Enhanced Lipid Synthesis Research Articles

Mitochondrial proliferation in the permanent vs. temporary cold: enzyme activities and mRNA levels in Antarctic and temperate zoarcid fish.

Adjustments in mitochondrial properties and capacities are crucial in acclimatization to seasonal cold and in evolutionary cold adaptation of marine ectotherms. Although long-term compensatory increments in aerobic capacity of fish tissues have frequently been described in response to cold, much less is known about transitional phases and gene expression patterns involved. We investigated the time course of adjustment to acute cold in liver of eurythermal eelpout Zoarces viviparus. Whereas citrate synthase (CS) activity rose progressively in liver, cytochrome c oxidase (COX) activity was not altered during cold acclimation. Species-specific RNA probes were used to determine mRNA levels. CS mRNA (nuclear encoded) displayed a delayed, transient increase in response to cold, such that transcript levels did not parallel the change in enzyme activity. The enzyme activities and mRNA levels in the confamilial Antarctic Pachycara brachycephalum indicate cold compensation of CS activity in this cold-adapted species. The ratio of CS and COX activities was elevated in acclimation and adaptation to cold, indicating enhanced citrate synthesis over respiratory chain capacities in cold-adapted liver mitochondria. This may support enhanced lipid synthesis typically found in cold. The ratio of enzyme activity and transcript levels differed largely between Z. viviparus populations from the Baltic and North Seas, indicating the influence of unidentified parameters other than temperature. Transcript levels may not be tightly correlated with enzyme activities during thermal adaptation and thereafter. The time course of the acclimation process indicates that regulation at the translational and posttranslational levels predominates in adjustment to moderate thermal challenges.

Implementation of fatty acid carriers to skin irritation and the epidermal barrier.

Acute perturbations are followed by barrier repair and enhanced lipid synthesis, as well as cellular fatty acid trafficking, yet irritation of the skin may be induced by repeat disturbance of barrier function. Recently, new insights in cellular fatty acid transport and metabolism have evolved with respect to skin irritation and barrier disturbances: (1) Employing sodium dodecyl sulfate, skin irritation is accompanied by the induction of an epidermal (E) cytosolic fatty acid binding protein (FABP) associated with enhanced barrier repair. Whether E-FABP contributes to the water barrier function in normal skin remains to be elucidated; (2) Cutaneous inflammation, as it occurs in irritant contact dermatitis, can be reduced by peroxisome proliferating activated receptor (PPAR) agonists, such as linoleic acid, with clinical effects comparable to that of glucocorticoids; (3) PPARalpha agonists accelerate barrier recovery and enhance lamellar body synthesis, neutral lipid synthesis, in particular that of ceramides and cholesterol; (4) PPARalpha agonists increase the minimal erythema dose in UVB-irradiated human skin. This review provides a brief overview of the current understanding of mammalian fatty acid (FA) metabolism with respect to epidermal barrier abrogation and repair, including new insights into cellular FA transport and metabolism.

Fasting and Postprandial Overproduction of Intestinally Derived Lipoproteins in an Animal Model of Insulin Resistance: EVIDENCE THAT CHRONIC FRUCTOSE FEEDING IN THE HAMSTER IS ACCOMPANIED BY ENHANCED INTESTINAL DE NOVO LIPOGENESIS AND ApoB48-CONTAINING LIPOPROTEIN OVERPRODUCTION

Insulin-resistant states are characterized by hypertriglyceridemia, predominantly because of overproduction of hepatic very low density lipoprotein particles. The additional contribution of intestinal lipoprotein overproduction to the dyslipidemia of insulin-resistant states has not been previously appreciated. Here, we have investigated intestinal lipoprotein production in a fructose-fed hamster model of insulin resistance previously documented to have whole body and hepatic insulin resistance, and hepatic very low density lipoprotein overproduction. Chronic fructose feeding for 3 weeks induced significant oversecretion of apolipoprotein B48 (apoB48)-containing lipoproteins in the fasting state and during steady state fat feeding, based on (a) in vivo Triton WR1339 studies of apoB48 production as well as (b) ex vivo pulse-chase labeling of intestinal enterocytes from fasted and fed hamsters. ApoB48 particle overproduction was accompanied by increased intracellular apoB48 stability, enhanced lipid synthesis, higher abundance of microsomal triglyceride transfer protein mass, and a significant shift toward the secretion of larger chylomicron-like particles. ApoB48 particle overproduction was not observed with short-term fructose feeding or in vitro incubation of enterocytes with fructose. Secretion of intestinal apoB48 and triglyceride was closely linked to intestinal enterocyte de novo lipogenesis, which was up-regulated in fructose-fed hamsters. Inhibition of fatty acid synthesis by cerulenin, a fatty acid synthase inhibitor, resulted in a dose-dependent decrease in intestinal apoB48 secretion. Overall, these findings further suggest that intestinal overproduction of apoB48 lipoproteins should also be considered as a major contributor to the fasting and postprandial dyslipidemia observed in response to chronic fructose feeding and development of an insulin-resistant state.

Diminished Hepatic Response to Fasting/Refeeding and Liver X Receptor Agonists in Mice with Selective Deficiency of Sterol Regulatory Element-binding Protein-1c

Two treatments, fasting/refeeding and administration of liver X receptor (LXR) agonists, elevate the mRNA for sterol regulatory element-binding protein-1c (SREBP-1c) and enhance lipid synthesis in liver. These treatments do not affect the mRNA for SREBP-1a, an alternative transcript from the same gene. Through homologous recombination, we eliminated the exon encoding SREBP-1c from the mouse genome, leaving the SREBP-1a transcript intact. On a normal diet, livers of SREBP-1c(-/-) mice manifested reductions in multiple mRNAs encoding enzymes of fatty acid and triglyceride synthesis, including acetyl-CoA carboxylase (ACC) and fatty acid synthase (FAS). In contrast, SREBP-1c(-/-) livers showed a compensatory increase in hepatic SREBP-2 mRNA, accompanied by increased mRNA levels for cholesterol biosynthetic enzymes. In fasted/refed animals, ACC and FAS mRNAs rose, but not to the same extent as in wild-type livers. The refeeding-induced increase in SREBP-1c(-/-) mice was greater than in mice lacking SREBP cleavage-activating protein (SCAP), in which all nuclear SREBPs are absent. Thus, SREBP-2 and/or SREBP-1a can substitute partially for SREBP-1c in permitting an insulin-mediated increase in ACC and FAS mRNAs. In contrast, mRNAs for several other lipogenic enzymes (glucose-6-phosphate dehydrogenase, malic enzyme, glycerol-3-phosphate acyltransferase, and stearoyl-CoA desaturase-1) showed a complete failure of the normal inductive response to refeeding, indicating specific reliance on SREBP-1c. Moreover, these mRNAs, as well as multiple other lipogenic mRNAs, showed a markedly blunted response to the LXR agonist T090137, indicating an essential role of SREBP-1c in the LXR response.

Characterization of acetate metabolism in tumor cells in relation to cell proliferation: Acetate metabolism in tumor cells

To reveal the metabolic fate of acetate in neoplasms that may characterize the accumulation patterns of [1- 11C]acetate in tumors depicted by positron emission tomography. Four tumor cell lines (LS174T, RPMI2650, A2780, and A375) and fibroblasts in growing and resting states were used. In uptake experiments, cells were incubated with[1- 14C]acetate for 40 min. [ 14C]CO 2 was measured in the tight-air chamber, and the metabolites in cells were identified by thin layer chromatography and paper chromatography. The glucose metabolic rate of each cell line was measured with [2,6- 3H]2-deoxy-glucose (DG), and the growth activity of each cell line was estimated by measuring the incorporation of [ 3H]methyl thymidine into DNA. Compared with resting fibroblasts, all four tumor cell lines showed higher accumulation of 14C activity from [1- 14C]acetate. These tumor-to-normal ratios of [1- 14C]acetate were larger than those of DG. Tumor cells incorporated 14C activity into the lipid-soluble fraction, mostly of phosphatidylcholine and neutral lipids, more prominently than did fibroblasts. The lipid-soluble fraction of 14C accumulation in cells showed a positive correlation with growth activity, whereas the water-soluble and CO 2 fractions did not. These findings suggest that the high tumor-to-normal ratio of [1- 14C]acetate is mainly due to the enhanced lipid synthesis, which reflects the high growth activity of neoplasms. This in vitro study suggests that [1- 11C]acetate is appropriate for estimating the growth activity of tumor cells.

Primary production, calcification and macromolecular synthesis in a bloom of the coccolithophore Emiliania huxleyi in the North Sea

Photosynthesis, calcification and the patterns of carbon (C) incorporation into different biomolecules were investigated during a bloom of the coccolithophore Emiliania huxleyi in the North Sea during June and July 1994. The bloom was confined to an area of ca 3000 km2 centred at 59'50' N. 00°42'E and characterized by enhanced thermal stability and low nitrate concentrations in the upper mlxed layer. Surface E. huxleyi densities within the bloom area ranged between 1 and 6 X 106 cells I-' The mesoscale distribution of E. huxleyi abundance suggested that the bloom formation was related to the presence of low concentrations of nitrate rather than phosphate. The bloom was sampled during an early stage of its development, as indicated by the low calcite-C levels (c50 my C m-3], the low calciteC to particulate organlc carbon (POC) ratlo ( ~ 0 . 2 5 ) and the low density of detached coccoliths (2 to 3 X 10' m1 l ) . Reduced levels of chlorophyll a (<45 mg m-2) and productivity (<1.2 g C m-L d-') were measured in the coccolithophore-rich waters as compared to stations outside the bloom area. Typical calcification rates within the bloom were 135 mg C m-' d-', representing up to 20% of the total C incorporation. Relative C incorporation into lipids in coccolithophore-rich waters was 1.5 times higher than outside the bloom area. Most of the recently synthesized lipid (70 to 90%) belonged to the neutral lipid fraction. The enhanced lipid synthesis resulted in a higher lipid content of the particulate matter. The relationships between irradiance and photosynthate partitioning consistently showed that C is preferentially channeled into the protein fraction at low light levels, whereas the relative synthesis of lipids increases only at high irradiances. These metabolic responses are discussed in relation to the development of E. huxleyi-rich assemblages. It is concluded that the observed patterns of C incorporation represent a general strategy of energy utilization that is also attributable to other groups of marine phytoplankton.

Progressive differentiation of human sebocytes in vitro is characterized by increasing cell size and altering antigen expression and is regulated by culture duration and retinoids

Increasing cell size, lipid accumulation, and altered antigen expression are features of sebaceous differentiation in vivo. Enhanced lipid synthesis with progressive differentiation is also present in cultured human sebocytes. This study was conducted to investigate the evolution of cell size and antigen expression of human sebocytes with progressive differentiation in vitro. Subconfluent human sebocyte cultures were examined for sebocyte differentiation evaluated on cytocentrifuge preparations by light microscopy and classified in stages according to morphological criteria described for sebocytes in vivo. Rates of 5.1 +/- 2.2% undifferentiated sebocytes, 29.2 +/- 4.9% early differentiated, 20.7 +/- 4.1% advanced differentiated, 37.6 +/- 6.4% fully differentiated, and 5.9 +/- 1.9% mature sebocytes were calculated in secondary cultures. The size of cultured sebocytes measured by computer-assisted planimetry significantly increased with progressive differentiation up to 4-5.5 times. The low rates of mature sebocytes and the only moderate increase of their size with progressive differentiation indicate an incomplete terminal differentiation in vitro. Sebocytes were subsequently stained with a series of monoclonal antibodies (mAb) to determine antigen expression using the alkaline phosphatase anti-alkaline phosphatase technique. The number of sebocytes labeled with the anti-keratin mAb CK8.12 and KL1, and the mAb 34D11 (82 kD protein) increased with progressive differentiation; significant differences were found after comparing early and advanced differentiated sebocytes. Sebocytes were positively stained with the anti-keratin mAb 6B10 (K 4), RPN1162 (K 7), CK13 (K 13), RPN1165 (K 19), CK8.60, and the mAb 115F5 (MAM-6c), OM-1 (sebaceous gland antigen), and 24F10 (basic polypeptides) only at late-stage differentiation. The expression of keratins 4, 7, 13, and 19 was confirmed by gel electrophoresis and immunoblotting. The data obtained were used to study the effects of the duration of cultivation and of the retinoids isotretinoin and tretinoin on sebocyte differentiation in vitro. Subcultivation of sebocytes upregulated, and treatment with isotretinoin but not with tretinoin downregulated labeling with mAb which recognize indicating progressive and late-stage differentiation.

Oligodendrocyte-substratum adhesion activates the synthesis of specific lipid species involved in cell signaling.

Ovine oligodendrocytes (OLGs) undergo biochemical and morphological changes following attachment to polylysine. Autoradiographs of two-dimensional thin-layer chromatograms of [14C]Gal-labeled OLG cultures revealed that attachment of OLGs to a polylysine substratum and their subsequent morphological differentiation is accompanied by an increased synthesis of multiple forms of galactosylceramide, sulfogalactosylceramide, and both sulfogalactosyl- and galactosyl-diglycerides, together with an array of complex sialoglycosphingolipids, predominantly GM2 ganglioside. As previously reported, overall lipid synthesis measured by [14C]acetate incorporation into glycerophosphatides, sphingomyelin, and neutral lipids also increased dramatically for up to 60 days (last time point examined) following OLG-substratum adhesion, reflecting membrane growth. Attachment was associated with a rapid augmentation in the synthesis of ethanolamine plasmalogen from 12 to 27% within 24 hr to reach a 35% plateau at 30 days and remain constant thereafter. In contrast, the plasmalogen content of phosphatidylcholine remained constant at 3-5%. This rapid increase in lipid synthesis (especially in the ethanolamine plasmalogen content following attachment) closely paralleled increased diacylglycerol (DAG) production and protein kinase C-dependent phosphorylation of both myelin basic protein and 2',3'-cyclic nucleotide phosphohydrolase. Labeling studies indicated that the major source of [3H]arachidonate-labeled DAG following attachment was from phosphatidylinositol turnover (and to a lesser extent phosphatidylcholine) rather than polyphosphoinositides or plasmalogens. Enhanced lipid synthesis is not only required for the production of membranes in these myelin-producing cells but is also a source of second messengers required in the posttranslational modification of key myelin and cellular proteins.

Dimethyl sulfoxide enhances lipid synthesis and secretion by long-term cultures of adult rat hepatocytes

Dimethyl sulfoxide (DMSO) was tested for its effects on lipid metabolism of long-term cultures of adult rat hepatocytes. The addition of 1% DMSO to 3T3-hepatocyte cultures was not toxic to cells and in fact treated cultures maintained better their characteristic morphology for up to 14 days of exposure. DMSO treatment increased 2–3 fold the de novo synthesis of total lipids from [ 14C]acetate. The analysis by thin layer chromatography of cellular and secreted lipids revealed that DMSO increased the levels of cellular triglycerides, phospholipides and free and sterified cholesterol at 7 days of exposure while at 14 days there was also a 2–3-fold increase in medium secreted lipids. Additionally, DMSO increased the activity of glycerol-phosphate dehydrogenase, a marker enzyme of glycerolipid synthesis, by > 50% at either 7 or 14 days of exposure. These results show that 1% DMSO not only is not detrimental to cultured hepatocytes but also enhances lipid synthesis and secretion, both hepatic-differentiated functions.

Stimulation of fatty acid uptake and triglyceride synthesis in human cultured skin fibroblasts and adipocytes by a serum protein

Lipoprotein deficient serum has been shown to enhance lipid synthesis in cultured normal human skin fibroblasts incubated in the presence of oleate-albumin. The factor responsible is nondialyzable and trypsin sensitive. The stimulation is proportional to the concentration of lipoprotein deficient serum in the media and is present at all oleate concentrations and incubation times assayed. The protein has been partially purified by column chromatography to yield a Peak II fraction which stimulates triglyceride synthesis in both fibroblasts and isolated human adipocytes. The stimulation is dependent on the concentration of protein fraction and increases to an apparent saturation level of 200% in fibroblasts. Triglyceride synthesis, however, increases to a much greater extent in adipocytes and did not demonstrate saturation at the maximum Peak II protein concentration assayed. These results suggest that human serum contains a protein which stimulates fatty acid uptake and esterification by adipose tissue.

Vitamin E prevents the platelet abnormalities induced by estrogen in rat

Platelet lipid biosynthesis in relation to aggregation has been studied in female rats treated with ethynylestradiol and fed laboratory chow or a vitamin E-deficient diet. In both normal and vitamin E-deficient rats, administration of ethynylestradiol highly significantly (p < .001) increased the biosynthesis of total lipids but mostly of lanosterol (+ dihydrolanosterol) by thirteen-fold in normal rats and by nine-fold in vitamin E-deficient rats. The increased lipid synthesis was associated with a higher response of platelets to thrombin-induced aggregation. Concomitant administration of α-tocopherol acetate in both normal and vitamin E-deficient rats depressed markedly the enhanced lipid synthesis and aggregation induced by estrogen. Administration of ethynylestradiol lowered considerably the level of vitamin E in plasma but not in platelets. Treatment by tocopherol partly corrected the low plasma level of vitamin E resulting from estrogen administration. In vitro addition of lanosterol to platelets highly significantly increased the response of platelets to thrombin- and ADP-induced aggregation. This hyperaggregability was almost entirely inhibited by preincubation of platelets with tocopherol acetate. In the present in vivo and in vitro studies, α-tocopherol was able to neutralize most of the adverse effects of estrogen on blood platelets.

Effect of β-aminopropionitrile on lipid metabolism by bone cell cultures: Biochemistry and morphology

The effects of β-aminopropionitrile (BAPN) on [ 14C]acetate monitored by bone cells derived from newborn rat calvaria were examined by biochemical techniques and light and electron microscopic procedures. The effects of BAPN depends upon both concentration and time of exposure to the lathyrogen. BAPN tends to stimulate the incorporation of [ 14C]acetate into most lipids at lathyrogen concentrations up to 40 m M. At 100 m M the incorporation is significantly reduced. Prolonging the time of exposure generally enhances lipid synthesis. The major exception is the reduced synthesis of cholesterol esters. At BAPN concentrations higher than 5 m M, vesicles, which appear to be dilated endoplasmic reticulum, are formed in the cell cytoplasm. Further, the BAPN treated cells do not appear to adhere to each other in the same manner as untreated cells suggesting changes in the cell glycocalyx. This study demonstrates BAPN induced alterations in lipogenesis in cells which have the potential to calcify. These alterations suggest an increase in cell membrane area giving rise to the vesicles and dilated endoplasmic reticulum.

Stimulation of Synthesis and Release of Lipids in Tumor Cells Under Attack by Antibody Plus Complement

Abstract Resistance to antibody-complement- (Ab-C) mediated killing of guinea pig hepatoma cells (line-1 and line-10) correlates with the ability of the cells to synthesize lipids, but not DNA, RNA, protein, or complex carbohydrate. We therefore studied the synthesis and release of macromolecules (especially lipids) by tumor cells under attack by Ab plus C. Ab-sensitized tumor cells (TA) incorporated increased amounts of acetate, glycerol, or fatty acids into complex lipids compared to control cells within 5 to 10 min after the addition of guinea pig C (GPC) to the cells (TAC). TA, TC, or TAΔC (Ab plus heat-inactivated GPC) did not show this effect. No enhancement of DNA, RNA, or protein synthesis in TAC as compared with control cells was noted. Line-1 cells, which are killed by GPC when sensitized with excess anti-Forssman Ab (TAxs), demonstrated enhanced lipid synthesis within 1 to 3 min after the addition of GPC to TAxs, before measurable killing of the cells had occurred.

Stimulation of the Synthesis and Release of Lipids in Tumor Cells under Attack by Antibody and C

Antibody-sensitized line-1 or line-10 tumor cells treated with GPC (TAC) incorporated fatty acids into complex cellular lipids and released increased amounts of fatty acids within 5 to 10 min after the addition of GPC as compared to control cells. This effect was dependent on the concentration of GPC used; however, under conditions where the cells were not killed, the enhanced synthesis and release of lipids were not dependent on the antibody concentration used to sensitize the cells. Treatment of the cells with antibody alone, GPC alone, or antibody plus heat-inactivated GPC did not result in enhanced synthesis or release of lipids. No enhancement in DNA, RNA, or protein synthesis in TAC was noted. Line-1 cells, which can be killed by GPC when sensitized with excess anti-Forssman IgM antibody, demonstrated enhanced lipid synthesis within 1 to 3 min after the addition of GPC to the antibody-sensitized cells, before measurable killing of the cells had occurred. This effect persisted in the surviving cells when tested 5 and 10 min after the formation of TAC. Addition of GPC deficient in C4 to antibody-sensitized cells did not result in enhanced lipid synthesis or release. These data suggest that the synthesis of macromolecules of which lipids are a major component is of central importance for the ability of the cells to resist antibody-GPC mediated attack.

Studies of membrane formation in Tetrahymena pyriformis: VI. Variations in the rate of lipid synthesis during division of heat-synchronized cells

Cells of Tetrahymena pyriformis, strain GL, induced to divide synchronously by alternating 30 min periods at 34 ° and 28 °C, exhibited an increased rate of phospholipid synthesis during the initial stage of cytokinesis. This sharp peak of biosynthesis was evident from labeling studies with either 32P-orthophosphate or 14C-acetate. However, the period of enhanced lipid synthesis could be dissociated from cytokinesis by altering incubation conditions and was apparently associated more closely with recovery from the heat shocks than with the division process. Cells synchronized by a different procedure involving a single 30 min heat shock per generation showed no peak of labeling with 32P i but retained the peak when tested with 14C-acetate. It appears that this latter perturbation was triggered by cytokinesis.

Inhibition of lipid synthesis by clofibrate: comparative study of human skin, rat skin, and rat liver in vitro

The effects of clofibrate (ethyl p-chlorophenoxyisobutyrate) on lipid synthesis by human skin were studied in vitro. The drug was found to inhibit lipid synthesis from [1-(14)C]acetate or [U-(14)C]glucose. While the synthesis of all classes of lipids was suppressed, inhibition of sterol synthesis was more pronounced than that of fatty acids and glycerides. By comparison, sodium p-chlorophenoxyisobutyrate was less effective as an inhibitor. The addition of glucose to the incubation medium enhanced lipid synthesis from both [1-(14)C]acetate and [U-(14)C]glucose. The inhibitory effect of clofibrate could be partially reversed by increasing the glucose concentration in the incubation medium. Rat skin and rat liver were studied similarly, using [1-(14)C]-acetate as a tracer for lipid synthesis, and the inhibitory effect of clofibrate was also demonstrated. Of the three tissues studied, human skin was the most sensitive to the drug and yielded more reproducible results.

Changes in lipid content during the reproductive cycle of Leucophaea maderae and effects of the juvenile hormone on lipid metabolism in vitro

1. 1. Lipid is stored in both the ovaries and fat body of female Leucophaea maderae during the 20-day period of oögenesis and is a major substrate for the developing embryos during the ensuing 2 months of embryogenesis. 2. 2. The lipid picture for the period of embryogenesis is characterized by a decreasing content of embryonic triglyceride and an increasing proportion of phospholipid. 3. 3. Both the fat body and ovaries (maturing oöcytes) are capable of incorporating labelled palmitate into glyceride and phospholipid. 4. 4. In vitro studies demonstrated the transfer of labelled lipid from the fat body to ovaries and it is likely that this takes place in vivo as well. The metabolic activity of the fat body is influenced by the stage of the reproductive cycle, particularly in reference to lipid metabolism. 5. 5. In vitro studies with active corpora allata suggest that one reason the juvenile (gonadotropic) hormone is indispensable for oöcyte maturation is its ability to enhance lipid synthesis in the ovary (maturing oöcytes) during oögenesis. 6. 6. These studies suggest that the juvenile hormone may act to depress the lipid synthetic ability of the fat body at this stage and thus make more substrate available to the developing oöcytes.