The intake of high-fat diets induces an obesogenic-like gene expression profile in peripheral blood mononuclear cells, which is reverted by dieting.

Peripheral blood mononuclear cells (PBMC) are increasingly used for nutrigenomic studies. In this study, we aimed to identify whether these cells could reflect the development of an obesogenic profile associated with the intake of high-fat (HF) diets. We analysed, by real-time RT-PCR, the dietary response of key genes related to lipid metabolism, obesity and inflammation in PBMC of control rats, rats fed a cafeteria or a commercial HF diet and rats fed a control diet after the intake of a cafeteria diet (post-cafeteria model). Cafeteria diet intake, which resulted in important overweight and related complications, altered the expressions of most of the studied genes in PBMC, evidencing the development of an obesogenic profile. Commercial HF diet, which produced metabolic alterations but in the absence of noticeably increased body weight, also altered PBMC gene expression, inducing a similar regulatory pattern as that observed for the cafeteria diet. Regulation of carnitine palmitoyltransferase I (Cpt1a) mRNA expression was of special interest; its expression reflected metabolic alterations related to the intake of both obesogenic diets (independently of increased body weight) even at an early stage as well as metabolic recovery in post-cafeteria animals. Thus, PBMC constitute an important source of biomarkers that reflect the increased adiposity and metabolic deregulation associated with the intake of HF diets. In particular, we propose an analysis of Cpt1a expression as a good biomarker to detect the early metabolic alterations caused by the consumption of hyperlipidic diets, and also as a marker of metabolic recovery associated to weight loss.

Gene expression of peripheral blood mononuclear cells is affected by cold exposure.

Because of the discovery of brown adipose tissue (BAT) in humans, there is increased interest in the study of induction of this thermogenic tissue as a basis to combat obesity and related complications. Cold exposure is one of the strongest stimuli able to activate BAT and to induce the appearance of brown-like (brite) adipocytes in white fat depots (browning process). We analyzed the potential of peripheral blood mononuclear cells (PBMCs) to reflect BAT and retroperitoneal white adipose tissue (rWAT) response to 1-wk cold acclimation (4°C) at different ages of rat development (1, 2, 4, and 6 mo). As expected, cold exposure increased fatty acid ß-oxidation capacity in BAT and rWAT (increased Cpt1a expression), explaining increased circulating nonesterified free fatty acids and decreased adiposity. Cold exposure increased expression of the key thermogenic gene, Ucp1, in BAT and rWAT, but only in 1-mo-old animals. Additionally, other brown/brite markers were affected by cold during the whole developmental period studied in BAT. However, in rWAT, cold exposure increased studied markers mainly at early age. PBMCs did not express Ucp1, but expressed other brown/brite markers, which were cold regulated. Of particular interest, PBMCs reflected adipose tissue-increased Cpt1a mRNA expression in response to cold (in older animals) and browning induction occurring in rWAT of young animals (1 mo) characterized by increased Cidea expression and by the appearance of a high number of multilocular CIDE-A positive adipocytes. These results provide evidence pointing to PBMCs as an easily obtainable biological material to be considered to perform browning studies with minimum invasiveness.

Cold exposure down-regulates immune response pathways in ferret aortic perivascular adipose tissue.

Perivascular adipose tissue (PVAT) surrounds blood vessels and releases paracrine factors, such as cytokines, which regulate local inflammation. The inflammatory state of PVAT has an important role in vascular disease; a pro-inflammatory state has been related with atherosclerosis development, whereas an anti-inflammatory one is protective. Cold exposure beneficially affects immune responses and, could thus impact the pathogenesis of cardiovascular diseases. In this study, we investigated the effects of one-week of cold exposure at 4°C of ferrets on aortic PVAT (aPVAT) versus subcutaneous adipose tissue. Ferrets were used because of the similarity of their adipose tissues to those of humans. A ferret-specific Agilent microarray was designed to cover the complete ferret genome and global gene expression analysis was performed. The data showed that cold exposure altered gene expression mainly in aPVAT. Most of the regulated genes were associated with cell cycle, immune response and gene expression regulation, and were mainly down-regulated. Regarding the effects on immune response, cold acclimation decreased the expression of genes involved in antigen recognition and presentation, cytokine signalling and immune system maturation and activation. This immunosuppressive gene expression pattern was depot-specific, as it was not observed in the inguinal subcutaneous depot. Interestingly, this depression in immune response related genes was also evident in peripheral blood mononuclear cells (PBMC). In conclusion, these results reveal that cold acclimation produces an inhibition of immune response-related pathways in aPVAT, reflected in PBMC, indicative of an anti-inflammatory response, which can potentially be exploited for the enhancement or maintenance of cardiovascular health.

Specific Features of the Hypothalamic Leptin Signaling Response to Cold Exposure Are Reflected in Peripheral Blood Mononuclear Cells in Rats and Ferrets.

Objectives: Cold exposure induces hyperphagia to counteract fat loss related to lipid mobilization and thermogenic activation. The aim of this study was investigate on the molecular mechanisms involved in cold-induced compensatory hyperphagia. Methods: We analyzed the effect of cold exposure on gene expression of orexigenic and anorexigenic peptides, and of leptin signaling-related genes in the hypothalamus of rats at different ages (1, 2, 4, and 6 months), as well as in ferrets. We also evaluated the potential of peripheral blood mononuclear cells to reflect hypothalamic molecular responses. Results: As expected, cold exposure induced hypoleptinemia in rats, which could be responsible for the increased ratio of orexigenic/anorexigenic peptides gene expression in the hypothalamus, mainly due to decreased anorexigenic gene expression, especially in young animals. In ferrets, which resemble humans more closely, cold exposure induced greater changes in hypothalamic mRNA levels of orexigenic genes. Despite the key role of leptin in food intake control, the effect of cold exposure on the expression of key hypothalamic leptin signaling cascade genes is not clear. In our study, cold exposure seemed to affect leptin signaling in 4-month-old rats (increased Socs3 and Lepr expression), likely associated with the smaller-increase in food intake and decreased body weight observed at this particular age. Similarly, cold exposed ferrets showed greater hypothalamic Socs3 and Stat3 gene expression. Interestingly, peripheral blood mononuclear cells (PBMC) mimicked the hypothalamic increase in Lepr and Socs3 observed in 4-month-old rats, and the increased Socs3 mRNA expression observed in ferrets in response to cold exposure. Conclusions: The most outstanding result of our study is that PBMC reflected the specific modulation of leptin signaling observed in both animal models, rats and ferrets, which points forwards PBMC as easily obtainable biological material to be considered as a potential surrogate tissue to perform further studies on the regulation of hypothalamic leptin signaling in response to cold exposure.

Sustained exposure to diets with an unbalanced macronutrient proportion alters key genes involved in energy homeostasis and obesity-related metabolic parameters in rats.

We have investigated the effects of long term intake of two unbalanced diets (rich in fat -HF- or protein -HP-) administered under isocaloric conditions to a control balanced diet (pair-feeding) to adult rats. Isocaloric intake of a HF diet did not affect the body weight but increased adiposity, liver-fat deposition, and induced insulin resistance. Gene expression changes in the liver and adipose tissue (increased lipolytic and decreased lipogenic gene expression) could try to compensate for increased adiposity. The HP diet decreased caloric intake, the body weight, the size of subcutaneous adipocytes, and circulating cholesterol. Higher insulin levels apparently not related to insulin resistance were observed. Changes at the gene expression level reflected an adaptation to lower diet carbohydrate content and to the use of amino acids as the energy source. The kidney size increased in HP-fed animals but serum creatinine was not affected. Circulating TNF-alpha levels were higher in both dietary models. Thus, a long-term increase in dietary fat proportion produces alterations related to metabolic syndrome even in the absence of increased body weight, whereas an increase in diet protein content reduces the body weight but alters metabolic parameters and kidney size which could be linked to an increased risk of suffering from different pathologies.

Reversion to a control balanced diet is able to restore body weight and to recover altered metabolic parameters in adult rats long-term fed on a cafeteria diet.

The increased intake of fat-rich foods is one of the causes of the increasing incidence of obesity. However, there are controversial data on the reversibility of diet-induced obesity and its metabolic complications when adopting a control energy-balanced diet. Our aim was to evaluate the ability to reset not only body weight but also metabolic disorders caused by a highly palatable high fat diet, cafeteria diet, administered to adult rats, when replaced by a control diet (post-cafeteria model). Four-months of cafeteria diet-feeding produced important metabolic alterations in comparison to a commercial purified high fat diet: a rapid, drastic increase in body weight, adiposity and related complications such as insulin resistance, decreased glucose tolerance and development of hepatic steatosis. At gene level, decreased lipogenic and increased lipolytic gene expression in key energy homeostatic tissues as a physiological adaptation to increased fat intake was observed. In addition, fasting response of serum parameters and of key genes in lipid metabolism was impaired in cafeteria-fed animals. Contrary to what we have previously described if cafeteria diet is administered early in life, when administered to adult animals, its replacement with a balanced diet is able to restore body weight. Cafeteria diet withdrawal also allows recovery from metabolic damage, gene expression regulation and fasting response, the degree of which is dependent on the time of exposure to the cafeteria diet. In conclusion, adherence to an ad libitum intake of a balanced standard diet can enable the recovery of healthy status in animals which were previously exposed to an unhealthy cafeteria diet in adult age.

Decreased RB1 mRNA, protein, and activity reflect obesity-induced altered adipogenic capacity in human adipose tissue.

Retinoblastoma (Rb1) has been described as an essential player in white adipocyte differentiation in mice. No studies have been reported thus far in human adipose tissue or human adipocytes. We aimed to investigate the possible role and regulation of RB1 in adipose tissue in obesity using human samples and animal and cell models. Adipose RB1 (mRNA, protein, and activity) was negatively associated with BMI and insulin resistance (HOMA-IR) while positively associated with the expression of adipogenic genes (PPARγ and IRS1) in both visceral and subcutaneous human adipose tissue. BMI increase was the main contributor to adipose RB1 downregulation. In rats, adipose Rb1 gene expression and activity decreased in parallel to dietary-induced weight gain and returned to baseline with weight loss. RB1 gene and protein expression and activity increased significantly during human adipocyte differentiation. In fully differentiated adipocytes, transient knockdown of Rb1 led to loss of the adipogenic phenotype. In conclusion, Rb1 seems to play a permissive role for human adipose tissue function, being downregulated in obesity and increased during differentiation of human adipocytes. Rb1 knockdown findings further implicate Rb1 as necessary for maintenance of adipogenic characteristics in fully differentiated adipocytes.