The mode of action of a lipid mobilising factor in cancer cachexia

Cachexia is characterised by a progressive weight loss due to depletion of both skeletal muscle and adipose tissue. The loss of adipose tissue is due to the production of a tumour-derived lipid mobilising factor (LMF), which has been shown to directly induce lipolysis in isolated epididymal murine white adipocytes. The administration of LMF to a non-tumour bearing mice produced a rapid weight loss, with a specific reduction in carcass lipid with also some redistribution of lipid with the accumulation of lipid in the liver. There was also up-regulation of uncoupling protein-1 and -2 mRNA and protein expression in brown adipose tissue, suggesting that an adaptive process occurs due to increased energy mobilisation. There was also up-regulation of UCP-2 in the livers of LMF treated mice, suggesting a protective mechanism to the build up of lipid in the livers, which would produce free radical by-products. LMF was also shown to stimulate cyclic AMP production in CHO-K1 cells transfected with human -3 adrenergic receptors and inhibited by the -β3 antagonist SR59230A. LMF binding was also inhibited by SR59230A in isolated receptors. This suggests that LMF mediates its effects through a β3 adrenergic receptor. There were also changes in glucose and fatty acid uptake in LMF treated mice, which suggests metabolic changes are occurring. The study suggests that a tumour derived lipolytic factor acts through the 3 adrenoceptor producing effects on lipid mobilisation, energy expenditure and glucose metabolism.

Studies on the antiobesity effect of zinc-α2-glycoprotein in the ob/ob mouse

OBJECTIVE: To investigate the mechanism of the lipid depletion by zinc-a(2)-glycoprotein (ZAG). DESIGN: Studies were conducted in the ob/ob mouse, or on isolated adipocytes from these animals or their lean counterparts. RESULTS: Treatment of these animals for 15 days with ZAG (100? µg, intravenously, daily) resulted in a reduction of body weight of 6.55? g compared with phosphate-buffered saline-treated controls, without a change in food or water intake, but with a 0.4?°C rise in rectal temperature. ZAG-treated mice had a 30% reduction in carcass fat mass and a twofold increase in weight of brown adipose tissue. Epididymal adipocytes from ZAG-treated mice showed an increased expression of ZAG and hormone-sensitive lipase (HSL), and this was maintained for a further 3 days in the absence of ZAG. There was an increased lipolytic response to isoproterenol, which was retained for 3 days in vitro in the absence of ZAG. Expression of HSL was also increased in subcutaneous and visceral adipose tissue, as was also adipose triglyceride lipase (ATGL). There was a rapid loss of labelled lipid from epididymal adipose tissue of ZAG-treated mice, but not from the other depots, reflecting the difference in sensitivity to lipolytic stimuli. The increased expression of HSL and ATGL may involve the extracellular signal-regulated kinase (ERK) pathway, as the active (phospho) form was upregulated in all adipose depots after ZAG administration, whereas in vitro studies showed induction of HSL and ATGL by ZAG to be attenuated by PD98059, an inhibitor of the ERK pathway. CONCLUSION: These results suggest that ZAG not only induces direct lipolysis, but also sensitizes adipose tissue to other lipolytic stimuli.

Studies on the anti-obesity activity of zinc-α2-glycoprotein in the rat

OBJECTIVE: To investigate the anti-obesity effect of the adipokine zinc-a(2)-glycoprotein (ZAG) in rats and the mechanism of this effect. SUBJECTS: Mature male Wistar rats (540 ± 83 g) were administered human recombinant ZAG (50 µg per 100 g body weight given intravenously daily) for 10 days, while control animals received an equal volume of phosphate-buffered saline (PBS). RESULTS: Animals treated with ZAG showed a progressive decrease in body weight, without a decrease in food and water intake, but with a 0.4 °C rise in body temperature. Body composition analysis showed loss of adipose tissue, but an increase in lean body mass. The loss of fat was due to an increase in lipolysis as shown by a 50% elevation of plasma glycerol, accompanied by increased utilization of non-esterified fatty acids, as evidenced by the 55% decrease in plasma levels. Plasma levels of glucose and triglycerides were also reduced by 36-37% and there was increased expression of the glucose transporter 4 in both skeletal muscle and adipose tissue. Expression of the lipolytic enzymes adipose triglyceride lipase and hormone-sensitive lipase in the white adipose tissue (WAT) were increased twofold after ZAG administration. There was almost a twofold increased expression of uncoupling proteins 1 and 3 in brown adipose tissue and WAT, which would contribute to increased substrate utilization. Administration of ZAG increased ZAG expression twofold in the gastrocnemius muscle, BAT and WAT, which was probably necessary for its biological effect. CONCLUSION: These results show that ZAG produces increased lipid mobilization and utilization in the rat.

Induction of lipolysis in vitro and loss of body fat in vivo by zinc-alpha(2)-glycoprotein

Loss of adipose tissue in cancer cachexia has been associated with tumour production of a lipid-mobilizing factor (LMF) which has been shown to be homologous with the plasma protein zinc-a2-glycoprotein (ZAG). The aim of this study was to compare the ability of human ZAG with LMF to stimulate lipolysis in vitro and induce loss of body fat in vivo, and to determine the mechanisms involved. ZAG was purified from human plasma using a combination of Q Sepharose and Superdex 75 chromatography, and was shown to stimulate glycerol release from isolated murine epididymal adipocytes in a dose-dependent manner. The effect was enhanced by the cyclic AMP phosphodiesterase inhibitor Ro20-1724, and attenuated by freeze/thawing and the specific ß3-adrenoreceptor antagonist SR59230A. In vivo ZAG caused highly significant, time-dependent, decreases in body weight without a reduction in food and water intake. Body composition analysis showed that loss of body weight could be attributed entirely to the loss of body fat. Loss of adipose tissue may have been due to the lipolytic effect of ZAG coupled with an increase in energy expenditure, since there was a dose-dependent increase in expression of uncoupling protein-1 (UCP-1) in brown adipose tissue. These results suggest that ZAG may be effective in the treatment of obesity.

Effect of a tumour-derived lipid-mobilising factor on glucose and lipid metabolism in vivo

Treatment of ex-breeder male NMRI mice with lipid mobilising factor isolated from the urine of cachectic cancer patients, caused a significant increase in glucose oxidation to CO2, compared with control mice receiving phosphate buffered saline. Glucose utilisation by various tissues was determined by the 2-deoxyglucose tracer technique and shown to be elevated in brain, heart, brown adipose tissue and gastrocnemius muscle. The tissue glucose metabolic rate was increased almost three-fold in brain, accounting for the ability of lipid mobilising factor to decrease blood glucose levels. Lipid mobilising factor also increased overall lipid oxidation, as determined by the production of 14CO2 from [14C carboxy] triolein, being 67% greater than phosphate buffered saline controls over a 24 h period. There was a significant increase in [14C] lipid accumulation in plasma, liver and white and brown adipose tissue after administration of lipid mobilising factor. These results suggest that changes in carbohydrate metabolism and loss of adipose tissue, together with an increased whole body fatty acid oxidation in cachectic cancer patients, may arise from tumour production of lipid mobilising factor. © 2002 Cancer Research UK.

Expression of uncoupling proteins-1,-2 and-3 mRNA is induced by an adenocarcinoma-derived lipid-mobilizing factor

The abnormalities of lipid metabolism observed in cancer cachexia may be induced by a lipid-mobilizing factor produced by adenocarcinomas. The specific molecules and metabolic pathways that mediate the actions of lipid-mobilizing factor are not known. The mitochondrial uncoupling proteins-1, -2 and -3 are suggested to play essential roles in energy dissipation and disposal of excess lipid. Here, we studied the effects of lipid-mobilizing factor on the expression of uncoupling proteins-1, -2 and -3 in normal mice. Lipid-mobilizing factor isolated from the urine of cancer patients was injected intravenously into mice over a 52-h period, while vehicle was similarly given to controls. Lipid-mobilizing factor caused significant reductions in body weight (-10%, P=0.03) and fat mass (-20%, P<0.01) accompanied by a marked decrease in plasma leptin (-59%, P<0.01) and heavy lipid deposition in the liver. In brown adipose tissue, uncoupling protein-1 mRNA levels were elevated in lipid-mobilizing factor-treated mice (+96%, P<0.01), as were uncoupling proteins-2 and -3 (+57% and +37%, both P<0.05). Lipid-mobilizing factor increased uncoupling protein-2 mRNA in both skeletal muscle (+146%, P<0.05) and liver (+142%, P=0.03). The protein levels of uncoupling protein-1 in brown adipose tissue and uncoupling protein-2 in liver were also increased with lipid-mobilizing factor administration (+49% and +67%, both P=0.02). Upregulation by lipid-mobilizing factor of uncoupling proteins-1, -2 and -3 in brown adipose tissue, and of uncoupling protein-2 in skeletal muscle and liver, suggests that these uncoupling proteins may serve to utilize excess lipid mobilized during fat catabolism in cancer cachexia.

Effect of maternal protein restriction during pregnancy and postweaning high-fat feeding on diet-induced thermogenesis in adult mouse offspring

Purpose: Prenatal undernutrition followed by postweaning feeding of a high-fat diet results in obesity in the adult offspring. In this study, we investigated whether diet-induced thermogenesis is altered as a result of such nutritional mismatch. Methods: Female MF-1 mice were fed a normal protein (NP, 18 % casein) or a protein-restricted (PR, 9 % casein) diet throughout pregnancy and lactation. After weaning, male offspring of both groups were fed either a high-fat diet (HF; 45 % kcal fat) or standard chow (C, 7 % kcal fat) to generate the NP/C, NP/HF, PR/C and PR/HF adult offspring groups (n = 7-11 per group). Results: PR/C and NP/C offspring have similar body weights at 30 weeks of age. Postweaning HF feeding resulted in significantly heavier NP/HF offspring (P <0.01), but not in PR/HF offspring, compared with their chow-fed counterparts. However, the PR/HF offspring exhibited greater adiposity (P <0.01) v the NP/HF group. The NP/HF offspring had increased energy expenditure and increased mRNA expression of uncoupling protein-1 and β-3 adrenergic receptor in the interscapular brown adipose tissue (iBAT) compared with the NP/C mice (both at P <0.01). No such differences in energy expenditure and iBAT gene expression were observed between the PR/HF and PR/C offspring. Conclusions: These data suggest that a mismatch between maternal diet during pregnancy and lactation, and the postweaning diet of the offspring, can attenuate diet-induced thermogenesis in the iBAT, resulting in the development of obesity in adulthood. © 2014 Springer-Verlag Berlin Heidelberg.

Caractérisation d'IGFBP-2 comme biomarqueur intégrateur de la santé cardiométabolique

La protéine de liaison aux facteurs de croissance analogues à l’insuline (IGFBP)-2 est une protéine circulante fortement associée à la résistance à l’insuline qui module les effets métaboliques d’IGF-I et IGF-II en s’y associant directement, et qui exerce aussi des actions IGF-indépendantes via sa liaison à la matrice extracellulaire et aux intégrines. Chez l’homme, de faibles niveaux d’IGFBP-2 sont associés à un profil lipidique délétère, ainsi qu’à une augmentation de la masse grasse et de la résistance à l’insuline. Les travaux décrits dans cette thèse montrent chez l’humain et la souris que les niveaux d’IGFBP-2 sont associés de manière indépendante aux composantes du risque cardiométabolique. Chez l’homme, de faibles niveaux d’IGFBP-2 sont associés à la dyslipidémie athérogène. Une valeur seuil d’IGFBP-2 de 221.5 ng/mL a permis de discriminer entre les sujets métaboliquement sains et ceux répondant aux critères du syndrome métabolique. En plus de son association avec la résistance à l’insuline et les composantes du profil lipidique, de faibles niveaux d’IGFBP-2 sont associés à une fonction cardiaque diminuée chez les patients atteints de sténose aortique, tel qu’évaluée par le volume d’éjection indexé, un indice de fonction global du ventricule gauche qui intègre la fonction pompe et le remodelage du tissu. Chez l’homme, des niveaux d’IGFBP-2 élevés sont associés à un tissu adipeux brun plus volumineux ainsi qu’à une activité métabolique plus importante de ce dernier. Ces observations, telles qu’évaluées par PET/CT, sont aussi validées chez les souris surexprimant la forme humaine d’IGFBP-2. Nos travaux démontrent que les niveaux d’IGFBP-2 sont fortement associés au métabolisme des lipoprotéines et des lipides, à la fonction cardiaque ainsi qu’à l’activité du tissu adipeux brun. L’influence des niveaux d’IGFBP-2 par différentes altérations métaboliques menant à l’augmentation du risque cardiométabolique pourrait faire de ce dernier un biomarqueur précoce et intégrateur. Les travaux exposés dans la présente thèse soulignent aussi un rôle mécanistique potentiel pour IGFBP-2 dans la protection contre certaines altérations du métabolisme.

Efeito do GQ-02, derivado de tiazolidinadionas, sobre a aquisição de fenótipo termogênico pelo adipócito in vivo e em cultura de células

Dissertação (mestrado)—Universidade de Brasília, Faculdade em Ciências da Saúde, Programa de Pós-Graduação em Ciências da Saúde, 2016.

Chondrogenic potential of injectable κ-carrageenan hydrogel with encapsulated adipose stem cells for cartilage tissue-engineering applications

Due to the limited self-repair capacity of cartilage, regenerative medicine therapies for the treatment of cartilage defects must use a significant amount of cells, preferably applied using a hydrogel system that can promise their delivery and functionality at the specific site. This paper discusses the potential use of k-carrageenan hydrogels for the delivery of stem cells obt ained from adipose tissue in the treatment of cartilage tissue defects. The developed hydrogels were produced by an ionotropic gelation met hod and human adipose stem cells (hASCs) were encapsulated in 1.5% w/v k-carrageenan solution at a cell density of 5  10 6 cells/ml. The results from the analysis of the cell-encapsulating hydrogels, cultured for up to 21 days, indicated that k-carrageenan hydrogels support the viability, proliferation and chondrogenic differentiation of hASCs. Additionally, the mec hanical analysis demonstrated an increase in stiffness and viscoelastic properties of k-carrageenan gels with their encapsulated cells with increasing time in culture with chondrogenic medium. These results allowed the conclusion that k-carrageenan exhibits properties t hat enable the in vitro functionality of encapsulated hASCs and thus may provide the basis for new successful approaches for the treatment of cartilage defects.

New genetic loci link adipose and insulin biology to body fat distribution.

Body fat distribution is a heritable trait and a well-established predictor of adverse metabolic outcomes, independent of overall adiposity. To increase our understanding of the genetic basis of body fat distribution and its molecular links to cardiometabolic traits, here we conduct genome-wide association meta-analyses of traits related to waist and hip circumferences in up to 224,459 individuals. We identify 49 loci (33 new) associated with waist-to-hip ratio adjusted for body mass index (BMI), and an additional 19 loci newly associated with related waist and hip circumference measures (P < 5 × 10(-8)). In total, 20 of the 49 waist-to-hip ratio adjusted for BMI loci show significant sexual dimorphism, 19 of which display a stronger effect in women. The identified loci were enriched for genes expressed in adipose tissue and for putative regulatory elements in adipocytes. Pathway analyses implicated adipogenesis, angiogenesis, transcriptional regulation and insulin resistance as processes affecting fat distribution, providing insight into potential pathophysiological mechanisms.

Aliskiren penetrates adipose and skeletal muscle tissue, and reduces Renin-Angiotensin System activity in obese hypertensive patients

Introduction: Tissue Renin-Angiotensin System activity is increased in obesity and may contribute to obesity-related hypertension and metabolic abnormalities. This open-label pilot study investigated the local effects of Aliskiren in adipose tissue and skeletal muscle.Methods: After a 1-2 week washout, 10 patients with hypertension and abdominal obesity received placebo for 2 weeks, then Aliskiren 300 mg once daily for 4 weeks, followed by a 4-week washout period and then another 4 weeks treatment period with Amlodipine 5 mg once daily. Drug concentrations and Renin-Angiotensin Systembiomarkers were measured in interstitial fluid employing the microdialysis zero-flow method, and in biopsies from abdominal subcutaneous adipose and skeletal muscle.Results: After 4 weeks treatment, microdialysate concentrations (mean±SD) of Aliskiren were 2.4±2.1 ng/ml in adipose tissue, and 7.1±4.2 ng/ml in skeletal muscle. These concentrations were similar to the mean plasma concentration of 8.4±4.4 ng/ml. Tissue concentrations (ng/g) of Aliskiren were 29.0±16.7 ng/g in adipose tissue, and 107.3±68.6 ng/g in skeletal muscle after 4 weeks treatment. Angiotensin II concentrations in microdialysates were below the lower limit of quantification in most patients, but pooled data from two patients suggested that Angiotensin II was reduced by Aliskiren and unchanged by Amlodipine. Aliskiren 300 mg significantly reduced mean plasma Renin activity by 68% and Angiotensin II by 61% (p<0.05 vs. baseline). Amlodipine 5 mg increased plasma Renin activity by 48% (p<0.05 vs. baseline), and non-significantly increased Angiotensin II by 60%. Both treatments increased plasma Renin concentration.Conclusion: Aliskiren 300 mg once daily penetrates adipose and skeletal muscle tissue at concentrations sufficient to reduce tissue Renin-Angiotensin System activity in obese patients with hypertension.

Ultrastructural and morphometrical analyses of the brown adipocyte nucleus in a hibernating dormouse.

The fine morphology, size, and perichromatin granule frequency were analysed in brown adipocyte nuclei from hibernating, arousing, and euthermic dormice, Muscardinus avellanarius. Unusual nuclear structural constituents such as nuclear amorphous bodies, coiled body-like constituents and bundles of nucleoplasmic filaments were described as typical of hibernating nuclei. Morphometrical findings showed significant difference in total nuclear and nucleolar size in the three physiological conditions investigated as well as decreasing frequency of perichromatin granules in nuclei of hibernating to arousing to euthermic animals. A possible involvement of these granules in the intranuclear transport or storage of pre-mRNA is discussed in the context of other experimental evidence.

Peroxisome proliferator-activated receptor gamma is required in mature white and brown adipocytes for their survival in the mouse.

The peroxisome proliferator-activated receptor gamma (PPARgamma) mediates the activity of the insulin-sensitizing thiazolidinediones and plays an important role in adipocyte differentiation and fat accretion. The analysis of PPARgamma functions in mature adipocytes is precluded by lethality of PPARgamma(-/-) fetuses and tetraploid-rescued pups. Therefore we have selectively ablated PPARgamma in adipocytes of adult mice by using the tamoxifen-dependent Cre-ER(T2) recombination system. We show that mature PPARgamma-null white and brown adipocytes die within a few days and are replaced by newly formed PPARgamma-positive adipocytes, demonstrating that PPARgamma is essential for the in vivo survival of mature adipocytes, in addition to its well established requirement for their differentiation. Our data suggest that potent PPARgamma antagonists could be used to acutely reduce obesity.

Adipose-derived stem cells and tissue revascularization: enhancing islet survival and performance for diabetes care.

Pancreatic islet transplantation represents a fascinating procedure that, at the moment, can be considered as alternative to standard insulin treatment or pancreas transplantation only for selected categories of patients with type 1 diabetes mellitus. Among the factors responsible for leading to poor islet engraftment, hypoxia plays an important role. Mesenchymal stem cells (MSCs) were recently used in animal models of islet transplantation not only to reduce allograft rejection, but also to promote revascularization. Currently adipose tissue represents a novel and good source of MSCs. Moreover, the capability of adipose-derived stem cells (ASCs) to improve islet graft revascularization was recently reported after hybrid transplantation in mice. Within this context, we have previously shown that hyaluronan esters of butyric and retinoic acids can significantly enhance the rescuing potential of human MSCs. Here we evaluated whether ex vivo preconditioning of human ASCs (hASCs) with a mixture of hyaluronic (HA), butyric (BU), and retinoic (RA) acids may result in optimization of graft revascularization after islet/stem cell intrahepatic cotransplantation in syngeneic diabetic rats. We demonstrated that hASCs exposed to the mixture of molecules are able to increase the secretion of vascular endothelial growth factor (VEGF), as well as the transcription of angiogenic genes, including VEGF, KDR (kinase insert domain receptor), and hepatocyte growth factor (HGF). Rats transplanted with islets cocultured with preconditioned hASCs exhibited a better glycemic control than rats transplanted with an equal volume of islets and control hASCs. Cotransplantation with preconditioned hASCs was also associated with enhanced islet revascularization in vivo, as highlighted by graft morphological analysis. The observed increase in islet graft revascularization and function suggests that our method of stem cell preconditioning may represent a novel strategy to remarkably improve the efficacy of islets-hMSCs cotransplantation.