Amyloid-ß reduces the expression of neuronal FAIM-L, thereby shifting the inflammatory response mediated by TNFα from neuronal protection to death.

The brains of patients with Alzheimer's disease (AD) present elevated levels of tumor necrosis factor-α (TNFα), a cytokine that has a dual function in neuronal cells. On one hand, TNFα can activate neuronal apoptosis, and on the other hand, it can protect these cells against amyloid-ß (Aß) toxicity. Given the dual behavior of this molecule, there is some controversy regarding its contribution to the pathogenesis of AD. Here we examined the relevance of the long form of Fas apoptotic inhibitory molecule (FAIM) protein, FAIM-L, in regulating the dual function of TNFα. We detected that FAIM-L was reduced in the hippocampi of patients with AD. We also observed that the entorhinal and hippocampal cortex of a mouse model of AD (PS1(M146L)xAPP(751sl)) showed a reduction in this protein before the onset of neurodegeneration. Notably, cultured neurons treated with the cortical soluble fractions of these animals showed a decrease in endogenous FAIM-L, an effect that is mimicked by the treatment with Aß-derived diffusible ligands (ADDLs). The reduction in the expression of FAIM-L is associated with the progression of the neurodegeneration by changing the inflammatory response mediated by TNFα in neurons. In this sense, we also demonstrate that the protection afforded by TNFα against Aß toxicity ceases when endogenous FAIM-L is reduced by short hairpin RNA (shRNA) or by treatment with ADDLs. All together, these results support the notion that levels of FAIM-L contribute to determine the protective or deleterious effect of TNFα in neuronal cells.

TNFα induces survival through the FLIP-L-dependent activation of the MAPK/ERK pathway.

Activation of tumor necrosis factor receptor-1 can trigger survival or apoptosis pathways. In many cellular models, including the neuronal cell model PC12, it has been demonstrated that inhibition of protein synthesis is sufficient to render cells sensitive to apoptosis induced by TNFα. The survival effect is linked to the translocation of the transcription factor nuclear factor-kappa B (NF-κB) to the nucleus and activation of survival-related genes such as FLICE-like inhibitory protein long form (FLIP-L) or IAPs. Nonetheless, we previously reported an NF-κB-independent contribution of Bcl-xL to cell survival after TNFα treatment. Here, we demonstrate that NF-κB-induced increase in FLIP-L expression levels is essential for mitogen-activated protein kinases/extracellular signal-regulated kinases (MAPK/ERK) activation. We demonstrate that FLIP-L behaves as a Raf-1 activator through both protein-protein interaction and Raf-1 kinase activation, without the requirement of the classical Ras activation. Importantly, prevention of FLIP-L increase by NF-κB inhibition or knockdown of endogenous FLIP-L blocks MAPK/ERK activation after TNFα treatment. From a functional point of view, we show that inhibition of the MAPK/ERK pathway and the NF-κB pathway are equally relevant to render PC12 cells sensitive to cell death induced by TNFα. Apoptosis induced by TNFα under these conditions is dependent on jun nuclear kinase1/2 JNK1/2-dependent Bim upregulation. Therefore, we report a previously undescribed and essential role for MAPK/ERK activation by FLIP-L in the decision between cell survival and apoptosis upon TNFα stimulation.

Serum haptoglobin: a novel marker of adiposity in humans.

Haptoglobin (Hp) is a glycoprotein involved in the acute phase response to inflammation. Our previous findings indicate that Hp mRNA and protein are present in the adipose tissue of rodents and that Hp gene expression is up-regulated in obese models. The aim of the present study was to establish whether Hp could be considered a marker of obesity in humans. In 312 subjects, serum Hp was correlated directly with body mass index (BMI), leptin, C-reactive protein (CRP), and age. In a multivariate stepwise regression analysis, BMI and CRP were independent determinants of serum Hp in females, with BMI having the strongest effect. CRP and age were independent determinants of serum Hp in males, although explaining only a modest percentage of the total variability. Serum Hp was positively associated with body fat, as assessed by dual-energy x-ray absorptiometry, both in female and in male groups. The level of significance improved when serum Hp was analyzed against fat mass adjusted for lean mass. Finally, Northern and Western blot analyses performed in biopsies of sc abdominal fat from 20 obese individuals showed the presence of Hp mRNA and protein in the human adipose tissue. In conclusion, serum Hp constitutes a novel marker of adiposity in humans, and the adipose tissue likely contributes to determine its levels.

Interleukin-15 mediates reciprocal regulation of adipose and muscle mass: a potential role in body weight control.

Interleukin (IL)-15 is a cytokine which is highly expressed in skeletal muscle. Cell culture studies have indicated that IL-15 may have an important role in muscle fiber growth and anabolism. However, data concerning the metabolic effects of this cytokine in vivo are lacking. In the present study, IL-15 was administered to adult rats for 7 days. While IL-15 did not cause changes in either muscle mass or muscle protein content, it induced significant changes in the fractional rates of both muscle protein synthesis and degradation, with no net changes in protein accumulation. Additionally, IL-15 administration resulted in a 33% decrease in white adipose tissue mass and a 20% decrease in circulating triacylglycerols; this was associated with a 47% lower hepatic lipogenic rate and a 36% lower plasma VLDL triacylglycerol content. The decrease in white fat induced by IL-15 was in adipose tissue. No changes were observed in the rate of lipolysis as a result of cytokine administration. These findings indicate that IL-15 has significant effects on both protein and lipid metabolism, and suggest that this cytokine may participate in reciprocal regulation of muscle and adipose tissue mass.

Interleukin-15 antagonizes muscle protein waste in tumour-bearing rats.

Tissue protein hypercatabolism (TPH) is an important feature in cancer cachexia, particularly with regard to the skeletal muscle. The Yoshida AH-130 rat ascites hepatoma is a model system for studying the mechanisms involved in the processes that lead to tissue depletion, since it induces in the host a rapid and progressive muscle wasting, primarily due to TPH. The present study was aimed at investigating if IL-15, which is known to favour muscle fibre hypertrophy, could antagonize the enhanced muscle protein breakdown in this cancer cachexia model. Indeed, IL-15 treatment partly inhibited skeletal muscle wasting in AH-130-bearing rats by decreasing (8-fold) protein degradative rates (as measured by 14C-bicarbonate pre-loading of muscle proteins) to values even lower than those observed in non-tumour-bearing animals. These alterations in protein breakdown rates were associated with an inhibition of the ATP-ubiquitin-dependent proteolytic pathway (35% and 41% for 2.4 and 1.2 kb ubiquitin mRNA, and 57% for the C8 proteasome subunit, respectively). The cytokine did not modify the plasma levels of corticosterone and insulin in the tumour hosts. The present data give new insights into the mechanisms by which IL-15 exerts its preventive effect on muscle protein wasting and seem to warrant the implementation of experimental protocols involving the use of the cytokine in the treatment of pathological states characterized by TPH, particularly in skeletal muscle, such as in the present model of cancer cachexia.

Leptin administration to lactating rats is unable to induce changes in lipid metabolism in white adipose tissue or mammary gland.

During lactation in the rat, despite hyperphagia, there are no changes in either the plasma levels or the gene expression of leptin. Removal of the litter, however, results in an important increase in the circulating concentration of leptin. Administration of leptin to lactating rats resulted in no changes in the in vivo lipogenic rate and lipoprotein lipase (LPL) activity in either adipose tissue or mammary gland, although there was an increase in insulin levels as a consequence of leptin administration. Conversely, litter removal resulted in an important decrease of LPL activity and lipogenic rate in the mammary gland while an increase in these parameters took place in adipose tissue. It is concluded that leptin is not the signal responsible for the changes in lipid metabolism that take place both in adipose tissue and mammary gland following litter removal.

Leptin and tumor growth in rats.

We have examined the role of leptin in tumor-induced anorexia in 2 different tumor models. In rats bearing the Yoshida AH-130 ascites hepatoma, the reduction in food intake becomes important from day 6 after tumor inoculation. Interestingly, at day 4, when the animals do not show any anorectic behavior, circulating leptin levels were already reduced. Indeed, in all the tumor-bearing groups studied the levels of leptin were lower than in control animals. Moreover, the changes in the circulating levels paralleled changes in adipose tissue leptin mRNA expression, even at early stages following tumor inoculation when neither food intake nor fat stores were modified by the presence of a tumor. Interestingly, 7-day pair-fed controls showed changes similar to those present in tumor-bearing rats. These results agree with previous observations relating fasting to decreased leptin expression. Similar results were observed in another tumor model, the mouse Lewis lung carcinoma; i.e., at day 8 after tumor inoculation (when the animals did not show anorexia) both the circulating levels and the adipose leptin mRNA expression were also reduced. Our results suggest that experimental cancer-induced anorexia is not related to leptin changes.

Leptin levels and gene expression during the perinatal phase in the rat.

The role of leptin in controlling food intake and adiposity has been the aim of many different investigations in the last 3 years. Pregnancy and lactation are two physiological situations associated with a clear hyperphagia (together with important changes in metabolism and adipose mass) to sustain the different and varying demands for foetal growth and milk production respectively. We therefore focused on the role of leptin in perinatal hyperphagia. The circulating leptin levels and leptin gene expression in adipose tissue of both pregnant and lactating rats were examined. Pregnant rats showed unchanged adipose tissue leptin mRNA levels but increased circulating leptin; this probably reflects the high fat carcass content characteristic of pregnancy. Conversely, lactating rats did not show any change either in circulating leptin or adipose tissue mRNA levels. Litter-removal caused a significant increase in both circulating leptin levels and gene expression. The results obtained permit us to suggest that leptin does not seem to have a role in controlling food intake during the perinatal phase.

Role of TNF receptor 1 in protein turnover during cancer cachexia using gene knockout mice.

The implantation of the Lewis lung carcinoma (a fast-growing mouse tumour that induces cachexia) to both wild-type and gene-deficient mice for the TNF-alpha receptor type I protein (Tnfr1 degree/Tnfr1 degree), resulted in a considerable loss of carcass weight in both groups. However, while in the wild-type mice there was a loss of both fat and muscle, in the gene-knockout mice muscle wastage was not affected to the same extent. In both groups, tumour burden resulted in significant increases in circulating TNF-alpha, a cytokine which, as we have previously demonstrated, can induce protein breakdown in skeletal muscle. Muscle wastage in wild-type mice was accompanied by an increase in the fractional rate of protein degradation, while no changes were observed in protein synthesis. The result is a decreased rate of protein accumulation that accounts for the muscle weight loss observed as a result of tumour burden. In contrast, gene knockout mice did not have significantly lower rates of protein accumulation as a result of tumour implantation. The increase in protein degradation in the tumour-bearing wild mice was accompanied by an enhanced expression of both ubiquitin and proteasome subunit genes, all of them related to the activation of the ATP-dependent proteolytic system in skeletal muscle. Tumour-bearing gene-deficient mice did not show any increase in gene expression. It is concluded that TNF-alpha (alone or in combination with other cytokines) is responsible for the activation of protein breakdown in skeletal muscle of tumour-bearing mice.

Protein turnover in skeletal muscle of tumour-bearing transgenic mice overexpressing the soluble TNF receptor-1.

The implantation of the Lewis lung carcinoma (a fast-growing mouse tumour that induces cachexia) to both wild-type and transgenic mice for the soluble TNF receptor type I protein (sTNF-R1) resulted in a considerable loss of carcass weight in both groups. However, while in the wild-type mice there was a loss of both fat and muscle, in the transgenic mice muscle waste was not affected to the same extent as in the wild-type group. Muscle waste in wild-type mice was accompanied by an increase in the fractional rate of protein degradation, while no changes were observed in protein synthesis. The result was a decreased rate of protein accumulation which accounted for the muscle weight loss observed as a result of the tumour burden. In contrast, transgenic mice did not have such low rates of protein accumulation after tumour implantation. The increase in protein degradation in the tumour-bearing transgenic mice was accompanied by a similar increase in protein synthesis which compensated for the loss of muscle protein by degradation. Both tumour-bearing groups showed an enhanced expression of ubiquitin and proteasome C8 subunit genes, all of them related to the activation of the ATP-dependent proteolytic system in skeletal muscle. It is suggested that TNF may, in part, be responsible for the loss of protein in skeletal muscle of tumour-bearing mice.

Short-term effects of leptin on lipid metabolism in the rat.

In this study, we have examined the short-term effects of leptin on lipid metabolism in the rat. Acute leptin administration induced hypertriglyceridaemia (31% increase in plasma triacylglycerols) which was not associated with changes in lipoprotein lipase activity in white adipose tissue. Surprisingly, leptin administration did not induce any changes in the lipogenic rate in either white adipose tissue or liver. Leptin administration caused a decreased tissue uptake of exogenous 14C-triacylglycerols. These data suggest that leptin induces important changes in lipid uptake in adipose tissue and skeletal muscle which could be responsible for the observed hypertriglyceridaemia.

Different cytokines modulate ubiquitin gene expression in rat skeletal muscle.

Intravenous administration of different cytokines caused important changes in the expression of ubiquitin genes in skeletal muscle. Tumour necrosis factor-alpha caused a 2.2- and 1.9-fold increase in the expression of the 2.4 and 1.2 kb transcripts, respectively. Administration of interferon-gamma also caused a 2.2- and 1.8-fold increase in the 2.4 and 1.2 kb transcripts, respectively. While administration of leukaemia inhibitory factor and interleukin-6 resulted in no changes in ubiquitin gene expression, interleukin-1 administration also caused an increase in both ubiquitin gene transcripts (2.8- and 1.9-fold for the 2.4 and 1.2 kb transcripts, respectively). The results suggest that some of the cytokine effects on the ubiquitin system gene expression could be related to the enhanced skeletal muscle proteolysis found during cancer cachexia and other pathological states.

Lipogenesis in rat tissues following carbohydrate refeeding: spleen lipogenesis is modulated by insulin.

Intraperitoneal administration of [1,2-14C]-acetate to Wistar rats was used to assess tissue lipogenic rates after estimating the incorporation of the label into the tissular lipid fractions. Refeeding the animals with glucose (after an overnight fast) induced an increase in white adipose tissue (4.5 fold), liver (4.1 fold), small intestine (1.9 fold), carcass (2.9 fold) and spleen (3.7 fold) lipogenesis (expressed as the radioactivity present in the lipid fraction corrected by the plasma circulating radioactivity). No changes were found following refeeding in either brain or brown adipose tissue. Administration of mannoheptulose (an inhibitor of insulin secretion) to refed rats completely abolished the increased lipogenesis in white adipose tissue, liver, carcass, spleen and small intestine, thus suggesting that insulin secretion is involved in this phenomenon. This is the first report showing that spleen lipogenesis may be modulated by refeeding via insulin secretion and suggests an important role of this organ on the in vivo lipogenic response of the organism after carbohydrate refeeding.

Lipid metabolism in tumour-bearing mice: studies with knockout mice for tumour necrosis factor receptor 1 protein.

The implantation of the Lewis lung carcinoma (a fast-growing mouse tumour that induces cachexia) to both wild-type and gene-deficient mice for the tumour necrosis factor (TNF) receptor type I protein (Tnfr1(0)/Tnfr1(0)), resulted in a considerable loss of carcass (26%) and white (77%) and brown adipose (37%) tissue weights in the wild-type mice, while it induced much less marked effects in the gene-deficient mice. Tumour burden also inflicted an important decrease in total lipoprotein lipase (LPL) activity in epididymal white adipose tissue (50%) in the wild-type mice while no changes were observed in the knockout mice. In addition, all tumour-bearing animals were clearly hypertriglyceridaemic (80% increase in circulating triacylglycerols in wild-type and 36% in knockout mice). It is concluded that although TNF seems to be to some extent responsible for adipose waste, LPL changes and hyperlipaemia (via receptor I), the role of other cytokines (alone or in combination with TNF) in promoting changes in lipid metabolism during cancer cachexia cannot be discarded.

Journey from cachexia to obesity by TNF.

Tumor necrosis factor-alpha (TNF-alpha) is a cytokine involved in the physiological and metabolic abnormalities found in cachectic states. Until very recently, it was inconceivable to think of TNF-alpha in obesity. However, recent studies have shown that TNF-alpha can also play a key role in obesity, the cytokine being overexpressed in adipose tissue of obese rodents and humans. The aim of this review is to reconcile the role of TNF-alpha in these two opposite metabolic situations: obesity and cachexia. It is suggested that TNF-alpha may have a key role in the control of body mass in normal weight-controlled situations and that abnormalities in either its production (during cachexia) or action (during obesity) are responsible for the lack of control of body weight.

Sequential changes in lipoprotein lipase activity and lipaemia induced by the Yoshida AH-130 ascites hepatoma in rats.

The implantation of the Yoshida AH-130 ascites hepatoma to rats resulted in an exponential growth of the tumour cells followed by a late stationary phase. The tumour burden was accompanied by a dramatic decrease in body weight. Tumour growth was associated with a marked hypertriglyceridaemia during the period of exponential growth, while in the stationary phase the plasma triacylglycerol concentration was similar to that observed in the non-tumour-bearing animals. Similar increases were observed, following tumour inoculation, in the plasma concentrations of non-esterified fatty acids and glycerol, suggesting an intense lipolytic activity. These changes in lipaemia were associated with a marked decrease in LPL activity in white adipose tissue; in contrast, LPL activity was increased in the tumour-bearing animals in brown adipose tissue at day 6 following inoculation and in the heart during most of the period studied. Although the presence of the tumour did not induce any changes in blood lactate concentrations, it caused a decrease in circulating glucose; conversely, the tumour induced an important increase in the concentration of circulating ketone bodies, suggesting a metabolic adaptation of the tumour-bearing rats to glucose sparing and alternative fuel utilization. It may be suggested that the hyperlipidaemia present in the Yoshida AH-130 bearing rats is partly due to a decreased LPL activity in white adipose tissue which does not seem to be influenced by changes in insulin circulating concentrations.

Comparative effects of beta2-adrenergic agonists on muscle waste associated with tumour growth.

The implantation of the Yoshida AH-130 ascites hepatoma (a fast growing tumour) to rats resulted in a dramatic loss of both white adipose tissue and muscle (skeletal and cardiac) mass. Administration of beta2-adrenergic agonists to tumour-bearing rats resulted in a partial recovery of skeletal muscle and heart mass. Treatment of the tumour-bearing animals with the different drugs (salbutamol, salmeterol and clenbuterol) did not influence tumour growth or food intake so it can be suggested that the effects were solely due to metabolic changes. In addition, while the three drugs had clear effects on gastrocnemius muscles, clenbuterol and salbutamol had also an effect on soleus, and salbutamol had a clear effect on cardiac muscle. It is suggested that any of the studied beta2-adrenergic agonists (but perhaps, particularly salmeterol) could be used clinically in the treatment of cancer cachexia.

Anti-TNF treatment does not reverse the abnormalities in lipid metabolism of the obese Zucker rat.

Because obesity, insulin resistance, and hyperlipidemia are often associated, and recent evidence suggests that the cytokine tumor necrosis factor-alpha (TNF) may influence the activity of insulin in various target tissues, the present study was designed to see whether TNF was also associated with the changes in lipid metabolism that lead to hyperlipidemia in the obese model of the Zucker rat. A polyclonal goat anti-rat TNF antibody was subcutaneously administered to Zucker rats for 4 days to block TNF actions. The results indicate that none of the alterations in lipid metabolism seen in the obese animals were reversed by the anti-TNF treatment. This was the case for the lipogenic rate in liver and adipose tissue, the disposal of an exogenous [14C]triolein load, adipose tissue lipoprotein lipase activity, and the hypertriglyceridemia. Measurements of lipolysis in adipose tissue slices from the anti-TNF-treated animals also did not show any significant effect of the treatment. In conclusion, TNF does not seem to be involved in the abnormalities of lipid metabolism observed in the obese Zucker rat.

Lipid metabolism in rats bearing the Yoshida AH-130 ascites hepatoma.

Rats bearing the Yoshida AH-130 ascites hepatoma showed important changes in lipid metabolism. The presence of this rapidly growing tumour induced a significant reduction in the intestinal absorption of an oral [14C]triolein load but without changes in whole body oxidation of the tracer to CO2. Both white (WAT) and brown (BAT) adipose tissue lipoprotein lipase (LPL) activities were increased at day 4 of tumour growth, changes that seem to be related with those observed in [14C]lipid accumulation; however, heart LPL activity was increased at day 7 but there was no change at day 4. In addition, there was a marked hyperlipemia in the tumour-bearing animals, whereas the blood ketone body concentrations were lower in these animals in comparison with the corresponding pair-fed group. The in vivo lipogenic rate was increased in liver of the tumour-bearing animals (day 4); conversely, it was decreased in WAT and skeletal muscle (day 4) and IBAT (day 7) of the AH-130-bearing rats. It may be suggested that the increased liver lipogenic rate associated with tumour burden is the main factor contributing to the hyperlipidaemia present in the Yoshida AH-130 bearing rats.

alpha-Adrenergic receptors may contribute to the hypertriglyceridemia associated with tumour growth.

The inoculation of the Yoshida AH-130 ascites hepatoma to rats resulted in an important loss of adipose tissue associated with a decrease in lipoprotein lipase (LPL) activity. Tumour burden also resulted in an important hyperlipidemia which affected both triglyceride and free fatty acids. Administration of phentolamine (an alpha-adrenergic antagonist) to tumour-bearing rats did not influence LPL activity, but it reversed the increase in plasma triglycerides associated with tumour burden. It is suggested that the hypertriglyceridemia associated with tumour growth may be, in part, a consequence of the effect of catecholamines on hepatic triglyceride secretion, via alpha-adrenergic receptors.