Anatase Titanium Dioxide Imparts Photoluminescent Properties to PA2200 Commercial 3D Printing Material to Generate Complex Optical Imaging Phantoms.

Selective laser sintering (SLS) is a prominent 3D printing modality that typically uses a polyamide (PA) powder as the substrate. One commercially available SLS material is known as PA2200, which is comprised of nylon 12 and titanium dioxide (TiO2) and is widely used to generate 3D-printed parts. Here, we report a unique optical photoluminescence (PL) characteristic of native, white PA2200, in which it yields a persistent, phosphorescence-type emission. An analysis of luminescence imaging data with emission measurements demonstrated that the anatase phase of the titanium dioxide additive is the source of the persistent PL properties. This characteristic of PA2200 enables advanced optical imaging applications, as demonstrated by luminescence imaging of an anatomical rat skeleton and a novel Derenzo-type phantom on a commercial image station. In summary, the light emission properties of PA2200 induced by the presence of anatase titanium dioxide open the door to a vast new array of complex optical applications, including the generation of imaging phantoms for training, calibration, and quality control.

Altered skeletal muscle glucose-fatty acid flux in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis is characterized by the degeneration of upper and lower motor neurons, yet an increasing number of studies in both mouse models and patients with amyotrophic lateral sclerosis suggest that altered metabolic homeostasis is also a feature of disease. Pre-clinical and clinical studies have shown that modulation of energy balance can be beneficial in amyotrophic lateral sclerosis. However, the capacity to target specific metabolic pathways or mechanisms requires detailed understanding of metabolic dysregulation in amyotrophic lateral sclerosis. Here, using the superoxide dismutase 1, glycine to alanine substitution at amino acid 93 (SOD1G93A) mouse model of amyotrophic lateral sclerosis, we demonstrate that an increase in whole-body metabolism occurs at a time when glycolytic muscle exhibits an increased dependence on fatty acid oxidation. Using myotubes derived from muscle of amyotrophic lateral sclerosis patients, we also show that increased dependence on fatty acid oxidation is associated with increased whole-body energy expenditure. In the present study, increased fatty acid oxidation was associated with slower disease progression. However, within the patient cohort, there was considerable heterogeneity in whole-body metabolism and fuel oxidation profiles. Thus, future studies that decipher specific metabolic changes at an individual patient level are essential for the development of treatments that aim to target metabolic pathways in amyotrophic lateral sclerosis.

Exosomes secreted by prostate cancer cells under hypoxia promote matrix metalloproteinases activity at pre-metastatic niches.

Approximately, 30 000 men die from prostate cancer (PCa) every year in the United States, mainly due to the metastasis. Thus, the key events associated with PCa metastasis are under rigorous investigation, with recent studies showing that preparation of pre-metastatic niches (PMN) in distant organs is an important step. However, the molecular basis for PMN preparation is still unclear. Hypoxia in primary tumors promotes aggressiveness; however, its precise role in metastasis is not clear. We recently reported that exosomes secreted by PCa cells under hypoxia promote stemness and invasiveness in naïve PCa cells; however, whether these extracellular vesicles also influence PMN remains unknown. In the present study, we isolated exosomes from human PCa PC3 cells under normoxic (21% O2 , exosomes secreted under normoxic condition [ExoNormoxic ]) and hypoxic (1% O2 , exosomes secreted under hypoxic condition [ExoHypoxic ]) conditions, and characterized their effect (10 µg exosomes, intraperitoneal (IP) treatment every 48 hours for 4 weeks) on key biomarkers associated with PMN in nude mice. Whole animal fluorescence imaging showed that ExoHypoxic treatment promotes matrix metalloproteinases (MMPs) activity in several putative metastatic sites. Histological studies confirmed that ExoHypoxic treatment enhanced the level of MMP2, MMP9, and extracellular matrix proteins (fibronectin and collagen) as well as increased the number of CD11b+ cells at selective PMN sites. Furthermore, proteomic profiling of exosomes by liquid chromatography/mass spectrometry identified cargo proteins in ExoNormoxic and ExoHypoxic as well as distinct canonical pathways targeted by them. These results suggest that exosomes secreted by PCa cells under hypoxia plausibly remodel distant PMN, and thus, could be a potential target to control metastatic PCa.

Non-invasive in vivo fluorescence imaging of apoptotic retinal photoreceptors.

Phosphatidylserine externalization is an early molecular signature for apoptosis. In many retinal degenerative diseases, photoreceptor neurons die by apoptosis. Here, we report utility of the phosphatidylserine-binding conjugate of Bis(zinc(II)-dipicolylamine (Zn-DPA) with Texas-red (PSVue-550) in transiently labeling apoptotic photoreceptors in living pigmented or albino rats and mice with retinal degeneration. Applying PSVue-550 as eyedrop is non-toxic and eliminates need for intraocular injection. PSVue-550 fluorescence specifically and transiently labeling dying retinal photoreceptors is detectable in anesthetized animals using standard retinal or whole small animal imaging systems. Importantly, prior PSVue-550 eyedrop administration and imaging does not affect repeat testing. Altogether, our results establish PSVue-550 imaging as a completely non-invasive method that provides the opportunity to longitudinally monitor retinal photoreceptor cell death in preclinical studies.

3D-printed Wash Station with Integrated Anesthesia Delivery Manifold for High-throughput Depilation of Laboratory Mice.

Depilation (that is, hair removal) is a necessary prerequisite for many small animal surgeries and optical imaging experiments. Over-the-counter depilatory creams are widely used, owing to their efficacy, safety, and low rates of skin irritation and infection. However, the use of these creams is generally messy and time-consuming and generates considerable waste. Furthermore, the process itself varies markedly among laboratories. Here we present a 3D-printed device that simplifies the depilation procedure by integrating 3 key elements: 1) a multiple-port, self-scavenging anesthesia manifold, 2) curved animal holders with flow-through slats, and 3) a removable waste collection tray. Reflecting insights gained from an international survey about depilatory lab procedures that highlighted the lack of standardized protocols, this apparatus is designed to improve the neatness, throughput, and safety of mouse depilation, resulting in efficient and repeatable processes that bolster the welfare of both researchers and subjects.

RIPK1-mediated induction of mitophagy compromises the viability of extracellular-matrix-detached cells.

For cancer cells to survive during extracellular matrix (ECM) detachment, they must inhibit anoikis and rectify metabolic deficiencies that cause non-apoptotic cell death. Previous studies in ECM-detached cells have linked non-apoptotic cell death to reactive oxygen species (ROS) generation, although the mechanistic underpinnings of this link remain poorly defined. Here, we uncover a role for receptor-interacting protein kinase 1 (RIPK1) in the modulation of ROS and cell viability during ECM detachment. We find that RIPK1 activation during ECM detachment results in mitophagy induction through a mechanism dependent on the mitochondrial phosphatase PGAM5. As a consequence of mitophagy, ECM-detached cells experience diminished NADPH production in the mitochondria, and the subsequent elevation in ROS levels leads to non-apoptotic death. Furthermore, we find that antagonizing RIPK1/PGAM5 enhances tumour formation in vivo. Thus, RIPK1-mediated induction of mitophagy may be an efficacious target for therapeutics aimed at eliminating ECM-detached cancer cells.

A novel method for intratracheal injection of infectious agents into mice.

Intratracheal injection is a traditional technique used in small animal studies of highly contagious airborne pathogens such as Mycobacterium tuberculosis. However, current techniques of intratracheal injection generally involve procedures that pose a risk of incident injury and infection for researchers, and may also cause collateral damage to experimental animals during the installation process. Here we describe an intratracheal injection method that was enabled by a three dimensional printing of a custom platform. This updated technique improved the overall ergonomics of intratracheal injection in mice, minimizing the risk of human injury and implementing the 3R (replacement, reduction and refinement) principle in mouse infection studies.

Patient Education for Endoscopic Sinus Surgery: Preliminary Experience Using 3D-Printed Clinical Imaging Data.

Within the Ear, Nose, and Throat (ENT) medical space, a relatively small fraction of patients follow through with elective surgeries to fix ailments such as a deviated septum or occluded sinus passage. Patient understanding of their diagnosis and treatment plan is integral to compliance, which ultimately yields improved medical outcomes and better quality of life. Here we report the usage of advanced, polyjet 3D printing methods to develop a multimaterial replica of human nasal sinus anatomy, derived from clinical X-ray computed tomography (CT) data, to be used as an educational aid during physician consultation. The final patient education model was developed over several iterations to optimize material properties, anatomical accuracy and overall display. A two-arm, single-center, randomized, prospective study was then performed in which 50 ENT surgical candidates (and an associated control group, n = 50) were given an explanation of their anatomy, disease state, and treatment options using the education model as an aid. Statistically significant improvements in patient ratings of their physician's explanation of their treatment options (p = 0.020), self-rated anatomical understanding (p = 0.043), self-rated understanding of disease state (p = 0.016), and effectiveness of the visualization (p = 0.007) were noted from the population that viewed the 3D education model, indicating it is an effective tool which ENT surgeons may use to educate and interact with patients.

Bismuth Infusion of ABS Enables Additive Manufacturing of Complex Radiological Phantoms and Shielding Equipment.

Radiopacity is a critical property of materials that are used for a range of radiological applications, including the development of phantom devices that emulate the radiodensity of native tissues and the production of protective equipment for personnel handling radioactive materials. Three-dimensional (3D) printing is a fabrication platform that is well suited to creating complex anatomical replicas or custom labware to accomplish these radiological purposes. We created and tested multiple ABS (Acrylonitrile butadiene styrene) filaments infused with varied concentrations of bismuth (1.2-2.7 g/cm³), a radiopaque metal that is compatible with plastic infusion, to address the poor gamma radiation attenuation of many mainstream 3D printing materials. X-ray computed tomography (CT) experiments of these filaments indicated that a density of 1.2 g/cm³ of bismuth-infused ABS emulates bone radiopacity during X-ray CT imaging on preclinical and clinical scanners. ABS-bismuth filaments along with ABS were 3D printed to create an embedded human nasocranial anatomical phantom that mimicked radiological properties of native bone and soft tissue. Increasing the bismuth content in the filaments to 2.7 g/cm³ created a stable material that could attenuate 50% of 99mTechnetium gamma emission when printed with a 2.0 mm wall thickness. A shielded test tube rack was printed to attenuate source radiation as a protective measure for lab personnel. We demonstrated the utility of novel filaments to serve multiple radiological purposes, including the creation of anthropomorphic phantoms and safety labware, by tuning the level of radiation attenuation through material customization.

Three-dimensional printing of X-ray computed tomography datasets with multiple materials using open-source data processing.

Advances in three-dimensional (3D) printing allow for digital files to be turned into a "printed" physical product. For example, complex anatomical models derived from clinical or pre-clinical X-ray computed tomography (CT) data of patients or research specimens can be constructed using various printable materials. Although 3D printing has the potential to advance learning, many academic programs have been slow to adopt its use in the classroom despite increased availability of the equipment and digital databases already established for educational use. Herein, a protocol is reported for the production of enlarged bone core and accurate representation of human sinus passages in a 3D printed format using entirely consumer-grade printers and a combination of free-software platforms. The comparative resolutions of three surface rendering programs were also determined using the sinuses, a human body, and a human wrist data files to compare the abilities of different software available for surface map generation of biomedical data. Data shows that 3D Slicer provided highest compatibility and surface resolution for anatomical 3D printing. Generated surface maps were then 3D printed via fused deposition modeling (FDM printing). In conclusion, a methodological approach that explains the production of anatomical models using entirely consumer-grade, fused deposition modeling machines, and a combination of free software platforms is presented in this report. The methods outlined will facilitate the incorporation of 3D printed anatomical models in the classroom. Anat Sci Educ 10: 383-391. © 2017 American Association of Anatomists.

Obesity Contributes to Ovarian Cancer Metastatic Success through Increased Lipogenesis, Enhanced Vascularity, and Decreased Infiltration of M1 Macrophages.

Epithelial ovarian cancer (EOC) is the leading cause of death from gynecologic malignancy, with high mortality attributable to widespread intraperitoneal metastases. Recent meta-analyses report an association between obesity, ovarian cancer incidence, and ovarian cancer survival, but the effect of obesity on metastasis has not been evaluated. The objective of this study was to use an integrative approach combining in vitro, ex vivo, and in vivo studies to test the hypothesis that obesity contributes to ovarian cancer metastatic success. Initial in vitro studies using three-dimensional mesomimetic cultures showed enhanced cell-cell adhesion to the lipid-loaded mesothelium. Furthermore, in an ex vivo colonization assay, ovarian cancer cells exhibited increased adhesion to mesothelial explants excised from mice modeling diet-induced obesity (DIO), in which they were fed a "Western" diet. Examination of mesothelial ultrastructure revealed a substantial increase in the density of microvilli in DIO mice. Moreover, enhanced intraperitoneal tumor burden was observed in overweight or obese animals in three distinct in vivo models. Further histologic analyses suggested that alterations in lipid regulatory factors, enhanced vascularity, and decreased M1/M2 macrophage ratios may account for the enhanced tumorigenicity. Together, these findings show that obesity potently affects ovarian cancer metastatic success, which likely contributes to the negative correlation between obesity and ovarian cancer survival.

Fluorescence Imaging of Interscapular Brown Adipose Tissue in Living Mice.

Brown adipose tissue (BAT) plays a key role in energy expenditure and heat generation and is a promising target for diagnosing and treating obesity, diabetes and related metabolism disorders. While several nuclear and magnetic resonance imaging methods are established for detecting human BAT, there are no convenient protocols for high throughput imaging of BAT in small animal models. Here we disclose a simple but effective method for non-invasive optical imaging of interscapular BAT in mice using a micellar formulation of the commercially available deep-red fluorescent probe, SRFluor680. Whole-body fluorescence imaging of living mice shows extensive accumulation of the fluorescent probe in the interscapular BAT and ex vivo analysis shows 3.5-fold selectivity for interscapular BAT over interscapular WAT. Additional imaging studies indicate that SRFluor680 uptake is independent of mouse species and BAT metabolic state. The results are consistent with an unusual pharmacokinetic process that involves irreversible translocation of the lipophilic SRFluor680 from the micelle nanocarrier into the adipocytes within the BAT. Multimodal PET/CT and planar fluorescence/X-ray imaging of the same living animal shows co-localization of BAT mass signal reported by the fluorescent probe and BAT metabolism signal reported by the PET agent, 18F-FDG. The results indicate a path towards a new, dual probe molecular imaging paradigm that allows separate and independent non-invasive visualization of BAT mass and BAT metabolism in a living subject.

Selective X-ray contrast enhancement of the spleen of living mice mediated by gold nanorods.

Gold nanomaterials (AuNPs) represent a promising new class of contrast agents for X-ray computed tomographic (CT) imaging in both research and clinical settings. These materials exhibit superior X-ray absorption properties compared with other iodinated agents, and thus require lower injection doses. Gold is nonimmunogenic and therefore contributes to safety profile in living specimens. Unfortunately, most reports on the use of AuNPs as X-ray CT enhancers only demonstrate marginal enhancement of the intended anatomical structure. In this study, we demonstrate the dramatic properties of gold nanorods (GNR) to serve as robust X-ray CT contrast-enhancing agent for selective imaging of the spleen. These organ-specific uptake properties were delineated by performing longitudinal CT imaging of living mice that were dosed with GNR at 2 day intervals. Rapid uptake in spleen was noted within 12 h of first systemic administration with a change in contrast enhancement of 90 Hounsfield units (ΔHU = 90) and with two subsequent injections a total contrast enhancement of over 200 HU was observed. The resulting images provide excellent contrast that will enable the detailed anatomical visualization and study of a range of pre-clinical models of spleen disease including infection and cancer.

Hepatic DNA deposition drives drug-induced liver injury and inflammation in mice.

UNLABELLED: Drug-induced liver injury (DILI) is an important cause of acute liver failure, with limited therapeutic options. During DILI, oncotic necrosis with concomitant release and recognition of intracellular content amplifies liver inflammation and injury. Among these molecules, self-DNA has been widely shown to trigger inflammatory and autoimmune diseases; however, whether DNA released from damaged hepatocytes accumulates into necrotic liver and the impact of its recognition by the immune system remains elusive. Here we show that treatment with two different hepatotoxic compounds (acetaminophen and thioacetamide) caused DNA release into the hepatocyte cytoplasm, which occurred in parallel with cell death in vitro. Administration of these compounds in vivo caused massive DNA deposition within liver necrotic areas, together with an intravascular DNA coating. Using confocal intravital microscopy, we revealed that liver injury due to acetaminophen overdose led to a directional migration of neutrophils to DNA-rich areas, where they exhibit an active patrolling behavior. DNA removal by intravenous DNASE1 injection or ablation of Toll-like receptor 9 (TLR9)-mediated sensing significantly reduced systemic inflammation, liver neutrophil recruitment, and hepatotoxicity. Analysis of liver leukocytes by flow cytometry revealed that emigrated neutrophils up-regulated TLR9 expression during acetaminophen-mediated necrosis, and these cells sensed and reacted to extracellular DNA by activating the TLR9/NF-κB pathway. Likewise, adoptive transfer of wild-type neutrophils to TLR9(-/-) mice reversed the hepatoprotective phenotype otherwise observed in TLR9 absence. CONCLUSION: Hepatic DNA accumulation is a novel feature of DILI pathogenesis. Blockage of DNA recognition by the innate immune system may constitute a promising therapeutic venue.

Correlation of X-ray computed tomography with quantitative nuclear magnetic resonance methods for pre-clinical measurement of adipose and lean tissues in living mice.

Numerous obesity studies have coupled murine models with non-invasive methods to quantify body composition in longitudinal experiments, including X-ray computed tomography (CT) or quantitative nuclear magnetic resonance (QMR). Both microCT and QMR have been separately validated with invasive techniques of adipose tissue quantification, like post-mortem fat extraction and measurement. Here we report a head-to-head study of both protocols using oil phantoms and mouse populations to determine the parameters that best align CT data with that from QMR. First, an in vitro analysis of oil/water mixtures was used to calibrate and assess the overall accuracy of microCT vs. QMR data. Next, experiments were conducted with two cohorts of living mice (either homogenous or heterogeneous by sex, age and genetic backgrounds) to assess the microCT imaging technique for adipose tissue segmentation and quantification relative to QMR. Adipose mass values were obtained from microCT data with three different resolutions, after which the data were analyzed with different filter and segmentation settings. Strong linearity was noted between the adipose mass values obtained with microCT and QMR, with optimal parameters and scan conditions reported herein. Lean tissue (muscle, internal organs) was also segmented and quantified using the microCT method relative to the analogous QMR values. Overall, the rigorous calibration and validation of the microCT method for murine body composition, relative to QMR, ensures its validity for segmentation, quantification and visualization of both adipose and lean tissues.

CAF-secreted IGFBPs regulate breast cancer cell anoikis.

UNLABELLED: Carcinoma-associated fibroblasts (CAFs) are now widely appreciated for their contributions to tumor progression. However, the ability of CAFs to regulate anoikis, detachment-induced cell death, has yet to be investigated. Here, a new role for CAFs in blocking anoikis in multiple cell lines, facilitating luminal filling in three-dimensional cell culture, and promoting anchorage-independent growth is defined. In addition, a novel mechanism underlying anoikis inhibition is discovered. Importantly, it was demonstrated that CAFs secrete elevated quantities of insulin-like growth factor-binding proteins (IGFBPs) that are both necessary for CAF-mediated anoikis inhibition and sufficient to block anoikis in the absence of CAFs. Furthermore, these data reveal a unique antiapoptotic mechanism for IGFBPs: the stabilization of the antiapoptotic protein Mcl-1. In aggregate, these data delineate a novel role for CAFs in promoting cell survival during detachment and unveil an additional mechanism by which the tumor microenvironment contributes to cancer progression. These results also identify IGFBPs as potential targets for the development of novel chemotherapeutics designed to eliminate detached cancer cells. IMPLICATIONS: The ability of CAF-secreted IGFBPs to block anoikis in breast cancer represents a novel target for the development of therapeutics aimed at specifically eliminating extracellular matrix-detached breast cancer cells.

Gold nanoparticle based X-ray contrast agent for tumor imaging in mice and dog: a potential nano-platform for computer tomography theranostics.

The goal of our study was to demonstrate the utility of nanocrystalline gold as an X-ray contrast agent for imaging tumor in living subjects. Even though significant progress has been achieved in this area by researchers, clinical translation remains challenging. Here, we investigated biocompatible gum Arabic stabilized gold nanocrystals (GA-AuNPs) as X-ray contrast agent in tumor bearing mice and dog. Single intratumoral injections of GA-AuNP resulted in X-ray contrast change of -26 HU in the tumor region after 1 hour post-injection period. Subsequently, five intratumoral injections were performed in the mice. The change in CT number in tumor region is not progressive; rather it reaches a saturation point after fourth injection. These data suggested that accumulation of GA-AuNP reaches a threshold limit within a short time period (5 h), and is retained in the tumor tissue for the rest of the period of investigation. A pilot study was conducted in a client-owned dog presented with collision tumor of thyroid carcinoma and osteosarcoma. In this study, GA-AuNP was injected intratumorally in dog and a contrast enhancement of 12 deltaHU was observed. The CT images of both mice and dog clearly demonstrated that GA-AuNP was effectively distributed and retained throughout the tumor site. The CT data obtained by the present study would provide the crucial dosimetry information for strategic therapy planning using this construct. Both mice and dog did not show any clinical changes, thereby confirming that GA-AuNP did not induce toxicity and can be explored for future clinical applications.

Preclinical anatomical, molecular, and functional imaging of the lung with multiple modalities.

In vivo imaging is an important tool for preclinical studies of lung function and disease. The widespread availability of multimodal animal imaging systems and the rapid rate of diagnostic contrast agent development have empowered researchers to noninvasively study lung function and pulmonary disorders. Investigators can identify, track, and quantify biological processes over time. In this review, we highlight the fundamental principles of bioluminescence, fluorescence, planar X-ray, X-ray computed tomography, magnetic resonance imaging, and nuclear imaging modalities (such as positron emission tomography and single photon emission computed tomography) that have been successfully employed for the study of lung function and pulmonary disorders in a preclinical setting. The major principles, benefits, and applications of each imaging modality and technology are reviewed. Limitations and the future prospective of multimodal imaging in pulmonary physiology are also discussed. In vivo imaging bridges molecular biological studies, drug design and discovery, and the imaging field with modern medical practice, and, as such, will continue to be a mainstay in biomedical research.

Non-invasive imaging and analysis of cerebral ischemia in living rats using positron emission tomography with 18F-FDG.

Stroke is the third leading cause of death among Americans 65 years of age or older(1). The quality of life for patients who suffer from a stroke fails to return to normal in a large majority of patients(2), which is mainly due to current lack of clinical treatment for acute stroke. This necessitates understanding the physiological effects of cerebral ischemia on brain tissue over time and is a major area of active research. Towards this end, experimental progress has been made using rats as a preclinical model for stroke, particularly, using non-invasive methods such as (18)F-fluorodeoxyglucose (FDG) coupled with Positron Emission Tomography (PET) imaging(3,10,17). Here we present a strategy for inducing cerebral ischemia in rats by middle cerebral artery occlusion (MCAO) that mimics focal cerebral ischemia in humans, and imaging its effects over 24 hr using FDG-PET coupled with X-ray computed tomography (CT) with an Albira PET-CT instrument. A VOI template atlas was subsequently fused to the cerebral rat data to enable a unbiased analysis of the brain and its sub-regions(4). In addition, a method for 3D visualization of the FDG-PET-CT time course is presented. In summary, we present a detailed protocol for initiating, quantifying, and visualizing an induced ischemic stroke event in a living Sprague-Dawley rat in three dimensions using FDG-PET.

Ablation of the ID2 gene results in altered circadian feeding behavior, and sex-specific enhancement of insulin sensitivity and elevated glucose uptake in skeletal muscle and brown adipose tissue.

Inhibitor of DNA binding 2 (ID2) is a helix-loop-helix transcriptional repressor rhythmically expressed in many adult tissues. Our earlier studies have demonstrated a role for ID2 in the input pathway, core clock function and output pathways of the mouse circadian system. We have also reported that Id2 null (Id2-/-) mice are lean with low gonadal white adipose tissue deposits and lower lipid content in the liver. These results coincided with altered or disrupted circadian expression profiles of liver genes including those involved in lipid metabolism. In the present phenotypic study we intended to decipher, on a sex-specific basis, the role of ID2 in glucose metabolism and in the circadian regulation of activity, important components of energy balance. We find that Id2-/- mice exhibited altered daily and circadian rhythms of feeding and locomotor activity; activity profiles extended further into the late night/dark phase of the 24-hr cycle, despite mice showing reduced total locomotor activity. Also, male Id2-/- mice consumed a greater amount of food relative to body mass, and displayed less weight gain. Id2-/- females had smaller adipocytes, suggesting sexual-dimorphic programing of adipogenesis. We observed increased glucose tolerance and insulin sensitivity in male Id2-/- mice, which was exacerbated in older animals. FDG-PET analysis revealed increased glucose uptake by skeletal muscle and brown adipose tissue of male Id2-/- mice, suggesting increased glucose metabolism and thermogenesis in these tissues. Reductions in intramuscular triacylglycerol and diacylglycerol were detected in male Id2-/- mice, highlighting its possible mechanistic role in enhanced insulin sensitivity in these mice. Our findings indicate a role for ID2 as a regulator of glucose and lipid metabolism, and in the circadian control of feeding/locomotor behavior; and contribute to the understanding of the development of obesity and diabetes, particularly in shift work personnel among whom incidence of such metabolic disorders is elevated.