Qualitative and quantitative preservation of the fine structure of absorptive cells in cultured biopsies of human small-intestine

The fine structure of the absorptive cells in human small-intestinal biopsies cultured for 6, 24, and 48 h was analyzed qualitatively and quantitatively. The findings show generally good preservation of the cultured absorptive cells and a normal distribution, size, and relative volume of their cell organelles, but there was a systematic decrease in the apical cell surface and an increase in the number of apical vesicles and tubules after culturing. Since the apical vesicles and tubules are thought to have a function in the transport of cell-coat material from the Golgi apparatus to the cell surface, these findings raise the question of whether a delayed transport or extrusion of cell surface material occurs. The diminished relative volume of the mitochondria and the increased signs of autophagy in some poorly preserved absorptive cells, are assumed to be an adaption to less favourable culture conditions.     

Temporal dynamics and regional distribution of [14C]serine uptake into mouse brain

In order to examine the uptake of L-serine into brain structures and brain metabolic compartments, L-[U-14C]serine was injected into tail vein of mice. The uptake was examined 30 min, 90 min, 3 h and 5 h after injection by both quantitative autoradiography of coronal brain sections and by biochemical analysis. Brain radioactivity was extracted and partitioned into protein associated pellets, metabolites soluble in aqueous phase and lipids soluble in the organic phase. Most of the radioactivity was found in the aqueous phase, about 10% was incorporated into lipids. Among phospholipids the highest label was found in phosphatidylserine, then in phosphatidylethanolamine and in phosphatidylcholine, it amounted to 52%, 30% and 18% of label by 90 min after injection, respectively. The brain distribution of L-serine uptake resembled that described for strychnine-insensitive [3H]glycine binding, with cortical structures being preferentially labelled.     

The importance of the Holter ECG monitoring of patients in the acute period of an ischemic stroke

10 guinea pigs of group 1 were injected subcutaneously kanamycin (400 mg/kg). The other 10 guinea pigs of group 2 received human ceruloplasmin as an intraperitoneal injection of 10% solution (0.25 ml of the protein preparation per 100 g body weight) 30 min before kanamycin administration. 7 days after the treatment the cochlease were isolated and examined electron microscopically. The Corti's organ was stained with alcyan blue and lanthanum hydroxide. In animals of group 1 there was mitochondrial vacuolization, enlargement of the cisterns of the endoplasmic network, Golgi apparatus, sub-superficial network; glycocalix layer on the surface of the receptor epithelium was unbroken but uneven, in the sensor cells of the Corti's organ of the group 2 animals only few mitochondria were vacuolized. A great number of ribosomes and mitochondrial contacts with membraneous cell structures were indicative of active protein synthesis and energetic processes. Glycocalix was less damaged. This was indicative of an otoprotective effect of ceruloplasmin which diminished kanamycin ototoxicity.     

Distribution of 2,2(1),4,4(1),5,5(1)-hexachlorobiphenyl in mice and Chinese hamsters: dose dependent accumulation in the mouse bronchial mucosa

The distribution of 2,2(1),4,4(1),5,5(1)-hexachlorobiphenyl-14C was studied in mice and Chinese Hamsters using whole body autoradiography and liquid scintillation counting. The mice exhibited a strong and persistent accumulation of radioactivity in the bronchial mucosa, and this accumulation was not fully developed until about 24 hours after an intravenous injection. The labelled substance passed to the fetuses of pregnant mice and was also concentrated in the fetal bronchi. Mice pretreated with a large dose of unlabelled PCB per os in peanut oil showed a completely different distribution pattern in the lungs--only traces of label being taken up by the bronchi. Quantitative measurements revealed a concomitant reduction of the total radioactivity retained by the lungs. Except for the lungs, however, no major differences in the distribution pattern were found at the various dose levels. The distribution in the Chinese hamsters equaled approximately that of the mice, but a very weak accumulation of label was observed in the hamster bronchi. The radioactivity in the mouse bronchi was considered as perhaps representing metabolized PCB.     

Effect of E4031, a class III antiarrhythmic drug, on ischemia- and reperfusion-induced arrhythmias in isolated rat hearts

With the onset of vitellogenesis, the follicular epithelium overlying the oocyte in stick insect ovarioles becomes highly polarized and patent by formation of wide intercellular spaces. The aim of the present study was to provide experimental support to the notion that the follicular epithelium in this insect species may be involved in transcytosis. Data demonstrate that the follicular epithelium carries out sulfo-conjugation of a 85 kDa fat body derived protein by allowing it ot transit from one cell pole to another. Along the basal end, follicle cells branch into a number of cytoplasmic finger-like projections. At the opposite end facing the oocyte they taper off into lance-head shapes. Different vesicular elements are evident at both these extremities. In vivo exposure to horseradish peroxidase shows that the vesicular elements present along the apical end provide an endocytic entry. In contrast, those present along the basal end are labeled with sodium [35S]-sulfate, suggesting that they may be exocytic vesicles containing a sulfo-conjugated secretory product. In vivo exposure to sodium [35S]-sulfate caused radioactivity to appear over the Golgi apparatus and some nearby vesicles of the follicle cell cytoplasm, including the exocytic vesicles. The intracellular pathway of the follicle cells was also examined by immunogold labeling using a monoclonal antibody raised against a 85 kDa fat body derived protein. Under these conditions, gold particles were consistently detected over the Golgi apparatus and the vesicular elements lying along both poles of the follicle cell membrane. Based on this evidence, it is concluded that follicular cells in stick insect ovarioles are endowed with the ability to undergo transcytosis by providing an endocytic entry along the apical end and by releasing exocytically a sulfo-conjugated 85 kDa protein along the baso-lateral domain of the follicle cell membrane.     

Clearance of chicken cystatin from the rat circulation

Chicken cystatin (mixed form) was prepared from egg white. Radioactively labeled preparations were administered intravenously to rats. 125I-cystatin disappeared from the rat circulation with a half-life of approximately 73 min. The radiolabeled inhibitor was rapidly taken up by the kidneys. Percoll density gradient analysis showed that it was incorporated into lysosomes. Within 24 hr after the injection of 125I-cystatin, 25% of the administered radioactivity was recovered in the urine, but only 2% was in the protein-bound form.     

Physiologic concentrations of zinc affect the kinetics of copper uptake and transport in the human intestinal cell model, Caco-2

Copper uptake was enhanced and copper transport was reduced in Caco-2 cells cultured in media containing high concentrations of zinc. Here we show that physiologic zinc concentrations also affect copper movement into and out of Caco-2 cells. Cells were seeded onto Falcon membranes with high pore density and maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum, nonessential amino acids, glucose and glutamine. In one experiment, the cells were exposed to media containing either 5 or 25 micromol zinc/L from d 14 to 21 after seeding. Then, copper uptake and transport, in both apical and basolateral directions, were measured by using 64Cu. Cells exposed to 25 micromol zinc/L had a 25% higher (P < 0.05) uptake of 64Cu from the apical side than those exposed to 5 micromol zinc/L. There was no effect of zinc on 64Cu uptake from the basolateral side, even though the amount of label taken up was as much as threefold higher (P > 0.05) than from the apical side. Transport of 64Cu across the cell layer in both directions was 50% less (P < 0.05) in cells exposed to 25 micromol zinc/L vs. 5 micromol zinc/L. In another experiment, zinc-exposed cells were labeled with 64Cu and efflux of the label to the apical and basolateral sides was measured over time. The rate of efflux to the apical side was linear and not affected by zinc. However, there was a 37% reduction (P < 0.05) in 64Cu efflux to the basolateral side by the higher zinc concentration. Curve-fit analysis showed that the basolateral efflux was made up of an exponential and a linear component. Cellular zinc concentrations were proportional to the zinc concentrations in the media. Although the data suggest that high media zinc inhibited the copper efflux transporter and enhanced the influx transporter, copper did not accumulate in the cell.     

The time sequence of response of the prolactin cells of the freshwater teleost, Poecilia reticulata, to an altered environmental salinity

The time sequence of the ultrastructural changes in the prolactin cells of the adult guppy, Poecilia reticulata, was studied in freshwater fish transferred to 1/3 seawater for 1, 4, 11 and 28 days. The morphological changes were slight and only detectable by quantitative (morphometric) procedures. The secretory granules showed a "biphasic' response to the altered environmental salinity, but the volume density of the Golgi apparatus progressively declined throughout the experiment. After an initial decrease in the volume density of the RER, that organelle regained its original prominence by 11 days in 1/3 seawater. The volume density of the nucleus was markedly higher in fish 28 days after transfer to 1/3 seawater than in any other group. Cell volume estimations showed that a shrinking of the cytoplasm rather than a swelling of the nucleus accounted for the high volume density figure. The changes in the volume density of the mitochondria closely paralleled those of the RER. Profiles of exocytosed granules were rarely found in any of the groups, but were least frequent in fish kept in 1/3 seawater for 28 days, when dense (lysosomal?) bodies were most abundant. The quantitative methods high-lighted some discrepancies in the rate and magnitude of the changes shown by some organelles and have led the authors to suggest that during adaptation of fish to dilute seawater, synthesis and release may not be coupled processes in the prolactin cell of P. reticulata.     

Serum malondialdehyde: possible use for the clinical management of chronic hepatitis C patients

BACKGROUND: A high risk of developing sludge or gallstones has been associated with pregnancy. The aim of this study was to relate the prevalence of sludge and gallstones during and shortly after pregnancy to fasting gallbladder volume as an indicator of gallbladder motility. METHODS: The population included 114 apparently healthy pregnant women from the Outpatient Clinic of Obstetrics of a large regional hospital and from the practices of regional midwives. Ultrasonography of the gallbladder was performed at weeks 15, 25, and 35 of gestation and at week 3 and month 6 postpartum. RESULTS: At gestational week 15, 3 women had gallstones and 10 had sludge (mean volume, 33.8 ml), and 99 women had a normal gallbladder (mean volume, 30.5 ml). At week 25, 1 woman with a normal gallbladder formed gallstones and underwent cholecystectomy shortly after, and 22 women had sludge, of whom 13 had a normal gallbladder at first examination (mean volume, 33.2 ml). In 88 women with normal gallbladders (of whom 2 had sludge at week 15) mean volume was 31.9 ml. At week 35, 2 women had gallstones, and 21 had sludge (mean volume, 30.5 ml). In the remaining 79 women the gallbladders were clear (mean volume, 29.5 ml). Eight women developed sludge and two women gallstones in normal gallbladders at week 25. Seven women with sludge at week 25 had a normal gallbladder at week 35. Three weeks postpartum only 10 of 100 women had sludge (mean volume, 29.1 ml). Of these 10, 9 women had a normal gallbladder at week 35. Twenty of 21 women with sludge at week 35 had normal gallbladders week 3 postpartum. Gallstones found at week 35 had disappeared. In the women with a normal gallbladder the mean volume was decreased to 19.7 ml (P < 0.0001). Six months postpartum, sludge was found in 6 (mean volume, 18.4 ml) of 93 women (mean volume, 20.3 ml), of whom 5 had a normal gallbladder at week 3 postpartum. Only 61 women showed a normal gallbladder at each examination of the study. No differences in patient characteristics were found between women with normal gallbladders and those with sludge or gallstones. CONCLUSIONS: Fasting gallbladder volume was increased in all pregnant women. This could not explain the formation of sludge or gallstones during gestation. Decrement of gallbladder volumes after delivery was faster in normal, clear gallbladders. More than a prerequisite, increased fasting gallbladder volume seemed to be a permissive factor of pregnancy-associated gallstone formation.     

Oxygen consumption, lactate metabolism, and gastric intramucosal pH in an experimental liver transplantation model

We have evaluated whether myocardial uptake of the fatty acid analog 123I-15-(p-iodophenyl)-3-R,S-methyl pentadecanoic acid (BMIPP) is dependent on the dietary state. METHODS: We compared the biodistribution of 150 MBq of 123I-BMIPP in six healthy volunteers in two states: after at least 12 hr of fasting and after oral glucose loading (75 g) 60 min before tracer administration, followed by a meal enriched in carbohydrates and protein. Planar and tomographic acquisitions were performed over a 4-hr time period after tracer injection; data were corrected for radioactive decay and injected dose. Radioactivity was measured in blood samples drawn at several points. RESULTS: Significant increases of glycemia and insulinemia and a significant drop in plasma nonesterified acids were documented after glucose loading. Half-time values for plasma radioactivity were significantly shorter in the glucose-loaded state than in the fasted state (4.3 +/- 1.4 min compared to 6.3 +/- 1.3 min, p < 0.05). Activity in the heart and liver tended to be higher in the glucose-loaded state than in the fasted state. SPECT images at 0.5 hr after tracer injection demonstrated that the myocardial wall-to-cavity ratio was higher after glucose than in the fasted state (2.53 +/- 0.59 compared to 2.11 +/- 0.21, p = 0.15). Washout from the liver between 1 and 4 hr after injection increased from 18.6% +/- 4.4% in the fasted study to 24.1% +/- 2.4% after glucose (p = 0.04). Washout from the myocardium between 0.5 and 3.5 hr after injection increased from 13.1% +/- 8.8% in the fasted study to 24.0% +/- 3.7% after glucose (p = 0.05). CONCLUSION: These results indicate that fasting before BMIPP scintigraphy is not mandatory to obtain adequate SPECT images. At the tire when SPECT is usually performed, glucose loading may provide improved ratios between myocardial and blood pool activity.     

Carbohydrate-mediated Golgi to cell surface transport and apical targeting of membrane proteins

Polarized expression of most epithelial plasma membrane proteins is achieved by selective transport from the Golgi apparatus or from endosomes to a specific cell surface domain. In Madin-Darby canine kidney (MDCK) cells, basolateral sorting generally depends on distinct cytoplasmic targeting determinants. Inactivation of these signals often resulted in apical expression, suggesting that apical transport of transmembrane proteins occurs either by default or is mediated by widely distributed characteristics of membrane glycoproteins. We tested the hypothesis of N-linked carbohydrates acting as apical targeting signals using three different membrane proteins. The first two are normally not glycosylated and the third one is a glycoprotein. In all three cases, N-linked carbohydrates were clearly able to mediate apical targeting and transport. Cell surface transport of proteins containing cytoplasmic basolateral targeting determinants was not significantly affected by N-linked sugars. In the absence of glycosylation and a basolateral sorting signal, the reporter proteins accumulated in the Golgi complex of MDCK as well as CHO cells, indicating that efficient transport from the Golgi apparatus to the cell surface is signal-mediated in polarized and non-polarized cells.     

Fermentation of dietary fibre by human colonic bacteria: disappearance of, short-chain fatty acid production from, and potential water-holding capacity of, various substrates

beta-CIT (2 beta-carbomethoxy-3 beta-(4-iodophenyl)tropane) is a cocaine analogue with a high affinity for the dopamine transporter. [11C] beta-CIT was prepared by N-methylation of nor-beta-CIT with [11C]methyl iodide. The total radiochemical yield of [11C] beta-CIT was 40-50% with an overall synthesis time of 35-40 min. The radiochemical purity was > 99% and the specific radioactivity at the time of injection was about 1000 Ci/mmol (37 GBq/mumol). Autoradiographic examination of [11C] beta-CIT binding in human brains post-mortem demonstrated a high level of specific binding in the striatum. PET examination of [11C] beta-CIT in a Cynomolgus monkey showed a marked accumulation of radioactivity in the striatum. The ratio of radioactivity in the striatum-to-cerebellum approached 5 after 87 min. In a displacement experiment, radioactivity in the striatum but not in the cerebellum, was markedly reduced after injection of unlabelled cocaine. [11C] beta-CIT has a potential as ligand for PET examination of cocaine effects in man.     

The effect of fasting and refeeding on total messenger ribonucleic acid content in the liver of chicks

1. The effect of 3-days fasting and the subsequent 3-days refeeding on total messenger ribonucleic acid (mRNA) content in the liver of chicks was investigated. 2. Body weight and liver weight were decreased by fasting, and were increased by refeeding to higher than initial values. 3. There was no influence of fasting and refeeding on RNA and mRNA contents per unit liver weight. 4. Total RNA content in the liver was decreased by fasting and was increased by refeeding to higher than the initial level. Fasting tended to reduce total mRNA content in the liver, and the following refeeding increased it significantly up to the initial level.     

The in vivo effect of colchicine on the addition of galactose and sialic acid to rat hepatic serum glycoproteins

Colchicine inhibits the secretion of plasma protein by rat hepatocytes and causes their intracellular accumulation in Golgi-derived secretory vesicles. This study examines whether colchicine affects secretion before or after galactose and sialic acid have been added to the secretory glycoproteins. D-[G-3H] Galactose was injected into rats and was found to be incorporated into serum glycoproteins contained within Golgi-derived secretory vesicles. The administration of colchicine (25 mumol/100 g, body weight), immediately before the injection of D-[G-3H] galactose, caused an increase in radioactivity of the serum glycoproteins in these cell fractions. D-[G-3H] Glucosamine was incorporated into serum glycoproteins contained within the rough and smooth endoplasmic reticulum and the Golgi cell fractions; however, its incorporation into the sialic acid moieties of these proteins only occurred in Golgi-derived cell fractions. Colchicine administration resulted in an increased incorporation of D-[G-3H] glucosamine into the sialic acid residues of serum glycoproteins contained within the Golgi cell fractions. These data indicate that colchicine inhibits secretion of serum proteins by rat liver after the addition of galactose and sialic acid to the secretory proteins has taken place.     

Aging rat vestibular ganglion: II. Quantitative electron microscopic evaluation

This laboratory has shown that age-related vestibular ganglion cell loss does not occur in the Wistar rat as it does in humans. However, in that study, intracellular changes were evident. The purpose of the present study was to quantitate some of these changes. The volume densities of mitochondria, rough endoplasmic reticulum (RER), Golgi apparatus, and aging pigment, as well as the diameter of the vestibular ganglion cells, of young (3 to 5 months) and old (24 to 31 months) female Wistar rats were determined by electron microscopy and stereological techniques. The data show a significant decrease in the volume densities of mitochondria (11.4%), Golgi apparatus (8.1%), and RER (8.9%), a significant increase in aging pigment (327%), and no change in mean profile diameter. These results suggest a decreased capacity for oxidative metabolism and protein synthesis that may reflect a decrease in the number of hair cells innervated by each ganglion cell and/or in the number of central connections. In either case, these findings suggest impaired metabolic and functional capabilities.     

Distribution of lead in lactating mice and suckling offspring with special emphasis on the mammary gland

The distribution of lead in lactating mice and suckling offspring was studied with whole body autoradiography at 4 and 24 h after a single intravenous injection of 203Pb (50 nmol Pb/kg) to the dams. In the lactating mice on day 14 of lactation, the highest uptake of radioactivity at 4 h after administration was recorded in renal cortex, skeleton and liver. A high uptake was also evident in the mammary gland. At 24 h after administration, the radioactivity had decreased in most organs except in the skeleton. In the suckling pups, exposed to lead only via dams' milk for 24 h, the highest level of radioactivity was present in the intestinal mucosa and a much lower level of radioactivity was present in the skeleton. The mammary glands from mice given three daily intravenous injections of 240 mumol Pb/kg were examined with X-ray microanalysis. At 4 h after the last injection, lead was found associated with casein micelles both inside the alveolar cell and in the milk lumen, indicating that lead is excreted into the milk, bound to casein, via the Golgi secretory system.     

A hydrophilic resin-embedding method for light and electron microscopic detection of tissue anionic sites with cationic colloidal iron: as applied to mouse Paneth cells

A cationic colloidal iron method was introduced for electron microscopic detection of anionic sites in hydrophilic resin-embedded specimens, and the method was applied to Paneth cells of the mouse jejunum. Mouse jejunal blocks were embedded in hydrophilic acrylic resin (LR White), cut into ultrathin sections, stained with the diluted cationic colloidal iron, and exposed to osmium vapor. The jejunal tissues, including the Paneth cells, embedded in hydrophilic resin were reactive to the fine cationic colloidal iron. At pH value 1.5, fine electron dense colloidal iron deposited along the rims of the secretory granules and the Golgi apparatus of the Paneth cell. Colloidal particles distributed on the osmiophilic reticular structures in the rim and in dot-like fashion lined the border between the granular core and rim. At pH value 4.0, ribosomes reacted to cationic colloidal iron particles in addition to the granular rims and Golgi apparatus. At pH 7.0, even the cores of the secretory granules were stained. Semi-thin sections prepared from the LR White-embedded specimens and stained at pH 1.5 with the diluted (1:3 in volume) cationic colloidal iron showed sufficient Prussian blue reaction for light microscopy in the rims of Paneth granules and mucus of goblet cells. This method is therefore useful for correlative light and electron microscopic detection of tissue anionic sites, including sulfate, carboxyl and phosphate groups, at various pH values.