. On the day before, the same procedure was applied with a placebo, and collected bile used as control. Pravastatin decreased both bile acid and phospholipid concentration to about 60% of basal values; this change was still significant after 10 h. Cholesterol concentration was also decreased to about 70% of basal values, but this change was significant only from the 5th to the 7th h. The per cent of cholic and chenodeoxycholic acid was not affected by the drug, but the ratio of glyco- to tauroconjugated bile acids was decreased to about half the initial values. Bilirubin concentration exhibited a late increase, suggesting a reduction in the bile flow. These results suggest that, in patients with interrupted enterohepatic circulation, biliary excretion of bile acids can be largely dependent on hepatic cholesterol synthesis.     

Effect of colectomy with ileo-anal anastomosis on the biliary lipids

Abstract. Total colectomy with ileo-anal anastomosis is an effective treatment for ulcerative colitis and familial adenomatous polyposis. The absence of the colon and the coexistence of bile acid malabsorption may increase bile lithogenicity, but data on biliary lipid composition in patients with this operation is lacking. Our aim was to assess bile lithogenicity, bile composition and mass of biliary lipids within the gallbladder. We studied 11 patients with total colectomy and ileo-anal anastomosis and 16 healthy controls. We measured the percentage composition of conjugated bile acids and the masses within the gallbladder of the three main biliary lipids. This method, in contrast with measurement of cholesterol saturation index, can determine the cause of bile lithogenicity in terms of absolute modifications of the biliary lipids. There was no difference in the cholesterol saturation index between patients and controls. Colectomy patients had reduced masses of all three biliary lipids (medians and ranges, mmol): cholesterol (Ml (0.03–0.24) vs. 0.36 (0.02–0.96), P <0.02; bile acid 1.62 (0.75–5.21) vs. 3.95 (1.27–8.70), P <;0.01; phospholipids 0.35 (0.07–0.69) vs. 1.14 (0.14–3.00), P >0.002. They also had reduced per cent deoxycholic acid: 3.8 (0.0–27.6) vs. 17.4 (6.4–44.7), P < 0.005, and increased percent cholic acid: 44.9 (23.3–71.4) vs. 34.3 (19.2–57.9), P <0.05. We conclude that, despite having bile acid malabsorption, patients with colectomy and ileo-anal anastomosis have a normal cholesterol saturation index, caused by a concomitant reduction in the masses of all three biliary lipids. The reduced per cent biliary deoxycholic acid may help explain the reduced cholesterol and phospholipid masses in these patients. Total colectomy with ileo-anal anastomosis does not seem to predispose to the formation of cholesterol gallstones.     

Effects of acipimox on plasma lipids and biliary lipids in healthy subjects

The effects of Acipimox (5-methylpyracine-2-carboxylic acid-4-oxide, 3 X 250 mg/day) on plasma lipids, lipoproteins, and biliary lipids were studied in 14 healthy male volunteers using a double blind cross-over design. There was a significant (P less than 0.05) rise of HDL-cholesterol by 9.8%, while effects on other lipids and lipoproteins were small and insignificant (total cholesterol minus 6.5%, LDL-cholesterol minus 11.8%, free cholesterol minus 4.2%, total triglycerides plus 13.5% and phospholipids plus 3.9%). There was a significant rise of the HDL-cholesterol/LDL-cholesterol ratio by 23.6% (P less than 0.02) and of the HDL-cholesterol/total cholesterol ratio by 16% (P less than 0.05). Apolipoproteins AI, AII, and B were not significantly affected. The ratio of HDL-cholesterol/Apo AI increased significantly (P less than 0.05), and the ratio of HDL-cholesterol/Apo AII rose from 1.22 to 1.32 (P less than 0.05), while the ratio LDL-cholesterol/Apo B fell from 1.96 to 1.73. The composition of HDL and LDL, therefore, must have been altered by Acipimox. The relative cholesterol concentration in bile was significantly (P less than 0.05) increased by treatment with Acipimox, while bile acids and phospholipids were not significantly affected. The lithogenic index rose significantly by 15.1% (P less than 0.02) as calculated according to Admirand and Small, while calculations according to Hegard and Dam yielded a slight insignificant rise (P less than 0.1). The findings suggest that treatment with Acipimox might be associated with an increased risk of cholelithiasis. However, only long-term epidemiologic studies can ultimately demonstrate whether or not Acipimox increases the risk of gallstone formation.     

Extrahepatic biliary obstruction by a common bile duct inflammatory polyp in association with a gallstone, and treatment by endoscopic sphincterotomy

After a two-year history of recurrent abdominal pain, an 84-year-old man presented with acute pancreatitis and obstructive jaundice. An endoscopic retrograde cholangiogram demonstrated two filling defects approximately 1.0 cm in diameter, in a dilated common bile duct. Endoscopic papillotomy was performed which resulted in a polypoid tumour delivering itself into the wound followed by a free flow of bile. In addition, a single 1.0 cm gallstone was removed from the common bile duct, above the tumour, using a Dormia basket. The patient recovered completely. Histological examination of biopsies of the tumour taken on three subsequent occasions showed it to consist only of inflammatory tissue (an inflammatory polyp) and later, regenerating bile duct mucosa. After six months this tumour had completely regressed.     

Estrogen-induced gallstone formation in males. Relation to changes in serum and biliary lipids during hormonal treatment of prostatic carcinoma.

To assess if and by which mechanisms pharmacological estrogen treatment induces gallstone disease, we examined patients with recently diagnosed prostatic cancer randomly allocated to estrogen therapy (n = 37) or orchidectomy (n = 35). According to gallbladder ultrasonography, after 1 yr new gallstones had developed in 5 of 28 estrogen-treated patients, compared with 0 of 26 orchidectomized patients (P = 0.03). Estrogen therapy for 3 mo increased the relative concentration of cholesterol and cholesterol saturation of bile by approximately 30% (n = 10). Serum LDL cholesterol was reduced by approximately 40%, and its relative change related inversely to that of bile cholesterol (Rs = -0.77). There were no changes in biliary or serum lipids after orchidectomy (n = 9). Secretion rates of biliary lipids were measured with a duodenal perfusion technique. Patients on chronic estrogen therapy (n = 5) had approximately 40% higher biliary excretion rates of cholesterol than age-matched controls (n = 7). Phospholipid secretion was also higher, but no difference in bile acid secretion was found. We conclude that an increased hepatic secretion of cholesterol results in increased cholesterol saturation of bile and an enhanced rate of gallstone formation during estrogen treatment. The changes in bile cholesterol seem to be related to the induced changes in serum lipoprotein metabolism.     

Effect of ciprofibrate treatment on biliary lipids in patients with hyperlipoproteinaemia

Abstract. Treatment with hypolipidaemic drugs such as clofibrate increases secretion of biliary cholesterol and induces supersaturation of bile, leading to an increased risk of gallstone formation. Ciprofibrate is a phenoxyisobutyrate derivative with lipid-lowering effects in hyperlipoproteinaemia. We analysed serum lipid levels and biliary lipid composition and cholesterol saturation of gallbladder bile in nineteen hyperli-poproteinaemic patients before and after 6 weeks treatment with ciprofibrate, 100 mg daily. In addition, hepatic secretion rates of biliary lipids were determined in eight of the patients. Ten of the patients were also studied after 1 year of treatment.
Short-term treatment reduced the serum cholesterol levels by about 20% ( P < 0·001) and the serum triglycerides by about 40% ( P < 0·001). The relative cholesterol concentration and cholesterol saturation of bile were not significantly increased for the group as a whole, nor in patients with familial hypercholesterolaemia ( n = 9), or with other types of hyperlipidaemia ( n = 10). During treatment, however, fourteen patients had saturated bile compared with nine before treatment. An increase in cholesterol saturation was the consequence of an increased hepatic secretion of cholesterol whereas the secretion rates of bile acids and phospholipids were unaffected.
After 1 year of treatment the serum lipid concentrations were reduced to about the same level as after 6 weeks, whereas biliary lipid composition and cholesterol saturation had returned to pre-treatment values.
In contrast to clofibrate ciprofibrate exerts hypolipidaemic effects without consistently increasing the relative cholesterol concentration in bile. In some patients it leads to a transient rise in cholesterol saturation of gallbladder bile. This effect is due to an enhanced secretion of cholesterol probably because of a mobilization of tissue cholesterol.     

Bezafibrate therapy and biliary lipids: effects of short-term and long-term treatment in patients with various forms of hyperlipoproteinaemia

Several hypolipidaemic drugs increase biliary cholesterol saturation and induce the formation of cholesterol gallstones. In order to determine the influence of bezafibrate, a clofibrate analogue with hypolipidaemic properties, on bile lipid composition, we studied twelve patients with various forms of hyperlipoproteinaemia (six type IIA, three type IIB and three type IV hyperlipoproteinaemia; six had a genetic diagnosis of familial hypercholesterolaemia and five familial combined hyperlipidaemia). After 4 weeks of therapy, when serum lipids were significantly reduced, the relative proportion of cholesterol in stimulated fasting duodenal bile was increased by 28% (P less than 0.001). Phospholipids were concomitantly increased and bile acids decreased, resulting in an increase in biliary cholesterol saturation from 89 +/- 4% to 105 +/- 8% (SEM, P less than 0.02). The changes induced were similar in patients with familial hypercholesterolaemia and familial combined hyperlipidaemia. After 1 year of continued treatment, serum lipid responses were unaltered. The changes in biliary lipids were also persistent as the relative cholesterol concentration remained increased by 33% (P less than 0.01) and cholesterol saturation averaged 106 +/- 8% (P less than 0.02). Although the effect on bile cholesterol appeared to be transient in some patients, the results of the present study suggest that the risk of cholesterol gallstone formation may be increased during bezafibrate therapy.     

Study of the long-term effects of selective biliary obstruction (SBO)

Selective biliary obstruction (SBO) is a model of partial cholestasis in the rat in which the bile duct draining the median lobe is ligated and transected; the remaining biliary tree remains intact. Other authors introduced this experimental model and studied morphological and biochemical modifications in the liver after 2 days from surgery. They suggest that an adaptation may occur. Choosing some markers of cholestasis and some other markers of various cytoplasmic organelles, we studied the long-term effects that occur in serum and in total liver homogenate of selectively obstructed rats as compared to controls. Alkaline phosphatase activity and bile acids content, which were significantly higher than controls in serum and in total liver homogenate of the median lobe after 2 and 8 days from SBO, returned to normal range values after 30 days. Cytochrome-oxidase and glucose-6-phosphatase activity in total homogenate of the SBO median lobe remains perfectly similar to the control values in time. Results, together with morphological observations, suggest that cholestasis is present immediately after operation, then decreases gradually and disappears finally. The obstructed median lobe seems to cope with cholestasis.     

The physical chemistry of cholesterol solubility in bile. Relationship to gallstone formation and dissolution in man.

We determined the maximum solubilities of cholesterol in aqueous conjugated bile salt-egg lecithin-cholesterol systems as a function of several physical-chemical variables including those of physiological importance employing phase equilibria techniques. Equilibration rates are influenced by time and the method of sample preparation in that metastable supersaturation is readily induced at high bile salt: lecithin ratios, and equilibrium saturation by dissolution is achieved sluggisly at low bile salt:lecithin ratios. Equilibrium values for cholesterol saturation vary with the bile salt species, bile salt: lecithin ratio, temperature, ionic strength, and, in particular, with the total concentration of biliary lipids. Within physiological bile salt:lecithin ratios at 37 degreesC the influence of bile salt type and ionic strength is small, whereas the effects of bile salt:lecithin ratio and the total lipid concentration are major factors. We plotted on triangular coordinates a family of cholesterol solubility curves for each total lipid concentration (0.30--30 g/dl) and computed fifth-degree polynomial equations for each curve. With both the curves and the polynomial equations the "per cent cholesterol saturation" of fasting gallbladder and hepatic biles from patients with and without gallstones was calculated and both methods gave similar values. These results deomonstrate that by employing cholesterol saturation values appropriate to the total lipid concentration (range 0.2--24.9 g/dl) of individual biles, all cholesterol stone patients have supersaturated gallbladder biles, (mean, 132% [normal weight individuals], and 199% [morbidly obese individuals]). With controls and pigment stone patients the mean values were 95 and 98%, respectively, and in both approximately 50% of biles were supersaturated. Fasting hepatic biles were significantly more supersaturated than gallbladder biles (means 228--273%). Cholesterol monohydrate crystals were found in the majority of gallbladder (83%) and hepatic (58%) biles of cholesterol gallstone patients but were not observed in pigment stone patients or controls. We conclude that of the several factors in addition to the bile salt:lecithin ratios which can influence the cholesterol saturation of bile the total lipid concentration is the predominant determinant physiologically. Our results demonstrate that (a) metastable supersaturation is frequent in both normal and abnormal biles, (b) cholesterol gallstone patients have supersaturated gallbladder and hepatic biles without exception, and (c) the predominant driving force for cholesterol precipitation appears to be the absolute degree of cholesterol supersaturation.     

血清铁与血脂在肾病综合征蛋白倒置患者中的关系

目的探讨肾病综合征（NS）蛋白倒置（白蛋白／球蛋白〈1）患者巾血清铁与血脂之间的关系。方法分别用比色法检测健康对照组和肾病综合征患者组血清铁（SI）以及血清总胆固醇（Tc）、甘油三酯（TG）、高密度胆固醇（HDLc）、低密度胆固醇（LDL-C）水平,用免疫比浊法检测ApoAI和ApoB。结果NS组：血清铁与ApoAI（r=0．693,P〈0．01）、与ApoB（r=0．539,P〈0．05）具有相关性,差异显著,具有统计学意义,而与Tc、TG、HDL-C、LDL—C则无相关性,差异无统计学意义（P〉0．05）;以SI为因变量y、血脂各项指标为自变量,采用Stepwise进行分析,其回归方程为：y=-3．36＋10．202&#215;ApoAI。健康对照组：血清铁与血脂诸项指标间均未有相关关系,差异无统计学意义（P〉0．05）。结论NS患者中APoAI的浓度与血清铁的水平有密切关系,其生物学意义尚待进一步研究。     

National status of the entry-level doctorate in occupational therapy (OTD).

A multifaceted survey was conducted to identify the factors that academic occupational therapy (OT) programs were considering in making decisions as to whether the entry-level clinical doctorate (OTD) is a viable alternative for their institutions. The survey was sent in the summer of 2004 to program directors of all (150) occupational therapy programs in the United States. Responses were received from 111 programs (response rate of 74%). Quantitative (demographic) and qualitative (factor identification) data were compiled and analyzed. Supporting factors for the development of entry-level OTD programs included (a) coexistence of physical therapy doctorate program, (b) enhanced preparation of graduates, and (c) improved student recruitment. Impeding factors included (a) limited resources, (b) philosophical objections, and (c) lack of demand. In addition, results suggested that overall there is greater support for the OTD as a postprofessional degree. The study provided a historical record of current decision making in occupational therapy academic programs. In addition, the results of the study suggest a need for the development of national consensus regarding the place of the OTD in occupational therapy education.