Nicola J. Lee

Tumor-induced anorexia and weight loss are mediated by the TGF-beta superfamily cytokine MIC-1.

Anorexia and weight loss are part of the wasting syndrome of late-stage cancer, are a major cause of morbidity and mortality in cancer, and are thought to be cytokine mediated. Macrophage inhibitory cytokine-1 (MIC-1) is produced by many cancers. Examination of sera from individuals with advanced prostate cancer showed a direct relationship between MIC-1 abundance and cancer-associated weight loss. In mice with xenografted prostate tumors, elevated MIC-1 levels were also associated with marked weight, fat and lean tissue loss that was mediated by decreased food intake and was reversed by administration of antibody to MIC-1. Additionally, normal mice given systemic MIC-1 and transgenic mice overexpressing MIC-1 showed hypophagia and reduced body weight. MIC-1 mediates its effects by central mechanisms that implicate the hypothalamic transforming growth factor-β receptor II, extracellular signal–regulated kinases 1 and 2, signal transducer and activator of transcription-3, neuropeptide Y and pro-opiomelanocortin. Thus, MIC-1 is a newly defined central regulator of appetite and a potential target for the treatment of both cancer anorexia and weight loss, as well as of obesity.

Peptide YY ablation in mice leads to the development of hyperinsulinaemia and obesity

Aims/hypothesisObese people exhibit reduced circulating peptide YY (PYY) levels, but it is unclear whether this is a consequence or cause of obesity. We therefore investigated the effect of Pyy ablation on energy homeostasis.MethodsBody composition, i.p. glucose tolerance, food intake and hypothalamic neuropeptide expression were determined in Pyy knock-out and wild-type mice on a normal or high-fat diet.ResultsPyy knock-out significantly increased bodyweight and increased fat mass by 50% in aged females on a normal diet. Male chow-fed Pyy−/− mice were resistant to obesity but became significantly fatter and glucose-intolerant compared with wild-types when fed a high-fat diet. Pyy knock-out animals exhibited significantly elevated fasting or glucose-stimulated serum insulin concentrations vs wild-types, with no increase in basal or fasting-induced food intake. Pyy knock-out decreased or had no effect on neuropeptide Y expression in the arcuate nucleus of the hypothalamus, and significantly increased proopiomelanocortin expression in this region. Male but not female knock-outs exhibited significantly increased growth hormone-releasing hormone expression in the ventromedial hypothalamus and significantly elevated serum IGF-I and testosterone levels. This sex difference in activation of the hypothalamo–pituitary somatotrophic axis by Pyy ablation may contribute to the resistance of chow-fed male knock-outs to late-onset obesity.Conclusions/interpretationPYY signalling is important in the regulation of energy balance and glucose homeostasis, possibly via regulation of insulin release. Therefore reduced PYY levels may predispose to the development of obesity, particularly with ageing or under conditions of high-fat feeding.

Novel Role of Y1 Receptors in the Coordinated Regulation of Bone and Energy Homeostasis

The importance of neuropeptide Y (NPY) and Y2 receptors in the regulation of bone and energy homeostasis has recently been demonstrated. However, the contributions of the other Y receptors are less clear. Here we show that Y1 receptors are expressed on osteoblastic cells. Moreover, bone and adipose tissue mass are elevated in Y1-/- mice with a generalized increase in bone formation on cortical and cancellous surfaces. Importantly, the inhibitory effects of NPY on bone marrow stromal cells in vitro are absent in cells derived from Y1-/- mice, indicating a direct action of NPY on bone cells via this Y receptor. Interestingly, in contrast to Y2 receptor or germ line Y1 receptor deletion, conditional deletion of hypothalamic Y1 receptors in adult mice did not alter bone homeostasis, food intake, or adiposity. Furthermore, deletion of both Y1 and Y2 receptors did not produce additive effects in bone or adiposity. Thus Y1 receptor pathways act powerfully to inhibit bone production and adiposity by nonhypothalamic pathways, with potentially direct effects on bone tissue through a single pathway with Y2 receptors.

Greater Bone Formation of Y2 Knockout Mice Is Associated with Increased Osteoprogenitor Numbers and Altered Y1 Receptor Expression

Germ line or hypothalamus-specific deletion of Y2 receptors in mice results in a doubling of trabecular bone volume. However, the specific mechanism by which deletion of Y2 receptors increases bone mass has not yet been identified. Here we show that cultured adherent bone marrow stromal cells from Y2-/- mice also demonstrate increased mineralization in vitro. Isolation of two populations of progenitor cell types, an immature mesenchymal stem cell population and a more highly differentiated population of progenitor cells, revealed a greater number of the progenitor cells within the bone of Y2-/- mice. Analysis of Y receptor transcripts in cultured stromal cells from wild-type mice revealed high levels of Y1 but not Y2, Y4, Y5, or y6 receptor mRNA. Interestingly, germ line Y2 receptor deletion causes Y1 receptor down-regulation in stromal cells and bone tissue possibly due to the lack of feedback inhibition of NPY release and subsequent overstimulation of Y1 receptors. Furthermore, deletion of Y1 receptors resulted in increased bone mineral density in mice. Together, these findings indicate that the greater number of mesenchymal progenitors and the altered Y1 receptor expression within bone cells in the absence of Y2 receptors are a likely mechanism for the greater bone mineralization in vivo and in vitro, opening up potential new treatment avenues for osteoporosis.

Neuropeptide Y Knockout Mice Reveal a Central Role of NPY in the Coordination of Bone Mass to Body Weight

Changes in whole body energy levels are closely linked to alterations in body weight and bone mass. Here, we show that hypothalamic signals contribute to the regulation of bone mass in a manner consistent with the central perception of energy status. Mice lacking neuropeptide Y (NPY), a well-known orexigenic factor whose hypothalamic expression is increased in fasting, have significantly increased bone mass in association with enhanced osteoblast activity and elevated expression of bone osteogenic transcription factors, Runx2 and Osterix. In contrast, wild type and NPY knockout (NPY −/−) mice in which NPY is specifically over expressed in the hypothalamus (AAV-NPY+) show a significant reduction in bone mass despite developing an obese phenotype. The AAV-NPY+ induced loss of bone mass is consistent with models known to mimic the central effects of fasting, which also show increased hypothalamic NPY levels. Thus these data indicate that, in addition to well characterized responses to body mass, skeletal tissue also responds to the perception of nutritional status by the hypothalamus independently of body weight. In addition, the reduction in bone mass by AAV NPY+ administration does not completely correct the high bone mass phenotype of NPY −/− mice, indicating the possibility that peripheral NPY may also be an important regulator of bone mass. Indeed, we demonstrate the expression of NPY specifically in osteoblasts. In conclusion, these data identifies NPY as a critical integrator of bone homeostatic signals; increasing bone mass during times of obesity when hypothalamic NPY expression levels are low and reducing bone formation to conserve energy under ‘starving’ conditions, when hypothalamic NPY expression levels are high.

PYY transgenic mice are protected against diet-induced and genetic obesity

The gut-derived hormone, peptide YY (PYY) reduces food intake and enhances satiety in both humans and animals. Obese individuals also have a deficiency in circulating peptide YY, although whether this is a cause or a consequence of obesity is unclear. Our aims were to determine whether peptide YY (PYY) over-expression may have therapeutic effects for the treatment of obesity by altering energy balance and glucose homeostasis. We generated PYY transgenic mice and measured body weight, food intake, temperature, adiposity, glucose tolerance, circulating hormone and lipid concentrations and hypothalamic neuropeptide levels (neuropeptide Y; proopiomelanocortin, and thyrotropin-releasing hormone) under chow and high-fat feeding and after crossing these mice onto the genetically obese leptin-deficient ob/ob mouse background. PYY transgenic mice were protected against diet-induced obesity in association with increased body temperature (indicative of increased thermogenesis) and sustained expression of thyrotropin-releasing hormone in the paraventricular nucleus of the hypothalamus. Moreover, PYY transgenic mice crossed onto the genetically obese ob/ob background had significantly decreased weight gain and adiposity, reduced serum triglyceride levels and improved glucose tolerance compared to ob/ob controls. There was no effect of PYY transgenic over expression on basal or fasting-induced food intake measured at 11-12 weeks of age. Together, these findings suggest that long-term administration of PYY, PYY-like compounds or agents that stimulate PYY synthesis in vivo can reduce excess adiposity and improve glucose tolerance, possibly via effects on the hypothalamo-pituitary-thyroid axis and thermogenesis.

Critical role for Y1 receptors in mesenchymal progenitor cell differentiation and osteoblast activity.

The neuropeptide Y (NPY) system has been implicated in the regulation of bone homeostasis and osteoblast activity, but the mechanism behind this is unclear. Here we show that Y1 receptor signaling is directly involved in the differentiation of mesenchymal progenitor cells isolated from bone tissue, as well as the activity of mature osteoblasts. Importantly, the mRNA levels of two key osteogenic transcription factors, runx2 and osterix, as well as the adipogenic transcription factor PPAR‐γ, were increased in long bones of Y1−/− mice compared with wild‐type mice. In vitro, bone marrow stromal cells (BMSCs) isolated from Y1−/− mice formed a greater number of mineralized nodules under osteogenic conditions and a greater number of adipocytes under adipogenic conditions than controls. In addition, both the number and size of fibroblast colony‐forming units formed in vitro by purified osteoprogenitor cells were increased in the absence of the Y1 receptors, suggestive of enhanced proliferation and osteogenesis. Furthermore, the ability of two specific populations of mesenchymal progenitor cells isolated from bone tissue, an immature mesenchymal stem cell population and a more committed osteoprogenitor cell population, to differentiate into osteoblasts and adipocytes in vitro was enhanced in the absence of Y1 receptor signaling. Finally, Y1 receptor deletion also enhanced the mineral‐producing ability of mature osteoblasts, as shown by increased in vitro mineralization by BMSCs isolated from osteoblast‐specific Y1−/− mice. Together these data demonstrate that the NPY system, via the Y1 receptor, directly inhibits the differentiation of mesenchymal progenitor cells as well as the activity of mature osteoblasts, constituting a likely mechanism for the high‐bone‐mass phenotype evident in Y1−/− mice.

NPY regulation of bone remodelling

Neuropeptide Y (NPY), a classic neuronal regulator of energy homeostasis, is now also known to be involved in the control of bone homeostasis. Of the five known Y receptors through which the NPY family of ligands signals, the Y1 and Y2 receptors have so far been implicated in the control of osteoblast activity and thus bone formation. Analysis of brain specific NPY overexpressing and Y receptor knockout models has revealed a powerful anabolic pathway likely involving hypothalamic Y2 receptors and osteoblastic Y1 receptors. Furthering our understanding of the mechanisms underlying the involvement of the NPY system in the control of bone could lead to the development of therapies to improve bone mass in patients with diseases such as osteoporosis.

Peripheral neuropeptide Y Y1 receptors regulate lipid oxidation and fat accretion

Objective:Neuropeptide Y and its Y receptors are important players in the regulation of energy homeostasis. However, while their functions in feeding regulation are well recognized, functions in other critical aspects of energy homeostasis are largely unknown. To investigate the function of Y1 receptors in the regulation of energy homeostasis, we examined energy expenditure, physical activity, body composition, oxidative fuel selection and mitochondrial oxidative capacity in germline Y1−/− mice as well as in a conditional Y1-receptor-knockdown model in which Y1 receptors were knocked down in peripheral tissues of adult mice.Results:Germline Y1−/− mice of both genders not only exhibit a decreased respiratory exchange ratio, indicative of increased lipid oxidation, but interestingly also develop late-onset obesity. However, the increased lipid oxidation is a primary effect of Y1 deletion rather than secondary to increased adiposity, as young Y1−/− mice are lean and show the same effect. The mechanism behind this is likely because of increased liver and muscle protein levels of carnitine palmitoyltransferase-1 (CPT-1) and maximal activity of key enzymes involved in β-oxidation; β-hydroxyacyl CoA dehydrogenase (βHAD) and medium-chain acyl-CoA dehydrogenase (MCAD), leading to increased mitochondrial capacity for fatty acid transport and oxidation. These effects are controlled by peripheral Y1-receptor signalling, as adult-onset conditional Y1 knockdown in peripheral tissues also leads to increased lipid oxidation, liver CPT-1 levels and βHAD activity. Importantly, these mice are resistant to diet-induced obesity.Conclusions:This work shows the primary function of peripheral Y1 receptors in the regulation of oxidative fuel selection and adiposity, opening up new avenues for anti-obesity treatments by targeting energy utilization in peripheral tissues rather than suppressing appetite by central effects.

Compensatory changes in [125I]-PYY binding in Y receptor knockout mice suggest the potential existence of further Y receptor(s).

Gene knockout approaches have helped to better understand the functions of the different Y receptors. However, some results obtained from these knockout mice are unexpected and differ from the results of pharmacological intervention experiments. One possible explanation for this is that germ-line gene deletion of a particular Y receptor can influence expression and function of the remaining Y receptors. Here we show that such compensation in mRNA and protein expression does occur in Y receptor single, double and triple knockout models. Radio-ligand binding experiments using [(125)I]-PYY revealed significant up- and down-regulation of remaining Y receptor binding sites in various Y receptor knockout models compared to results from control mice employing Y receptor preferring agonist or antagonists for displacement of the radio-ligand. The most obvious change can be seen in the hippocampus of Y(1) knockout mice, where the level of the remaining Y receptors is strongly down-regulated. In Y(2) knockout mice no such trend can be seen, however, the expression pattern is significantly changed with a strong up-regulation of [(125)I]-PYY specific binding in the dentate gyrus. Interestingly, this pattern was also seen in Y(1)Y(2)Y(4) triple knockout mice. Y(5) receptor mRNA was approximately 20% higher in the hippocampus and dentate gyrus in the triple knockout mice compared to wild-type controls, while Y(6) mRNA expression could not be detected. However, competition binding experiments in Y(1)Y(2)Y(4) triple knockout mice with the Y(5) receptor preferring ligands [Leu(31), Pro(34)] NPY and [A(31), Aib(32)] NPY were able to replace only approximately 50% of [(125)I]-PYY binding in the dentate gyrus suggesting the existence of further yet unidentified Y receptor(s).