In vitro age-dependent incorporation of [1-14C]acetate into lipid subclasses in rat ventral prostate

The [1-14C]acetate incorporation into different lipid subclasses by the rat prostate gland was lineal between 20 and 80 mg of wet tissue. The in vitro [1-14C]acetate incorporation into lipid subclasses was a development-dependent process. The highest values of [1-14C]acetate incorporation into triacylglycerols, free cholesterol and esterified cholesterol were observed at puberty, but radioactivity incorporation into phospholipids was similar in both prepuberty and puberty, then decreasing in maturity. The relationship between triacylglycerols, free cholesterol and esterified cholesterol with respect to total lipids was about 12, 10 and 3.5%, respectively, values being maintained during the animal development. The in vitro [1-14C]acetate incorporation into lipid subclasses in castrated rats decreased considerably as compared with normal rats.     

Seasonal changes in blood serum protein fractions and in activity of AspAT and AlAT in Arabian brood mares and their foals

In 34 pure breed Arabian horses divided into four groups (Gr. I--10 pregnant mares, Gr. II--7 barren mares, Gr. III--10 foals born in 1981, Gr. IV--7 foals born in 1982) seasonal changes in total blood serum protein, its electrophoretic fractions and the activity of AspAT and AlAT were studied. Seasonal cyclicity was found in all groups in the amount of total serum proteins, and alpha 2- and beta 1-globulin fractions. Cyclicity was found in the level of albumin and activity of AspAT in three groups, not Gr. II, and in gamma-globulin, not Gr. IV. beta 2-globulin and AlAT cyclicity was found in two groups and alpha 1-globulin cyclicity was found only in Gr. II. Out of nine indices studied, cyclicity was found in eight of them in pregnant mares, Gr. I; in seven in older foals, Gr. III; in six in the young foals, Gr. IV; and in five in barren mares, Gr. II.     

Incorporation of acetate-1-14C into lipids by the perfused liver of normal, X-irradiated, or partially hepatectomized rats

In order to study lipid metabolism in the liver without interference due to transport from and to the liver the isolated livers of normal, X-irradiated, and partially hepatectomized rats were perfused with acetate-1-(14)C and the distribution of radioactivity in total lipids, total fatty acids, individual lipids, and fatty acids of individual lipids was determined. In X-irradiated animals, an increased incorporation of acetate into many lipids, particularly into cholesterol, was observed. Lipids in the liver of the partially hepatectomized rats exhibited a marked increase in triglyceride content together with a decreased rate of incorporation into all but the phospholipid fractions. It is concluded that the increase usually observed in lipid pontent of the regenerating liver is due to the changes in transcort rather than to changes in synthesis. The changes observed in irradiated liver could be the result of alterations in the metabolism of precursors common to most lipids.     

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.     

Heat-induced changes in the incorporation of [H3]acetate in membrane lipids

The effects of heat treatments at temperatures from 42 to 47 degrees C on the rate of incorporation of [3H]acetate into different classes of lipids have been studied in V-79 Chinese hamster cells. Thermotolerance induction and subtoxic heat treatments decreased the incorporation of [3H]acetate into phospholipids and caused the ratio [3H]cholesterol/[3H]phospholipids to increase several fold, and a positive correlation between heat dose and the ratio [3H]cholesterol/[3H]phospholipids was obtained for subtoxic hyperthermic treatments. The duration of this hyperthermic effect on the incorporation of [3H]acetate into the different lipid fractions was followed in pulse-label experiments. The highest increase of the ratio [3H]cholesterol/[3H]phospholipids was obtained during the first 24 h, but a significant elevation was also present for the 24-72 h pulse-labelled group. Thermotolerance induction was maximal 24 h after the heat treatment and then declined during the next 24 h. The increased [3H]cholesterol/[3H]phospholipid ratio observed in response to hyperthermia resembles the processes that serve to provide homeoviscous adaptation to sustain thermosensitive membrane-located functional groups, in analogy with the mechanisms responsible for thermal adaptation. However, the lack of a positive correlation between thermotolerance induction and the changes in lipid synthesis, for the whole time interval studied, remains to be further explored before any mechanistic interpretation of the data can be found.     

Effects of cholesterol sulfate on lipid metabolism in cultured human keratinocytes and fibroblasts

Effects of cholesterol sulfate on acetate incorporation into lipid fractions were examined in normal human fibroblast and keratinocyte cultures. Inhibition of sterologenesis in normal fibroblast cultures by cholesterol sulfate was less profound than that produced by either lipoprotein-containing serum or 25-hydroxycholesterol. Cholesterol sulfate also inhibited sterologenesis in low density lipoprotein receptor-deficient fibroblasts and inhibited both sterologenesis and 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in keratinocytes. Cholesterol sulfate increased incorporation of acetate into fatty acid-containing lipids in preconfluent cultures of both cell types in lipoprotein-depleted media. Similar effects were not observed either in response to lipoprotein-containing serum or 25-hydroxycholesterol. Cholesterol sulfate had no effect on oleic acid incorporation into diglycerides, triglycerides, or phospholipid fractions; neither did it inhibit acid lipase activity; nor did it inhibit fatty acid oxidation, indicating that cholesterol sulfate does not inhibit catabolism of acyl lipids. Because cholesterol sulfate had similar effects on fatty acid metabolism in steroid sulfatase-deficient fibroblasts lines, desulfation to cholesterol is not a prerequisite. Cholesterol sulfate did not significantly affect incorporation of oleic acid into sterol esters in fibroblast cultures, but in contrast, inhibited sterol esterification in keratinocyte cultures. These data suggest a novel role for cholesterol sulfate as a modulator of cellular lipid biosynthesis.     

INCORPORATION OF LIPIDS INTO SUBCELLULAR FRACTIONS OF BRAIN OF QUAKING MUTANT

The synthesis of lipids and their assembly into subcellular membrane fractions of the myelin deficient Quaking mutant and control brains was studied in 18-, 24- and 41-day-old animals using a double label methodology with¹⁴C and ³H acetate as precursors. As a general procedure, Quaking mutants were injected intracranially with 50 μCi [¹⁴C]acetate and their littermate controls with 300 μCi [³H]acetate. The animals were killed 3 h post-injection, their brains were pooled and subcellular fractions prepared from the common homogenate. An 80-90% decrease in the incorporation of acetate into eleven lipids of myelin in the Quaking mutant was found. This occurred in the face of apparent normal incorporation (relative to microsomes) into lipids of the other main subcellular fractions (nuclear. mitochondrial and synaptosomal) with the exception of decreased incorporation into the myelin-like fraction at 18 and 24 days. Cholesterol and cerebroside were less readily incorporated into Quaking myelin than the other lipids. Although the microsomal synthesis of cholesterol and cerebroside was depressed by about 30% in the Quaking mutant, the incorporation of cholesterol into nuclear, synaptosomal and mitochondrial fractions was unaffected in the mutant. This indicates that sufficient cholesterol is synthesized for the normal assembly of these organelles. In contrast the incorporation of acetate into cholesterol and cerebroside of Quaking myelin was decreased much more than microsomal synthesis. This latter result is consistent with a defect in the process of myclin membrane assembly     

The effect of blood serum on synthesis of lipids in rat thymocytes

The increase in the incorporation of [2-14C]sodium acetate into cholesterol, free fatty acids and rat thymocyte phospholipids during incubation of rat cells in the absence of blood serum was demonstrated. The label incorporation into the thymocytes was found to be activated in an insignificant degree. Rat blood sera obtained 1 hour after gamma-irradiation of animals with doses of 4 and 10 Gr contained the same concentrations of cholesterol and its esters as the intact rat sera. Irradiation of animals did not affect the ability of the sera to inhibit lipid synthesis in thymocytes or the levels of total phospholipids, cholesterol and its esters. Incubation of cells with the blood sera from irradiated rats led to an increase in the cholesterol/cholesterol ester ratio in the incubation mixture.     

Regional adipose tissue metabolism in postmenopausal women after treatment with exogenous sex steroids

In order to evaluate the effect of exogenous sex steroids on adipose tissue metabolism, two groups of postmenopausal women were studied. In one of the groups, the effect of 50 micrograms ethinyl estradiol (EE) was investigated given orally alone and in combination with 10 mg norethisterone acetate (NET). This combination is reminiscent of an old high dose oral contraceptive. In the other group, the effect of 3 mg 17 beta-estradiol was evaluated when administered percutaneously alone and in combination with 300 mg micronized progesterone given orally. These substances and doses were chosen to provide a "physiological" hormonal influence. In the femoral region 50 micrograms EE induced an increase in LPL activity. This elevated LPL value was reversed with the addition of 10 mg NET. Moreover, during treatment with 50 micrograms EE, a decrease in norepinephrine stimulated lipolysis was seen in the abdominal region. The percutaneous administration of 17 beta-estradiol with or without micronized progesterone, however, was inert as regards subcutaneous adipose tissue metabolism. Our findings indicate, therefore, that EE in doses used in oral contraception might promote lipid accumulation in the femoral adipose tissue depot.     

Phospholipid synthesis in isolated alveolar type II cells exposed in vitro to paraquat and hyperoxia.

Isolated alveolar epithelial type II cells were exposed to paraquat and to hyperoxia by gas diffusion through the thin Teflon bottom of culture dishes. After exposure, type II cells were further incubated in the presence of labelled substrates to assess their capacity to synthesize lipids. Hyperoxia alone (90% O2; 5 h) had minor effects on lipid metabolism in the type II cells. At low paraquat concentrations (5 and 10 microM), hyperoxia enhanced the paraquat-induced decrease of [Me-14C]choline incorporation into phosphatidylcholines. The incorporation rates of [Me-14C]choline, [1-14C]palmitate, [1-14C]glucose and [1,3-3H]glycerol into various phospholipid classes and neutral lipids were decreased by paraquat, depending on the concentration and duration of the exposure. The incorporation of [1-14C]acetate into phosphatidylcholines, phosphatidylglycerols and neutral lipids appeared to be very sensitive to inactivation by paraquat. At 5 microM-paraquat the rate of [1-14C]acetate incorporation was decreased to 50% of the control values. The rate of [1-14C]palmitate incorporation into lipids was much less sensitive; it even increased at low paraquat concentrations. At 10 microM-paraquat both NADPH and ATP were significantly decreased. It is concluded that lipid synthesis in isolated alveolar type II cells is extremely sensitive to paraquat. At low concentrations of this herbicide, lipid synthesis, and particularly fatty acid synthesis, is decreased. The effects on lipid metabolism may be partly related to altered NADPH and ATP concentrations.     

Metabolic effects of combined cyproterone acetate and percutaneous 17 beta oestradiol after six and twelve months therapy in 61 patients

The metabolic effects of combined cyproterone acetate (50 mg) and percutaneous 17 beta oestradiol were studied during one year in 61 patients admitted for hyperandrogenia. Before treatment and at 6 and 12 months the following tests were performed: oral glucose tolerance test (OGTT) with insulinemia dosage, determination of total cholesterol, LDL, VLDL and HDL fractions, triglycerids, A1 and B apoproteins, liver function tests: bilirubinemia, alkaline phosphatases, transaminases and gamma glutamyl transferases. The patients' mean age was 27.0 +/- 6.8 years, the body mass index was 22.4 +/- 3.5 kg/m2. After one year of treatment the body mass index was not modified. Blood glucose slightly increased during OGTT; at 6 months this was significant at +30 minutes, at 12 months at +30, +60 and +90 minutes (P less than 0.05). There was no variation in insulinemia during OGTT. Total cholesterol decreased significantly at 6 and 12 months (P less than 0.001), this was associated with a decrease in HDL cholesterol, but without modification of the LDL + VLDL/HDL ratio. Decrease in HDL cholesterol was associated with a significant decrease in A1 apoproteins. No change in triglycerids and in liver function tests was observed at either date. In conclusion the metabolic effects of this association are described. These effects are minimal compared to those observed with cyproterone acetate and ethinyl oestradiol association in the literature. However, attention should be drawn to the possibility of glucose intolerance and decrease in HDL cholesterol and A1 apoproteins.     

Treatment of diminished sexual response associated with the use of oral contraceptives (author's transl)]

Loss of libido associated with the use of oral contraceptives has been studied in 113 women of reproductive age who had taken a combined pill for periods ranging from 1 to 3 years. The patients were divided in four groups. In the first group (I) of 24 women oral contraceptive treatment was discontinued and all women received in intra-uterine contraceptive device (IUCD). The second group (II) of 36 patients, the brand of oral contraceptive was changed. Women in group (III) had their oral contraceptive maintained receiving in addition a mixture of an androgen and an estrogen (methyltestosterone 4 mg and ethynilestradiol 0.002 mg) daily. To women of group (IV) the oral contraceptive was discontinued but the androgen-estrogen mixture was given. All women in this group received an IUCD. Evaluation of the psyco-sexual parameters included changes in libido, time to reach an orgasm, duration and intensity or orgasms. Frequency of intercourse and response to autostimulation was also investigated. Libido was restored in 94.2% of patients in group II, in 97.3% of group III and in 92.8% of group IV. In group I only 55.6% of patients had libido fully restored. In view of the prompt restoration of libido in all groups except in patients discontinuing oral contraceptive therapy, it is suggested that loss of libido in oral contraceptive users has an important physological component which can be overcome probably by psychotherapy. Short term treatment with a mixture of methyltestosterone and ethynilestradiol seems to be highly effective in restoring libido in all patients.     

Seasonal changes in the red blood cell indices in Arabian brood mares and their foals

In 34 pure-bred Arabian horses, divided into four groups (Gr. I, 10 pregnant mares; Gr. II, seven barren mares; Gr. III, 10 foals born in 1981; Gr. IV, seven foals born in 1982), seasonal changes in haemoglobin level, haematocrit value, sedimentation rate, red blood cell number and diameter, percentage of erythroblasts and reticulocytes, and index F were studied. Seasonal cyclicity was found in all groups in the haemoglobin level, haematocrit value and RBC diameter. It was also found in the sedimentation rate (PCV) and in index F, but not for the youngest foals (Gr. IV). For the RBC number the cyclicity is given in both groups of foals, and the erythroblast and reticulocyte percentage only in the older foals (Gr. III). Out of eight indices studied cyclicity was found in all foals in Gr. III, in five of the mares in Groups I and II, and in four of the youngest foals in Gr. IV. There is no difference in the cyclicity of indices studied between pregnant and barren mares.     

Influence of an hypolipemic agent on the incorporation of 14C-labelled precursors in hepatic and serous lipids of the rat]

The action of a new hypolipidemic agent, the calcium diacetone-(ceto 2)-gulonate (D.C.G.Ca), on cholesterol and other lipid compounds biosynthesis has been studied in rats. The incorporation of 14C labelled acetate and mevalonate in liver and serum lipids of control rats has been compared with that of rats previously treated with D.C.G.Ca. No action of this compound has been noted on cholesterol biosynthesis after the mevalonate stage, which is an important result in consideration of therapeutic risks. Otherwise, a significant decrease of 14C-acetate incorporation in triglycerides has been observed in treated rats.     

[14C]Acetate incorporation by cultured normal,familial hypercholesterolemia and Down's syndrome fibroblasts

Fibroblasts from patients with homozygous familial hypercholesterolemia (FH), a disease characterized by accelerated atherogenesis, are known to lack functional low-density-lipoprotein receptors, which ultimately results in increased cholesterol biosynthesis in the cultured cells. [¹⁴C]Acetate incorporation in these cells was compared to that of normal fibroblasts and to fibroblasts from patients with Down's syndrome, a disease in which atherosclerosis is rare. Total [¹⁴C]acetate incorporation did not differ significantly between normal and Down's fibroblasts, nor did its partitioning into the hexane-extractable and aqueous fractions of the cell hydrolysates. [¹⁴C]Acetate incorporation was much greater in FH cells in both the aqueous and hexane-extractable fractions. Preincubation in fetal bovine serum increased acetate incorporation only by FH cells, while 50 μg low-density lipoprotein/ml medium depressed acetate incorporation in all three groups. A C₂₇ sterol, identified by gas chromatography-mass spectrometry as a probable isomer of cholesterol, was present in small amounts in FH fibroblasts, but was not detectable in the normal or Down's cells. The absolute amounts of [¹⁴C]acetate incorporated into the non-sterol lipids were greater in the FH fibroblasts, indicating that these cells may have to synthesize, in addition to cholesterol, other required cellular lipids which are delivered to the normal cells by low-density lipoproteins.     

Anti-fertility effects of embelin in female Sprague-Dawley rats may be due to suppression of ovarian function

Effects of embelin on oestrous cycle, plasma levels of progesterone and oestradiol, and in vitro production of oestradiol and progesterone by mixed ovarian cells was studied. Forty adult (4 months old) regularly cycling female Sprague-Dawley rats were divided into four groups of 10 rats each. Groups I and II (controls) were given 1 ml/kg body weight of physiological saline or corn oil (vehicle). Groups III and IV received 10 mg/kg and 20 mg/kg body weight embelin in corn oil, respectively. Emberlin disrupted the oestrous cycles in Groups III and IV animals, and there was a significant depression in plasma oestradiol (p <0.05) and progesterone (p <0.02) at both 10 and 20 mg/kg body weights, respectively. Isolated mixed ovarian cells from embelin treated rats produced significantly less progesterone and estradiol than controls in vitro. It is concluded that embelin probably interferes with reproductive functions in female rats by suppressing ovarian production of sex steroid hormones.     

Regulation of lipid synthesis in soybeans by two benzoic Acid herbicides

The effects of 3-nitro-2,5-dichlorobenzoic acid (dinoben) and 3-amino-2,4-dichlorobenzoic acid (chloramben) on lipid formation and on the incorporation of various substrates into lipids by intact seeds and subcellular fractions of germinating soybean (Glycine max [L.] Merr. ;Amsoy') were studied. Dinoben (20 mug/ml) inhibited synthesis of total lipids 67%, neutral lipids 73%, glycolipids 51%, and phospholipids 39% in germinating seeds. When polar lipids were analyzed further, inhibition of individual lipid classes was also observed. Chloramben (20 mug/ml) stimulated total lipid synthesis 25%. With the exception of the mitochondrial fraction where malonate thiokinase was absent, dinoben inhibited up to 99% the incorporation of acetate and malonate into lipids, but did not inhibit acetyl-CoA and malonyl-CoA incorporation. Chloramben stimulated the incorporation of all substrates tested into lipids by all fractions except the mitochondrial fraction when malonate was the substrate. When dinoben and chloramben were used in combinations, chloramben did not reverse the inhibitory effect of dinoben.It is concluded that the dinoben inhibitory effect is specific and is associated with the acetate and malonate thiokinase systems. The chloramben effect is stimulatory to either acetyl-CoA carboxylase or fatty acid synthetase or both.     

Marrow culture studies in adult acute myeloid leukemia at diagnosis and during complete remission

We used the human placental conditioned medium stimulated single layer agar culture technique to study the in vitro growth of marrow cells from 62 adult patients with acute myeloid leukemia (AML). Bone marrow cells were cultured from 50 patients at the time of initial diagnosis, 19 patients in early remission and 20 patients during their full complete remission. Marrow cultures from untreated patients exhibited heterogeneous growth patterns ranging from complete growth failure to excessive microcluster formation. We classified the growth patterns into 4 groups: (1) Gr I: normal growth, (2) Gr II: no growth, (3) Gr III: decreased growth, (4) Gr IV: excessive growth of microclusters. At presentation, none had Gr I growth; Gr II growth was observed in 23; Gr III in 14 and Gr IV in 13. A predominance of no growth were seen in M1 and M3 subtypes, while Gr IV growth was more commonly observed in M2 or M4 subtype. We were unable to correlate the culture findings with age or white cell count. The present results not only indicated that AML at diagnosis was characterized by abnormal granulopoiesis but also demonstrated that leukemic progenitor cells were heterogeneous with different capacities to express their proliferating potential in vitro. Except few with decreased growth, the growth characteristics generally returned to normal with successful remission induction. Both Gr II and Gr IV growth patterns were not observed either in early remission or during full complete remission.     

Studies of hyperlipidemia in drug-induced diabetic rats by high-performance liquid chromatography

A hyperlipidemic condition is often associated with diabetes. The possibility that specific serum lipids (i.e., individual triglycerides or cholesterol esters) may be altered in the diabetic state was investigated. Serum lipids from both controls and streptozotocin- and alloxantreated rats were separated into approximately twenty chromatographic fractions by reversed-phase high-performance liquid chromatography; a number of individual triglycerides and cholesterol esters were identified. The methodology described allowed subtle changes in individual lipid components to be detected. Only minor variations in the cholesterol and cholesterol ester fractions were observed between the control and diabetic samples. While not uniform throughout, elevations in the triglyceride fractions occurred in the diabetics. Also, differences in triglyceride content were found to exist between the groups of streptozotocin- and alloxan-treated animals.     

Inhibition of lipid synthesis in isolated rat hepatocytes by serum lipoproteins

The incorporation of labeled acetate into lipids was studied in rat hepatocytes isolated after treatment of liver with collagenase and hyaluronidase. About 60% of the lipid radioactivity was in free cholesterol and 13% was in triglycerides. Acetate incorporation was markedly inhibited when human serum lipoproteins were present in the incubation medium. Very low, high, and low density lipoproteins, at concentrations of 1.0 mg/ml, inhibited acetate incorporation by 70, 55, and 35%, respectively. Chylomicrons, at similar concentrations, did not inhibit acetate incorporation. The distribution of radioactivity into lipid classes was unchanged by the addition of lipoproteins. Lipoproteins did not produce a nonspecific toxic effect on hepatocytes, since their addition did not alter the rate of leucine incorporation into protein. The addition of the delipidated protein from low density lipoprotein or of lecithin in amounts comparable to those present in inhibitory concentrations of lipoproteins failed to diminish acetate incorporation. Artificial cholesterol-lecithin emulsions containing small amounts of free cholesterol did not inhibit lipid synthesis. Although the mechanism for the inhibition of acetate incorporation by lipoproteins is unclear, such effects may play some physiological role in the control of lipid biosynthesis in the liver.