Short-term oleoyl-estrone treatment affects capacity to manage lipids in rat adipose tissue

Background: Short-term OE (oleoyl-estrone) treatment causes significant decreases in rat weight mainly due to adipose tissue loss. The aim of this work was to determine if OE treatment affects the expression of genes that regulate lipid metabolism in white adipose tissue. Results: Gene expression in adipose tissue from female treated rats (48 hours) was analysed by hybridization to cDNA arrays and levels of specific mRNAs were determined by real-time PCR. Treatment with OE decreased the expression of 232 genes and up-regulated 75 other genes in mesenteric white adipose tissue. The use of real-time PCR validate that, in mesenteric white adipose tissue, mRNA levels for Lipoprotein Lipase (LPL) were decreased by 52%, those of Fatty Acid Synthase (FAS) by 95%, those of Hormone Sensible Lipase (HSL) by 32%, those of Acetyl CoA Carboxylase (ACC) by 92%, those of Carnitine Palmitoyltransferase 1b (CPT1b) by 45%, and those of Fatty Acid Transport Protein 1 (FATP1) and Adipocyte Fatty Acid Binding Protein (FABP4) by 52% and 49%, respectively. Conversely, Tumour Necrosis Factor (TNF¿) values showed overexpression (198%). Conclusion: Short-term treatment with OE affects adipose tissue capacity to extract fatty acids from lipoproteins and to deal with fatty acid transport and metabolism.

Different modulation of adipocytokine expression in four main white adipose tissue sites in the rat: mesenteric, perigonadal, retroperitoneal and subcutaneous

White adipose tissue (WAT) is a disperse organ acting as energy storage depot and endocrine/paracrine controlling factor in the management of energy availability and inflammation. WAT sites response under energy-related stress is not uniform. In the present study we have analyzed how different WAT sites respond to limited food restriction as a way to better understand the role of WAT in the pathogenesis of the metabolic syndrome.

FiltekTM Silorane and FiltekTM Supreme XT resins: tissue reaction after subcutaneous implantation in isogenic mice

The aim of this study was to evaluate the tissue compatibility of a silorane-based resin system (FiltekTM Silorane) and a methacrylatebased nanoparticle resin (FiltekTM Supreme XT) after implantation in the subcutaneous connective tissue of isogenic mice. One hundred and thirty five male isogenic BALB/c mice were randomly assigned to 12 experimental and 3 control groups, according to the implanted material and the experimental period of 7, 21 and 63 days. At the end of each period, the animals were killed and the tubes with the surrounding tissues were removed and processed for microscopic analysis. Samples were subjected to a descriptive and a semi-quantitative analyses using a 4-point scoring system (0-3) to evaluate the collagen fiber formation and inflammatory infiltrate. Data were statistically analyzed using the Kruskal Wallis test (a=0.05). The results showed that there was no significant difference between the experimental and control groups considering the three evaluation periods (p>0.05). The silorane-based and the methacrylate-based nanoparticle resins presented similar tissue response to that of the empty tube (control group) after subcutaneous implantation in isogenic mice.

Selenium uptake by edible plants from enriched peat.

As a constituent of selenoproteins, selenium (Se) is considered an essential element for human health.The main way that Se enters the body is via the consumption of vegetables, whose concentration of thiselement depends on soil Se content. We grew cabbage, lettuce, chard and parsley, in peat enriched in Seby means of the additive Selcote Ultra®and Na2SeO3and Na2SeO4. Total Se in plants was determinedby acidic digestion and Se speciation by an enzymatic extraction. Both were measured by ICP/MS. Theconcentration ranges were between 0.1 mg Se kg−1and 30 mg Se kg−1for plants grown in Selcote Ultra®media, and between 0.4 mg Se kg−1and 1606 mg Se kg−1for those grown in peat enriched with Se sodiumsalts. We found Se (IV), Se (VI) and SeMet in all the extracts. Peat fortified with Selcote Ultra®gave slightlyhigher Se concentration than natural content values. For plants grown with selenium sodium salts, Secontent increases with the Se added and part of the inorganic Se was converted mainly to SeMet. A highSe fortification can damage or inhibit plant growth. Cabbage showed the greatest tolerance to Se.

Positional bias of general and tissue-specific regulatory motifs in mouse gene promoters

Background: The arrangement of regulatory motifs in gene promoters, or promoterarchitecture, is the result of mutation and selection processes that have operated over manymillions of years. In mammals, tissue-specific transcriptional regulation is related to the presence ofspecific protein-interacting DNA motifs in gene promoters. However, little is known about therelative location and spacing of these motifs. To fill this gap, we have performed a systematic searchfor motifs that show significant bias at specific promoter locations in a large collection ofhousekeeping and tissue-specific genes.Results: We observe that promoters driving housekeeping gene expression are enriched inparticular motifs with strong positional bias, such as YY1, which are of little relevance in promotersdriving tissue-specific expression. We also identify a large number of motifs that show positionalbias in genes expressed in a highly tissue-specific manner. They include well-known tissue-specificmotifs, such as HNF1 and HNF4 motifs in liver, kidney and small intestine, or RFX motifs in testis,as well as many potentially novel regulatory motifs. Based on this analysis, we provide predictionsfor 559 tissue-specific motifs in mouse gene promoters.Conclusion: The study shows that motif positional bias is an important feature of mammalianproximal promoters and that it affects both general and tissue-specific motifs. Motif positionalconstraints define very distinct promoter architectures depending on breadth of expression andtype of tissue.

Changes in lipoprotein lipase modulate tissular energy supply during stress

We studied the variations caused by stress in lipoprotein lipase (LPL) activity, LPL-mRNA, and local blood flow in LPL-rich tissues in the rat. Stress was produced by body immobilization (Immo): the rat's limbs were taped to metal mounts, and its head was placed in a plastic tube. Chronic stress (2 h daily of Immo) decreased total LPL activity in mesenteric and epididymal white adipose tissue (WAT) and was accompanied by a weight reduction of these tissues. In limb muscle, heart, and adrenals, total LPL activity and mRNA levels increased, and, in plasma, LPL activity and mass also increased. Acute stress (30-min Immo) caused a decrease in total LPL activity only in retroperitoneal WAT and an increase in preheparin plasma active LPL, but the overall weight of this tissue did not vary significantly. We propose an early release of the enzyme from this tissue into the bloodstream by some unknown extracellular pathways or other local mechanisms. These changes in this key energy-regulating enzyme are probably induced by catecholamines. They modify the flow of energy substrates between tissues, switching the WAT from importer to exporter of free fatty acids and favoring the uptake by muscle of circulating triacylglycerides for energy supply. Moreover, we found that acute stress almost doubled blood flow in all WAT studied, favoring the export of free fatty acids.

Nitric oxide and the release of lipoprotein lipase from white adipose tissue

Background/Aim: Lipoprotein lipase (LPL) is the main enzyme responsible for the distribution of circulating triacylglycerides in tissues. Its regulation via release from active sites in the vascular endothelium is poorly understood. In a previous study we reported that in response to acute immobilization (IMMO), LPL activity rapidly increases in plasma and decreases in white adipose tissue (WAT) in rats. In other stress situations IMMO triggers a generalized increase in nitric oxide (NO) production. Methods/Results: Here we demonstrate that in rats: 1) in vivo acute IMMO rapidly increases NO concentrations in plasma 2) during acute IMMO the WAT probably produces NO via the endothelial isoform of nitric oxide synthase (eNOS) from vessels, and 3) epididymal WAT perfused in situ with an NO donor rapidly releases LPL from the endothelium. Conclusion: We propose the following chain of events: stress stimulus / rapid increase of NO production in WAT (by eNOS) / release of LPL from the endothelium in WAT vessels. This chain of events could be a new mechanism that promotes the rapid decrease of WAT LPL activity in response to a physiological stimulus.

Tumor necrosis factor-alpha mediates changes in tissue protein turnover in a rat cancer cachexia model.

Rats bearing the Yoshida AH-130 ascites hepatoma showed enhanced fractional rates of protein degradation in gastrocnemius muscle, heart, and liver, while fractional synthesis rates were similar to those in non-tumor bearing rats. This hypercatabolic pattern was associated with marked perturbations of the hormonal homeostasis and presence of tumor necrosis factor in the circulation. The daily administration of a goat anti-murine TNF IgG to tumor-bearing rats decreased protein degradation rates in skeletal muscle, heart, and liver as compared with tumor-bearing rats receiving a nonimmune goat IgG. The anti-TNF treatment was also effective in attenuating early perturbations in insulin and corticosterone homeostasis. Although these results suggest that tumor necrosis factor plays a significant role in mediating the changes in protein turnover and hormone levels elicited by tumor growth, the inability of such treatment to prevent a reduction in body weight implies that other mediators or tumor-related events were also involved.

TRPM5-mediated calcium uptake regulates mucin secretion from human colon goblet cells

Mucin 5AC (MUC5AC) is secreted by goblet cells of the respiratory tract and, surprisingly, also expressed de novo in mucus secreting cancer lines. siRNA-mediated knockdown of 7343 human gene products in a human colonic cancer goblet cell line (HT29-18N2) revealed new proteins, including a Ca(2+)-activated channel TRPM5, for MUC5AC secretion. TRPM5 was required for PMA and ATP-induced secretion of MUC5AC from the post-Golgi secretory granules. Stable knockdown of TRPM5 reduced a TRPM5-like current and ATP-mediated Ca(2+) signal. ATP-induced MUC5AC secretion depended strongly on Ca(2+) influx, which was markedly reduced in TRPM5 knockdown cells. The difference in ATP-induced Ca(2+) entry between control and TRPM5 knockdown cells was abrogated in the absence of extracellular Ca(2+) and by inhibition of the Na(+)/Ca(2+) exchanger (NCX). Accordingly, MUC5AC secretion was reduced by inhibition of NCX. Thus TRPM5 activation by ATP couples TRPM5-mediated Na(+) entry to promote Ca(2+) uptake via an NCX to trigger MUC5AC secretion

Muscle transcriptomic profiles in pigs with divergent phenotypes for fatness traits

BACKGROUND: Selection for increasing intramuscular fat content would definitively improve the palatability and juiciness of pig meat as well as the sensorial and organoleptic properties of cured products. However, evidences obtained in human and model organisms suggest that high levels of intramuscular fat might alter muscle lipid and carbohydrate metabolism. We have analysed this issue by determining the transcriptomic profiles of Duroc pigs with divergent phenotypes for 13 fatness traits. The strong aptitude of Duroc pigs to have high levels of intramuscular fat makes them a valuable model to analyse the mechanisms that regulate muscle lipid metabolism, an issue with evident implications in the elucidation of the genetic basis of human metabolic diseases such as obesity and insulin resistance. RESULTS: Muscle gene expression profiles of 68 Duroc pigs belonging to two groups (HIGH and LOW) with extreme phenotypes for lipid deposition and composition traits have been analysed. Microarray and quantitative PCR analysis showed that genes related to fatty acid uptake, lipogenesis and triacylglycerol synthesis were upregulated in the muscle tissue of HIGH pigs, which are fatter and have higher amounts of intramuscular fat than their LOW counterparts. Paradoxically, lipolytic genes also showed increased mRNA levels in the HIGH group suggesting the existence of a cycle where triacylglycerols are continuously synthesized and degraded. Several genes related to the insulin-signalling pathway, that is usually impaired in obese humans, were also upregulated. Finally, genes related to antigen-processing and presentation were downregulated in the HIGH group. CONCLUSION: Our data suggest that selection for increasing intramuscular fat content in pigs would lead to a shift but not a disruption of the metabolic homeostasis of muscle cells. Future studies on the post-translational changes affecting protein activity or expression as well as information about protein location within the cell would be needed to to elucidate the effects of lipid deposition on muscle metabolism in pigs.

Treatment of rats with a self-selected hyperlipidic diet, increases the lipid content of the main adipose tissue sites in a proportion similar to that of the lipids in the rest of organs and tissues

Adipose tissue (AT) is distributed as large differentiated masses, and smaller depots covering vessels, and organs, as well as interspersed within them. The differences between types and size of cells makes AT one of the most disperse and complex organs. Lipid storage is partly shared by other tissues such as muscle and liver. We intended to obtain an approximate estimation of the size of lipid reserves stored outside the main fat depots. Both male and female rats were made overweight by 4-weeks feeding of a cafeteria diet. Total lipid content was analyzed in brain, liver, gastrocnemius muscle, four white AT sites: subcutaneous, perigonadal, retroperitoneal and mesenteric, two brown AT sites (interscapular and perirenal) and in a pool of the rest of organs and tissues (after discarding gut contents). Organ lipid content was estimated and tabulated for each individual rat. Food intake was measured daily. There was a surprisingly high proportion of lipid not accounted for by the main macroscopic AT sites, even when brain, liver and BAT main sites were discounted. Muscle contained about 8% of body lipids, liver 1-1.4%, four white AT sites lipid 28-63% of body lipid, and the rest of the body (including muscle) 38-44%. There was a good correlation between AT lipid and body lipid, but lipid in"other organs" was highly correlated too with body lipid. Brain lipid was not. Irrespective of dietary intake, accumulation of body fat was uniform both for the main lipid storage and handling organs: large masses of AT (but also liver, muscle), as well as in the"rest" of tissues. These storage sites, in specialized (adipose) or not-specialized (liver, muscle) tissues reacted in parallel against a hyperlipidic diet challenge. We postulate that body lipid stores are handled and regulated coordinately, with a more centralized and overall mechanisms than usually assumed.

Persistence of tracer in the application site -a potential confounding factor in nerve regeneration studies

Selective reinnervation of peripheral targets after nerve injury might be assessed by injecting a first tracer in a target before nerve injury to label the original neuronal population, and applying a second tracer after the regeneration period to label the regenerated population. However, altered uptake of tracer, fading, and cell death may interfere with the results. Furthermore, if the first tracer injected remains in the target tissue, available for 're-uptake' by misdirected regenerating axons, which originally innervated another region, then the identification of the original population would be confused. With the aim of studying this problem, the sciatic nerve of adult rats was sectioned and sutured. After 3 days, to allow the distal axon to degenerate avoiding immediate retrograde transport, one of the dyes: Fast Blue (FB), Fluoro-Gold (FG) or Diamidino Yellow (DY), was injected into the tibial branch of the sciatic nerve, or in the skin of one of the denervated digits. Rats survived 2-3 months. The results showed labelled dorsal root ganglion (DRG) cells and motoneurones, indicating that late re-uptake of a first tracer occurs. This phenomenon must be considered when the model of sequential labelling is used for studying the accuracy of peripheral reinnervation.

Tissue distribution of retinoids in common dolphins (Delphinus delphis).

Exposure to organochlorines induces retinoid deficiency in mammals; hence, retinoids are potential biomarkers of the impact of these pollutants. Appropriate target tissues to monitor retinoids in cetaceans have not been properly identified because of a lack of information on the contribution of each tissue to total body retinoids. Therefore, we have addressed this issue by studying the contribution of the main body tissues to retinoids in 21 common dolphins obtained from incidental catches and in apparent good health and nutritive condition. Although concentrations in the liver were highest, those in blubber were also high and accounted for 43% of the total retinoid load of the compartments examined. As blubber can be obtained using non-invasive biopsy techniques, this tissue is proposed as a reliable indicator of retinoid status in cetaceans. However, blubber topographical variation in structure and composition requires standardization of sampling sites. Retinoid concentrations did not differ significantly between sexes or with body size for any of the tissues, but the lipid content of blubber strongly influenced these concentrations. Biopsies from healthy, free-ranging individuals are preferred to samples from stranded animals. Further research on the influence of factors (age, sex, reproductive condition, diet) that potentially affect retinoid levels is required to implement the use of retinoids as biomarkers of pollutant exposure in cetaceans.

Effects of heparin on the uptake of lipoprotein lipase in rat liver

BACKGROUND: Lipoprotein lipase (LPL) is anchored at the vascular endothelium through interaction with heparan sulfate. It is not known how this enzyme is turned over but it has been suggested that it is slowly released into blood and then taken up and degraded in the liver. Heparin releases the enzyme into the circulating blood. Several lines of evidence indicate that this leads to accelerated flux of LPL to the liver and a temporary depletion of the enzyme in peripheral tissues. RESULTS: Rat livers were found to contain substantial amounts of LPL, most of which was catalytically inactive. After injection of heparin, LPL mass in liver increased for at least an hour. LPL activity also increased, but not in proportion to mass, indicating that the lipase soon lost its activity after being bound/taken up in the liver. To further study the uptake, bovine LPL was labeled with 125I and injected. Already two min after injection about 33 % of the injected lipase was in the liver where it initially located along sinusoids. With time the immunostaining shifted to the hepatocytes, became granular and then faded, indicating internalization and degradation. When heparin was injected before the lipase, the initial immunostaining along sinusoids was weaker, whereas staining over Kupffer cells was enhanced. When the lipase was converted to inactive before injection, the fraction taken up in the liver increased and the lipase located mainly to the Kupffer cells. CONCLUSIONS: This study shows that there are heparin-insensitive binding sites for LPL on both hepatocytes and Kupffer cells. The latter may be the same sites as those that mediate uptake of inactive LPL. The results support the hypothesis that turnover of endothelial LPL occurs in part by transport to and degradation in the liver, and that this transport is accelerated after injection of heparin.

Cardiac tissue engineering by electrical stimulation with subcutaneous and cardiac adipose-tissue derived progenitor cells (ATDPCs)

A major challenge of cardiac tissue engineering is directing cells to establish the physiological structure and function of the myocardium being replaced. In native heart, pacing cells generate electrical stimuli that spread throughout the heartcausing cell membrane depolarization and activation of contractile apparatus. We ought to examine whether electricalstimulation of adipose tissue-derived progenitor cells (ATDPCs) exerts phenotypic and genetic changes that enhance theircardiomyogenic potential.