A Mathematical Model of Murine Metabolic Regulation by Leptin: Energy Balance and Defense of a Stable Body Weight

We have developed a physiologically based mathematical model, with parameters derived from published experimental data, to simulate the regulatory effects of the leptin pathway on murine energy homeostasis. Model outcomes are consistent with data reported in the literature and reproduce key characteristics of the energy regulatory system, including compensatory responses that counteract changes in body weight and the failure of this ability when the leptin pathway is disrupted. Our model revealed the possibility of multiple steady states for body weight. It also provided a unified theoretical framework for two historically antagonistic hypotheses regarding body weight regulation (“set-point” versus “settling point”). Finally, our model has identified potential avenues for future investigations.National Institutes of Health (U.S.) (grant NIH CA80124)National Institutes of Health (U.S.) (grant NIH CA85140)National Institutes of Health (U.S.) (grant NIH CA96915)National Institutes of Health (U.S.) (grant NIH CA115767

Secreted Gaussia Luciferase as a Biomarker for Monitoring Tumor Progression and Treatment Response of Systemic Metastases

Background: Currently, only few techniques are available for quantifying systemic metastases in preclinical model. Thus techniques that can sensitively detect metastatic colonization and assess treatment response in real-time are urgently needed. To this end, we engineered tumor cells to express a naturally secreted Gaussia luciferase (Gluc), and investigated its use as a circulating biomarker for monitoring viable metastatic or primary tumor growth and their treatment responses. Methodology/Principal Findings: We first developed orthotopic primary and metastatic breast tumors with derivative of MDA-MB-231 cells expressing Gluc. We then correlated tumor burden with Gluc activity in the blood and urine along with bioluminescent imaging (BLI). Second, we utilized blood Gluc assay to monitor treatment response to lapatinib in an experimental model of systemic metastasis. We observed good correlation between the primary tumor volume and Gluc concentration in blood (R2 = 0.84) and urine (R2 = 0.55) in the breast tumor model. The correlation deviated as a primary tumor grew due to a reduction in viable tumor fraction. This was also supported by our mathematical models for tumor growth to compare the total and viable tumor burden in our model. In the experimental metastasis model, we found numerous brain metastases as well as systemic metastases including bone and lungs. Importantly, blood Gluc assay revealed early growth of metastatic tumors before BLI could visualize their presence. Using secreted Gluc, we localized systemic metastases by BLI and quantitatively monitored the total viable metastatic tumor burden by blood Gluc assay during the course of treatment with lapatinib, a dual tyrosine kinase inhibitor of EGFR and HER2. Conclusion/Significance: We demonstrated secreted Gluc assay accurately reflects the amount of viable cancer cells in primary and metastatic tumors. Blood Gluc activity not only tracks metastatic tumor progression but also serves as a longitudinal biomarker for tumor response to treatments

Correlative intravital imaging of cGMP signals and vasodilation in mice

Cyclic guanosine monophosphate (cGMP) is an important signaling molecule and drug target in the cardiovascular system. It is well known that stimulation of the vascular nitric oxide (NO)-cGMP pathway results in vasodilation. However, the spatiotemporal dynamics of cGMP signals themselves and the cGMP concentrations within specific cardiovascular cell types in health, disease, and during pharmacotherapy with cGMP-elevating drugs are largely unknown. To facilitate the analysis of cGMP signaling in vivo, we have generated transgenic mice that express fluorescence resonance energy transfer (FRET)-based cGMP sensor proteins. Here, we describe two models of intravital FRET/cGMP imaging in the vasculature of cGMP sensor mice: (1) epifluorescence-based ratio imaging in resistance-type vessels of the cremaster muscle and (2) ratio imaging by multiphoton microscopy within the walls of subcutaneous blood vessels accessed through a dorsal skinfold chamber. Both methods allow simultaneous monitoring of NO-induced cGMP transients and vasodilation in living mice. Detailed protocols of all steps necessary to perform and evaluate intravital imaging experiments of the vasculature of anesthetized mice including surgery, imaging, and data evaluation are provided. An image segmentation approach is described to estimate FRET/cGMP changes within moving structures such as the vessel wall during vasodilation. The methods presented herein should be useful to visualize cGMP or other biochemical signals that are detectable with FRET-based biosensors, such as cyclic adenosine monophosphate or Ca2+, and to correlate them with respective vascular responses. With further refinement and combination of transgenic mouse models and intravital imaging technologies, we envision an exciting future, in which we are able to “watch” biochemistry, (patho-)physiology, and pharmacotherapy in the context of a living mammalian organism

Blockade of VEGFR2 and Not VEGFR1 Can Limit Diet-Induced Fat Tissue Expansion: Role of Local versus Bone Marrow-Derived Endothelial Cells

Background: We investigated if new vessel formation in fat involves the contribution of local tissue-derived endothelial cells (i.e., angiogenesis) or bone marrow-derived cells (BMDCs, i.e. vasculogenesis) and if antiangiogenic treatment by blockade of vascular endothelial growth factor (VEGF) receptors can prevent diet-induced obesity (DIO). Methodology/Principal Findings: We performed restorative bone marrow transplantation into wild-type mice using transgenic mice expressing green fluorescent protein (GFP) constitutively (driven by β-actin promoter) or selectively in endothelial cells (under Tie2 promoter activation) as donors. The presence of donor BMDCs in recipient mice was investigated in fat tissue vessels after DIO using in vivo and ex vivo fluorescence microscopy. We investigated the roles of VEGF receptors 1 and 2 (VEGFR1/VEGFR2) by inducing DIO in mice and treating them with blocking monoclonal antibodies. We found only marginal (less than 1%) incorporation of BMDCs in fat vessels during DIO. When angiogenesis was inhibited by blocking VEGFR2 in mice with DIO, treated mice had significantly lower body weights than control animals. In contrast, blocking VEGFR1 had no discernable effect on the weight gain during DIO. Conclusions/Significance: Formation of new vessels in fat tissues during DIO is largely due to angiogenesis rather than de novo vasculogenesis. Antiangiogenic treatment by blockade of VEGFR2 but not VEGFR1 may limit adipose tissue expansion

Secreted Gaussia Luciferase as a Biomarker for Monitoring Tumor Progression and Treatment Response of Systemic Metastases

Currently, only few techniques are available for quantifying systemic metastases in preclinical model. Thus techniques that can sensitively detect metastatic colonization and assess treatment response in real-time are urgently needed. To this end, we engineered tumor cells to express a naturally secreted Gaussia luciferase (Gluc), and investigated its use as a circulating biomarker for monitoring viable metastatic or primary tumor growth and their treatment responses.We first developed orthotopic primary and metastatic breast tumors with derivative of MDA-MB-231 cells expressing Gluc. We then correlated tumor burden with Gluc activity in the blood and urine along with bioluminescent imaging (BLI). Second, we utilized blood Gluc assay to monitor treatment response to lapatinib in an experimental model of systemic metastasis. We observed good correlation between the primary tumor volume and Gluc concentration in blood (R(2) = 0.84) and urine (R(2) = 0.55) in the breast tumor model. The correlation deviated as a primary tumor grew due to a reduction in viable tumor fraction. This was also supported by our mathematical models for tumor growth to compare the total and viable tumor burden in our model. In the experimental metastasis model, we found numerous brain metastases as well as systemic metastases including bone and lungs. Importantly, blood Gluc assay revealed early growth of metastatic tumors before BLI could visualize their presence. Using secreted Gluc, we localized systemic metastases by BLI and quantitatively monitored the total viable metastatic tumor burden by blood Gluc assay during the course of treatment with lapatinib, a dual tyrosine kinase inhibitor of EGFR and HER2.We demonstrated secreted Gluc assay accurately reflects the amount of viable cancer cells in primary and metastatic tumors. Blood Gluc activity not only tracks metastatic tumor progression but also serves as a longitudinal biomarker for tumor response to treatments

Vascular diseases await translation of blood vessels engineered from stem cells

The discovery of human induced pluripotent stem cells (hiPSCs) opened new avenues for research and potential clinical applications for tissue regeneration and engineering. In particular,hiPSCs may offer a long sought solution for obtaining large numbers of autologous cells sufficient for tissue engineering. For vascular tissue engineering, several methods of generating endothelial cells or perivascular cells from hiPSCs in vitro have been reported. We review the current developments in the generation of vascular progenitor cells from hiPSCs and their functional capacity in vivo, and discuss the opportunities and challenges ahead for clinical translation and modeling vascular diseases using hiPSC-derived vascular cells with a focus on diabetes

Arginine dependence of tumor cells: targeting a chink in cancer’s armor

Arginine, one among the twenty most common natural amino acids, plays a pivotal role in cellular physiology as it is being involved in numerous cellular metabolic and signaling pathways. Dependence on arginine is diverse for both tumor and normal cells. Due to decreased expression of argininosuccinate synthetase (ASS) and/or ornithine transcarbamoylase (OTC), several types of tumor are auxotrophic for arginine. Deprivation of arginine exploits a significant vulnerability of these tumor cells and leads to their rapid demise. Hence, enzyme-mediated arginine depletion is a potential strategy for the selective destruction of tumor cells. Arginase, arginine deiminase (ADI) and arginine decarboxylase (ADC) are potential enzymes that may be used for arginine deprivation therapy. These arginine catabolizing enzymes not only reduce tumor growth but also make them susceptible to concomitantly administered anti-cancer therapeutics. Most of these enzymes are currently under clinical investigations and if successful will potentially be advanced as anti-cancer modalities

A study of the X-rayed outflow of APM 08279+5255 through photoionization codes

We present new results from our study of the X-rayed outflow of the z = 3.91 gravitationally lensed broad absorption line (BAL) quasar APM 08279+5255. These results are based on spectral fits to all the long exposure observations of APM 08279+5255 using a new quasar-outflow model. This model is based on cloudy simulations of a near-relativistic quasar outflow. The main conclusions from our multi-epoch spectral re-analysis of Chandra, XMM-Newton and Suzaku observations of APM 08279+5255 are: 1) In every observation we confirm the presence of two strong features, one at rest-frame energies between 1-4 keV, and the other between 7-18 keV. 2) We confirm that the low-energy absorption (1-4 keV rest-frame) arises from a low-ionization absorber with logNH~23 and the high-energy absorption (7-18 keV rest-frame) arises from highly ionized (3>log xi>4; where xi is the ionization parameter) iron in a near-relativistic outflowing wind. Assuming this interpretation, we find that the velocities on the outflow could get up to ~0.7c. 3) We confirm a correlation between the maximum outflow velocity and the photon index and find possible trends between the maximum outflow velocity and the X-ray luminosity, and between the total column density and the photon index. We performed calculations of the force multipliers of material illuminated by absorbed power laws and a Mathews-Ferland SED. We found that variations of the X-ray and UV parts of the SEDs and the presence of a moderate absorbing shield will produce important changes in the strength of the radiative driving force. These results support the observed trend found between the outflow velocity and X-ray photon index in APM 08279+5255. If this result is confirmed it will imply that radiation pressure is an important mechanism in producing quasar outflows.Comment: Paper accepted in the Astrophysical journa

Micelle-Encapsulated Quantum Dot-Porphyrin Assemblies as

Micelles have been employed to encapsulate the supramolecular assembly of quantum dots with palladium(II) porphyrins for the quantification of O₂ levels in aqueous media and in vivo. Förster resonance energy transfer from the quantum dot (QD) to the palladium porphyrin provides a means for signal transduction under both one- and two-photon excitation. The palladium porphyrins are sensitive to O₂ concentrations in the range of 0–160 Torr. The micelle-encapsulated QD-porphyrin assemblies have been employed for in vivo multiphoton imaging and lifetime-based oxygen measurements in mice with chronic dorsal skinfold chambers or cranial windows. Our results establish the utility of the QD-micelle approach for in vivo biological sensing applications.National Cancer Institute (U.S.) (R01- CA126642)International Society for Neurochemistry (W911NF-07-D-0004)United States. Dept. of Energy. Office of Basic Energy Sciences (DE-SC0009758