Physical-chemical behavior of dietary and biliary lipids during intestinal digestion and absorption. 2. Phase analysis and aggregation states of luminal lipids during duodenal fat digestion in healthy adult human beings.

Following the feeding of a triacylglycerol-rich meal to healthy adult human beings, duodenal contents were aspirated for ex vivo chemical and physical-chemical analyses. The aspirates were collected during established lipid digestion and absorption into a "cocktail" of chemical inhibitors that rapidly inhibited ex vivo lipolysis. Following ultracentrifugation, the lipids separated into a floating oil layer, several interfacial layers, a "clear" or turbid "subphase", and a precipitated "pellet". By chemical and phase analyses, the floating layer was composed of oil-in-water emulsion particles with cores of triacylglycerol (TG), diacylglycerols (DG), and cholesteryl esters (CE) emulsified with a surface coat of partially ionized fatty acids (FA), monoacylglycerols (MG), diacylphosphatidylcholine (PL), and bile salts (BS). The interfacial layers contained similar emulsion particles dispersed among excess emulsifier which adopted a lamellar liquid-crystalline structure. Precipitated pellets were composed principally of emulsifying lipids, with smaller amounts of crystalline calcium soaps and BS. Relative lipid compositions of all but three subphases fell within a two-phase region of the condensed ternary phase diagram (Staggers et al., 1990, companion paper) where saturated mixed micelles composed of BS, FA "acid-soaps", MG, PL, cholesterol (Ch), and traces of DG (and TG) coexisted with unilamellar liquid-crystalline vesicles composed of the same lipids. Attempts to achieve clean separation of vesicles from micelles by repeat ultracentrifugation failed. Compared with the structure and sizes of lipid particles in equilibrated model systems (Staggers et al., 1990), quasielastic light scattering (QLS) analysis revealed that ex vivo micellar sizes (mean hydrodynamic radii, Rh) were similar (less than or equal to 40 A), whereas unilamellar vesicle sizes (Rh = 200-600 A) were appreciably smaller. Two-component QLS analysis of the subphases showed that much larger proportions of lipids were solubilized by micelles than were dispersed as unilamellar vesicles. When followed as functions of time, vesicles frequently dissolved spontaneously into mixed micelles, indicating that, in the nonequilibrium in vivo conditions, the constituent micellar phase was often unsaturated with lipids. These results are consistent with the hypothesis that, during hydrolysis of emulsified DG and TG by luminal lipases, unilamellar vesicles originate in lamellar liquid crystals that form at emulsion-water interfaces in the upper small intestine. In a BS-replete environment, unilamellar vesicles probably represent the primary dispersed product phase of human fat digestion and facilitate the dissolution of lipolytic products into unsaturated mixed micelles.(ABSTRACT TRUNCATED AT 400 WORDS)

Physical-chemical behavior of dietary and biliary lipids during intestinal digestion and absorption. 1. Phase behavior and aggregation states of model lipid systems patterned after aqueous duodenal contents of healthy adult human beings.

We developed equilibrium phase diagrams corresponding to aqueous lipid compositions of upper small intestinal contents during lipid digestion and absorption in adult human beings. Ternary lipid systems were composed of a physiological mixture of bile salts (BS), mixed intestinal lipids (MIL), principally partially ionized fatty (oleic) acid (FA) plus racemic monooleylglycerol (MG), and cholesterol (Ch), all at fixed aqueous-electrolyte concentrations, pH, temperature, and pressure. The condensed phase diagram for typical physiological conditions (1 g/dL total lipids, FA:MG molar ratio of 5:1, pH 6.5, 0.15 M Na+ at 37 degrees C) was similar to that of a dilute model bile [BS/lecithin (PL)/Ch] system [Carey, M. C., & Small, D. M. (1978) J. Clin. Invest. 61, 998-1026]. We identified two one-phase zones composed of mixed micelles and lamellar liquid crystals, respectively, and two two-phase zones, one composed of Ch monohydrate crystals and Ch-saturated micelles and the other of physiologic relevance composed of Ch- and MIL-saturated mixed micelles and unilamellar vesicles. A single large three-phase zone in the system was composed of Ch-saturated micelles, Ch monohydrate crystals, and liquid crystals. Micellar phase boundaries for otherwise typical physiological conditions were expanded by increases in total lipid concentration (0.25-5 g/dL), pH (5.5-7.5), and FA:MG molar ratio (5-20:1), resulting in a reduction of the size of the physiological two-phase zone. Mean particle hydrodynamic radii (Rh), measured by quasielastic light scattering (QLS), demonstrated an abrupt increase from micellar (less than 40 A) to micelle plus vesicle sizes (400-700 A) as this two-phase zone was entered. With relative lipid compositions within this zone, unilamellar vesicles formed spontaneously following coprecipitation, and their sizes changed markedly as functions of time, reaching equilibrium values only after 4 days. Further, vesicle Rh values were influenced appreciably by MIL:mixed bile salt (MBS) ratio, pH, total lipid concentration, and FA:MG ratio, but not by Ch content. In comparison, micellar systems equilibrated rapidly, and their Rh values only slightly influenced by physical-chemical variables of physiological importance. In contrast to the BS-PL-Ch system [Mazer, N. A., & Carey, M. C. (1983) Biochemistry 22, 426-442], no divergence in micellar sizes occurred as the micellar phase boundary was approached. The ionization state of FA at simulated "intestinal" pH values (5.5-7.5) in the micellar and physiologic two-phase zones was principally that of 1:1 sodium hydrogen dioleate, an insoluble swelling "acid soap" compound. By phase separation and analysis, tie-lines for the constituent phase in the two-phase zone demonstrated that the mixed micelles were saturated with MIL and Ch and the coexisting vesicles were saturated with MBS, but not with Ch.(ABSTRACT TRUNCATED AT 400 WORDS)

Immunohistochemical localization of proteoglycans in interstitial elements of human pancreas and biliary system.

The immunohistochemical localization of large proteoglycan and small proteoglycan was observed, using antibodies 2B1 and 6B6 (Sobue et al., 1988, 1989a), in fetal and adult pancreas and biliary system as well as in tumour tissues, obtained from 11 autopsies and 74 biopsies. The distribution of chondroitin 4- and 6-sulphate side chains, type I and IV collagen and elastin were also studied. In adult pancreas and all the biliary tracts examined, periductal fibrous tissues consisted mainly of dermatan sulphate small proteoglycan with networks of fibrous elements, which were composed of large proteoglycan, elastin, type I collagen and type IV collagen. In the interstitial components of cystadenoma of pancreas and biliary duct carcinoma, similar small proteoglycan-rich components were relatively abundant, although large proteoglycan was present in much larger amounts than that in non-neoplastic adult tissues. In some cholangiomas, the extra- and intracellular hyaline globules formed by the carcinoma cells were found to contain chondroitin sulphate large proteoglycan, laminin and fibronectin. The distribution of proteoglycans was observed to be different in the arterial walls of the interlobular tissues of the adult and the fetal pancreas. The biological significance of large and small proteoglycans in the interstitial connective tissues was discussed.

[Cultured biliary tract epithelial cells/alkaline and neutral elution as a model for the detection of carcinogens to the biliary tract epithelium].

Alkaline and neutral elution techniques were applied to detect chemical carcinogens in biliary tract cancer using biliary tract epithelial cells in culture. Since biliary tract epithelial cells actively grow in culture, DNA breaks induced by carcinogens in the [14C] thymidine-prelabeled DNA could be detected as radioactivities in the eluted fractions. Our study demonstrated that aflatoxin B1, 20-methylcholanthrene and dimethylnitrosamine induced DNA single-strand breaks in the biliary tract epithelial cells by the use of this system. Cultured biliary tract epithelial cells/alkaline and neutral elution offers a sensitive and organ-specific model for the detection of chemical carcinogens to the biliary tract epithelium.

Mercury-selenium interaction: distribution and excretion of 203Hg2+ in rats after simultaneous administration of selenite or selenate.

In female rats intravenously injected with 203HgCl2 (0.6 mg Hg2+ per kg body wt.) the effect of intraperitoneal administration of selenite or selenate (0.525 mg Se per kg body wt.) on distribution and excretion of 203Hg was studied. The content of 203Hg was lower in kidney and higher in liver and blood in the groups treated with selenate or selenite when compared with rats which received only mercury. The brain content of 203Hg was significantly increased in rats injected with selenite. Both selenium compounds injected immediately after mercury significantly decreased urinary as well as biliary excretion of 203Hg. A transient increase in the rate of biliary excretion of 203Hg during the first 2 h after administration was observed in rats treated with selenate. This finding seems to support the idea that the reduction of selenate to selenite in the body is not rapid but takes at least several hours.

Calcitonin gene-related peptide immunoreactivity in the biliary pathway and liver of the guinea-pig: distribution and colocalization with substance P.

Calcitonin gene-related peptide immunoreactivity was localized immunohistochemically in nerve fibers innervating the biliary pathway and liver of the guinea-pig. Immunoreactive fibers are present in all layers of the gallbladder and biliary tract and are particularly numerous around blood vessels. In the liver, immunoreactive processes are usually restricted to the interlobular space and porta hepatis, and only a few, very thin, beaded processes were observed in the hepatic parenchyma. A rich innervation is also associated with the vena portae. Positive ganglion cell bodies were not visualized within the ganglionated plexus of the biliary system, whereas they were found in the myenteric and submucosal plexus in the cranial portion of the duodenum corresponding to the sphincter of Oddi. The vast majority, if not all, of calcitonin gene-related peptide-immunoreactive fibers contain substance P immunoreactivity; however, there are some substance P-containing fibers lacking calcitonin gene-related peptide immunoreactivity. The lack of co-occurrence of calcitonin gene-related peptide and substance P immunoreactivities in intrinsic ganglion cells suggests that these two peptides are coexpressed in the extrinsic component of the innervation of the hepatobiliary system.

Pancreaticobiliary factors in the modulation of small intestinal enterokinase in the rat.

Chronic pancreaticobiliary diversion was employed to study the modulation of enterokinase in the small intestine of adult rats. Diversion resulted in apparent trophic changes of the proximal bypassed portion of the intestinal mucosa. An almost complete loss of mucosal enterokinase activity in the proximal duodenum but no increase of enterokinase in the segments distal to reentry of the common duct was found in the pancreaticobiliary-diverted rats. The effect on the enterokinase activity in the proximal segment was specific in that no other brush-border enzymes measured in that segment were decreased. The decrease in enterokinase was partially prevented by dietary supplementation with pancreatic trypsinogen and completely avoided with the addition of a combination of bile acids and trypsinogen. Supplementation with bile acid alone did not preserve the enterokinase levels in the bypassed rats. The results suggested that trypsinogen is the primary factor responsible for modulating enterokinase levels in the proximal small intestine, with bile acids acting as a modifier.

Peptide immunoreactive nerves and cells of the guinea pig gall bladder and biliary pathways.

Using the methods of immunocytochemistry and radioimmunoassay, five peptides (vasoactive intestinal polypeptide (VIP), substance P, somatostatin, met-enkephalin, and bombesin) have been found in the gall bladder and the biliary tracts of guinea pig and each of them possesses a characteristic distribution pattern. Networks of nerves containing each peptide were found in the smooth muscle, around blood vessels and, occasionally, in the lamina propria. The distribution of the peptide immunoreactive nerves in the gall bladder and biliary tract is similar to those found in the gut. Vasoactive intestinal polypeptide (11 +/- 1.5 pmol/g in the sphincters, mean +/- SEM) and substance P (21.5 +/- 1.8 pmol/g in the common bile duct) were found to be the most abundant peptides and a few VIP and substance P immunoreactive neurones were localised in the ganglionated plexus. Bombesin immunoreactive nerves were mainly seen in the sphincter of Oddi, where the mean concentration of extractable bombesin was 14.6 +/- 2 pmol/g. Somatostatin immunoreactive mucosal endocrine cells were identified in the epithelium of the common bile duct and the sphincter. The extractable somatostatin in these regions were 76 +/- 19 pmol/g and 162 +/- 30 pmol/g respectively.

[Clinical studies on PC-904 in biliary tract diseases (author's transl)].

The clinical effects of PC-904, a new semisynthetic penicillin, were studied in patients with biliary tract diseases, and the results were as follows: 1) PC-904 showed an average peak serum level of 40.7 +/- 11.6 microgram/ml 2 hours after an intravenous drip infusion of 1 g of the agent. The biliary level showed a peak value of 126.5 +/- 85.4 microgram/ml 2 hours to 3 hours after the infusion. 2) Isolated organisms from bile before the treatment were E. coli, Klebsiella, Proteus, Pseudomonas, Enterococcus and Bacteroides. MIC of PC-904 on 18 strains of isolated organisms was almost 6.25 microgram/ml or less. All isolated organisms except one strain of Klebsiella oxytoca disappeared after the treatment. 3) Six patients with cholelithiasis were medicated with PC-904 to prevent post-operative infections. The clinical effects were good in 4, poor in 1 and unknown in 1 case. 4) As to side effects no adverse reactions and allergic reactions were noted. Also no significant abnormalities of laboratory findings were observed.

Possible dual role for vasoactive intestinal peptide as gastrointestinal hormone and neurotransmitter substance.

Vasoactive intestinal peptide (V.I.P.) has been found in high concentrations both in the gastrointestinal tract and, unexpectedly, in the central nervous system. Immunocytochemical studies have demonstrated V.I.P. in nerve-fibers. These findings challenge the concept of V.I.P. as a simple gastrointestinal hormone and suggest a possible neurotransmitter function.