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1.
IEEE Open J Eng Med Biol ; 1: 265-267, 2020.
Article En | MEDLINE | ID: mdl-33748768

GOAL: To develop a micron-scale device that can operate as an MRI-based reporter for the presence of SARS-CoV-2 virus. METHODS: Iron rod microdevices were constructed via template-guided synthesis and suspended in phosphate buffered saline (PBS). Heat-inactivated SARS-CoV-2 viruses were added to the samples and imaged with low-field MRI. RESULTS: MRI of microdevices and viruses showed decreased signal intensity at low concentrations of viruses that recovered at higher concentrations. Electron micrographs suggest that reduced MRI intensity may be due to concentration-dependent shielding of water protons from local magnetic inhomogeneities caused by the iron microdevices. CONCLUSIONS: The preliminary results presented in this letter provide justification for further studies exploring the potential diagnostic role of magnetic microdevices in assessing the presence and concentration of SARS-CoV-2 viruses.

2.
Mol Neurodegener ; 13(1): 17, 2018 04 04.
Article En | MEDLINE | ID: mdl-29618365

BACKGROUND: Traumatic Brain Injury (TBI) is a major cause of disability and mortality, to which there is currently no comprehensive treatment. Blood Brain Barrier (BBB) dysfunction is well documented in human TBI patients, yet the molecular mechanisms that underlie this neurovascular unit (NVU) pathology remains unclear. The apolipoprotein-E (apoE) protein has been implicated in controlling BBB integrity in an isoform dependent manner, via suppression of Cyclophilin A (CypA)-Matrix metallopeptidase-9 (MMP-9) signaling cascades, however the contribution of this pathway in TBI-induced BBB permeability is not fully investigated. METHODS: We exposed C57Bl/6 mice to controlled cortical impact and assessed NVU and BBB permeability responses up to 21 days post-injury. We pharmacologically probed the role of the CypA-MMP-9 pathway in BBB permeability after TBI using Cyclosporin A (CsA, 20 mg/kg). Finally, as the apoE4 protein is known to be functionally deficient compared to the apoE3 protein, we used humanized APOE mice as a clinically relevant model to study the role of apoE on BBB injury and repair after TBI. RESULTS: In C57Bl/6 mice there was an inverse relationship between soluble apoE and BBB permeability, such that damaged BBB stabilizes as apoE levels increase in the days following TBI. TBI mice displayed acute pericyte loss, increased MMP-9 production and activity, and reduced tight-junction expression. Treatment with the CypA antagonist CsA in C57Bl/6 mice attenuates MMP-9 responses and enhances BBB repair after injury, demonstrating that MMP-9 plays an important role in the timing of spontaneous BBB repair after TBI. We also show that apoe mRNA is present in both astrocytes and pericytes after TBI. We report that APOE3 and APOE4 mice have similar acute BBB responses to TBI, but APOE3 mice display faster spontaneous BBB repair than APOE4 mice. Isolated microvessel analysis reveals delayed pericyte repopulation, augmented and sustained MMP-9 expression at the NVU, and impaired stabilization of Zonula Occludens-1, Occludin and Claudin-5 expression at tight junctions in APOE4 mice after TBI compared to APOE3 mice. CONCLUSIONS: These data confirm apoE as an important modulator of spontaneous BBB stabilization following TBI, and highlights the APOE4 allele as a risk factor for poor outcome after TBI.


Apolipoprotein E4/metabolism , Blood-Brain Barrier/pathology , Brain Injuries, Traumatic/metabolism , Brain Injuries, Traumatic/pathology , Animals , Apolipoprotein E3/metabolism , Capillary Permeability/physiology , Humans , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic
3.
Neurobiol Dis ; 111: 80-90, 2018 03.
Article En | MEDLINE | ID: mdl-29274430

Acoustically evoked seizures (e.g., audiogenic seizures or AGS) are common in models of inherited epilepsy and occur in a variety of species including rat, mouse, and hamster. Two models that have been particularly well studied are the genetically epilepsy prone rat (GEPR-3) and the Wistar Audiogenic Rat (WAR) strains. Acute and repeated AGS, as well as comorbid conditions, displays a close phenotypic overlap in these models. Whether these similarities arise from convergent or divergent structural changes in the brain remains unknown. Here, we examined the brain structure of Sprague Dawley (SD) and Wistar (WIS) rats, and quantified changes in the GEPR-3 and WAR, respectively. Brains from adult, male rats of each strain (n=8-10 per group) were collected, fixed, and embedded in agar and imaged using a 7 tesla Bruker MRI. Post-acquisition analysis included voxel-based morphometry (VBM), diffusion tensor imaging (DTI), and manual volumetric tracing. In the VBM analysis, GEPR-3 displayed volumetric changes in brainstem structures known to be engaged by AGS (e.g., superior and inferior colliculus, periaqueductal grey) and in forebrain structures (e.g., striatum, septum, nucleus accumbens). WAR displayed volumetric changes in superior colliculus, and a broader set of limbic regions (e.g., hippocampus, amygdala/piriform cortex). The only area of significant overlap in the two strains was the midline cerebellum: both GEPR-3 and WAR showed decreased volume compared to their control strains. In the DTI analysis, GEPR-3 displayed decreased fractional anisotropy (FA) in the corpus callosum, posterior commissure and commissure of the inferior colliculus (IC). WAR displayed increased FA only in the commissure of IC. These data provide a biological basis for further comparative and mechanistic studies in the GEPR-3 and WAR models, as well as provide additional insight into commonalities in the pathways underlying AGS susceptibility and behavioral comorbidity.


Brain/diagnostic imaging , Epilepsy/diagnostic imaging , Magnetic Resonance Imaging , Rats, Sprague-Dawley , Rats, Wistar , Animals , Brain/pathology , Disease Models, Animal , Epilepsy/pathology , Image Processing, Computer-Assisted , Male , Organ Size , Phenotype , Species Specificity
4.
IEEE Int Conf Rehabil Robot ; 2017: 346-351, 2017 07.
Article En | MEDLINE | ID: mdl-28813843

To restore locomotor function following spinal cord injury the disrupted descending supraspinal drive needs to be re-connected to regions caudal to the injury. Robotic gait training aims to facilitate recovery by stimulating the proprioceptive networks of the legs in a coordinated walking pattern while the descending supraspinal connections are re-established. In incomplete injuries, it is believed that the interneuronal networks near the injury site form relay circuits to reroute the supraspinal signals through the spared tissue. Stimulating these neurons should lead to increased arborization, and more robust relay circuits. We set out to show that robotic gait training is more effective when it is paired with training that activates interneurons at the level of the injury. Following a C4/5 over-hemisection injury the addition of skilled forelimb training actually reduced the gains found with robotic gait training alone. Not just gains within the training device, but also in independent overground locomotion. MRI imaging shows that skilled forelimb training reduces cellular activity, and reduces the organization of the axonal tracks at the injury site. This may be evidence that spared interneurons have limited resources and/or ability in establishing relay circuits to route supraspinal drive around the injury site. Skilled forelimb training and robotic gait training may not be synergistic, but competitive rehabilitative tasks.


Exercise Therapy , Forelimb/physiopathology , Gait/physiology , Robotics/methods , Spinal Cord Injuries , Animals , Female , Rats , Rats, Sprague-Dawley , Recovery of Function/physiology , Spinal Cord Injuries/physiopathology , Spinal Cord Injuries/rehabilitation
5.
Am J Pathol ; 186(11): 3040-3053, 2016 11.
Article En | MEDLINE | ID: mdl-27743558

Neuroblastoma (NB) is a pediatric malignant neoplasm of sympathoadrenal origin. Challenges in its management include stratification of this heterogeneous disease and a lack of both adequate treatments for high-risk patients and noninvasive biomarkers of disease progression. Our previous studies have identified neuropeptide Y (NPY), a sympathetic neurotransmitter expressed in NB, as a potential therapeutic target for these tumors by virtue of its Y5 receptor (Y5R)-mediated chemoresistance and Y2 receptor (Y2R)-mediated proliferative and angiogenic activities. The goal of this study was to determine the clinical relevance and utility of these findings. Expression of NPY and its receptors was evaluated in corresponding samples of tumor RNA, tissues, and sera from 87 patients with neuroblastic tumors and in tumor tissues from the TH-MYCN NB mouse model. Elevated serum NPY levels correlated with an adverse clinical presentation, poor survival, metastasis, and relapse, whereas strong Y5R immunoreactivity was a marker of angioinvasive tumor cells. In NB tissues from TH-MYCN mice, high immunoreactivity of both NPY and Y5R marked angioinvasive NB cells. Y2R was uniformly expressed in undifferentiated tumor cells, which supports its previously reported role in NB cell proliferation. Our findings validate NPY as a therapeutic target for advanced NB and implicate the NPY/Y5R axis in disease dissemination. The correlation between elevated systemic NPY and NB progression identifies serum NPY as a novel NB biomarker.


Neuroblastoma/metabolism , Neuropeptide Y/metabolism , Adolescent , Animals , Biomarkers/metabolism , Cell Proliferation , Child , Child, Preschool , Disease Progression , Female , Humans , Infant , Infant, Newborn , Male , Mice , Neuroblastoma/drug therapy , Neuroblastoma/pathology , Neuropeptide Y/genetics , Receptors, Neuropeptide Y/genetics , Receptors, Neuropeptide Y/metabolism
6.
J Vis Exp ; (118)2016 12 09.
Article En | MEDLINE | ID: mdl-28060251

Hypoxia has been implicated in the metastasis of Ewing sarcoma (ES) by clinical observations and in vitro data, yet direct evidence for its pro-metastatic effect is lacking and the exact mechanisms of its action are unclear. Here, we report an animal model that allows for direct testing of the effects of tumor hypoxia on ES dissemination and investigation into the underlying pathways involved. This approach combines two well-established experimental strategies, orthotopic xenografting of ES cells and femoral artery ligation (FAL), which induces hindlimb ischemia. Human ES cells were injected into the gastrocnemius muscles of SCID/beige mice and the primary tumors were allowed to grow to a size of 250 mm3. At this stage either the tumors were excised (control group) or the animals were subjected to FAL to create tumor hypoxia, followed by tumor excision 3 days later. The efficiency of FAL was confirmed by a significant increase in binding of hypoxyprobe-1 in the tumor tissue, severe tumor necrosis and complete inhibition of primary tumor growth. Importantly, despite these direct effects of ischemia, an enhanced dissemination of tumor cells from the hypoxic tumors was observed. This experimental strategy enables comparative analysis of the metastatic properties of primary tumors of the same size, yet significantly different levels of hypoxia. It also provides a new platform to further assess the mechanistic basis for the hypoxia-induced alterations that occur during metastatic tumor progression in vivo. In addition, while this model was established using ES cells, we anticipate that this experimental strategy can be used to test the effect of hypoxia in other sarcomas, as well as tumors orthotopically implanted in sites with a well-defined blood supply route.


Hypoxia/pathology , Neoplasm Metastasis/pathology , Sarcoma, Ewing/pathology , Animals , Disease Models, Animal , Humans , Mice , Mice, SCID , Neoplasm Transplantation
7.
Aging (Albany NY) ; 7(10): 854-68, 2015 Oct.
Article En | MEDLINE | ID: mdl-26540407

Medulloblastoma (MB), a primitive neuroectodermal tumor, is the most common malignant childhood brain tumor and remains incurable in about a third of patients. Currently, survivors carry a significant burden of late treatment effects. The p53 tumor suppressor protein plays a crucial role in influencing cell survival in response to cellular stress and while the p53 pathway is considered a key determinant of anti-tumor responses in many tumors, its role in cell survival in MB is much less well defined. Herein, we report that the experimental drug VMY-1-103 acts through induction of a partial DNA damage-like response as well induction of non-survival autophagy. Surprisingly, the genetic or chemical silencing of p53 significantly enhanced the cytotoxic effects of both VMY and the DNA damaging drug, doxorubicin. The inhibition of p53 in the presence of VMY revealed increased late stage apoptosis, increased DNA fragmentation and increased expression of genes involved in apoptosis, including CAPN12 and TRPM8, p63, p73, BIK, EndoG, CIDEB, P27Kip1 and P21cip1. These data provide the groundwork for additional studies on VMY as a therapeutic drug and support further investigations into the intriguing possibility that targeting p53 function may be an effective means of enhancing clinical outcomes in MB.


Adenine/analogs & derivatives , Antineoplastic Agents/pharmacology , Dansyl Compounds/pharmacology , Medulloblastoma/drug therapy , Tumor Suppressor Protein p53/antagonists & inhibitors , Adenine/pharmacology , Adenine/therapeutic use , Antineoplastic Agents/therapeutic use , Cell Death/drug effects , Cell Line, Tumor , Dansyl Compounds/therapeutic use , Drug Evaluation, Preclinical , Humans , Signal Transduction/drug effects
8.
Oncotarget ; 5(6): 1683-98, 2014 Mar 30.
Article En | MEDLINE | ID: mdl-24742967

Strigolactones are a novel class of plant hormones produced in roots and regulate shoot and root development. We have previously shown that synthetic strigolactone analogues potently inhibit growth of breast cancer cells and breast cancer stem cells. Here we show that strigolactone analogues inhibit the growth and survival of an array of cancer-derived cell lines representing solid and non-solid cancer cells including: prostate, colon, lung, melanoma, osteosarcoma and leukemic cell lines, while normal cells were minimally affected. Treatment of cancer cells with strigolactone analogues was hallmarked by activation of the stress-related MAPKs: p38 and JNK and induction of stress-related genes; cell cycle arrest and apoptosis evident by increased percentages of cells in the sub-G1 fraction and Annexin V staining. In addition, we tested the response of patient-matched conditionally reprogrammed primary prostate normal and cancer cells. The tumor cells exhibited significantly higher sensitivity to the two most potent SL analogues with increased apoptosis confirmed by PARP1 cleavage compared to their normal counterpart cells. Thus, Strigolactone analogues are promising candidates for anticancer therapy by their ability to specifically induce cell cycle arrest, cellular stress and apoptosis in tumor cells with minimal effects on growth and survival of normal cells.


Apoptosis/drug effects , JNK Mitogen-Activated Protein Kinases/metabolism , Lactones/pharmacology , Neoplasms/pathology , Neoplastic Stem Cells/pathology , Prostatic Neoplasms/pathology , p38 Mitogen-Activated Protein Kinases/metabolism , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Blotting, Western , Cell Cycle/drug effects , Cell Cycle Checkpoints/drug effects , Cell Proliferation/drug effects , Gene Expression Profiling , Heterocyclic Compounds, 3-Ring/pharmacology , Humans , JNK Mitogen-Activated Protein Kinases/genetics , Male , Neoplasms/drug therapy , Neoplasms/metabolism , Neoplastic Stem Cells/drug effects , Neoplastic Stem Cells/metabolism , Oligonucleotide Array Sequence Analysis , Prostate/drug effects , Prostate/metabolism , Prostatic Neoplasms/drug therapy , Prostatic Neoplasms/metabolism , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction/drug effects , Tumor Cells, Cultured , p38 Mitogen-Activated Protein Kinases/genetics
9.
Oncotarget ; 5(5): 1212-25, 2014 Mar 15.
Article En | MEDLINE | ID: mdl-24681808

Mutations of the p53 gene hallmark many human cancers. Several p53 mutant proteins acquire the capability to promote cancer progression and metastasis, a phenomenon defined as Gain of Oncogenic Function (GOF). The downstream targets by which GOF p53 mutants perturb cellular programs relevant to oncogenesis are only partially known. We have previously demonstrated that SLC25A1 (CIC) promotes tumorigenesis, while its inhibition blunts tumor growth. We now report that CIC is a direct transcriptional target of several p53 mutants. We identify a novel interaction between mutant p53 (mutp53) and the transcription factor FOXO-1 which is responsible for regulation of CIC expression levels. Tumor cells harboring mutp53 display higher CIC levels relative to p53 null or wild-type tumors, and inhibition of CIC activity blunts mutp53-driven tumor growth, partially overcoming GOF activity. CIC inhibition also enhances the chemotherapeutic potential of platinum-based agents. Finally, we found that elevated CIC levels predict poor survival outcome in tumors hallmarked by high frequency of p53 mutations. Our results identify CIC as a novel target of mutp53 and imply that the employment of CIC inhibitors may improve survival rates and reduce chemo-resistance in tumors harboring these types of mutations, which are among the most intractable forms of cancers.


Anion Transport Proteins/genetics , Biomarkers, Tumor/genetics , Breast Neoplasms/genetics , Carcinogenesis/genetics , Forkhead Transcription Factors/metabolism , Gene Expression Regulation, Neoplastic/genetics , Lung Neoplasms/genetics , Mitochondrial Proteins/genetics , Ovarian Neoplasms/genetics , Tumor Suppressor Protein p53/genetics , Animals , Anion Transport Proteins/antagonists & inhibitors , Antineoplastic Agents/pharmacology , Benzene Derivatives/pharmacology , Carcinogenesis/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cell Proliferation/genetics , Cisplatin/pharmacology , Drug Resistance, Neoplasm/genetics , Female , Forkhead Box Protein O1 , Humans , Kaplan-Meier Estimate , Mice , Mitochondrial Proteins/antagonists & inhibitors , Mutation , Organic Anion Transporters , Ovarian Neoplasms/drug therapy , Prognosis , Transcription, Genetic , Tricarboxylic Acids/pharmacology , Tumor Suppressor Protein p53/metabolism
10.
Article En | MEDLINE | ID: mdl-25750594

Ever growing "omics" data and continuously accumulated biological knowledge provide an unprecedented opportunity to identify molecular biomarkers and their interactions that are responsible for cancer phenotypes that can be accurately defined by clinical measurements such as in vivo imaging. Since signaling or regulatory networks are dynamic and context-specific, systematic efforts to characterize such structural alterations must effectively distinguish significant network rewiring from random background fluctuations. Here we introduced a novel integration of network biology and imaging to study cancer phenotypes and responses to treatments at the molecular systems level. Specifically, Differential Dependence Network (DDN) analysis was used to detect statistically significant topological rewiring in molecular networks between two phenotypic conditions, and in vivo Magnetic Resonance Imaging (MRI) was used to more accurately define phenotypic sample groups for such differential analysis. We applied DDN to analyze two distinct phenotypic groups of breast cancer and study how genomic instability affects the molecular network topologies in high-grade ovarian cancer. Further, FDA-approved arsenic trioxide (ATO) and the ND2-SmoA1 mouse model of Medulloblastoma (MB) were used to extend our analyses of combined MRI and Reverse Phase Protein Microarray (RPMA) data to assess tumor responses to ATO and to uncover the complexity of therapeutic molecular biology.


Computational Biology/methods , Gene Regulatory Networks/physiology , Neoplasms , Signal Transduction/physiology , Animals , Magnetic Resonance Imaging , Mice , Molecular Imaging , Neoplasms/classification , Neoplasms/metabolism , Neoplasms/pathology , Phenotype
11.
ACS Nano ; 7(10): 9040-8, 2013 Oct 22.
Article En | MEDLINE | ID: mdl-24047405

Metal-oxo clusters have been used as building blocks to form hybrid nanomaterials and evaluated as potential MRI contrast agents. We have synthesized a biocompatible copolymer based on a water stable, nontoxic, mixed-metal-oxo cluster, Mn8Fe4O12(L)16(H2O)4, where L is acetate or vinyl benzoic acid, and styrene. The cluster alone was screened by NMR for relaxivity and was found to be a promising T2 contrast agent, with r1 = 2.3 mM(-1) s(-1) and r2 = 29.5 mM(-1) s(-1). Initial cell studies on two human prostate cancer cell lines, DU-145 and LNCap, reveal that the cluster has low cytotoxicity and may be potentially used in vivo. The metal-oxo cluster Mn8Fe4(VBA)16 (VBA = vinyl benzoic acid) can be copolymerized with styrene under miniemulsion conditions. Miniemulsion allows for the formation of nanometer-sized paramagnetic beads (~80 nm diameter), which were also evaluated as a contrast agent for MRI. These highly monodispersed, hybrid nanoparticles have enhanced properties, with the option for surface functionalization, making them a promising tool for biomedicine. Interestingly, both relaxivity measurements and MRI studies show that embedding the Mn8Fe4 core within a polymer matrix decreases r2 effects with little effect on r1, resulting in a positive T1 contrast enhancement.


Contrast Media , Magnetic Resonance Imaging/methods , Magnetics , Nanoparticles , Cell Line, Tumor , Humans , Magnetic Resonance Spectroscopy , Male , Microscopy, Electron, Transmission
12.
EMBO Mol Med ; 5(10): 1569-85, 2013 10.
Article En | MEDLINE | ID: mdl-24014378

Absence of dystrophin makes skeletal muscle more susceptible to injury, resulting in breaches of the plasma membrane and chronic inflammation in Duchenne muscular dystrophy (DMD). Current management by glucocorticoids has unclear molecular benefits and harsh side effects. It is uncertain whether therapies that avoid hormonal stunting of growth and development, and/or immunosuppression, would be more or less beneficial. Here, we discover an oral drug with mechanisms that provide efficacy through anti-inflammatory signaling and membrane-stabilizing pathways, independent of hormonal or immunosuppressive effects. We find VBP15 protects and promotes efficient repair of skeletal muscle cells upon laser injury, in opposition to prednisolone. Potent inhibition of NF-κB is mediated through protein interactions of the glucocorticoid receptor, however VBP15 shows significantly reduced hormonal receptor transcriptional activity. The translation of these drug mechanisms into DMD model mice improves muscle strength, live-imaging and pathology through both preventive and post-onset intervention regimens. These data demonstrate successful improvement of dystrophy independent of hormonal, growth, or immunosuppressive effects, indicating VBP15 merits clinical investigation for DMD and would benefit other chronic inflammatory diseases.


Anti-Inflammatory Agents/pharmacology , Myoblasts/drug effects , Pregnadienediols/pharmacology , Animals , Anti-Inflammatory Agents/toxicity , Cell Line , Cell Membrane/drug effects , Cell Membrane/physiology , Immunosuppressive Agents/pharmacology , Immunosuppressive Agents/toxicity , Lasers , Mice , Mice, Inbred mdx , Muscular Dystrophies/metabolism , Muscular Dystrophies/pathology , Myoblasts/cytology , Myoblasts/radiation effects , NF-kappa B/antagonists & inhibitors , NF-kappa B/metabolism , Necrosis/etiology , Phenotype , Prednisolone/pharmacology , Prednisolone/toxicity , Pregnadienediols/toxicity , Protein Interaction Maps/drug effects , Receptors, Glucocorticoid/genetics , Receptors, Glucocorticoid/metabolism , Signal Transduction/drug effects , T-Lymphocytes/drug effects , T-Lymphocytes/immunology , Transcription, Genetic/drug effects
13.
Aging (Albany NY) ; 5(8): 607-22, 2013 Aug.
Article En | MEDLINE | ID: mdl-23928451

Despite recent epidemiological evidences linking radiation exposure and a number of human ailments including cancer, mechanistic understanding of how radiation inflicts long-term changes in cerebral cortex, which regulates important neuronal functions, remains obscure. The current study dissects molecular events relevant to pathology in cerebral cortex of 6 to 8 weeks old female C57BL/6J mice two and twelve months after exposure to a γ radiation dose (2 Gy) commonly employed in fractionated radiotherapy. For a comparative study, effects of 1.6 Gy heavy ion 56Fe radiation on cerebral cortex were also investigated, which has implications for space exploration. Radiation exposure was associated with increased chronic oxidative stress, oxidative DNA damage, lipid peroxidation, and apoptosis. These results when considered with decreased cortical thickness, activation of cell-cycle arrest pathway, and inhibition of DNA double strand break repair factors led us to conclude to our knowledge for the first time that radiation caused aging-like pathology in cerebral cortical cells and changes after heavy ion radiation were more pronounced than γ radiation.


Cellular Senescence/radiation effects , Cerebral Cortex/radiation effects , DNA Repair/radiation effects , Animals , Apoptosis/radiation effects , Cell Cycle Checkpoints/radiation effects , Cerebral Cortex/pathology , Cerebral Cortex/physiology , DNA Damage/radiation effects , Dose-Response Relationship, Radiation , Female , Gamma Rays , Glial Fibrillary Acidic Protein , Lipid Peroxidation/radiation effects , Mice , Mice, Inbred C57BL , Nerve Tissue Proteins/metabolism , Nestin/metabolism , Oxidative Stress/radiation effects , Radiation, Ionizing , Reactive Oxygen Species/metabolism , Up-Regulation/radiation effects
14.
Am J Pathol ; 182(2): 312-8, 2013 Feb.
Article En | MEDLINE | ID: mdl-23219428

Biologically accurate mouse models of human cancer have become important tools for the study of human disease. The anatomical location of various target organs, such as brain, pancreas, and prostate, makes determination of disease status difficult. Imaging modalities, such as magnetic resonance imaging, can greatly enhance diagnosis, and longitudinal imaging of tumor progression is an important source of experimental data. Even in models where the tumors arise in areas that permit visual determination of tumorigenesis, longitudinal anatomical and functional imaging can enhance the scope of studies by facilitating the assessment of biological alterations, (such as changes in angiogenesis, metabolism, cellular invasion) as well as tissue perfusion and diffusion. One of the challenges in preclinical imaging is the development of infrastructural platforms required for integrating in vivo imaging and therapeutic response data with ex vivo pathological and molecular data using a more systems-based multiscale modeling approach. Further challenges exist in integrating these data for computational modeling to better understand the pathobiology of cancer and to better affect its cure. We review the current applications of preclinical imaging and discuss the implications of applying functional imaging to visualize cancer progression and treatment. Finally, we provide new data from an ongoing preclinical drug study demonstrating how multiscale modeling can lead to a more comprehensive understanding of cancer biology and therapy.


Magnetic Resonance Imaging/methods , Neoplasms/diagnosis , Research , Systems Biology/methods , Animals , Antineoplastic Agents/therapeutic use , Computational Biology , Humans , Neoplasms/drug therapy
15.
Cell Cycle ; 11(20): 3801-9, 2012 Oct 15.
Article En | MEDLINE | ID: mdl-22983062

The development of new small molecule-based therapeutic drugs requires accurate quantification of drug bioavailability, biological activity and treatment efficacy. Rapidly measuring these endpoints is often hampered by the lack of efficient assay platforms with high sensitivity and specificity. Using an in vivo model system, we report a simple and sensitive liquid chromatography-tandem mass spectrometry assay to quantify the bioavailability of a recently developed novel cyclin-dependent kinase inhibitor VMY-1-103, a purvalanol B-based analog whose biological activity is enhanced via dansylation. We developed a rapid organic phase extraction technique and validated wide and functional VMY-1-103 distribution in various mouse tissues, consistent with its enhanced potency previously observed in a variety of human cancer cell lines. More importantly, in vivo MRI and single voxel proton MR-Spectroscopy further established that VMY-1-103 inhibited disease progression and affected key metabolites in a mouse model of hedgehog-driven medulloblastoma.


Adenine/analogs & derivatives , Antineoplastic Agents/pharmacology , Cerebellar Neoplasms/drug therapy , Cyclin-Dependent Kinases/antagonists & inhibitors , Dansyl Compounds/pharmacology , Medulloblastoma/drug therapy , Protein Kinase Inhibitors/pharmacology , Adenine/chemistry , Adenine/pharmacokinetics , Adenine/pharmacology , Animals , Antineoplastic Agents/pharmacokinetics , Biological Availability , Cell Cycle/drug effects , Cerebellar Neoplasms/genetics , Cerebellar Neoplasms/metabolism , Chromatography, Liquid , Cyclin-Dependent Kinases/genetics , Cyclin-Dependent Kinases/metabolism , Dansyl Compounds/pharmacokinetics , Humans , Magnetic Resonance Imaging , Medulloblastoma/genetics , Medulloblastoma/metabolism , Mice , Protein Kinase Inhibitors/pharmacokinetics , Tandem Mass Spectrometry
16.
Toxicol Sci ; 122(1): 26-37, 2011 Jul.
Article En | MEDLINE | ID: mdl-21507987

Our studies found that BRCA1 levels negatively correlate with DNA adducts induced by Benzo(a)pyrene (BaP). Pulse-chase experiments showed that the increase in BaP-induced DNA adducts in BRCA1 knockdown cells may not be associated with BRCA1's function in nucleotide excision repair activity; rather, it may be associated with its function in modulating transcriptional regulation. BRCA1 knockdown in MCF-10A cells significantly attenuated the induction of CYP1A1 following BaP treatment indicating that the increase in BaP-induced adducts in BRCA1 knockdown cells is not CYP1A1 dependent. However, our study shows that BRCA1 defective cells may still be able to biotransform BaP by regulating other CYP enzymes, including CYP1B1. Knockdown of BRCA1 also severely affected the expression levels of two types of uridine diphosphate glucorunyltransferase (UGT1A1 and UGT1A9) and NRF2. Both UGTs are known as BaP-specific detoxification enzymes, and NRF2 is a master regulator of antioxidant and detoxification genes. Thus, we concluded that the increased amount of BaP-induced DNA adducts in BRCA1 knockdown cells is strongly associated with its loss of functional detoxification. Chromatin immunoprecipitation assay revealed that BRCA1 is recruited to the promoter/enhancer sequences of UGT1A1, UGT1A9, and NRF2. Regulation of UGT1A1 and UGT1A9 expression showed that the induction of DNA adducts by BaP is directly affected by their expression levels. Finally, overexpression of UGTs, NRF2, or ARNT significantly decreased the amount of BaP-induced adducts in BRCA1-deficient cells. Overall, our results suggest that BRCA1 protects cells by reducing the amount of BaP-induced DNA adducts possibly via transcriptional activation of detoxification gene expression.


BRCA1 Protein/metabolism , Benzo(a)pyrene/toxicity , DNA Adducts/analysis , Inactivation, Metabolic , Animals , Aryl Hydrocarbon Hydroxylases/genetics , Aryl Hydrocarbon Hydroxylases/metabolism , BRCA1 Protein/genetics , Biotransformation , Blotting, Western , Carcinogens, Environmental/toxicity , Cell Line , Chromatin Immunoprecipitation , Cytochrome P-450 CYP1A1/genetics , Cytochrome P-450 CYP1A1/metabolism , Cytochrome P-450 CYP1B1 , Female , Gene Expression Regulation , Gene Knockdown Techniques , Glucuronosyltransferase/genetics , Glucuronosyltransferase/metabolism , Humans , Mice , Mice, Knockout , NF-E2-Related Factor 2/genetics , NF-E2-Related Factor 2/metabolism , RNA, Small Interfering , Xenobiotics/toxicity
17.
Cell Cycle ; 9(9): 1824-9, 2010 May.
Article En | MEDLINE | ID: mdl-20404514

Diet and obesity, and their associated metabolic alterations, are some of the fastest-growing causes of disease and death in America. Findings from epidemiological studies correlating obesity, the sources of dietary fat and prostate cancer (PCa) are conflicting. We have previously shown that 15% of PB-ErbB-2 x pten(+/-) mice developed PCa and exhibited increased phosphorylated 4E-BP1, but not the key PI3-kinase intermediary phospho-protein, mTOR, when maintained on unrefined mouse chow. We report herein that 100% of animals fed refined, westernized AIN-93-based diets containing corn oil developed PCa by 12 months of age. Increases in visceral fat and mTO R activation in the tumors were also observed. Furthermore, nuclear cyclin E levels were significantly induced by the AIN-93-corn oil-based diets versus chow. Replacing 50% of the corn oil with menhaden oil, with 21% of its triglycerides being n-3 PUFA's, had no effect on tumorigenesis, fat deposition, cyclin E or mTOR. Phosphorylated BAD levels were similar in the tumors of mice in all three diets. Our data demonstrated that in the context of our preclinical model, components of crude chow, but not dietary n-3 PUFAs, protect against PCa progression. In addition, these data establish phosphorylated mTOR, nuclear cyclin E and visceral fat deposits as possible biomarkers of increased dietary risk for PCa.


Fatty Acids, Omega-3/therapeutic use , PTEN Phosphohydrolase/genetics , Prostatic Neoplasms/prevention & control , Receptor, ErbB-2/genetics , Adaptor Proteins, Signal Transducing , Animals , Carrier Proteins/metabolism , Cell Cycle Proteins , Cyclin E/metabolism , Disease Models, Animal , Disease Progression , Eukaryotic Initiation Factors , Intracellular Signaling Peptides and Proteins/metabolism , Male , Mice , PTEN Phosphohydrolase/metabolism , Phosphoproteins/metabolism , Phosphorylation , Prostatic Neoplasms/pathology , Protein Serine-Threonine Kinases/metabolism , Receptor, ErbB-2/metabolism , TOR Serine-Threonine Kinases , bcl-Associated Death Protein/metabolism
18.
Am J Pathol ; 174(6): 2051-60, 2009 Jun.
Article En | MEDLINE | ID: mdl-19443706

Loss of function at the Pten tumor-suppressor locus is a common genetic modification found in human prostate cancer. While recent in vivo and in vitro data support an important role of aberrant ErbB-2 signaling to clinically relevant prostate target genes, such as cyclin D1, the role of Pten in ErbB-2-induced prostate epithelial proliferation is not well understood. In the Pten-deficient prostate cancer cell line, LNCaP, restoration of Pten was able to inhibit ErbB-2- and heregulin-induced cell cycle progression, as well as cyclin D1 protein levels and promoter activity. Previously, we established that probasin-driven ErbB-2 transgenic mice presented with high-grade prostate intraepithelial neoplasia and increased nuclear cyclin D1 levels. We show that mono-allelic loss of pten in the probasin-driven-ErbB-2 model resulted in increased nuclear cyclin D1 and proliferating cell nuclear antigen levels and decreased disease latency compared to either individual genetic model and, unlike the probasin-driven-ErbB-2 mice, progression to adenocarcinoma. Activated 3-phosphoinositide-dependent protein kinase-1 was observed during cancer initiation combined with the activation of p70S6K (phospho-T389) and inactivation of the 4E-binding protein-1 (phosphorylated on T37/46) and was primarily restricted to those cases of prostate cancer that had progressed to adenocarcinoma. Activation of mTOR was not seen. Our data demonstrates that Pten functions downstream of ErbB-2 to restrict prostate epithelial transformation by blocking full activation of the PDK1 signaling cascade.


Adenocarcinoma/metabolism , PTEN Phosphohydrolase/metabolism , Prostatic Neoplasms/metabolism , Protein Serine-Threonine Kinases/metabolism , Receptor, ErbB-2/metabolism , Signal Transduction/physiology , 3-Phosphoinositide-Dependent Protein Kinases , Adenocarcinoma/genetics , Animals , Blotting, Western , Cell Line, Tumor , Flow Cytometry , Humans , Image Processing, Computer-Assisted , Immunohistochemistry , Male , Mice , Mice, Transgenic , Prostatic Neoplasms/genetics , Receptor, ErbB-2/genetics
19.
Cell Cycle ; 6(15): 1946-50, 2007 Aug 01.
Article En | MEDLINE | ID: mdl-17671425

Pheochromocytoma (PCC) is a rare catecholamine-producing tumor that arises from the adrenal medulla and is often familial. The genetic basis for familial PCC involves mutations of RET, VHL, SHDx or NF-1 in more than 20% of cases. Additional genes may be important in pathogenesis of both familial and sporadic PCC. ErbB-2/Her2/Neu is a growth factor receptor tyrosine kinase that is frequently overexpressed in tumors and there is clinical evidence suggesting that enhanced ErbB-2 growth factor receptor signaling may play a role in PCC. In the present study, ectopic expression of an activated ErbB-2 transgene resulted in bilateral adrenal PCC. Analyses of tumor samples and normal adrenal tissue revealed that levels of the Pten tumor suppressor protein were greatly reduced in PCCs, while levels of the cell cycle regulatory protein cyclin D1 were usually increased. In addition, levels of phospo-AKT were increased in PCCs versus normal adrenal tissue. Biochemical analyses established that PCC's were functionally active, producing abundant levels of the catecholamines, epinephrine and norepinephrine. These data establish that increased ErbB-2 growth factor receptor signaling in the adrenal medulla can lead to PCC through combined influences on Pten, AKT andcyclin D1.


Adrenal Gland Neoplasms/metabolism , Adrenal Gland Neoplasms/pathology , Cell Transformation, Neoplastic/metabolism , Cell Transformation, Neoplastic/pathology , Pheochromocytoma/metabolism , Pheochromocytoma/pathology , Receptor, ErbB-2/metabolism , Adrenal Gland Neoplasms/genetics , Animals , Cell Transformation, Neoplastic/genetics , Cyclin D1/metabolism , Gene Expression Regulation, Neoplastic , Immunohistochemistry , Mice , PTEN Phosphohydrolase/genetics , Pheochromocytoma/genetics , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction
20.
Mol Biol Cell ; 18(10): 3860-72, 2007 Oct.
Article En | MEDLINE | ID: mdl-17652459

In migrating cells, actin polymerization promotes protrusion of the leading edge, whereas actomyosin contractility powers net cell body translocation. Although they promote F-actin-dependent protrusions of the cell periphery upon adhesion to fibronectin (FN), Abl family kinases inhibit cell migration on FN. We provide evidence here that the Abl-related gene (Arg/Abl2) kinase inhibits fibroblast migration by attenuating actomyosin contractility and regulating focal adhesion dynamics. arg-/- fibroblasts migrate at faster average speeds than wild-type (wt) cells, whereas Arg re-expression in these cells slows migration. Surprisingly, the faster migrating arg-/- fibroblasts have more prominent F-actin stress fibers and focal adhesions and exhibit increased actomyosin contractility relative to wt cells. Interestingly, Arg requires distinct functional domains to inhibit focal adhesions and actomyosin contractility. The kinase domain-containing Arg N-terminal half can act through the RhoA inhibitor p190RhoGAP to attenuate stress fiber formation and cell contractility. However, Arg requires both its kinase activity and its cytoskeleton-binding C-terminal half to fully inhibit focal adhesions. Although focal adhesions do not turn over efficiently in the trailing edge of arg-/- cells, the increased contractility of arg-/- cells tears the adhesions from the substrate, allowing for the faster migration observed in these cells. Together, our data strongly suggest that Arg inhibits cell migration by restricting actomyosin contractility and regulating its coupling to the substrate through focal adhesions.


Actomyosin/metabolism , DNA-Binding Proteins/metabolism , Fibronectins/metabolism , Focal Adhesions/enzymology , GTPase-Activating Proteins/metabolism , Protein-Tyrosine Kinases/metabolism , Repressor Proteins/metabolism , Animals , Cell Adhesion , Cell Movement , Fibroblasts/cytology , Fibroblasts/enzymology , Mice , Mutant Proteins/metabolism , Protein Structure, Tertiary , Protein-Tyrosine Kinases/chemistry , Protein-Tyrosine Kinases/deficiency , Stress Fibers/enzymology , rho GTP-Binding Proteins/metabolism
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