Cell-density-dependent release of hepatic lipase from cultured rat hepatocytes

Cultured rat hepatocytes release the enzyme hepatic lipase. In this study we investigated the effect of cell density on this metabolic function under a variety of experimental conditions. The release of hepatic lipase from cultured rat hepatocytes exhibits a cell-density dependence, the secretion per mg cell protein being increased with increasing cell density. When cell density dependence was taken into consideration no significant effect of insulin on the release of hepatic lipase from cultured hepatocytes was observed, whereas glucagon suppressed the release. Glucose stimulating the release of the enzyme, especially in cultures with high cell density.     

Essential fatty acid deficiency in mice is associated with hepatic steatosis and secretion of large VLDL particles

Essential fatty acid (EFA) deficiency in mice decreases plasma triglyceride (TG) concentrations and increases hepatic TG content. We evaluated in vivo and in vitro whether decreased hepatic secretion of TG-rich very low-density lipoprotein (VLDL) contributes to this consequence of EFA deficiency. EFA deficiency was induced in mice by feeding an EFA-deficient (EFAD) diet for 8 wk. Hepatic VLDL secretion was quantified in fasted EFAD and EFA-sufficient (EFAS) mice using the Triton WR-1339 method. In cultured hepatocytes from EFAD and EFAS mice, VLDL secretion into medium was measured by quantifying [(3)H]-labeled glycerol incorporation into TG and phospholipids. Hepatic expression of genes involved in VLDL synthesis and clearance was measured, as were plasma activities of lipolytic enzymes. TG secretion rates were quantitatively similar in EFAD and EFAS mice in vivo and in primary hepatocytes from EFAD and EFAS mice in vitro. However, EFA deficiency increased the size of secreted VLDL particles, as determined by calculation of particle diameter, particle sizing by light scattering, and evaluation of the TG-to-apoB ratio. EFA deficiency did not inhibit hepatic lipase and lipoprotein lipase activities in plasma, but increased hepatic mRNA levels of apoAV and apoCII, both involved in control of lipolytic degradation of TG-rich lipoproteins. EFA deficiency does not affect hepatic TG secretion rate in mice, but increases the size of secreted VLDL particles. Present data suggest that hypotriglyceridemia during EFA deficiency is related to enhanced clearance of altered VLDL particles.     

Lipid metabolism response to a single, prolonged bout of endurance exercise in healthy young men

To discover the alterations in lipid metabolism linked to postexercise hypotriglyceridemia, we measured lipid kinetics, lipoprotein subclass distribution and lipid transfer enzymes in seven healthy, lean, young men the day after 2 h of cycling and rest. Compared with rest, exercise increased fatty acid rate of appearance and whole body fatty acid oxidation by approximately 65 and 40%, respectively (P < 0.05); exercise had no effect on VLDL-triglyceride (TG) secretion rate, increased VLDL-TG plasma clearance rate by 40 +/- 8%, and reduced VLDL-TG mean residence time by approximately 40 min and VLDL-apolipoprotein B-100 (apoB-100) secretion rate by 24 +/- 8% (all P < 0.05). Exercise also reduced the number of VLDL but almost doubled the number of IDL particles in plasma (P < 0.05). Muscle lipoprotein lipase content was not different after exercise and rest, but plasma lipoprotein lipase concentration increased by approximately 20% after exercise (P < 0.05). Plasma hepatic lipase and lecithin:cholesterol acyltransferase concentrations were not affected by exercise, whereas cholesterol ester transfer protein concentration was approximately 10% lower after exercise than after rest (P = 0.052). We conclude that 1) greater fatty acid availability after exercise does not stimulate VLDL-TG secretion, probably because of the increase in fatty acid oxidation and possibly also fatty acid use for restoration of tissue TG stores; 2) reduced secretion of VLDL-apoB-100 lowers plasma VLDL particle concentration; and 3) increased VLDL-TG plasma clearance maintains low plasma TG concentration but is not accompanied by similar increases in subsequent steps of the delipidation cascade. Acutely, therefore, the cardioprotective lowering of plasma TG and VLDL concentrations by exercise is counteracted by a proatherogenic increase in IDL concentration.     

Overexpression of human apolipoprotein A-II in mice induces hypertriglyceridemia due to defective very low density lipoprotein hydrolysis

Two lines of transgenic mice, hAIItg-delta and hAIItg-lambda, expressing human apolipoprotein (apo)A-II at 2 and 4 times the normal concentration, respectively, displayed on standard chow postprandial chylomicronemia, large quantities of very low density lipoprotein (VLDL) and low density lipoprotein (LDL) but greatly reduced high density lipoprotein (HDL). Hypertriglyceridemia may result from increased VLDL production, decreased VLDL catabolism, or both. Post-Triton VLDL production was comparable in transgenic and control mice. Postheparin lipoprotein lipase (LPL) and hepatic lipase activities decreased at most by 30% in transgenic mice, whereas adipose tissue and muscle LPL activities were unaffected, indicating normal LPL synthesis. However, VLDL-triglyceride hydrolysis by exogenous LPL was considerably slower in transgenic compared with control mice, with the apparent Vmax of the reaction decreasing proportionately to human apoA-II expression. Human apoA-II was present in appreciable amounts in the VLDL of transgenic mice, which also carried apoC-II. The addition of purified apoA-II in postheparin plasma from control mice induced a dose-dependent decrease in LPL and hepatic lipase activities. In conclusion, overexpression of human apoA-II in transgenic mice induced the proatherogenic lipoprotein profile of low plasma HDL and postprandial hypertriglyceridemia because of decreased VLDL catabolism by LPL.     

Effects of infection and inflammation on lipid and lipoprotein metabolism: mechanisms and consequences to the host

Infection and inflammation induce the acute-phase response (APR), leading to multiple alterations in lipid and lipoprotein metabolism. Plasma triglyceride levels increase from increased VLDL secretion as a result of adipose tissue lipolysis, increased de novo hepatic fatty acid synthesis, and suppression of fatty acid oxidation. With more severe infection, VLDL clearance decreases secondary to decreased lipoprotein lipase and apolipoprotein E in VLDL. In rodents, hypercholesterolemia occurs attributable to increased hepatic cholesterol synthesis and decreased LDL clearance, conversion of cholesterol to bile acids, and secretion of cholesterol into the bile. Marked alterations in proteins important in HDL metabolism lead to decreased reverse cholesterol transport and increased cholesterol delivery to immune cells. Oxidation of LDL and VLDL increases, whereas HDL becomes a proinflammatory molecule. Lipoproteins become enriched in ceramide, glucosylceramide, and sphingomyelin, enhancing uptake by macrophages. Thus, many of the changes in lipoproteins are proatherogenic. The molecular mechanisms underlying the decrease in many of the proteins during the APR involve coordinated decreases in several nuclear hormone receptors, including peroxisome proliferator-activated receptor, liver X receptor, farnesoid X receptor, and retinoid X receptor. APR-induced alterations initially protect the host from the harmful effects of bacteria, viruses, and parasites. However, if prolonged, these changes in the structure and function of lipoproteins will contribute to atherogenesis.     

Acute regulation of hepatic lipase secretion by rat hepatocytes.

Hepatic lipase is involved in cholesterol uptake by the liver. Although it is known that catecholamines are responsible for the daily variation of enzyme activity, the mechanisms involved are poorly understood. Rat hepatocytes incubated with adrenaline or other Ca(2+)-mobilizing hormones were used as an experimental model. Adrenaline reduced in a similar proportion the secretion of both hepatic lipase and albumin. The effect of adrenaline disappeared completely in cells exposed to cycloheximide. Adrenaline decreased incorporation of [35S]Met into cellular and secreted proteins, but it affected neither degradation of [35S]Met-prelabeled proteins nor the abundance of total and specific (albumin, hepatic lipase, beta-actin) mRNA. Other Ca(2+)-mobilizing agents had the opposite effect on hepatic lipase secretion: it was decreased by vasopressin but was increased by epidermal growth factor. Vasopressin and epidermal growth factor had the opposite effect on [35S]Met incorporation into cellular and secreted proteins, but neither affected hepatic lipase mRNA. The acute effect of adrenaline, vasopressin, and epidermal growth factor on hepatic lipase secretion is the consequence of the effect of these hormones on protein synthesis and is therefore nonspecific.     

Acute inhibition of hepatic lipase and increase in plasma lipoproteins after alcohol intake

Chronic alcohol intake is associated with an increase in fasting plasma high density lipoproteins (HDL). To study alcohol's acute effects on plasma lipoproteins, we measured plasma lipoprotein concentrations and activities of postheparin plasma lipases in nine normolipemic males after ingestion of 40 g of ethanol (as whiskey). After alcohol there was no change in lipoprotein lipase activity but hepatic lipase was decreased to 67% of baseline at 6 hr. There were associated increases in HDL phospholipids (12 mg/dl) and cholesterol (10 mg/dl) resulting in prominence of larger, lipid-enriched HDL particles. Changes were most pronounced in the HDL3 and HDL2a subclasses. Very low density lipoprotein (VLDL) phospholipids and cholesterol were also increased by 13 and 9 mg/dl, respectively, with no significant change in triglycerides. Changes in lipoproteins and lipase were largely reversed 10 hr after alcohol intake. The transient increases in VLDL and HDL lipids after alcohol may result in part from acute inhibition of hepatic lipase activity. The results suggest a role of hepatic lipase in the catabolism of phospholipids of VLDL and possibly HDL.     

PPARalpha activation increases triglyceride mass and adipose differentiation-related protein in hepatocytes

Adipose differentiation-related protein (ADRP) is a lipid droplet-associated protein that is expressed in various tissues. In mice treated with the peroxisome proliferator-activated receptor alpha (PPARalpha) agonist Wy14,643 (Wy), hepatic mRNA and protein levels of ADRP as well as hepatic triglyceride content increased. Also in primary mouse hepatocytes, Wy increased ADRP expression and intracellular triglyceride mass. The triglyceride mass increased in spite of unchanged triglyceride biosynthesis and increased palmitic acid oxidation. However, Wy incubation decreased the secretion of newly synthesized triglycerides, whereas apolipoprotein B secretion increased. Thus, decreased availability of triglycerides for VLDL assembly could help to explain the cellular accumulation of triglycerides after Wy treatment. We hypothesized that this effect could be mediated by increased ADRP expression. Similar to PPARalpha activation, adenovirus-mediated ADRP overexpression in mouse hepatocytes enhanced cellular triglyceride mass and decreased the secretion of newly synthesized triglycerides. In ADRP-overexpressing cells, Wy incubation resulted in a further decrease in triglyceride secretion. This effect of Wy was not attributable to decreased cellular triglycerides after increased fatty acid oxidation because the triglyceride mass in Wy-treated ADRP-overexpressing cells was unchanged. In summary, PPARalpha activation prevents the availability of triglycerides for VLDL assembly and increases hepatic triglyceride content in part by increasing the expression of ADRP.     

Prostaglandins suppress VLDL secretion in primary rat hepatocyte cultures: relationships to hepatic calcium metabolism.

In primary cultures of rat hepatocytes, prostaglandin E2 and prostaglandin D2 (PGE2 and PGD2) inhibited the secretion of very low density lipoprotein (VLDL)-associated apoB, triacylglycerol, and cholesterol. These effects were concentration-dependent and remained apparent for at least 3 days of culture without an effect on the apoB/triacylglycerol ratio of the secreted VLDL. Prostaglandins had no effect on the overall synthesis of triacylglycerol but triacylglycerol accumulated within the cells, without intracellular accumulation of apoB. PGE2, when added to the medium together with glucagon, increased the inhibition of VLDL secretion, compared to that observed with glucagon alone. However, PGE2 did not increase the stimulatory effect of glucagon on ketogenesis. Unlike glucagon, the prostaglandins did not inhibit fatty acid synthesis nor did they stimulate ketogenesis or production of cAMP. Thus, of all the parameters of hepatic lipid metabolism studied, PGE2 and PGD2 selectively affected VLDL. Selective inhibition of VLDL secretion was also observed with the calcium antagonist verapamil. The divalent cation ionophore A23187 also inhibited VLDL release but, in contrast, also inhibited fatty acid and cholesterol synthesis. The results suggest that VLDL secretion is modulated at some optimal cell calcium concentration that may be mediated selectively by agents such as prostaglandins.     

Novel roles of hepatic lipase and phospholipid transfer protein in VLDL as well as HDL metabolism

Objective

Elevated plasma phospholipid transfer protein (PLTP) expression may increase atherosclerosis in mice by reducing plasma HDL and increasing hepatic VLDL secretion. Hepatic lipase (HL) is a lipolytic enzyme involved in several aspects of the same pathways of lipoprotein metabolism. We investigated whether the effects of elevated PLTP activity are compromised by HL deficiency.

Methods and results

HL deficient mice were crossbred with PLTP transgenic (PLTPtg) mice and studied in the fasted state. Plasma triglycerides were decreased in HL deficiency, explained by reduced hepatic triglyceride secretion. In PLTPtg mice, a redistribution of HL activity between plasma and tissue was evident and plasma triglycerides were also decreased. HL deficiency mitigated or even abolished the stimulatory effect of elevated PLTP activity on hepatic triglyceride secretion. HL deficiency had a modest incremental effect on plasma HDL, which remained present in PLTP transgenic/HL^−/− mice, thereby partially compensating the decrease in HDL caused by elevation of PLTP activity. HDL decay experiments showed that the fractional turnover rate of HDL cholesteryl esters was delayed in HL deficient mice, increased in PLTPtg mice and intermediate in PLTPtg mice in an HL^−/− background.

Conclusions

HL affects hepatic VLDL. Elevated PLTP activity lowers plasma HDL-cholesterol by stimulating the plasma turnover and hepatic uptake of HDL cholesteryl esters. HL is not required for the increase in hepatic triglyceride secretion or for the lowering of HDL-cholesterol induced by PLTP overexpression.     

Effects of hormones and pyruvate on the rates of secretion of very-low-density lipoprotein triacylglycerol and cholesterol by rat hepatocytes

The secretion of very-low-density lipoprotein (VLDL) triacylglycerol and cholesterol was determined under various conditions in hepatocytes prepared from rats maintained on a controlled lighting and feeding schedule. The rate of lipogenesis in hepatocytes prepared from rats during the feeding period was 2-3-fold higher than that in cells prepared immediately before the animals had access to food. However, there were no corresponding changes in the rates of secretion of triacylglycerol and cholesterol. Pyruvate alone stimulated triacylglycerol secretion but had no effect on the secretion of cholesterol. Despite its stimulation of lipogenesis, insulin suppressed the secretion of both triacylglycerol and cholesterol. This effect on triacylglycerol secretion was more pronounced when lipogenesis was enhanced in the presence of pyruvate. Thus, insulin may act to alleviate hypertriglyceridaemia, which may arise during periods of increased hepatic lipogenesis. The inhibitory effect of glucagon on cholesterol secretion was much less pronounced than that on the secretion of triacylglycerol. The inhibitory effects of glucagon were reversed by pyruvate on cholesterol secretion differed according to whether glucagon was present or absent. These results suggest that the rate of hepatic VLDL triacylglycerol secretion is not necessarily coupled to the rate of lipogenesis in the liver; nor is there any obligatory coupling between the output of triacylglycerol and cholesterol associated with VLDL.     

ApoB100 is required for increased VLDL-triglyceride secretion by microsomal triglyceride transfer protein in ob/ob mice

Microsomal triglyceride transfer protein (Mttp) is a key player in the assembly and secretion of hepatic very low density lipoproteins (VLDL). Here we determined the effects of Mttp overexpression on hepatic triglyceride (TG) and VLDL secretion in leptin-deficient (ob/ob) mice, specifically in relation to apolipoproteinB (apoB) isoforms. We crossed Apobec1(-/-) mice with congenic ob/ob mice to generate apoB100-only ob/ob mice (A-ob/ob). The obesity phenotype in both genotypes was similar, but A-ob/ob mice had greater hepatic TG content. Administration of recombinant adenovirus expressing murine Mttp cDNA (Ad-mMTP) increased hepatic Mttp content and activity and increased hepatic VLDL-TG secretion in A-ob/ob mice. However, despite equivalent overexpression of Mttp, there was no change in VLDL-TG secretion in ob/ob mice in a wild-type Apobec1 background. Metabolic labeling studies in primary hepatocytes from A-ob/ob mice demonstrated that Ad-mMTP increased triglyceride secretion without changing the synthesis and secretion of apoB100, suggesting greater incorporation of TG into existing VLDL particles rather than increased particle number. Ad-mMTP administration failed to increase hepatic VLDL secretion in lean Apobec1(-/-) mice or controls. By contrast, VLDL secretion increased and hepatic TG content decreased following Ad-mMTP administration to human APOB transgenic mice crossed into the Apobec1(-/-) line. These findings demonstrate that Ad-mMTP increases murine hepatic VLDL-TG secretion only in the apoB100 background, and even then only in situations with either increased hepatic TG accumulation or increased apoB100 expression.     

Increased hepatic lipase activity and increased direct removal of very-low-density lipoprotein remnants in Watanabe heritable hyperlipidaemic (WHHL) rabbits treated with ethinyl oestradiol.

We studied the effects of ethinyl oestradiol on the serum concentrations and metabolism of very-low- and low-density lipoproteins (VLDL and LDL) in Watanabe heritable hyperlipidaemic (WHHL) homozygous rabbits, an animal model for familial hypercholesterolaemia. The results were compared with those in untreated homozygotes as well as in heterozygotes treated or not with ethinyl oestradiol. The gain in body weight was similar in all groups. Treatment with ethinyl oestradiol resulted in the homozygotes in an approx. 80% decrease in the concentrations of lipids and apoprotein B in the d less than 1.019 lipoprotein fraction; those in the LDL fraction did not change. In the heterozygotes, basal serum lipids and apoprotein B levels in the d less than 1.019 fraction were low; ethinyl oestradiol treatment especially affected the LDL fraction (cholesterol -84%, apoprotein B -64%). Turnover experiments with 125I-labelled VLDL revealed that, on treatment with ethinyl oestradiol, the fractional catabolic rate in homozygous rabbits increased 2-fold. The secretion rates of lipids and protein in the d less than 1.019 fraction as estimated after injection of Triton WR-1339 was not decreased. In homozygotes and heterozygotes increases in post-heparin hepatic lipase activity of 62 and 80% respectively were observed, with no changes in lipoprotein lipase activity. We conclude that ethinyl oestradiol induces in homozygous WHHL rabbits a direct removal of VLDL and VLDL remnants from the plasma, apparently due to an increase in hepatic lipase activity.     

Lipoprotein lipase and hepatic lipase activities are differentially regulated in isolated hepatocytes from neonatal rats.

Lipoprotein lipase and hepatic lipase are members of the lipase gene family sharing a high degree of homology in their amino acid sequences and genomic organization. We have recently shown that isolated hepatocytes from neonatal rats express both enzyme activities. We show here that both enzymes are, however, differentially regulated. Our main findings are: (i) fasting induced an increase of the lipoprotein lipase activity but a decrease of the hepatic lipase activity in whole liver, being in both cases the vascular (heparin-releasable) compartment responsible for these variations. (ii) In isolated hepatocytes, secretion of lipoprotein lipase activity was increased by adrenaline, dexamethasone and glucagon but was not affected by epidermal growth factor, insulin or triiodothyronine. On the contrary, secretion of hepatic lipase activity was decreased by adrenaline but was not affected by other hormones. (iii) The effect of adrenaline on lipoprotein lipase activity appeared to involve beta-adrenergic receptors, but stimulation of both beta- and alpha 1-receptors seemed to be required for the effect of this hormone on hepatic lipase activity. And (iv), increased secretion of lipoprotein lipase activity was only observed after 3 h of incubation with adrenaline and was blocked by cycloheximide. On the contrary, decreased secretion of hepatic lipase activity was already significant after 90 min of incubation and was not blocked by cycloheximide. We suggest that not only synthesis of both enzymes, but also the posttranslational processing, are under separate control in the neonatal rat liver.     

The ACAT inhibitor avasimibe increases the fractional clearance rate of postprandial triglyceride-rich lipoproteins in miniature pigs

Previously, we have shown, in vivo, that the acyl coenzyme A: cholesterol acyltransferase (ACAT) inhibitor avasimibe decreases hepatic apolipoprotein (apo) B secretion into plasma. To test the hypothesis that avasimibe modulates postprandial triglyceride-rich lipoprotein (TRL) metabolism in vivo, an oral fat load (2 g fat/kg) containing retinol was given to 9 control miniature pigs and to 9 animals after 28 days treatment with avasimibe (10 mg/kg/day, n=5; 25 mg/kg/day, n=4). The kinetic parameters for plasma retinyl palmitate (RP) metabolism were determined by multi-compartmental modeling using SAAM II. Avasimibe decreased the 2-h TRL (d<1.006 g/mL; S(f)>20) triglyceride concentrations by 34%. The TRL triglyceride 0-12 h area under the curve (AUC) was decreased by 21%. In contrast, avasimibe had no effect on peak TRL RP concentrations, time to peak, or its rate of appearance into plasma, however, the TRL RP 0-12 h AUC was decreased by 17%. Analysis of the RP kinetic parameters revealed that the TRL fractional clearance rate (FCR) was increased 1.4-fold with avasimibe. The TRL RP FCR was negatively correlated with very low density lipoprotein (VLDL) apoB production rate measured in the fasting state (r=-0.504). No significant changes in total intestinal lipid concentrations were observed. Thus, although avasimibe had no effect on intestinal TRL secretion, plasma TRL clearance was significantly increased; an effect that may relate to a decreased competition with hepatic VLDL for removal processes.     

Atherosclerosis is enhanced by testosterone deficiency and attenuated by CETP expression in transgenic mice

In this work, we investigated the impact of testosterone deficiency and cholesteryl ester transfer protein (CETP) expression on lipoprotein metabolism and diet-induced atherosclerosis. CETP transgenic mice and nontransgenic (nTg) littermates were studied 4 weeks after bilateral orchidectomy or sham operation. Castrated mice had an increase in the LDL fraction (+36% for CETP and +79% for nTg mice), whereas the HDL fraction was reduced (-30% for CETP and -11% for nTg mice). Castrated mice presented 1.7-fold higher titers of anti-oxidized LDL (Ox-LDL) antibodies than sham-operated controls. Plasma levels of CETP, lipoprotein lipase, and hepatic lipase were not changed by castration. Kinetic studies showed no differences in VLDL secretion rate, VLDL-LDL conversion rate, or number of LDL and HDL receptors. Competition experiments showed lower affinity of LDL from castrated mice for tissue receptors. Diet-induced atherosclerosis studies showed that testosterone deficiency increased by 100%, and CETP expression reduced by 44%, the size of aortic lesion area in castrated mice. In summary, testosterone deficiency increased plasma levels of apolipoprotein B-containing lipoproteins (apoB-LPs) and anti-OxLDL antibodies, decreased LDL receptor affinity, and doubled the size of diet-induced atherosclerotic lesions. The expression of CETP led to a milder increase of apoB-LPs and reduced atherosclerotic lesion size in testosterone-deficient mice.     

Head group specificity in the requirement of phosphatidylcholine biosynthesis for very low density lipoprotein secretion from cultured hepatocytes

We have demonstrated that hepatic very low density lipoprotein (VLDL) secretion requires active phosphatidylcholine (PC) synthesis via either the CDP-choline pathway or phosphatidylethanolamine (PE) methylation pathway (Yao, Z., and Vance, D.E. (1988) J. Biol. Chem. 263, 2998-3004). In the present work, the head group specificity of phospholipid synthesis required for lipoprotein secretion was investigated in cultured hepatocytes isolated from choline-deficient rats. When N-monomethylethanolamine (0.1 mM) or N,N-dimethylethanolamine (0.1 mM) was added to the culture medium, the cells synthesized correspondingly phosphatidylmonomethylethanolamine (PMME) or phosphatidyldimethylethanolamine (PDME). However, the synthesis of PDME could correct the impaired VLDL secretion only to a limited extent, whereas the synthesis of PMME inhibited VLDL secretion. Although dimethylethanolamine did not promote VLDL secretion as well as choline, dimethylethanolamine altered the increased triacylglycerol synthesis in the choline-deficient cells as effectively as choline. Supplementation of the culture medium with ethanolamine (0.1 mM) had little effect on cellular PE or PC levels, nor was normal VLDL secretion resumed. However, the amounts of cellular PC and PE were both decreased when the medium was supplemented with N-monomethylethanolamine or N,N-dimethylethanolamine. These results suggest that the choline head group moiety of PC is specifically required for normal VLDL secretion and cannot be replaced with ethanolamine, monomethylethanolamine, or dimethylethanolamine. In addition, the impaired VLDL secretion from the choline-deficient hepatocytes could also be corrected by supplementation of betaine (0.2 mM) and homocysteine (0.2 mM), indicating the utilization of a methyl group from betaine for PC formation via methylation of PE.     

Effects of threonine-poor apolipoproteins on post-heparin plasma hepatic triacylglycerol lipase and lipoprotein lipase

Whole-irradiated rabbit pre-heparin plasma had an important inhibitory effect on hepatic triacylglycerol lipase and lipoprotein lipase activities, whereas control rabbit pre-heparin plasma slightly inhibited hepatic triacylglycerol lipase activity at a high concentration and enhanced lipoprotein lipase activity. As some apolipoproteins were known to modulate these two lipolytic enzymes, the inhibitory effects of irradiated rabbit plasma were investigated in apolipoproteins. Three apolipoproteins, with isoelectric points of about 6.58, 6.44 and 6.12, characterized by their low content in threonine (threonine-poor apolipoproteins) were produced in high concentrations in rabbit VLDL and HDL after irradiation. The effects of these apolipoproteins on control rabbit post-heparin plasma hepatic triacylglycerol lipase and extrahepatic lipoprotein lipase were studied. Threonine-poor apolipoproteins substantially inhibited the hepatic triacylglycerol lipase activity and enhanced the apolipoprotein C-II-stimulated activity of lipoprotein lipase. The amounts of these apolipoproteins in triacylglycerol-rich lipoprotein particles may determine the lipolytic activity of lipoprotein lipase and hepatic triacylglycerol lipase in triacylglycerol hydrolysis. The existence of another inhibitor of lipoprotein lipase remains to be determined.     

Loss of the lipoprotein lipase activating ability of rat serum after administration of some fatty liver inducing drugs.

The effects of the administration of different fatty liver inducing drugs on the serum lipoprotein lipase activating ability was investigated in rats. Addition of serum from 2-mercaptoethanol-, 2-mercaptoacetate-, ethionine- or D-galactosamine- treated rats failed to activate heart and adipose tissue lipoprotein lipase from control rats. The activating effect of serum was only slightly reduced in isopropanol-treated rats, whereas it was found unaffected in ethanol-treated ones. Electrophoresis of the lipoproteins and of the very low density lipoproteins (VLDL) fraction of sera from 2-mercaptoethanol-, 2-mercaptoacetate-, isopropanol-, ethionine- and D-galactosamine-treated rats suggest that the lack of lipoprotein lipase activation ability of these sera is most probably related to the impairing effects of these drugs upon VLDL metabolism, i.e. reduction of VLDL secretion in the case of 2-mercaptoethanol, 2-mercaptoacetate and isopropanol, production of abnormal VLDL in the case of D-galactosamine and both decreased VLDL secretion and production of abnormal VLDL in the case of ethionine.     

Cholesterol is required for secretion of very-low-density lipoprotein by rat liver.

To study potential effects of hepatic cholesterol concentration on secretion of very-low-density lipoprotein (VLDL) by the liver, male rats were fed on unsupplemented chow, chow with lovastatin (0.1%), or chow with lovastatin (0.1%) and cholesterol (0.1%) for 1 week. Livers were isolated from these animals and perfused in vitro, with a medium containing [2-14C]acetate, bovine serum albumin and glucose in Krebs-Henseleit buffer, and with an oleate-albumin complex. With lovastatin feeding, the hepatic concentrations of cholesteryl esters and triacylglycerols before perfusion were decreased, although free cholesterol was unchanged. However, hepatic secretion of all the VLDL lipids was decreased dramatically by treatment with lovastatin. Although total secretion of VLDL triacylglycerol, phospholipid, cholesterol and cholesteryl esters was decreased, the decrease in triacylglycerol was greater than that in free cholesterol or cholesteryl esters, resulting in secretion of a VLDL particle enriched in sterols relative to triacylglycerol. In separate studies, the uptake of VLDL by livers from control animals or animals treated with lovastatin was measured. Uptake of VLDL was estimated by disappearance of VLDL labelled with [1-14C]oleate in the triacylglycerol moiety, and was observed to be similar in both groups. During perfusion, triacylglycerol accumulated to a greater extent in livers from lovastatin-fed rats than in control animals. The depressed output of VLDL triacylglycerols and the increase in triacylglycerol in the livers from lovastatin-treated animals was indicative of a limitation in the rate of VLDL secretion. Addition of cholesterol (either free cholesterol or human low-density lipoprotein) to the medium perfusing livers from lovastatin-fed rats, or addition of cholesterol to the diet of lovastatin-fed rats, increased the hepatic concentration of cholesteryl esters and the output of VLDL lipids. The concentration of cholesteryl esters in the liver was correlated with the secretion of VLDL by the liver. These data suggest that cholesterol is an obligate component of the VLDL required for its secretion. It is additionally suggested that cholesteryl esters are in rapid equilibrium with a small pool of free cholesterol which comprises a putative metabolic pool available and necessary for the formation and secretion of the VLDL. Furthermore, the specific radioactivity (d.p.m./mumol) of the secreted VLDL free cholesterol was much greater than that of hepatic free cholesterol, suggesting that the putative hepatic metabolic pool is only a minor fraction of total hepatic free cholesterol.