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Biomedical subjects

J S Patton

Publications and source records attributed to J S Patton.

At least 37 records · Page 2Linked to original sources
PubMed1987

Possible relationships between in vivo antitumour activity and toxicity of tumour necrosis factor-alpha.

The discoveries of a tumour necrosis-inducing substance in sera of experimental mice and of cytotoxic factor(s) in cultures of stimulated lymphoid cells triggered intense research efforts which have culminated in the production of two distinct but related recombinant materials, human tumour necrosis factors TNF-alpha (cachectin) and TNF-beta (lymphotoxin). The necrosis of tumours by TNF is but one feature of high doses of these immune system hormones that possess numerous biological activities. Apart from their direct cytotoxic/cytostatic activities against tumours in vitro and in vivo, the in vivo antitumour activities of TNF-alpha or TNF-beta may involve the following biological activities: the induction of interleukin 1 production; activation of polymorphonuclear neutrophil functions; modulation of endothelial cell functions; and augmentation of specific immune functions. Many of these activities are associated with an irreversible acute inflammation which appears to be the immediate lethal effect of TNF on transplantable tumours in mice. This inflammation leads to thrombosis, disruption of the tumour's blood supply and, finally, tumour death. Inflammatory effects of high doses of TNF are also seen in the rodent gastrointestinal tract but here the inflammation seems to be reversible.

Animals↗
PubMed1987

Bile salt absorption in killifish intestine.

Bile salt absorption was examined in vitro using entire small intestines from the killifish Fundulus heteroclitus. Intestines were everted over a glass rod and incubated in solutions containing 10 nM to 20 mM bile salts. After rinsing and correcting for the adherent fluid space, uptake rates and bile salt concentrations in the tissue were determined. The distal intestine contained a Na+-dependent active transport system for bile salt uptake with an apparent Vmax for taurocholate and cholate of 1.4 and 2.3 nmol.min-1.mg dry wt-1 (Km = 117 and 357 microM), respectively. At low concentrations (10 nM to 500 microM), absorption occurred almost exclusively (greater than 84%) in the distal intestine. However, at concentrations of 1 mM and above, bile salt absorption in the middle and proximal regions equaled that in the distal intestine. Thus, although an active transport system makes the distal intestine more efficient in absorbing bile salts, passive absorption appears to account for a significant amount of bile salt uptake at concentrations above the critical micellar concentration. The presence of oleic acid did not significantly affect bile salt uptake.

Animals↗
PubMed1987

Development of partial tolerance to the gastrointestinal effects of high doses of recombinant tumor necrosis factor-alpha in rodents.

Treatment of healthy rats and mice with a single intravenous injection of recombinant human tumor necrosis factor-alpha (rHuTNF-alpha) caused a dose-dependent gastrointestinal inflammation. Within 30 min gastric emptying was blocked and tissue edema occurred in the small and large intestine. In the cecum hemorrhage occurred after 4 h at doses greater than or equal to 250 micrograms/kg. The cecum exhibited an acute inflammatory response following rHuTNF-alpha treatment similar to that seen in tumor necrosis at the same dose. The vascular endothelium became swollen, increased numbers of neutrophils and other leukocytes attached to and penetrated the endothelium, and finally hemorrhage occurred. Treatment of rats with daily injections of rHuTNF-alpha (250 micrograms/kg per d) for 3 wk failed to produce cachexia. Within 24-48 h rats became resistant to the hemorrhagic effect of rHuTNF-alpha, however, the cytokine still caused a transitory block of gastric emptying after 10 d of treatment. Treatment at 5- or 10-d intervals produced results similar to the initial injection. These results suggest that maximum hemorrhagic response will occur when rHuTNF-alpha is administered at intervals of 5-10 d rather than daily.

Animals↗
PubMed1987

Dietary fat inhibits the intestinal metabolism of the carcinogen benzo[a]pyrene in fish.

Following the intestinal absorption of dietary benzo[a]pyrene (BP) by the killifish, this compound becomes incorporated along with dietary triglycerides into membrane-bound fat vacuoles within the intestinal epithelial cell (1985. J. Lipid Res. 26: 428-434). These vacuoles, arising from the smooth endoplasmic reticulum, are important transient structures involved in both the uptake and metabolism of dietary BP and, presumably, other lipophilic toxicants as well. In the present study we used subcellular fractions isolated from the intestines of spot (Leiostomus xanthurus), a teleost fish, to study factors that influence the metabolism of BP in a fat vacuole/microsomal system. Triglyceride-solubilized BP is capable of diffusion from fat vacuoles to microsomal enzymes. Increases in the concentration of fat vacuoles decrease the availability of BP to microsomal BP hydroxylase. The effect of fat vacuoles on the activity of BP hydroxylase becomes more pronounced as the concentration of BP in our test system decreases. Addition of cytosolic glutathione transferases to the fat vacuole/microsomal system enhances the activity of BP hydroxylase. Examination of binding of 3H-labeled BP to killifish (Fundulus heteroclitus) intestinal cytosolic proteins in vivo indicated that a large fraction of the radioactivity was associated within glutathione transferase. These results suggest that dietary fat inhibits metabolism of low levels of BP in the intestine. A consequence of this would be greater exposure of peripheral tissues to dietary carcinogens.

Animals↗
PubMed1986

Interferons and tumor necrosis factors have similar catabolic effects on 3T3 L1 cells.

The effect of a variety of cytokines on lipid metabolism in 3T3 L1 mouse fibroblasts and adipocytes was studied. Uptake of [3H]acetate by adipocytes and heparin-releasable lipoprotein lipase activity was inhibited after treatments of the cells with picomolar concentrations of recombinant human tumor necrosis factor alpha (rHuTNF-alpha), human tumor necrosis factor beta (rHuTNF-beta, also called lymphotoxin), murine interferon-gamma (rMuIFN-gamma), and a human hybrid interferon-alpha [rHuIFN-alpha 2/alpha 1 (Bgl II)]. Recombinant human interferon-gamma (rHuIFN-gamma), natural human colony-stimulating factor (HuCSF), and human interleukin 2 (HuIL-2) had no effect. Similar though less-marked suppression of [3H]acetate uptake by cytokines was seen in 3T3 L1 fibroblasts. Cytokines inhibited the incorporation of [3H]acetate into both membrane and storage lipids in the adipocytes. In addition to blocking lipid uptake and synthesis, rHuTNF-alpha and -beta, and rMuIFN-gamma stimulated the release of free fatty acid into the medium from adipocytes. Binding studies suggest that rHuTNF-alpha and rHuTNF-beta compete for the same cell-surface receptor on 3T3 L1 adipocytes, while rMuIFN-gamma binds to a separate receptor. The binding of rTNF-alpha to both adipocytes and fibroblasts can be significantly enhanced by preexposure of the cells to rMuIFN-gamma. There appear to be both high- and low-affinity receptors for rHuTNF-alpha on adipocytes, whereas fibroblasts exhibit a single class of high-affinity receptors. These results suggest that a variety of structurally distinct cytokines possess lipid mobilization activity, which may be of critical importance to the host in defense against infection or malignancy.

Acetates↗
PubMed1986

Visualization by freeze fracture, in vitro and in vivo, of the products of fat digestion.

The technique of freeze fracture was used to visualize triglyceride (TG) hydrolysis and the production of lipolytic products (LPs) in vitro and in vivo in the presence of bile salts (BS). Three systems were investigated: pure lipolytic products (oleic acid and monoolein) in the presence of a pure bile salt (taurodeoxycholate (TDC)), lipolytic products produced from TG by pancreatic lipase in the presence of a variety of bile salts, and lipolytic products produced in the intestine of the killifish, Fundulus heteroclitus, after fat feeding. In vitro, lamellae (4-5 nm thick with 0-8-nm water spacings) appeared on the surface of TG droplets in all preparations with LP/BS molar ratios of 1.5 or greater and spherical vesicles (diameter range, 20-130 nm) were produced from these lamellae. With model killifish bile (taurocholate-cholate 1:1) at LP/BS ratios between 1.5 and 4, homogeneous vesicles or particles (mean diameter, 23.8 nm) were produced by lipase at pH 6.9. In vivo, lamellar product phases also occurred after fat feeding. The smallest visible LP/BS structures by freeze fracture electron microscopy were approximately 20 nm globular particles. Large disc-shaped micelles either were not present or were below the resolution limit of the replica (approximately 10 nm). The dominant aggregated lipolytic product phase was composed of multiple layers of rough-textured lamellae. No evidence of cubic structure was seen. These results show that lamellar and vesicular lipolytic product phases can be intermediates in intestinal fat digestion. However, no evidence for the direct endocytotic absorption of these product phases by the intestinal microvillus membrane was found.

Animals↗
PubMed1985

Intramembranous particles are clustered on microvillus membrane vesicles.

Many intramembranous particles in pig jejunal microvillus membranes cluster during cell disruption and membrane vesiculation with the MgCl2 aggregation technique (Hauser, H., Howell, K., Dawson, R.M.C. and Bowyer, D.E. (1980) Biochim. Biophys. Acta 602, 567-577). Isolated brush borders and purified microvillus membrane vesicles were jet-frozen and examined by freeze-fracture electron microscopy. From 30 to 60% of purified vesicles exhibited no intramembranous particles on their fracture face and 22-39% exhibited clustered or aggregated intramembranous particles. Only 6-15% of the vesicles exhibited the random distribution of intramembranous particles that is characteristic of intact enterocytes. Aggregation was not reversed after dialysis to remove divalent cations. Prior freezing of tissue or vesicles (-70 degrees C) gave the same results as fresh unfrozen material. Heterogeneity of microvillus vesicles may occur among the vesicles generated from a single microvillus.

Animals↗
PubMed1985

The use of the Iatroscan TH-10 analyzer to quantify total lipids in a variety of sample types and lipid classes in human gallbladder bile.

Two methods for the measurement of total lipid weight in biological and geological samples and the major lipid classes in human gallbladder bile using the Iatroscan TH-10 analyzer are described. Total lipid determination involves the application of small (5 microliter) volumes to Chromarods, focusing of the sample into one band by partial development in chloroform-methanol (1:1), and quantification by flame ionization detection (FID). The response variation between different sample types did not affect the linearity of response, allowing a reproducibility of +/- 10% of the mean or better for samples ranging from 0.5 to 32 micrograms. Total lipid determinations in 10 samples could be performed in 30 min. The three major components of human gallbladder bile (cholesterol, phospholipids and bile acids) also were quantified with the Iatroscan. Samples focused on Chromarods were separated using a double development scheme in two solvent systems. All three components exhibited a linear response over the range of 0.25 to 8 micrograms. The repeated scanning of rods required at concentrations greater than 3 micrograms did not affect linearity of response. Samples from 10 patients could be processed in less than one hr. Several techniques are discussed to increase reproducibility when performing quantitative lipid analysis with the Iatroscan.

Bile↗
PubMed1985

Dietary fat assimilation and bile salt absorption in the killifish intestine.

Radiolabeled taurocholate (TC) and triolein were used to study fat assimilation and bile salt absorption in the stomachless saltwater killifish, Fundulus heteroclitus. Fat absorption occurred primarily in the proximal intestine with approximately 87% of a single dose (9 mg fat/8 g fish) absorbed in 2 h. Luminal triolein hydrolysis and enterocyte triolein resynthesis were tightly coupled. Killifish gallbladder bile contains taurocholate and cholate in an equal molar ratio at a combined concentration of 237 +/- 25 mM (n = 10) in 24-h-fasted fish. During fat assimilation luminal bile salt and fatty acid concentrations ranged between 10 and 30 mM. Between and during meals the total concentration of bile salts in the intestinal tissue remained roughly constant (4-6 mM) with the proximal one-third of the intestine containing 40% of the total and the remainder equally distributed between the mid and distal regions. All three regions of the intestine rapidly incorporated ingested TC in vivo, with the amount incorporated proportional to the pool size. In contrast, in vitro at low TC concentrations (60 nM), the distal one-third of the intestine incorporated 10 times as much TC in 2-min uptake experiments as the proximal and mid regions. Although there are many similarities between fat and bile salt assimilation in killifish and mammals, overall the processes are much simpler in killifish.

Animals↗
PubMed1985

Coassimilation of dietary fat and benzo(a)pyrene in the small intestine: an absorption model using the killifish.

Benzo(a)pyrene (BP) was dissolved in dietary fat and fed in a single dose to killifish (Fundulus heteroclitus). Fluorescence microscopic examinations of small intestinal content and frozen sections of whole small intestine revealed that during fat digestion BP was codispersed in liquid crystalline product phases produced during lipolysis (1979. Patton, J. S., and M. C. Carey, Science. 204: 145-148) and then coabsorbed with dietary lipid followed by its reappearance in intracellular fat droplets. During the time that the absorbed fat remained in the enterocytes, BP fluorescence was initially concentrated in the intracellular fat droplets and then spread throughout the cytosol of the enterocytes. Tissue analyses showed that BP was rapidly metabolized in the intestine and transported to the gallbladder. These studies show that separation of a dissolved hydrophobic carcinogen from dietary fat occurs primarily after the fat has been digested, dispersed, absorbed, and reassembled in the enterocyte. The inability of the enterocyte to discriminate between dietary fat and dissolved carcinogenic compounds may be a partial explanation of the observed link between high fat diets and the incidence of some cancers. In vertebrates, the intestine and not the liver, appears to be the major site of metabolism of dietary polycyclic aromatic hydrocarbons (PAHs).

Animals↗
PubMed1984

Similar bioavailability and lymphatic transport of benzo(a)pyrene when administered to rats in different amounts of dietary fat.

This study assessed the effect of concomitant lipid absorption on the bioavailability and lymphatic transport of benzo(a)pyrene (BP), a carcinogenic polycyclic aromatic hydrocarbon (PAH). Conscious, male Sprague-Dawley rats, equipped with biliary and mesenteric lymphatic catheters received intraduodenally a dose of 0.4 mumoles 3H-labeled BP completely dissolved in either 50 mumoles or 500 mumoles of olive oil. Diversion of mesenteric lymph allowed biliary and urinary excretion of 3H to be used as an indirect measurement of relative 3H portal transport. Total radiolabel recovered in a 24-hr period in each group was 20.0 +/- 2.6% of the 3H dose given in 50 mumoles of oil, and 17.0 +/- 1.0% of the 3H dose administered in 500 mumoles of oil. In animals receiving the low-fat test meal, 79.4 +/- 1.4% of the recovered radiolabel was found in bile; the corresponding value for the high fat dose was 78.5 +/- 2.6%. Thus a tenfold variation in the mass of the carrier vehicle (triglyceride oil) did not significantly effect the disposition of BP, and portal, not lymphatic transport, was the major route of post-absorptive transport. Although the chylomicrons produced from both fat doses were initially contaminated with BP, within 1-1.5 hr the radioactivity in lymph began to drop such that by 3 hr in the animals fed high fat, the chylomicrons were essentially free of BP. These results show that the rat enterocyte quickly adapts to PAH-contaminated dietary fat, even during the assimilation of a single dose of fat. Presumably, during the post-absorptive synthesis of chylomicrons from pre-chylomicrons, BP is metabolized and removed from the triglyceride oil droplets.

Animals↗
PubMed1984

A simple inexpensive cryogenic storage device for microscopy specimens.

Cryogenic storage devices for microscopy specimens are costly and may require a particular size or type of dewar. Described here is a simple inexpensive storage container for small specimens (less than 5 mm in diameter) which can be constructed in less than 15 min from common laboratory materials. The unit is modular in design, can fit into any thermos type dewar and is reusable.

Freeze Fracturing↗
PubMed1984

Solubility of fatty acids and other hydrophobic molecules in liquid trioleoylglycerol.

The fat solubilities of some long chain fatty acids, alcohols, alkanes, and triacyglycerols, and of some aromatic, chlorinated aromatic, and chlorinated aliphatic hydrocarbons were measured in trioleoylglycerol. Above their melting temperature, all test compounds are theoretically miscible with liquid fat. Below their melting temperature the solubility of all test compounds can be estimated by the equation: log (mole fraction solubility) = (Formula: see text) where delta Sf, the entropy of fusion, can be estimated from chemical structure according to Yalkowsky and Valvani (J. Pharm. Sci. 1980. 69:912-922), and the melting point (Tm) is either known or experimentally determined. For long chain compounds, solubility in trioleoylglycerol dropped precipitously with an increase in melting point. For the aromatic and chlorinated compounds, the drop was more gradual. Since the entropy of fusion of rigid aromatic compounds is approximately 13.5 e.u. at room temperature, their solubility in triacylglycerol is a linear function of melting point.

Alcohols↗
PubMed1983

The production of liquid crystalline product phases by pancreatic lipase in the absence of bile salts. A freeze-fracture study.

The hydrolysis of gum arabic-stabilized trioleylglycerol emulsions by pancreatic lipase was examined by freeze-fracture electron microscopy in the absence of bile salts. A sequence of liquid crystalline product phases was produced during the non-equilibrium conditions of hydrolysis. The morphology of the product phases were pH- and droplet size-dependent. At pH 8.3 the initial product phase was composed of homogeneous spherical vesicles regardless of trioleylglycerol drop size. As the reaction progressed the partially hydrolyzed droplets showed a crystalline 'crust' and a true lamellar phase which was often swollen, giving an isotropic appearance to this phase. Some droplets demonstrated a possible transitory hexagonal phase composed of tubular-lamellar elements in close association with the oil phase. These tubular-lamellar elements graded into a lamellar phase at the aqueous/product interface. A cubic phase was not discernible. At pH 7.0 a single phase was seen which covered the drop surface with an amorphous layered 'crust'. The significance of these phases is discussed in relation to those produced by pure and mixed lipids under equilibrium conditions.

Animals↗
PubMed1982

Hydrolysis of triacylglycerol emulsions by lingual lipase. A microscopic study.

The effect of lingual lipase on four different triacylglycerol emulsions was observed by light microscopy at pH 5-6. The extent of hydrolysis on the microscope slide was determined with the aid of radioactive emulsions or by analyzing the products by gas-liquid chromatography. Artificial emulsions that had been stabilized with amphiphilic lipids gradually coalesced during the unstirred lipase reactions. Gum arabic-stabilized emulsions and human milk fat droplets did not stick to each other or coalesce during lingual lipase hydrolysis. No visible liquid-crystalline product phases, as are seen with pancreatic lipase (Patton, J.S. and Carey, M.C. (1979) Science 204, 145-148), were observed with lingual lipase. The products of lingual lipase activity, protonated fatty acid and diacylglycerol, appear to remain dissolved in the oil phase of the triacylglycerol particle.

Animals↗