Klára Kitajka

The role of n-3 polyunsaturated fatty acids in brain: Modulation of rat brain gene expression by dietary n-3 fatty acids

Rats were fed either a high linolenic acid (perilla oil) or high eicosapentaenoic + docosahexaenoic acid (fish oil) diet (8%), and the fatty acid and molecular species composition of ethanolamine phosphoglycerides was determined. Gene expression pattern resulting from the feeding of n-3 fatty acids also was studied. Perilla oil feeding, in contrast to fish oil feeding, was not reflected in total fatty acid composition of ethanolamine phosphoglycerides. Levels of the alkenylacyl subclass of ethanolamine phosphoglycerides increased in response to feeding. Similarly, levels of diacyl phosphatidylethanolamine molecular species containing docosahexaenoic acid (18:0/22:6) were higher in perilla-fed or fish oil-fed rat brains whereas those in ethanolamine plasmalogens remained unchanged. Because plasmalogen levels in the brains of rats fed a n-3 fatty acid-enriched diet increased, it is plausible, however, that docosahexaenoic acid taken up from the food or formed from linolenic acid was deposited in this phospholipid subclass. Using cDNA microarrays, 55 genes were found to be overexpressed and 47 were suppressed relative to controls by both dietary regimens. The altered genes included those controlling synaptic plasticity, cytosceleton and membrane association, signal transduction, ion channel formation, energy metabolism, and regulatory proteins. This effect seems to be independent of the chain length of fatty acids, but the n-3 structure appears to be important. Because n-3 polyunsaturated fatty acids have been shown to play an important role in maintaining normal mental functions and docosahexaenoic acid-containing ethanolamine phosphoglyceride (18:0/22:6) molecular species accumulated in response to n-3 fatty acid feeding, a casual relationship between the two events can be surmised.

Short-term administration of omega 3 fatty acids from fish oil results in increased transthyretin transcription in old rat hippocampus

Reduced brain levels of long chain polyunsaturated fatty acids [arachidonic acid and docosahexanoic acid (DHA)] are observed in elderly subjects and patients with Alzheimers disease. To determine the effects of n-3 fatty acids on aged rat brain, 2-year-old rats were fed fish oil (27% DHA content) for 1 month, and gene expression analysis and fatty acid and molecular species composition of the major phospholipid species were assessed. No significant alteration could be observed in the fatty acid composition of ethanolamine phosphoglycerides and phosphatidylserines with the exception of DHA, which was slightly higher in brains of rats receiving fish oil. However, a drastic reduction in arachidonic acid in phosphatidylinositoles was observed. The expression of 23 genes was altered in response to fish oil feeding in the hippocampus. The transcription of transthyretin (TTR) was induced by 10-fold as evidenced by microarray analysis and confirmed by real-time quantitative RT-PCR. Expression of IL-1 and NO synthase, which has been implicated in the prevention of neurological diseases, was unaltered. TTR is an amyloid β protein scavenger, so an increase in its expression could prevent amyloid aggregate formation. We believe the beneficial effects of fish oil might be common to other agents, i.e., induce TTR expression, like nicotine and Ginkgo biloba extract.

Gene expression and molecular composition of phospholipids in rat brain in relation to dietary n-6 to n-3 fatty acid ratio

Rats were fed from conception till adulthood either with normal rat chow with a linoleic (LA) to linolenic acid (LNA) ratio of 8.2:1 or a rat chow supplemented with a mixture of perilla and soy bean oil giving a ratio of LA to LNA of 4.7:1. Fat content of the feed was 5%. Fatty acid and molecular species composition of ethanolamine phosphoglyceride was determined. Effect of this diet on gene expression was also studied. There was an accumulation of docosahexaenoic (DHA) and arachidonic acids (AA) in brains of the experimental animals. Changes in the ratio sn-1 saturated, sn-2 docosahexaenoic to sn-1 monounsaturated, sn-2 docosahexaenoic were observed. Twenty genes were found overexpressed in response to the 4.7:1 mixture diet and four were found down-regulated compared to normal rat chow. Among them were the genes related to energy household, lipid metabolism and respiration. The degree of up-regulation exceeded that observed with perilla with a ratio of LA to LNA 8.2:1 [Proc. Natl. Acad. Sci. U. S. A. 99 (2002) 2619]. It was concluded that brain sensitively reacts to the fatty acid composition of the diet. It was suggested that alteration in membrane architecture and function coupled with alterations in gene expression profiles may contribute to the observed beneficial impact of n-3 type polyunsaturated fatty acids on cognitive functions.

Gene expression profile analysis of lymphocytes from Alzheimer's patients.

Since the function and metabolism of peripheral lymphocytes is known to be altered in Alzheimers disease (AD), a pilot study was carried out to examine differences in gene expression profiles of these cells in 16 AD patients and aged control probands. Using a cDNA microarray representing 3200 distinct human genes, we identified 20 candidate genes whose expression is altered in AD lymphocytes compared with the control probands. Among these were the &agr;2C-adrenoreceptor gene, known to regulate blood pressure and learning, the defensin, histocompability complex enhancer-binding protein, carboxypeptidase M, and the Fc fragment of IgE known to be involved in cellular and humoral immune responses. Others, like human cell death protein, TRAIL, and galectin-4 participate in the regulation of apoptosis. Real-time quantitative reverse transcription-polymerase chain reaction analysis was performed in order to confirm the expression changes in AD lymphocytes, and it could detect down-regulation of defensin and &agr;2c-adrenoceptor genes, while other genes seemed unaltered in their expression, including heat-shock protein (hsp90), cholesteryl ester transfer protein, and apolipoprotein B100 (apoB). The altered expression profile of these genes might be connected with the previously reported AD-specific lymphocyte abnormalities. It remains to be elucidated, however, how these genes are related to the pathomechanism of dementia and whether the gene expression differences of AD lymphocytes reflect disease traits or stage processes.

Molecular architecture and biophysical properties of phospholipids during thermal adaptation in fish: An experimental and model study

Phospholipids from livers of carps (Cyprinus carpio L.) adapted to winter (5°C) and summer (25°C) temperatures were isolated, and the fatty acid composition of total phospholipids, as well as molecular species composition of diacyl phosphatidylcholines and ethanolamines, were determined. Order parameter of 5-doxyl stearic acid and steady-state fluorescence anisotropy of different anthroyloxy fatty acids—[2-, 12(N-9-anthroyloxy)stearic acid and 16(N-9-anthroyloxy)palmitic acid—embedded in native and synthetic (16∶0/16∶0, 16∶0/22∶6, 18∶0/22∶6, 18∶1/22∶6, 20∶4/20∶4, 22∶6/22∶6 phosphatidylcholines and 16∶0/18∶1, 18∶1/22∶6 phosphatidylethanolamines) phospholipid vesicles was also determined between −30 and 30°C and 5 and 30°C, respectively. There is an accumulation of 1-monoenoic, 2-polyenoic diacyl phosphatidylcholine and ethanolamine with a concomitant reduction of 1-stearoyl,2-docosahexaenoyl species in the cold-adapted state. Despite a 30% accumulation of long-chain polyunsaturated fatty acids in phospholipids in cold, there is only a 5°C downshift in the solid-gel to liquid-crystalline phase transition temperature (−8 vs. −13°C). Vesicles from total phospholipids of cold-adapted fish proved to be more disordered in all segments than from the warmadapted ones when assayed using 2,12-(N-9-anthroyloxy)stearic and 16-(N-9-anthroyloxy)palmitic acid. Vesicles made from purified phosphatidylcholines showed the same pattern, but they were more disordered than the corresponding total phospholipids. This could be modelled using mixed phospholipid vesicles made of synthetic 16∶0/22∶6 phosphatidylcholine (75%) and either 18∶1/22∶6 phosphatidylethanolamine (25%) vs. 16∶0/18∶1 phosphatidylethanolamine (25%) and comparison of the anisotropy parameters of 100% 16∶0/22∶6 and 100% 18∶1/22∶6 phosphatidylcholine vesicles. Mixing either 16∶0/18∶1 (25%) or 18∶1/22∶6 (25%) phosphatidylethanolamines to 18∶0/22∶6 (75%) phosphatidylcholine shifted down or up, respectively, the transition temperature of vesicles compared to 100% 18∶0/22∶6 vesicles assayed by electron spin resonance spectroscopy using 5-doxylstearic acid. It is concluded that it is not the gross amount of long-chain polyunsaturated fatty acids in phospholipids, but rather their specific combination withcis Δ9 monounsaturated fatty acids in the positionsn-1, especially in phosphatidylethanolamines, that is important in determining the physical properties of biomembranes in relation to adaptational temperature.

MicroRNA profile of polyunsaturated fatty acid treated glioma cells reveal apoptosis-specific expression changes

BackgroundPolyunsaturated fatty acids (PUFAs) such as γ-linolenic acid (GLA), arachidonic acid (AA) and docosahexaenoic acid (DHA) have cytotoxic action on glioma cells.ResultsWe evaluated the cytotoxic action of GLA, AA and DHA on glioma cells with specific reference to the expression of miRNAs. Relative expression of miRNAs were assessed by using high throughput nanocapillary real-time PCR. Most of the miRNA target genes that showed altered expression could be classified as apoptotic genes and were up-regulated by PUFA or temozolomide treatment, while similar treatments resulted in repression of the corresponding mRNAs, such as cox2, irs1, irs2, ccnd1, itgb3, bcl2, sirt1, tp53inp1 and k-ras.ConclusionsOur results highlight involvement of miRNAs in the induction of apoptosis in glioma cells by fatty acids and temozolomide.

Cholesterol diet‐induced hyperlipidemia influences gene expression pattern of rat hearts: a DNA microarray study

To profile gene expression patterns involved in the direct myocardial effect of cholesterol‐enriched diet‐induced hyperlipidemia, we monitored global gene expression changes by DNA microarray analysis of 3200 genes in rat hearts. Twenty‐six genes exhibited significant up‐regulation and 25 showed down‐regulation in hearts of rats fed a 2% cholesterol‐enriched diet for 8 weeks as compared to age‐matched controls. The expression changes of 12 selected genes were also assessed by real‐time quantitative polymerase chain reaction. Genes with altered expression in the heart due to hyperlipidemia included procollagen type III, cofilin/destrin, tensin, transcription repressor p66, synaptic vesicle protein 2B, Hsp86, chaperonin subunit 5ϵ, metallothionein, glutathione S‐transferase, protein kinase C inhibitor, ATP synthase subunit c, creatine kinase, chloride intracellular channel 4, NADH oxidoreductase and dehydrogenase, fibronectin receptor β chain, CD81 antigen, farnesyltransferase, calreticulin, disintegrin, p120 catenin, Smad7, etc. Although some of these genes have been suspected to be related to cardiovascular diseases, none of the genes has been previously shown to be involved in the mechanism of the cardiac effect of hyperlipidemia.

Melatonin-induced gene expression changes and its preventive effects on adriamycin-induced lipid peroxidation in rat liver

Abstract:  Adriamycin (ADR) provokes lipid peroxidation process, while melatonin (MEL) is a free radical scavenger that has been found to protect against lipid peroxidation in vitro and in many experimental models. In the present study, the effects of ADR and the combination of ADR and MEL were analyzed on the modulation of fatty acid composition, lipid peroxidation and gene expression in rat liver. Sixty genes were selected for the study of relative gene expression changes in the liver. ADR treatment decreased the polyunsaturated fatty acids C22:6 n‐3 and C20:4 n‐6 in rat liver mitochondria. When the treatment of ADR was followed by MEL, decrease in these fatty acids could not be detected. A significant increase in lipid peroxidation was observed after administration of ADR, which was restored to control values by post‐treatment with MEL. Gene expression profiles of ADR‐ versus ADR + MEL‐treated rat livers indicated that both treatments induced significant changes. Quantitative real‐time polymerase chain reaction analysis of 60 genes involved in oxidative stress revealed that cyp1b1, which is involved in electron transport, cyclin‐dependent kinase inhibitor 1a that possesses cyclin‐dependent protein kinase inhibitor activity, was induced at a more pronounced level in the ADR + MEL‐treated samples than in the ADR‐treated ones. Several genes having roles in heat‐shock response were downregulated in MEL‐treated animals, such as hsp40, hsp70 and hsp90 proteins reflecting the reduced oxidative stress in these animals. Global gene expression analysis will highlight the gene expression changes accompanying oxidative damage and its prevention in more details.

Gene expression profile analysis of the rat cortex following treatment with imipramine and citalopram

The effect of antidepressants is the culmination of a series of molecular actions occurring in the brain. These events are thought to lead to changes in the expression level of numerous, but as yet unknown genes that result in different cellular functions. In our present study we addressed this issue by establishing gene expression profiles of the rat brain after treatment with imipramine and citalopram at therapeutic doses. After 96 h and 4 wk, fronto-temporal cortices from controls and each treated strain were prepared and total RNA was isolated, and assessed using a cDNA microarray system containing 3200 clones. The expression of 6 genes was decreased and 8 were over-expressed by imipramine, whereas 27 were repressed and 7 were up-regulated by citalopram. Members of signal transduction (e.g. phosphatidylinositol transfer protein), structural elements (e.g. tubulin, fibronectin), factors related to protein metabolism in general (e.g. proteasomal subunits, ubiquitin-like proteins, polyadenylation sites), components involved in cell survival (e.g. midkine, stress-inducible protein), and determinants of membrane conductance and ion transport (e.g. vacuolar H+-ATPase), and basics of nuclear functions (e.g. translin, basal transcription factor 3), were some of the genes with altered expression. These data demonstrate that antidepressants interfere with the expression of a large array of genes involved in signalling, survival and protein metabolism. Our results demonstrate for the first time that antidepressants specifically regulate neuronal plasticity through induction of a highly specific transcriptional programme in brain cells.

The Effect of Citalopram on Gene Expression Profile of Alzheimer Lymphocytes

Antidepressants are widely used in the treatment of mood disorders associated with dementia, however little information is available on their effect at the molecular level. In certain neurodegenerative disorders, such as in Alzheimers disease, lymphocytes have been used to assess mirror changes that thought to occur in the brain. Gene expression profiles of lymphocytes from Alzheimer patients have been shown to differ from that seen with controls. To address this issue in light of antidepressant treatment, we used lymphocytes derived from Alzheimers disease patients and control individuals to assess the impact of the selective serotonine reuptake inhibitor citalopram on gene expression using a cDNA microarray representing 3200 distinct human genes. Sequences that are differentially regulated after treatment with citalopram were identified and categorized based on similarities in biological functions. This analysis revealed that the overexpression of genes in control and Alzheimer white blood cells by citalopram are implicated in cell survival. Apart from this, citalopram did not markedly alter genes involved in other molecular functions in control cells. In contrast, alteration of genes implicated in ionic currents, cell-adhesion, immune mechanism, and adrenergic functions, were also observed in Alzheimer lymphocytes. The expression of genes of Alzheimer lymphocytes by citalopram is modulated differently which may correlate with the pathology.