Mechanism of carbon tetrachloride-induced hepatotoxicity. Hepatocellular damage by reactive carbon tetrachloride metabolites.

CCl4-induced liver damage was modeled in monolayer cultures of rat primary hepatocytes with a focus on involvement of covalent binding of CCl4 metabolites to cell components and/or peroxidative damage as the cause of injury. (1) Covalent binding of 14C-labeled metabolites was detected in hepatocytes immediately after exposure to CCl4. (2) Low oxygen partial pressure increased the reductive metabolism of CCl4 and thus covalent binding. (3) [14C]-CCl4 was bound to lipids and to proteins throughout subcellular fractions. Binding occurred preferentially to triacylglycerols and phospholipids, with phosphatidylcholine containing the highest amount of label. (4) The lipid peroxidation potency of CCl4 revealed subtle differences compared to other peroxidative substances, viz., ADP-Fe3+ and cumol hydroperoxide, respectively. (5) CCl4, but not the other peroxidative substances, decreased the rate of triacylglycerol secretion as very low density lipoproteins. (6) The anti-oxidant vitamin E (alpha-tocopherol) blocked lipid peroxidation, but not covalent binding, and secretion of lipoproteins remained inhibited. (7) The radical scavenger piperonyl butoxide prevented CCl4-induced lipid peroxidation as well as covalent binding of CCl4 metabolites to cell components, and also restored lipoprotein metabolism. The results confirm that covalent binding of the CCl3* radical to cell components initiates the inhibition of lipoprotein secretion and thus steatosis, whereas reaction with oxygen, to form CCl3-OO*, initiates lipid peroxidation. The two processes are independent of each other, and the extent to which either process occurs depends on partial oxygen pressure. The former process may result in adduct formation and, ultimately, cancer initiation, whereas the latter results in loss of calcium homeostasis and, ultimately, apoptosis and cell death.

Hepatocyte damage induced by carbon tetrachloride: inhibited lipoprotein secretion and changed lipoprotein composition.

Changes of lipoprotein secretion and composition in response to CCl4 treatment were studied in monolayer cultures of rat primary hepatocytes. (1) CCl4 decreased secretion of very low density lipoproteins (VLDL) by about 85%, while high density lipoprotein (HDL) secretion was less affected (about 40%). The effect was concentration-dependent. (2) CCl4 significantly inhibited secretion of VLDL- and HDL-associated triglycerides and cholesterol esters. VLDL- and HDL-associated cholesterol was not affected, while secretion of phospholipids was increased. (3) Hepatocytes secreted the apolipoproteins B48, B100, E, C, and A-I. CCl4 reduced secretion of apoproteins associated with VLDL by almost 20%, and by about 75% when associated with HDL. The de novo synthesis of apolipoproteins was attenuated by CCl4. (4) CCl4 caused variations in the apolipoprotein composition in VLDL and HDL. CCl4 intoxication of the liver affected the morphology and/or function of the lipoproteins, which drastically impaired their ability to act as transport vehicles for lipids from the liver to the circulation.

Pathogenesis of carbon tetrachloride-induced hepatocyte injury bioactivation of CCI4 by cytochrome P450 and effects on lipid homeostasis.

The CCl4-induced development of liver damage was studied in monolayer cultures of primary rat hepatocytes: (1) CCl4 caused accumulation of triglycerides in hepatocytes following cytochrome P450 induction with beta-naphthoflavone or metyrapone. Ethanol or a high dose of insulin plus triiodothyronine had the same effect. (2) CCl4 increased the synthesis of fatty acids and triglycerides and the rate of lipid esterification. Cholesterol and phospholipid synthesis from acetate was also increased. (3) CCl4 reduced beta-oxidation of fatty acids as assessed by CO2-release and ketone body formation. Hydrolysis of triglycerides was also reduced. (4) The content of unsaturated fatty acids in microsomal lipids was decreased by almost 50% after incubation with CCl4, while saturated fatty acids increased slightly. (5) CCl4 exerted a pronounced inhibitory effect on the exocytosis of macromolecules (albumin), but did not affect secretion of bile acids from hepatocytes.

In vivo and in vitro studies on the regulatory link between 3-hydroxy-3-methylglutaryl coenzyme A reductase and cholesterol 7 alpha-hydroxylase in rat liver.

The activities of 3-hydroxy-3-methylglutaryl CoA reductase (HMGCoA reductase; EC 1.1.1.34), rate-limiting enzyme of cholesterol biosynthesis, and cholesterol 7 alpha-hydroxylase (EC 1.14.13.17), key enzyme of the neutral bile acid synthesis pathway, were measured in the microsomal fraction of rat liver and in rat liver cells to investigate the coordinate regulation of the two pathways. Both enzyme activities exhibited the same diurnal rhythm and responded in a coordinate fashion to fasting or bile acid-feeding (decrease) and to cholestyramine-feeding (increase). Cholesterol-feeding decreased the activity of HMGCoA reductase, increased that of cholesterol 7 alpha-hydroxylase, and concomitantly increased free cholesterol in microsomes. In an ex vivo setting using primary hepatocytes from animals fed a high cholesterol diet the activity of HMGCoA reductase was initially low and that of cholesterol 7 alpha-hydroxylase was elevated. Release of cholesterol into the medium with ongoing incubation caused HMGCoA reductase activity to increase, and that of cholesterol 7 alpha-hydroxylase to decline. Incubation of hepatocytes with a cholesterol-containing lipoprotein fraction stimulated the activity of cholesterol 7 alpha-hydroxylase, but left HMGCoA reductase activity unaffected. The results confirm the idea of a joint regulation of the two key enzymes of cholesterol metabolism in response to the levels of substrate and metabolites, and support the notion that with respect to bile acid and cholesterol levels, respectively, regulation of HMGCoA reductase activity may be secondary to that of cholesterol 7 alpha-hydroxylase. The in vitro studies supply evidence that the effects of cholesterol and bile acid excess or deficiency are direct and do not involve accessory changes of hormone levels or mediators.

Heteroclitic immunization induces tumor immunity.

In tumor transplantation models in mice, cytotoxic T lymphocytes (CTLs) are typically the primary effector cells. CTLs recognize major histocompatibility complex (MHC) class I-associated peptides expressed by tumors, leading to tumor rejection. Peptides presented by cancer cells can originate from viral proteins, normal self-proteins regulated during differentiation, or altered proteins derived from genetic alterations. However, many tumor peptides recognized by CTLs are poor immunogens, unable to induce activation and differentiation of effector CTLs. We used MHC binding motifs and the knowledge of class I:peptide:TCR structure to design heteroclitic CTL vaccines that exploit the expression of poorly immunogenic tumor peptides. The in vivo potency of this approach was demonstrated using viral and self-(differentiation) antigens as models. First, a synthetic variant of a viral antigen was expressed as a tumor antigen, and heteroclitic immunization with peptides and DNA was used to protect against tumor challenge and elicit regression of 3-d tumors. Second, a peptide from a relevant self-antigen of the tyrosinase family expressed by melanoma cells was used to design a heteroclitic peptide vaccine that successfully induced tumor protection. These results establish the in vivo applicability of heteroclitic immunization against tumors, including immunity to poorly immunogenic self-proteins.

Tumor immunity and autoimmunity induced by immunization with homologous DNA.

The immune system can recognize self antigens expressed by cancer cells. Differentiation antigens are prototypes of these self antigens, being expressed by cancer cells and their normal cell counterparts. The tyrosinase family proteins are well characterized differentiation antigens recognized by antibodies and T cells of patients with melanoma. However, immune tolerance may prevent immunity directed against these antigens. Immunity to the brown locus protein, gp75/ tyrosinase-related protein-1, was investigated in a syngeneic mouse model. C57BL/6 mice, which are tolerant to gp75, generated autoantibodies against gp75 after immunization with DNA encoding human gp75 but not syngeneic mouse gp75. Priming with human gp75 DNA broke tolerance to mouse gp75. Immunity against mouse gp75 provided significant tumor protection. Manifestations of autoimmunity were observed, characterized by coat depigmentation. Rejection of tumor challenge required CD4(+) and NK1.1(+) cells and Fc receptor gamma-chain, but depigmentation did not require these components. Thus, immunization with homologous DNA broke tolerance against mouse gp75, possibly by providing help from CD4(+) T cells. Mechanisms required for tumor protection were not necessary for autoimmunity, demonstrating that tumor immunity can be uncoupled from autoimmune manifestations.

The enzyme inducers 3-methylcholanthrene and phenobarbital affect the activities of glucocorticoid hormone-regulated enzymes in rat liver and kidney.

3-Methylcholanthrene, an inducer of P448-type cytochromes (mostly 1A1 and 1A2), and phenobarbital, an inducer of P450-type cytochromes (mostly 2B1 and 2B2), are prototypical for the actions of many xenobiotics. They cause endocrine disruption by affecting, among others, steroid hormone levels. Rats were treated with single bolus doses of 3-methylcholanthrene or phenobarbital, and enzyme activities that are controlled by glucocorticoids were measured in liver and kidney. The activities of the cytosolic enzymes L-alanine aminotransferase, indoleamine 2,3-dioxygenase (L-tryptophan pyrrolase), phosphoenolpyruvate carboxykinase, L-serine dehydratase and L-tyrosine aminotransferase were affected in a similar fashion: an initial activity reduction followed by two overshoots of activity 1 and 2 days after dosing. 3-Hydroxy-3-methylglutaryl coenzyme A reductase, the microsomal key enzyme of sterol synthesis, responded with a temporary reduction of activity only and evidently lost its diurnal rhythm. The time course of these changes is most likely caused by a combination of sub-physiological levels of glucocorticoids plus changes of other regulatory hormones elicited by feed intake, postprandial state, etc. A possible role for a combined action of the arylhydrocarbon (Ah) and glucocorticoid receptors in the effects of 3-methylcholanthrene is also suggested.

Polychlorinated biphenyls affect the activities of gluconeogenic and lipogenic enzymes in rat liver: is there an interference with regulatory hormone actions?

1. The effects of dietary polychlorinated biphenyls (PCBs) (30-2000 ppm) on activities of gluconeogenic (phosphoenolpyruvate carboxykinase-PEPCK, and fructose 1,6-bisphosphatase-FdPase) and lipogenic enzymes (fatty acid synthase-FAS, ATP citrate lyase-ACL, malic enzyme-ME, glucose 6-phosphate dehydrogenase-G6PDH, and 6-phosphogluconate dehydrogenase-PGDH) were studied in livers of the female Sprague-Dawley and Wistar rat. 2. PCB amounts accumulating in the liver reflected the extent of dietary exposure. The Wistar strain was more sensitive to PCBs than the Sprague-Dawley strain. Of the Clophentype PCBs those containing 60 and 64% chlorine displayed the most pronounced effects. 3. Activities of gluconeogenic enzymes (PEPCK and FdPase) were dose-dependently decreased by PCBs, PEPCK being considerably more sensitive. This decrease was also found under conditions where the activity of PEPCK was induced (administration of adrenalin, glucagon or cAMP, feeding high protein diets, starvation). 4. Activities of lipogenic enzymes were induced by PCBs. The increase was much greater with ME, G6PDH and PGDH (up to 10-fold) than with FAS and ACL (approximately 2-fold). PCB effects were dose-dependent, but transient. 5. In cultured hepatocytes basal activities of lipogenic enzymes were induced by PCBs in the absence of hormones. With saturating levels of insulin or triiodothyronine, enzyme activities were also induced, but addition of PCBs resulted in an additive effect. 6. These results suggest that in the female rat PCBs can mimic the actions of certain hormones by affecting either hormone levels, hormone receptor systems or regulatory systems.

Co-expression of transforming growth factor beta and interferon tau during peri-implantation period in the ewe.

The transforming growth factor beta (TGFbeta) family is known to control cell migration, growth, differentiation, function and regulation of extracellular matrix, all of which are required for the process of implantation. Expression of TGFbeta by the conceptus and endometrium was studied during the period of implantation in the ewe. A total of thirty-four ewes were hysterectomized on day 12, 14, 16, 18 or 20 of pregnancy (day 0 = day of estrus). Conceptus (200 mg wet weight) and endometrial (300 mg wet weight) tissues were cultured in vitro in 7 and 10 ml Eagle's minimal essential medium, respectively. The culture media were subjected to a bioassay to determine concentrations of TGFbeta. Conceptus culture media (CCM) were also analyzed for contents of ovine interferon-tau (oIFNr), low molecular weight acidic protein, produced by the trophectoderm between days 8 and 21 of pregnancy. Whole uteri including conceptus(es) and conceptuses (day 16) only were fixed and subjected to immunohistochemical and in situ hybridization studies. Levels of oIFNr produced by conceptuses were the highest on day 16 at 4.4 microg/ml. Concentrations of TGFbeta in day 12, 14, 16, 18 and 20 CCM were 38+/-19, 102+/-56, 862+/-152, 728+/-191 and 336+/-106 pg/ml, respectively, and approximately 90% of TGFbeta activity in CCM was due to TGFbeta1 whereas less than 10% was due to TGFbeta3 based on neutralization with TGFbeta subtype-specific antibodies. Immunohistochemical studies revealed that day 16 conceptuses displayed major staining for TGFbeta1, no beta2 staining and minor staining for beta3. In situ hybridization studies also revealed that day 16 trophectoderm possessed most TGFbeta1 mRNA while day 14 trophectoderm and day 20 chorion/amnion displayed weaker staining for TGFbeta1 mRNA. TGFbeta in day 12, 14, 16, 18 and day 20 endometrial culture media was 156+/-37, 129+/-33, 49+/-22, 62+/-23 and 179+/-40 pg/ml, respectively, and approximately 65% and 35% of the activities were due to TGFbeta1 and beta2, respectively. These results indicate that TGFbeta production by the conceptus coincides with the time when oIFNtau production starts to decline. These observations support the postulate that TGFbeta may play an important role in implantation in the ovine species.

The toxicity of brominated and mixed-halogenated dibenzo-p-dioxins and dibenzofurans: an overview.

Brominated dibenzo-p-dioxins and dibenzofurans can be formed under laboratory conditions by pyrolysis of flame retardants based on polybrominated biphenyls and biphenyl ethers. Their occurrence in the environment, however, is due to combustion processes such as municipal waste incineration and internal combustion engines. As these processes generally take place in the presence of an excess of chlorine, predominantly mixed brominated and chlorinated compounds have been identified so far in environmental samples. Brominated dibenzo-p-dioxins or dibenzofurans bind to the cytosolic Ah receptor about as avidly as their chlorinated congeners and induce hepatic microsomal enzymes with comparable potency. The same holds true for mixed brominated-chlorinated compounds. Gross pathologic symptoms-hypothyroidism, thymic atrophy, wasting of body mass, lethality-also occur at doses that, on a molar concentration basis, are virtually identical to those seen with the chlorinated compounds. Their potency to induce malformations in mice following prenatal exposure is equivalent to that of chlorinated dibenzo-p-dioxins and dibenzofurans. Possible activities as (co)carcinogens and endocrine disrupters have not been evaluated, but are likely to exist. Considering the overall similarity in action of chlorinated and brominated dibenzo-p-dioxins and dibenzofurans, environmental and health assessments should be based on molar body burdens without discrimination for the nature of the halogen.

Priming for T-cell-mediated rejection of established tumors by cutaneous DNA immunization.

DNA immunization has been shown to elicit both antibody and CTL responses against antigens expressed by infectious organisms. Because CTL responses have been implicated in rejection of cancer, we investigated whether DNA immunization by particle bombardment using a gene gun could induce CTL responses that were capable of rejecting tumors in mice. DNA immunization by particle bombardment using genes encoding beta-galactosidase and ovalbumin primed mice to generate CTLs in two genetic backgrounds (DBA/2 and C57BL/6 strains, respectively). DNA immunization was more potent in inducing CTLs than immunization with an optimized regimen of ovalbumin peptide plus immune adjuvant. Immunity induced by DNA immunization protected mice against s.c. challenge with tumors expressing the beta-galactosidase antigen. Tumors were rejected even when DNA immunization was started 3 or 7 days after tumor challenge as tumors were becoming established. Tumor rejection required CD8(+) T cells, confirming a role for CTLs in vivo. These studies show that DNA immunization by particle bombardment can efficiently induce CTL responses that are capable of rejecting even established tumors.

Nutritional regulation of the activities of lipogenic enzymes of rat liver and brown adipose tissue.

Nutrition-induced effects on the activity of enzymes of lipogenesis, fatty acid synthase (FAS: EC 2.3.1.85), ATP citrate lyase (ACL: EC 4.1.3.8), malic enzyme (ME; EC 1.1.1.40), glucose-6-phosphate dehydrogenase (G6PDH: EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (PGDH; EC 1.1.1.44) were investigated in liver and interscapular brown adipose tissue (BAT) of rats. The lipogenic enzymes could be grouped into two categories according to their response to dietary manipulations: FAS and ACL, both key enzymes of lipogenesis, responded fast and strongly to dietary manipulations. ME, G6PDH and PGDH, enzymes which also contribute to metabolic pathways other than lipogenesis, responded in a more sustained and less pronounced fashion. Feed deprivation caused the specific activities of lipogenic enzymes to decline several-fold. Refeeding of previously fasted (up to 3 days) animals increased the activities dramatically (10-to 25-fold) to far above pre-fasting levels ("overshoot"). Repetition of the fasting/refeeding regimen increasingly impaired the ability of both tissues to synthesize overshooting enzyme activities in the subsequent refeeding period. The fasting-induced decline of the activities was prevented when sugars were provided to the animals via drinking water. The sugars displayed different effectivities: sucrose = glucose > fructose > maltose > > lactose. Sugars as the sole nutrient after fasting were also able to induce overshooting enzyme activities. Again, activities of FAS and ACL responded in a more pronounced fashion than the other three enzymes. Transition from feeding one diet to feeding a new diet of different composition led to adaptation of the lipogenic enzyme activities to levels characteristic for the new diet. Replacing a low-carbohydrate with a high-carbohydrate diet proceeded with major alterations of enzyme activities. This process of attaining a new level took up to 20 days and involved pronounced oscillations of the specific activities. In contrast, when a high-carbohydrate diet was replaced with another diet. particular one high in fat, transition to new enzyme activities was completed within 2-3 days and proceeded without oscillations. All dietary manipulations caused more pronounced responses in young (35d-old) than in adult (180d-old) animals.

The response of rat serum lipids to diets of varying composition or contaminated with organochlorine pesticides.

The effects of different diets (high carbohydrate, high protein, high fat) and diets contaminated with polychlorinated biphenyls (PCBs) and/or gamma-hexachlorocyclohexane (lindane) on the levels of serum triglycerides, cholesterol and phospholipids were investigated in Wistar rats. Serum triglyceride levels differed significantly among the diets, while those of cholesterol and phospholipids were much less affected by the diet composition. A change in diet composition resulted in a gradual adaptation to the lipid levels characteristic of the new diet with major variations including oscillations. There was, however, no specific component of a diet that could be associated with any specific change in serum lipids. While feed deprivation decreased the serum lipids (40-65% in 3 days), refeeding the starved animals caused pronounced increases of the lipids that were different among the diets. The response of the triglyceride levels was the strongest (up to 10 times the starvation levels) followed by those of the phospholipids (4-fold) and cholesterol (2.5-fold). Response of the triglyceride levels peaked within 1 or 2 days of refeeding, whereas those of cholesterol and phospholipids took 4 days to reach the maximum. Feeding PCB-contaminated diets increased the serum lipids in a dose-dependent manner (15-250 ppm). Higher PCB concentrations were increasingly inhibitory (350 ppm) or overtly toxic (> 400 ppm). Elevated lipids returned to the starting levels immediately after peaking (triglycerides) or only after several days (cholesterol, phospholipids) but with an earlier onset at lower PCB concentrations. Refeeding starved animals with PCB-contaminated diets also increased the serum lipids dose-dependently. Feeding lindane-containing diets (50-150 ppm) as well as refeeding animals with lindane diets resulted in a considerable increase of the triglyceride levels, while cholesterol and phospholipids increased much less. Higher lindane concentrations (250 ppm) were inhibitory. The outcome on serum lipid levels on feeding diets contaminated with both PCBs and lindane was basically additive.

Toxicokinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin in female Sprague-Dawley rats including placental and lactational transfer to fetuses and neonates.

The toxicokinetics of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in virgin female Sprague-Dawley (S-D) rats, the effects of pregnancy, parturition, and lactation on the distribution and/or redistribution of TCDD, and placental and lactational transfer to fetuses and neonates were investigated. Doses of 5.6 micrograms/kg of 14C-labeled TCDD were given i.v. either to virgin rats or to pregnant rats on Day 18 of gestation and 1 day postparturition, respectively. Virgin females were terminated on Day 1, 2, 4, 8, 16, or 32, pregnant rats on Day 1, 2, 4, or 8, after dosing to collect tissues. Two groups of neonates, which were born either to TCDD-treated or nontreated dams were cross-fostered beginning on the first day after birth to simulate exposure to TCDD either by lactational transfer only or by both placental and lactational transfer. Serum and 18 different tissues were collected from virgin rats to evaluate the kinetic profile of TCDD. Serum and tissue samples from liver, kidney, brown, and white adipose tissue were collected from pregnant and postparturition rats. Liver samples from fetuses and neonates were obtained on Gestational Days 19 and 20, or postnatally on Days 1 and 5. TCDD equivalents were calculated from measurement of radioactivity. The results show that the profile of TCDD distribution in virgin female rats was similar to that in male rats but that the concentration of TCDD in most tissues was higher in females than in males.(ABSTRACT TRUNCATED AT 250 WORDS)

Correlation between toxicity and effects on intermediary metabolism in 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated male C57BL/6J and DBA/2J mice.

Male mice were treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) by gavage. C57BL/6J (C57) mice received 0.03 to 235 micrograms/kg, DBA/2J (DBA) mice 1 to 3295 micrograms/kg. On Day 8 after dosing blood was collected, and livers and kidneys were removed. Body weights and feed intake were not much affected until Day 8 after exposure. Hepatomegaly developed at doses above 3 and 97.5 micrograms/kg in C57 and DBA mice, respectively. Ethoxyresorufin O-deethylase activity was induced in liver with an ED50 of 1.1 and 16 micrograms/kg and in kidney with an ED50 of 65 and 380 micrograms/kg in C57 and DBA mice, respectively. The activity of phosphoenolpyruvate carboxykinase (PEPCK) in livers of both mouse strains was reduced over the entire dose range, displaying a plateau in the dose response at the onset of acute toxicity of TCDD. This enzyme activity was decreased by as much as 80% at the respective lethal doses. PEPCK activity in kidney was not affected. Glucose-6-phosphatase activity (G-6-Pase) in liver was altered only in the lethal dose range with a maximum reduction of about 50%. Serum glucose concentration was reduced over the entire dose range, but the reduction was significant only at doses in which G-6-Pase activity was affected, reaching levels as low as 3 mmol/liter in DBA mice. Tryptophan 2,3-dioxygenase activity was not lowered at any dose of TCDD in either mouse strain, and no increase in serum tryptophan levels was observed. Serum levels of thyroxine (T4) and triiodothyronine (T3) were dose dependently decreased over most of the dose range administered, with T3 levels exactly paralleling T4 levels in both mouse strains. It is concluded that TCDD causes acute toxicity in male C57 and DBA mice by a severe reduction of gluconeogenesis, but, in contrast to rats, it does not affect tryptophan homeostasis. Following administration of TCDD serum T3 levels in the mouse appear to correlate with T4 levels, whereas in the rat they are independent of each other.

Incorporation of first-order uptake rate constants from simple mammillary models into blood-flow limited physiological pharmacokinetic models via extraction efficiencies.

Incorporation of First-Order Uptake Rate Constants from Simple Mammillary Models into Blood-Flow Limited Physiological Pharmacokinetic Models via Extraction Efficiencies. W. L. Roth, L. W. D. Weber, and K. Rozman (1995). Pharm. Res. 263-269. First-order rate constants obtained from classical pharmacokinetic models correspond to mammillary systems in which all of the blood (or plasma) is assumed to be located in a central compartment. In such models the rate at which chemicals are transported out of this pool and into another compartment is the product of the mass of chemical in the central compartment multiplied by a rate constant, which is not limited in magnitude by the blood flow, or the rate at which chemicals from the blood are delivered to the peripheral compartment. Most of the physiologically-based models published to date dispense with some of the information available from mammillary models by assuming that all of the chemical delivered by the flow of blood rapidly equilibrates and can be taken up by the tissue under the control of a "partition coefficient" (Rij = Cj/Ci). We show that the partition coefficient alone does not retain the uptake rate (kji) information available from a classical mammillary model, but that the uptake rate information can be incorporated via unitless extraction efficiency parameters, epsilon j.

The effect of gamma-hexachlorocyclohexane (lindane) on the activities of liver lipogenic enzymes and on serum lipids in rats.

The effect of dietary gamma-hexachlorocyclohexane (lindane) (50-350 ppm, 0.17-1.19 mumol/kg chow) on the activity of enzymes of lipogenesis, viz., fatty acid synthase (FAS; EC 2.3.1.85), citrate cleavage enzyme (CCE; EC 4.1.3.8), malic enzyme (ME; EC 1.1.1.40), glucose-6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (PGDH; EC 1.1.1.44), and on serum lipid levels, was investigated in livers of 35-day-old male Wistar rats. Lindane (150 ppm) caused a substantial decline of enzyme activities within the first 24 h of treatment. The decrease was transient, however, and enzyme activities subsequently recovered despite continuation of lindane feeding. The recovery of enzyme activities was comparatively fast in the case of ME, G6PDH and PGDH, but very slow with FAS and CCE. Activities of lipogenic enzymes decrease when animals are starved, and increase much beyond prestarvation levels upon subsequent refeeding. Lindane in the refeeding diet blunted this overshoot of FAS and CCE activities in a dose-dependent manner. In contrast, activities of Me, G6PDH and PGDH responded to low dietary lindane concentrations with a substantial stimulation of the increase of activity, whereas at high lindane concentrations the overshoot was inhibited. According to their responses to lindane exposure, liver lipogenic enzymes could be grouped into 2 categories with FAS and CCE representing one and ME, G6PDH and PGDH representing the other group. Polychlorinated biphenyls (PCBs) in the diet caused basically opposite changes of the activities of the lipogenic enzymes. Co-administration of lindane and PCBs resulted in an apparent cancellation of effects, suggesting that lindane and PCBs affect fatty acid synthesis at opposite points.(ABSTRACT TRUNCATED AT 250 WORDS)

Lipogenic enzymes of rat liver and adipose tissue. Dietary variations and effect of polychlorinated biphenyls.

The lipogenic enzymes fatty acid synthase (FAS; EC 2.3.1.85), citrate cleavage enzyme (CCE; EC 4.1.3.8), malic enzyme (ME; EC 1.1.1.40), glucose-6-phosphate dehydrogenase (G6PDH; EC 1.1.1.49) and 6-phosphogluconate dehydrogenase (PGDH; EC 1.1.1.44) were investigated in liver and in brown adipose tissue (BAT) of Wistar rats under various dietary conditions and in the presence of 15 to 250 ppm (approximately 0.045-0.75 mumol/kg chow) polychlorinated biphenyls (PCBs). In response to refeeding starved animals, enzyme activities in both tissues increased to above normal levels and thereafter exhibited pronounced oscillations of their activities. The extent of increase depended on the carbohydrate and fat content of the diet. The lipogenic enzymes could be grouped in two categories according to their sensitivity to dietary carbohydrate: FAS and CCE responded faster to smaller changes in dietary composition, while ME, G6PDH and PGDH required larger changes and more time to respond. Diet-induced alterations of enzyme activities were of the same order of magnitude in liver and BAT. They were age-dependent, being more pronounced in young animals. Independent of the type of dietary manipulations, activities changed in a coordinate fashion, i.e., the changes of the activities of all 5 enzymes occurred at similar ratios to each other with an identical time course. Feeding PCB-containing diets resulted in a considerable increase of the activities of the lipogenic enzymes in liver, which was significantly greater with ME, G6PDH and PGDH. The effect was dose-dependent but transient. In liver the response to PCB feeding was identical in male and female animals, whereas in BAT lipogenic activities increased in females, but decreased in males. Refeeding starved animals with a PCB-containing diet led to an additional stimulation of the normal refeeding-induced increase of the enzyme activities in liver and BAT. This PCB-induced increase was 2-fold for FAS and CCE, but up to 15-fold for the other enzymes. All PCB-induced effects were significantly less pronounced in old than in young animals. In primary hepatocytes activities increased in hormone-free medium in the presence of PCBs. While activity was induced in insuline- and triiodothyronine-containing medium, this increase was significantly greater with PCBs present.

A pharmacodynamically responsive model of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) transfer between liver and fat at low and high doses.

We have constructed a pharmacokinetic model for TCDD in the rat, emphasizing its transfer between plasma, lipid, and cytoplasmic compartments of the liver, white adipose tissue (WAT), and brown adipose tissue (BAT). Volumes of the lipid pools are controlled by a submodel of triglyceride (TG) metabolism and transport that responds via a receptor for TCDD in WAT cytoplasm with a Kd of about 2 nM. This submodel, and one for cytochrome P450IA2 induction, allowed us to simulate binding of TCDD to the induced P450IA2 binding sites at low doses (1 ng/kg to 10 micrograms/kg) independently of the decreased feed intake and hyperlipidemia associated with higher doses (20 to 120 micrograms/kg). In low-dose simulations, the induction of cytochrome P450IA2 binding sites for TCDD dominated the redistribution of TCDD between WAT and liver. When simulations were performed using a partitioning model with constant gastrointestinal flows, increased WAT lipolysis driven by reduced feed intake (from 10 to 120 micrograms/kg) is predicted to result in a decreased WAT volume and a sharp drop in the mass of WAT-associated TCDD, while initially increasing the levels of TCDD in liver. However, the observed concentration of TCDD in WAT increased in rats treated with a high dose (72 micrograms/kg) of TCDD. The rise in tissue concentrations could not be explained without incorporating decreased intestinal flows into the gastrointestinal absorption process, which increased the resorption of TCDD. TCDD concentrations in tissue increased only when the relative tissue volumes decreased more rapidly than the whole-body TCDD elimination rate.