Regina Maria Vieira da Costa Guaragna

SIRT1 Transcription Is Decreased in Visceral Adipose Tissue of Morbidly Obese Patients with Severe Hepatic Steatosis

BackgroundVisceral adipose tissue is known to release greater amounts of adipokines and free fatty acids into the portal vein, being one of the most predictive factors of nonalcoholic fatty liver disease (NAFLD). Our study has the purpose to evaluate sirtuin 1 (SIRT1), adiponectin, Forkhead/winged helix (FOXO1), peroxisome proliferator-activated receptor (PPAR)γ1–3, and PPARβ/δ mRNA expression in morbidly obese patients in three different lipid depots: visceral (VAT), subcutaneous (SAT), and retroperitoneal (RAT). Recent studies suggest that SIRT1, a NAD+-dependent deacetylase, protects rats from NAFLD.MethodsWe divided the patients in two groups: those with slight or moderate steatosis (hepatic steatosis, HS) and other comprising individuals with severe steatosis associated or not with necroinflammation and fibrosis (severe hepatic steatosis, SHS). The adipose tissue depots were obtained during bariatric surgery. Total RNAs were extracted using TRIzol. The amount of genes of interest was determined by quantitative real-time polymerase chain reaction.ResultsWhen comparing the two groups of patients, a decrease in SIRT1 was observed in VAT of morbidly obese patients in SHS group (p = 0.006). The mRNA expression of the other genes showed no differences in VAT. No difference was found either in SAT or in RAT for all genes in the study. In addition, the homeostasis model assessment for insulin resistance (HOMA-IR) value was higher in SHS group compared to HS (p = 0.006). Also, our results show that the mRNA expression of SIRT1 and the value of HOMA-IR were positively correlated in VAT of SHS patients (r = 0.654; p = 0.048).ConclusionsDownregulation of SIRT1 mRNA expression in VAT of SHS could be possible impairing mitochondria biogenesis and fatty acid oxidation, promoting severe steatosis in obese patients. Our results provide a possible proof of SIRT1 protective potential in VAT against NAFLD in humans.

Resveratrol Upregulated SIRT1, FOXO1, and Adiponectin and Downregulated PPARγ1–3 mRNA Expression in Human Visceral Adipocytes

BackgroundThe SIRT1 enzyme is involved in adipose tissue (AT) lipolysis. FOXO1 is a protein that plays a significant role in regulating metabolism. Adiponectin is an adipokine, secreted by the AT, which has been considered to have an antiobesity function. PPARγ is one of the key actors in adipocytes differentiation. This study was undertaken to investigate whether resveratrol can regulate SIRT1, FOXO1, adiponectin, PPARγ1–3, and PPARβ/δ in human AT.MethodsThe effects of resveratrol were analyzed in freshly isolated adipocytes prepared from visceral fat tissue samples obtained during bariatric surgery. Genes messenger ribonucleic acid (mRNA) levels were determined by qRT-PCR.ResultsOurs results show that resveratrol modulates the studied genes, increasing SIRT1 (p = 0.021), FOXO1 (p = 0.001), and adiponectin (p = 0.025) mRNA expression and decreasing PPARγ1–3 (p = 0.003) mRNA in human visceral adipocytes.ConclusionsResveratrol, in vitro and at low concentration, modulates genes that are related to lipid metabolism, possibly preventing metabolic disease in human visceral adipose tissue (VAT).

Relationship between oxidative stress levels and activation state on a hepatic stellate cell line

Abstract: Background/Aims: Oxidative stress plays an important role in liver fibrosis. Under pathological conditions, hepatic stellate cells (HSC) undergo an activation process, developing a myofibroblast‐like phenotype from the lipocyte phenotype. In this study, we determined the levels of oxidative stress and proliferation in different activation states of an experimental model of mouse HSC, the GRX cell line. These cells can be induced in vitro to display a more activated state or a quiescent phenotype.

β‐Carotene storage, conversion to retinoic acid, and induction of the lipocyte phenotype in hepatic stellate cells

Hepatic stellate cells (HSCs) are the major site of retinol (ROH) metabolism and storage. GRX is a permanent murine myofibroblastic cell line, derived from HSCs, which can be induced to display the fat‐storing phenotype by treatment with retinoids. Little is known about hepatic or serum homeostasis of β‐carotene and retinoic acid (RA), although the direct biogenesis of RA from β‐carotene has been described in enterocytes. The aim of this study was to identify the uptake, metabolism, storage, and release of β‐carotene in HSCs. GRX cells were plated in 25 cm2 tissue culture flasks, treated during 10 days with 3 μmol/L β‐carotene and subsequently transferred into the standard culture medium. β‐Carotene induced a full cell conversion into the fat‐storing phenotype after 10 days. The total cell extracts, cell fractions, and culture medium were analyzed by reverse phase high‐performance liquid chromatography for β‐carotene and retinoids. Cells accumulated 27.48 ± 6.5 pmol/L β‐carotene/106 cells, but could not convert it to ROH nor produced retinyl esters (RE). β‐Carotene was directly converted to RA, which was found in total cell extracts and in the nuclear fraction (10.15 ± 1.23 pmol/L/106 cells), promoting the phenotype conversion. After 24‐h chase, cells contained 20.15 ± 1.12 pmol/L β‐carotene/106 cells and steadily released β‐carotene into the medium (6.69 ± 1.75 pmol/ml). We conclude that HSC are the site of the liver β‐carotene storage and release, which can be used for RA production as well as for maintenance of the homeostasis of circulating carotenoids in periods of low dietary uptake.

Hepatic stellate cell line modulates lipogenic transcription factors

Background/Aims: Pre‐adipocyte differentiation into adipocyte is a terminal differentiation process triggered by a cascade of transcription factors. Conversely, hepatic stellate cells (HSC) can switch between lipid storing and the myofibroblast phenotype in association with liver fibrotic processes. Here, adipogenic/lipogenic‐related transcription factors and downstream‐regulated genes were evaluated in a murine HSC cell line. GRX‐HSC cells are transitional myofibroblasts that differentiate into lipocytes following retinol or indomethacin treatment.

Higher Content of Trans Fatty Acids in Abdominal Visceral Fat of Morbidly Obese Individuals undergoing Bariatric Surgery compared to Non-Obese Subjects

Background: The purpose of this study was to determine the total content of trans fatty acids (TFA) in subcutaneous, retroperitoneal and visceral fat of morbidly obese and non-obese patients submitted to bariatric surgery or plastic and abdominal surgery. Methods: The adipose tissues were obtained by surgery; lipids were extracted, saponified and esterified. TFA were measured by FTIR-ATR spectroscopy. Results: The TFA average in obese patients was 6.3% for retroperitoneal and 8.7% for visceral fat. For non-obese patients, the figures were 6.9% (subcutaneous) and 9.3% (visceral). There was no difference between the groups. However, the TFA depot in visceral fat was higher than other fatty tissues for morbidly obese (P<0.001) and non-obese (P<0.05) patients. Conclusions: Our values for TFA content in all adipose tissues analyzed are higher than reported in other countries (3-6%). We showed more TFA in visceral adipose tissue than in other abdominal fat (subcutaneous and retroperitoneal) stores. The visceral adipose tissue level is worrisome because the higher rate of lipolysis in this tissue appears to be an important indicator of metabolic alterations and the levels of TFA found in adipose tissue presumably reflect the higher dietary intake of TFA by Brazilians.

Changes of sphingolipid species in the phenotype conversion from myofibroblasts to lipocytes in hepatic stellate cells

Sphingolipids play a relevant role in cell–cell interaction, communication, and migration. We studied the sphingolipid content in the murine hepatic stellate cell line GRX, which expresses the myofibroblast phenotype, and can be induced in vitro to display the fat‐storing phenotype. Lipid modifications along this induction were investigated by labeling sphingolipids with [14C]galactose, [14C]serine, or [14C]choline, and determination of fatty acid composition of sphingomyelin. The total ganglioside content and the GM2 synthase activity were lower in myofibroblasts. Both phenotypes presented similar gangliosides of the a‐pathway: GM2, GM1, and GD1a as well as their precursor GM3. Sphingomyelin and all the gangliosides were expressed as doublets; the upper/lower band ratio increased in lipocytes, containing more long‐chain fatty acids in retinol‐induced lipocytes as compared to the insulin/indomethacin induced ones. Time‐course experiments indicated a transfer of metabolic precursors from phosphatidylcholine to sphingomyelin in the two phenotypes. Taken together, these results indicate that myofibroblast and lipocytes can use distinct ceramide pools for sphingolipid synthesis. Differential ganglioside expression and presence of the long‐chain saturated fatty acids suggested that they may participate in formation of distinct membrane microdomains or rafts with specific functions on the two phenotypes of GRX‐cells.

Lipid metabolism during in vitro induction of the lipocyte phenotype in hepatic stellate cells

Molecular mechanisms of lipid synthesis and their controls in hepatic stellate cells are not known. We have previously proposed that, in contrast to other fat storing cells, hepatic stellate cells are not involved in energy storage, but they represent a particular cell population specialized in storage of lipid-soluble substances, the major one being probably retinol. In agreement with this hypothesis, induction of the lipocyte phenotype in stellate cells is not under the control of insulin, but responds to retinoids and other molecules that modify the gene expression program in these cells. In the present study we have monitored the activity of the two major enzymes involved in lipid synthesis during the induction of the lipocyte phenotype in hepatic stellate cells: glycerol-3-phosphate dehydrogenase (GPDH) that mediates the de novo lipid synthesis, and lipoprotein lipase that mediates incorporation of plasma lipids. In early stages of lipocyte induction, both pathways of lipid synthesis are activated. When lipocytes have already constituted the lipid droplets, lipoprotein lipase pathway is downregulated, while GPDH activity remains high. Adult liver has been reported to lack lipoprotein lipase, but under stress, lipase activity was detected around and at the surface of the intrahepatic vasculature. We have now shown that the lipase activity can be induced in the hepatic stellate cells, located in the Disses space. The high lipoprotein lipase activity under acute induction of lipocyte phenotype, followed by the low activity under conditions of metabolic equilibrium, are in compass with the increased activity of this enzyme under stress, and its low activity in adult liver parenchyma under normal conditions.

Parallel Down-Regulation of FOXO1, PPARγ and Adiponectin mRNA Expression in Visceral Adipose Tissue of Class III Obese Individuals

Objective: Adipose tissue is responsible for secretion of several cytokines that mediate systemic effects on obesity and insulin resistance. Subcutaneous abdominal adipose tissue (SAT) and visceral adipose tissue (VAT) are metabolically different and have differences in their gene expression profile. Our study evaluated the expression of adiponectin, FOXO1, PPARγ, and SIRT1 in VAT and SAT of non-obese and class III obese subjects. Methods: The adipose tissue samples were obtained by surgery. Reverse transcripts of studied genes were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Results: Comparing the different lipid depots, adiponectin expression was lower only in VAT of obese individuals (p = 0.043); FOXO1 and PPARγ levels were decreased in VAT of both groups. When non-obese and obese were compared, only adiponectin expression was lower in SAT and in VAT of obese subjects (p = 0.004 and p = 0.002, respectively). No difference was found with regard to SIRT1 levels in VAT or SAT in both groups. FOXO1 expression in SAT of obese subjects had a negative correlation with age (r = –0.683; p = 0.029) and triglyceride serum levels (r = –0.794; p = 0.006). Conclusion: The decrease mRNA expression of this genes in VAT, responsible for central adiposity, may be associated with an increased risk of obesity and co-morbidities.

Lipídio: fator de risco e prevenção do câncer de mama

The hypothesis that a high fat diet promotes the development of postmenopausal breast cancer is supported by case-controls studies that show a strong correlation between fat intake and breast cancer rates. On the other hand, high fat intake is not correlated with breast cancer in cohort studies. In view of these conflicting findings, it has been difficult to propose nutritional recommendations for the prevention of breast cancer. Animal studies and recent observations in humans have shown that a diet high in polyunsaturated fatty acids stimulates several stages in the development of mammary cancer. Some evidences show that fish oil-derived w-3 fatty acids seem to prevent cancer by influencing the activity of enzymes and proteins related to cell proliferation. Thus, epidemiological studies that integrate the interactions of specific fatty acids with hormone catabolism, protective nutritional factors and risk factors associated with breast cancer are needed. In this article we will discuss protective factors, risk factors and qualitative and quantitative implications of dietary fatty acids in breast cancer.