Marta Gil-Ortega

High-fat diets impair spatial learning in the radial-arm maze in mice

It has been suggested that hyperglycemia and insulin resistance triggered by energy-dense diets can account for hippocampal damage and deficits of cognitive behaviour. We wonder if the impairment of learning and memory processes detected in diet-induced obese (DIO) mice is linked to diet composition itself. With this purpose we have evaluated learning performance in mice undergoing a short-term high-fat (HF) treatment, which leads to a pre-obese state characterized by increased adiposity without significant changes of glucose and insulin plasma levels. C57BL/6J mice were fed either a HF (45 kcal% from fat) or control diet (10 kcal% from fat) during 8 weeks. Learning performance was evaluated by using the four-arm baited version of the eight-arm radial maze test (RAM). Mice were trained to learn the RAM protocol and then memory was tested at different time-points. Time spent to consume food placed in baited arms and errors committed to find them were measured in all sessions. DIO mice significantly spent more time in learning the task and made a greater number of errors than controls. Moreover, retention tests revealed that both working and total memory errors were also more numerous in DIO mice. The current results show that short-term DIO impairs spatial learning and suggest that impairment of hippocampal learning elicited by HF diets might be perceptible before metabolic alterations linked to obesity develop.

Adaptative Nitric Oxide Overproduction in Perivascular Adipose Tissue during Early Diet-Induced Obesity

Perivascular adipose tissue (PVAT) plays a paracrine role in regulating vascular tone. We hypothesize that PVAT undergoes adaptative mechanisms during initial steps of diet-induced obesity (DIO) which contribute to preserve vascular function. Four-week-old male C57BL/6J mice were assigned either to a control [low-fat (LF); 10% kcal from fat] or to a high-fat diet (HF; 45% kcal from fat). After 8 wk of dietary treatment vascular function was analyzed in the whole perfused mesenteric bed (MB) and in isolated mesenteric arteries cleaned of PVAT. Relaxant responses to acetylcholine (10(-9)-10(-4) m) and sodium nitroprusside (10(-12)-10(-5) m) were significantly ameliorated in the whole MB from HF animals. However, there was no difference between HF and LF groups in isolated mesenteric arteries devoid of PVAT. The enhancement of relaxant responses detected in HF mice was not attributable to an increased release of nitric oxide (NO) from the endothelium nor to an increased sensitivity and/or activity of muscular guanilylcyclase. Mesenteric PVAT of HF animals showed an increased bioavailability of NO, detected by 4,5-diaminofluorescein diacetate (DAF2-DA) staining, which positively correlated with plasma leptin levels. DAF-2DA staining was absent in PVAT from ob/ob mice but was detected in these animals after 4-wk leptin replacement. The main finding in this study is that adaptative NO overproduction occurs in PVAT during early DIO which might be aimed at preserving vascular function.

Anticontractile Effect of Perivascular Adipose Tissue and Leptin are Reduced in Hypertension

Leptin causes vasodilatation both by endothelium-dependent and -independent mechanisms. Leptin is synthesized by perivascular adipose tissue (PVAT). The hypothesis of this study is that a decrease of leptin production in PVAT of spontaneously hypertensive rats (SHR) might contribute to a diminished paracrine anticontractile effect of the hormone. We have determined in aorta from Wistar-Kyoto (WKY) and SHR (i) leptin mRNA and protein levels in PVAT, (ii) the effect of leptin and PVAT on contractile responses, and (iii) leptin-induced relaxation and nitric oxide (NO) production. Leptin mRNA and protein expression were significantly lower in PVAT from SHR. Concentration-response curves to angiotensin II were significantly blunted in presence of PVAT as well as by exogenous leptin (10−9 M) only in WKY. This anticontractile effect was endothelium-dependent. Vasodilatation induced by leptin was smaller in SHR than in WKY, and was also endothelium-dependent. Moreover, release of endothelial NO in response to acute leptin was higher in WKY compared to SHR, but completely abolished in the absence of endothelium. In conclusion, the reduced anticontractile effect of PVAT in SHR might be attributed to a reduced PVAT-derived leptin and to an abrogated effect of leptin on endothelial NO release probably due to an impaired activation of endothelial NO synthase.

Regional differences in perivascular adipose tissue impacting vascular homeostasis

Perivascular adipose tissue (PVAT) releases several important vasoactive factors with physiological and pathophysiological paracrine effects. A large body of evidence suggests regional phenotypic and functional differences among PVAT depots, depending on the specific vascular bed or different regions in the vascular bed where the PVAT is located. These non-uniform and separate PVATs exert various paracrine effects on vascular structure and function that largely impact disease states, such as endothelial dysfunction, atherosclerosis, or insulin resistance. This emerging view of PVAT function requires considering heterogeneous PVAT as a specialized organ that can differentially regulate vascular function depending on its anatomical location. In this context, the adipose-vascular axis may represent a novel target for pharmacological intervention in vasculopathy in cardiometabolic disorders.

Mechanisms of Perivascular Adipose Tissue Dysfunction in Obesity

Most blood vessels are surrounded by adipose tissue. Similarly to the adventitia, perivascular adipose tissue (PVAT) was considered only as a passive structural support for the vasculature, and it was routinely removed for isolated blood vessel studies. In 1991, Soltis and Cassis demonstrated for the first time that PVAT reduced contractions to noradrenaline in rat aorta. Since then, an important number of adipocyte-derived factors with physiological and pathophysiological paracrine vasoactive effects have been identified. PVAT undergoes structural and functional changes in obesity. During early diet-induced obesity, an adaptative overproduction of vasodilator factors occurs in PVAT, probably aimed at protecting vascular function. However, in established obesity, PVAT loses its anticontractile properties by an increase of contractile, oxidative, and inflammatory factors, leading to endothelial dysfunction and vascular disease. The aim of this review is to focus on PVAT dysfunction mechanisms in obesity.

Imbalance between pro and anti-oxidant mechanisms in perivascular adipose tissue aggravates long-term high-fat diet-derived endothelial dysfunction.

Background The hypothesis of this study is that long-term high-fat diets (HFD) induce perivascular adipose tissue (PVAT) dysfunction characterized by a redox imbalance, which might contribute to aggravate endothelial dysfunction in obesity. Methods and Results C57BL/6J mice were fed either control or HFD (45% kcal from fat) for 32 weeks. Body weight, lumbar and mesenteric adipose tissue weights were significantly higher in HFD animals compared to controls. The anticontractile effect of PVAT in mesenteric arteries (MA) was lost after 32 week HFD and mesenteric endothelial-dependent relaxation was significantly impaired in presence of PVAT in HFD mice (Emax = 71.0±5.1 vs Emax = 58.5±4.2, p<0.001). The inhibitory effect of L-NAME on Ach-induced relaxation was less intense in the HFD group compared with controls suggesting a reduction of endothelial NO availability. Expression of eNOS and NO bioavailability were reduced in MA and almost undetectable in mesenteric PVAT of the HFD group. Superoxide levels and NOX activity were higher in PVAT of HFD mice. Apocynin only reduced contractile responses to NA in HFD animals. Expression of ec-SOD and total SOD activity were significantly reduced in PVAT of HFD mice. No changes were observed in Mn-SOD, Cu/Zn-SOD or catalase. The ratio [GSSG]/([GSH]+[GSSG]) was 2-fold higher in the mesenteric PVAT from HFD animals compared to controls. Conclusions We suggest that the imbalance between pro-oxidant (NOX, superoxide anions, hydrogen peroxide) and anti-oxidant (eNOS, NO, ecSOD, GSSG) mechanisms in PVAT after long-term HFD might contribute to the aggravation of endothelial dysfunction.

Native Adipose Stromal Cells Egress from Adipose Tissue In Vivo: Evidence During Lymph Node Activation†‡§

Adipose tissue (AT) has become accepted as a source of multipotent progenitor cells, the adipose stromal cells (ASCs). In this regard, considerable work has been performed to harvest and characterize this cell population as well as to investigate the mechanisms by which transplanted ASCs mediate tissue regeneration. In contrast the endogenous release of native ASCs by AT has been poorly investigated. In this work, we show that native ASCs egress from murine AT. Indeed, we demonstrated that the release of native ASCs from AT can be evidenced both using an ex vivo perfusion model that we set up and in vivo. Such a mobilization process is controlled by CXCR4 chemokine receptor. In addition, once mobilized from AT, circulating ASCs were found to navigate through lymph fluid and to home into lymph nodes (LN). Therefore, we demonstrated that, during the LN activation, the fat depot encapsulating the activated LN releases native ASCs, which in turn invade the activated LN. Moreover, the ASCs invading the LN were visualized in close physical interaction with podoplanin and ER‐TR7 positive structures corresponding to the stromal network composing the LN. This dynamic was impaired with CXCR4 neutralizing antibody. Taken together, these data provide robust evidences that native ASCs can traffic in vivo and that AT might provide stromal cells to activated LNs. STEM Cells2013;31:1309–1320

Circadian Feeding Drive of Metabolic Activity in Adipose Tissue and not Hyperphagia Triggers Overweight in Mice: Is There a Role of the Pentose-Phosphate Pathway?

High-fat (HF) diets trigger an increase in adipose tissue and body weight (BW) and disordered eating behavior. Our study deals with the hypothesis that circadian distribution of energy intake is more relevant for BW dynamics than diet composition. Four-week-old mice were exposed for 8 wk to a HF diet and compared with animals receiving control chow. HF mice progressively increased BW, decreased the amount of nocturnal (1800-0900 h) calories (energy or food intake) (30%) and increased diurnal (0900-1800 h) caloric intake (energy or food intake), although total daily intake was identical between groups. Animals were killed at 3-h intervals and plasma insulin, leptin, corticosterone, glucose, and fatty acid levels quantified. Adipose tissue was weighed, and enzymatic activities integral to the pentose phosphate pathway (PPP) assayed in lumbar adipose tissue. Phosphorylated AMP-dependent protein kinase and fatty acid synthase were quantified by Western blotting. In HF mice, there was a shift in the circadian oscillations of plasma parameters together with an inhibition of PPP activity and a decrease in phosphorylated AMP-dependent protein kinase and fatty acid synthase. In a second experiment, HF mice were forced to adhere to a circadian pattern of food intake similar to that in control animals. In this case, BW, adipose tissue, morning plasma parameters and PPP activity appeared to be normal. These data indicate that disordered feeding behavior can trigger BW gain independently of food composition and daily energy intake. Because PPP is the main source of reduced nicotinamide adenine dinucleotide phosphate, we suggest that PPP inhibition might be an early marker of adipose dysfunction in diet-induced obesity.

Leptin resistance develops spontaneously in mice during adult life in a tissue-specific manner. Consequences for hepatic steatosis.

Leptin is an adipocyte-derived hormone which stimulates β-oxidation in peripheral tissues and prevents steatosis. Because leptin production naturally increases during adult life, we have hypothesized that leptin receptors might undergo a physiological and gradual desensitization during ageing. Therefore we have characterized in three- five- and ten-month old mice i) the weight of different white adipose pads, heart and liver, ii) lipid content in these tissues/organs, and iii) responsiveness to acute leptin, measured in terms of phosphorylation of signal transducer and activator of transcription 3 (STAT3) and protein kinase B (Akt). In this study we have detected that leptin-mediated STAT3 phosphorylation appears to be preserved in cardiac tissue even in 10-month old animals but not in adipose tissue and liver of five- and ten-month old mice, respectively. Nevertheless, leptin increased pAkt content in the liver of these mice. In a parallel study we have analyzed the functionality of leptin signalling pathways in 10-month old obese mice and we have observed that the STAT3 pathway appears to be only operative in the heart whereas the Akt pathway remains functional both in heart and liver. Nevertheless, hepatic lipids increased almost 300% compared to age-matched lean controls. Our data demonstrate that during adult life there is a lost of leptin receptor functionality which is tissue-dependent and mainly affects the STAT3 pathway. Otherwise we demonstrate that the antisteatotic effect of leptin is independent of the Akt signalling pathway.

Mild caloric restriction reduces blood pressure and activates endothelial AMPK-PI3K-Akt-eNOS pathway in obese Zucker rats.

Genetic obesity models exhibit endothelial dysfunction associated to adenosine monophosphate-activated protein kinase (AMPK) dysregulation. This study aims to assess if mild short-term caloric restriction (CR) restores endothelial AMPK activity leading to an improvement in endothelial function. Twelve-week old Zucker lean and obese (fa/fa) male rats had access to standard chow either ad libitum (AL, n=8) or 80% of AL (CR, n=8) for two weeks. Systolic blood pressure was significantly higher in fa/fa AL rats versus lean AL animals, but was normalized by CR. Endothelium-dependent relaxation to acetylcholine (ACh, 10(-9) to 10(-4) M) was reduced in fa/fa AL compared to control lean AL rats (p<0.001), and restored by CR. The AMPK activator AICAR (10(-5) to 8·10(-3) M) elicited a lower relaxation in fa/fa AL rings that was normalized by CR (p<0.001). Inhibition of PI3K (wortmannin, 10(-7) M), Akt (triciribine, 10(-5) M), or eNOS (L-NAME, 10(-4) M) markedly reduced AICAR-induced relaxation in lean AL, but not in fa/fa AL rats. These inhibitions were restored by CR in Zucker fa/fa rings. These data show that mild short-term CR improves endothelial function and lowers blood pressure in obesity due to the activation of the AMPK-PI3K-Akt-eNOS pathway.