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Sample records for vivo oxygen-sensing applications

  1. Vitamin C is dispensable for oxygen sensing in vivo

    PubMed Central

    Nytko, Katarzyna J.; Maeda, Nobuyo; Schläfli, Philipp; Spielmann, Patrick; Wenger, Roland H.

    2011-01-01

    Prolyl-4-hydroxylation is necessary for proper structural assembly of collagens and oxygen-dependent protein stability of hypoxia-inducible transcription factors (HIFs). In vitro function of HIF prolyl-4-hydroxylase domain (PHD) enzymes requires oxygen and 2-oxoglutarate as cosubstrates with iron(II) and vitamin C serving as cofactors. Although vitamin C deficiency is known to cause the collagen-disassembly disease scurvy, it is unclear whether cellular oxygen sensing is similarly affected. Here, we report that vitamin C–deprived Gulo−/− knockout mice show normal HIF-dependent gene expression. The systemic response of Gulo−/− animals to inspiratory hypoxia, as measured by plasma erythropoietin levels, was similar to that of animals supplemented with vitamin C. Hypoxic HIF induction was also essentially normal under serum- and vitamin C–free cell-culture conditions, suggesting that vitamin C is not required for oxygen sensing in vivo. Glutathione was found to fully substitute for vitamin C requirement of all 3 PHD isoforms in vitro. Consistently, glutathione also reduced HIF-1α protein levels, transactivation activity, and endogenous target gene expression in cells exposed to CoCl2. A Cys201Ser mutation in PHD2 increased basal hydroxylation rates and conferred resistance to oxidative damage in vitro, suggesting that this surface-accessible PHD2 cysteine residue is a target of antioxidative protection by vitamin C and glutathione. PMID:21346252

  2. Lifetime-based photoacoustic oxygen sensing in vivo

    NASA Astrophysics Data System (ADS)

    Ray, Aniruddha; Rajian, Justin Rajesh; Lee, Yong-Eun Koo; Wang, Xueding; Kopelman, Raoul

    2012-05-01

    The determination of oxygen levels in blood and other tissues in vivo is critical for ensuring proper body functioning, for monitoring the status of many diseases, such as cancer, and for predicting the efficacy of therapy. Here we demonstrate, for the first time, a lifetime-based photoacoustic technique for the measurement of oxygen in vivo, using an oxygen sensitive dye, enabling real time quantification of blood oxygenation. The results from the main artery in the rat tail indicated that the lifetime of the dye, quantified by the photoacoustic technique, showed a linear relationship with the blood oxygenation levels in the targeted artery.

  3. Oxygen Sensing Mesenchymal Progenitors Promote Neo-Vasculogenesis in a Humanized Mouse Model In Vivo

    PubMed Central

    Hofmann, Nicole A.; Ortner, Anna; Jacamo, Rodrigo O.; Reinisch, Andreas; Schallmoser, Katharina; Rohban, Rokhsareh; Etchart, Nathalie; Fruehwirth, Margareta; Beham-Schmid, Christine; Andreeff, Michael; Strunk, Dirk

    2012-01-01

    Despite insights into the molecular pathways regulating hypoxia-induced gene expression, it is not known which cell types accomplish oxygen sensing during neo-vasculogenesis. We have developed a humanized mouse model of endothelial and mesenchymal progenitor co-transplantation to delineate the cellular compartments responsible for hypoxia response during vasculogenesis. Mesenchymal stem/progenitor cells (MSPCs) accumulated nuclear hypoxia-inducible transcription factor (HIF)-1α earlier and more sensitively than endothelial colony forming progenitor cells (ECFCs) in vitro and in vivo. Hypoxic ECFCs showed reduced function in vitro and underwent apoptosis within 24h in vivo when used without MSPCs. Surprisingly, only in MSPCs did pharmacologic or genetic inhibition of HIF-1α abrogate neo-vasculogenesis. HIF deletion in ECFCs caused no effect. ECFCs could be rescued from hypoxia-induced apoptosis by HIF-competent MSPCs resulting in the formation of patent perfused human vessels. Several angiogenic factors need to act in concert to partially substitute mesenchymal HIF-deficiency. Results demonstrate that ECFCs require HIF-competent vessel wall progenitors to initiate vasculogenesis in vivo and to bypass hypoxia-induced apoptosis. We describe a novel mechanistic role of MSPCs as oxygen sensors promoting vasculogenesis thus underscoring their importance for the development of advanced cellular therapies. PMID:22970226

  4. Early non-destructive biofouling detection and spatial distribution: Application of oxygen sensing optodes.

    PubMed

    Farhat, N M; Staal, M; Siddiqui, A; Borisov, S M; Bucs, Sz S; Vrouwenvelder, J S

    2015-10-15

    Biofouling is a serious problem in reverse osmosis/nanofiltration (RO/NF) applications, reducing membrane performance. Early detection of biofouling plays an essential role in an adequate anti-biofouling strategy. Presently, fouling of membrane filtration systems is mainly determined by measuring changes in pressure drop, which is not exclusively linked to biofouling. Non-destructive imaging of oxygen concentrations (i) is specific for biological activity of biofilms and (ii) may enable earlier detection of biofilm accumulation than pressure drop. The objective of this study was to test whether transparent luminescent planar O2 optodes, in combination with a simple imaging system, can be used for early non-destructive biofouling detection. This biofouling detection is done by mapping the two-dimensional distribution of O2 concentrations and O2 decrease rates inside a membrane fouling simulator (MFS). Results show that at an early stage, biofouling development was detected by the oxygen sensing optodes while no significant increase in pressure drop was yet observed. Additionally, optodes could detect spatial heterogeneities in biofouling distribution at a micro scale. Biofilm development started mainly at the feed spacer crossings. The spatial and quantitative information on biological activity will lead to better understanding of the biofouling processes, contributing to the development of more effective biofouling control strategies.

  5. Optical oxygen sensing systems for drug discovery applications: Respirometric Screening Technology (RST)

    NASA Astrophysics Data System (ADS)

    Papkovsky, Dmitri B.; Hynes, James; Fernandes, Richard

    2005-11-01

    Quenched-fluorescence oxygen sensing allows non-chemical, reversible, real-time monitoring of molecular oxygen and rates of oxygen consumption in biological samples. Using this approach we have developed Respirometric Screening Technology (RST); a platform which facilitates the convenient analysis of cellular oxygen uptake. This in turn allows the investigation of compounds and processes which affect respiratory activity. The RST platform employs soluble phosphorescent oxygen-sensitive probes, which may be assessed in standard microtitter plates on a fluorescence plate reader. New formats of RST assays and time-resolved fluorescence detection instrumentation developed by Luxcel provide improvements in assay sensitivity, miniaturization and overall performance. RST has a diverse range of applications in drug discovery area including high throughput analysis of mitochondrial function; studies of mechanisms of toxicity and apoptosis; cell and animal based screening of compound libraries and environmental samples; and, sterility testing. RST has been successfully validated with a range of practical targets and adopted by several leading pharmaceutical companies.

  6. Oxygen Sensing and Homeostasis

    PubMed Central

    Semenza, Gregg L.

    2015-01-01

    The discovery of carotid bodies as sensory receptors for detecting arterial blood oxygen levels, and the identification and elucidation of the roles of hypoxia-inducible factors (HIFs) in oxygen homeostasis have propelled the field of oxygen biology. This review highlights the gas-messenger signaling mechanisms associated with oxygen sensing, as well as transcriptional and non-transcriptional mechanisms underlying the maintenance of oxygen homeostasis by HIFs and their relevance to physiology and pathology. PMID:26328879

  7. Oxygen sensing and signaling.

    PubMed

    van Dongen, Joost T; Licausi, Francesco

    2015-01-01

    Oxygen is an indispensable substrate for many biochemical reactions in plants, including energy metabolism (respiration). Despite its importance, plants lack an active transport mechanism to distribute oxygen to all cells. Therefore, steep oxygen gradients occur within most plant tissues, which can be exacerbated by environmental perturbations that further reduce oxygen availability. Plants possess various responses to cope with spatial and temporal variations in oxygen availability, many of which involve metabolic adaptations to deal with energy crises induced by low oxygen. Responses are induced gradually when oxygen concentrations decrease and are rapidly reversed upon reoxygenation. A direct effect of the oxygen level can be observed in the stability, and thus activity, of various transcription factors that control the expression of hypoxia-induced genes. Additional signaling pathways are activated by the impact of oxygen deficiency on mitochondrial and chloroplast functioning. Here, we describe the molecular components of the oxygen-sensing pathway.

  8. Hydrogen sulfide and oxygen sensing in the cardiovascular system.

    PubMed

    Olson, Kenneth R; Whitfield, Nathan L

    2010-05-15

    Vertebrate cardiorespiratory homeostasis is inextricably dependent upon specialized cells that provide feedback on oxygen status in the tissues, blood, and on occasion, environment. These "oxygen sensing" cells include chemoreceptors and oxygen-sensitive chromaffin cells that initiate cardiorespiratory reflexes, vascular smooth muscle cells that adjust perfusion to metabolism or ventilation, and other cells that condition themselves in response to episodic hypoxia. Identification of how these cells sense oxygen and transduce this into the appropriate physiological response has enormous clinical applicability, but despite intense research there is no consensus regarding the initial hypoxia-effector coupling mechanism. This review examines an alternative mechanism of oxygen sensing using oxidation of endogenously produced hydrogen sulfide (H(2)S) as the O(2)-sensitive couple. Support for this hypothesis includes the similarity of effects of hypoxia and H(2)S on a variety of tissues, augmentation of hypoxic responses by precursors of H(2)S production and their inhibition by inhibitors of H(2)S synthesis, and the rapid consumption of H(2)S by O(2) in the range of intracellular/mitochondrial Po(2). These studies also indicate that, under normoxic conditions, it is doubtful that free H(2)S has longer than a transient existence in tissue or extracellular fluid.

  9. A rhenium complex doped in a silica molecular sieve for molecular oxygen sensing: Construction and characterization

    NASA Astrophysics Data System (ADS)

    Yang, Xiaozhou; Li, Yanxiao

    2016-01-01

    This paper reported a diamine ligand and its Re(I) complex for potential application in oxygen sensing. The novelty of this diamine ligand localized at its increased conjugation chain which had a typical electron-withdrawing group of 1,3,4-oxadiazole. Electronic distribution of excited electrons and their lifetime were supposed to be increased, favoring oxygen sensing collision. This hypothesis was confirmed by single crystal analysis, theoretical calculation and photophysical measurement. It was found that this Re(I) complex had a long-lived emission peaking at 545 nm, favoring sensing application. By doping this complex into a silica matrix MCM-41, oxygen sensing performance and mechanism of the resulting composites were discussed in detail. Non-linear Stern-Volmer working curves were observed with maximum sensitivity of 5.54 and short response time of 6 s.

  10. Evolution and physiology of neural oxygen sensing

    PubMed Central

    Costa, Kauê M.; Accorsi-Mendonça, Daniela; Moraes, Davi J. A.; Machado, Benedito H.

    2014-01-01

    Major evolutionary trends in animal physiology have been heavily influenced by atmospheric O2 levels. Amongst other important factors, the increase in atmospheric O2 which occurred in the Pre-Cambrian and the development of aerobic respiration beckoned the evolution of animal organ systems that were dedicated to the absorption and transportation of O2, e.g., the respiratory and cardiovascular systems of vertebrates. Global variations of O2 levels in post-Cambrian periods have also been correlated with evolutionary changes in animal physiology, especially cardiorespiratory function. Oxygen transportation systems are, in our view, ultimately controlled by the brain related mechanisms, which senses changes in O2 availability and regulates autonomic and respiratory responses that ensure the survival of the organism in the face of hypoxic challenges. In vertebrates, the major sensorial system for oxygen sensing and responding to hypoxia is the peripheral chemoreflex neuronal pathways, which includes the oxygen chemosensitive glomus cells and several brainstem regions involved in the autonomic regulation of the cardiovascular system and respiratory control. In this review we discuss the concept that regulating O2 homeostasis was one of the primordial roles of the nervous system. We also review the physiology of the peripheral chemoreflex, focusing on the integrative repercussions of chemoreflex activation and the evolutionary importance of this system, which is essential for the survival of complex organisms such as vertebrates. The contribution of hypoxia and peripheral chemoreflex for the development of diseases associated to the cardiovascular and respiratory systems is also discussed in an evolutionary context. PMID:25161625

  11. Role Of Hif2α Oxygen Sensing Pathway In Bronchial Epithelial Club Cell Proliferation

    PubMed Central

    Torres-Capelli, Mar; Marsboom, Glenn; Li, Qilong Oscar Yang; Tello, Daniel; Rodriguez, Florinda Melendez; Alonso, Tamara; Sanchez-Madrid, Francisco; García-Rio, Francisco; Ancochea, Julio; Aragonés, Julián

    2016-01-01

    Oxygen-sensing pathways executed by the hypoxia-inducible factors (HIFs) induce a cellular adaptive program when oxygen supply becomes limited. However, the role of the HIF oxygen-sensing pathway in the airway response to hypoxic stress in adulthood remains poorly understood. Here we found that in vivo exposure to hypoxia led to a profound increase in bronchial epithelial cell proliferation mainly confined to Club (Clara) cells. Interestingly, this response was executed by hypoxia-inducible factor 2α (HIF2α), which controls the expression of FoxM1, a recognized proliferative factor of Club cells. Furthermore, HIF2α induced the expression of the resistin-like molecules α and β (RELMα and β), previously considered bronchial epithelial growth factors. Importantly, despite the central role of HIF2α, this proliferative response was not initiated by in vivo Vhl gene inactivation or pharmacological inhibition of prolyl hydroxylase oxygen sensors, indicating the molecular complexity of this response and the possible participation of other oxygen-sensing pathways. Club cells are principally involved in protection and maintenance of bronchial epithelium. Thus, our findings identify a novel molecular link between HIF2α and Club cell biology that can be regarded as a new HIF2α-dependent mechanism involved in bronchial epithelium adaptation to oxygen fluctuations. PMID:27150457

  12. Quality assessment of packaged foods by optical oxygen sensing

    NASA Astrophysics Data System (ADS)

    Papkovsky, Dmitri B.; O'Mahony, Fiach C.; Kerry, Joe P.; Ogurtsov, Vladimir I.

    2005-11-01

    A phase-fluorometric oxygen sensor system has been developed, which allows non-destructive measurement of residual oxygen levels in sealed containers such as packaged foods. It operates with disposable solid-state sensors incorporated in each pack, and a portable detector which interrogates with the sensors through a (semi)transparent packaging material. The system has been optimized for packaging applications and validated in small and medium scale trials with different types of food, including MAP hams, cheese, convenience foods, smoked fish, bakery. It has demonstrated high efficiency in monitoring package integrity, oxygen profiles in packs, performance of packaging process and many other research and quality control tasks, allowing control of 100% of packs. The low-cost batch-calibrated sensors have demonstrated reliability, safety, stability including direct contact with food, high efficiency in the low oxygen range. Another system, which also employs the fluorescence-based oxygen sensing approach, provides rapid assessment of microbial contamination (total viable counts) in complex samples such as food homogenates, industrial waste, environmental samples, etc. It uses soluble oxygen-sensitive probes, standard microtitter plates and fluorescence measurements on conventional plate reader to monitor growth of aerobic bacteria in small test samples (e.g. food homogenates) via their oxygen respiration. The assay provides high sample through put, miniaturization, speed, and can serve as alternative to the established methods such as agar plate colony counts and turbidimetry.

  13. High performance oxygen sensing nanofibrous membranes of Eu(III) complex/polystyrene prepared by electrospinning

    NASA Astrophysics Data System (ADS)

    Yingkui, Li

    2011-07-01

    In this paper, we report the synthesis, characterization, crystal structure, and photophysical properties of a Eu 3+ complex of Eu(TTA) 3Phen, where TTA = 2-thenoyltrifluoroacetonate, and Phen = 1,10-phenanthroline. Its elementary application for oxygen-sensing application is also investigated by doping it into a polymer matrix of polystyrene (PS). Experimental data suggest that the 3 wt% doped Eu(TTA) 3Phen nanofibrous membrane exhibits a high sensitivity of 3.4 towards oxygen with a good linear relationship of R2 = 0.996. In addition, the 3 wt% doped Eu(TTA) 3Phen nanofibrous membrane owns a quick response of 9 s towards molecular oxygen, along with its excellent atmosphere insensitivity and photobleaching resistance. All these results suggest that both Eu(TTA) 3Phen and Eu(TTA) 3Phen/PS system are promising candidates for oxygen-sensing optical sensors.

  14. L-type calcium channel: Clarifying the "oxygen sensing hypothesis".

    PubMed

    Cserne Szappanos, Henrietta; Viola, Helena; Hool, Livia C

    2017-03-18

    The heart is able to respond acutely to changes in oxygen tension. Since ion channels can respond rapidly to stimuli, the "ion channel oxygen sensing hypothesis" has been proposed to explain acute adaptation of cells to changes in oxygen demand. However the exact mechanism for oxygen sensing continues to be debated. Mitochondria consume the lion's share of oxygen in the heart, fuelling the production of ATP that drives excitation and contraction. Mitochondria also produce reactive oxygen species that are capable of altering the redox state of proteins. The cardiac L-type calcium channel is responsible for maintaining excitation and contraction. Recently, the reactive cysteine on the cardiac L-type calcium channel was identified. These data clarified that the channel does not respond directly to changes in oxygen tension, but rather responds to cellular redox state. This leads to acute alterations in cell signalling responsible for the development of arrhythmias and pathology.

  15. Multifunctional mesoporous nanocomposites with magnetic, optical, and sensing features: synthesis, characterization, and their oxygen-sensing performance.

    PubMed

    Wang, Yanyan; Li, Bin; Zhang, Liming; Song, Hang

    2013-01-29

    In this paper, the fabrication, characterization, and application in oxygen sensing are reported for a novel multifunctional nanomaterial of [Ru(bpy)(2)phen-MMS] (bpy, 2,2'-bipyridyl; phen, phenathrolin) which was simply prepared by covalently grafting the ruthenium(II) polypyridyl compounds into the channels of magnetic mesoporous silica nanocomposites (MMS). Scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, X-ray diffraction, N(2) adsorption-desorption, a superconducting quantum interference device, UV-vis spectroscopy, and photoluminescence spectra were used to characterize the samples. The well-designed multifunctional nanocomposites show superparamagnetic behavior and ordered mesoporous characteristics and exhibit a strong red-orange metal-to-ligand charge transfer emission. In addition, the obtained nanocomposites give high performance in oxygen sensing with high sensitivity (I(0)/I(100) = 5.2), good Stern-Volmer characteristics (R(2) = 0.9995), and short response/recovery times (t↓ = 6 s and t↑ = 12 s). The magnetic, mesoporous, luminescent, and oxygen-sensing properties of this multifunctional nanostructure make it hold great promise as a novel multifunctional oxygen-sensing system for chemical/biosensor.

  16. Ceramide Mediates Acute Oxygen Sensing in Vascular Tissues

    PubMed Central

    Moreno, Laura; Moral-Sanz, Javier; Morales-Cano, Daniel; Barreira, Bianca; Moreno, Enrique; Ferrarini, Alessia; Pandolfi, Rachele; Ruperez, Francisco J.; Cortijo, Julio; Sanchez-Luna, Manuel; Villamor, Eduardo; Perez-Vizcaino, Francisco

    2014-01-01

    Abstract Aims: A variety of vessels, such as resistance pulmonary arteries (PA) and fetoplacental arteries and the ductus arteriosus (DA) are specialized in sensing and responding to changes in oxygen tension. Despite opposite stimuli, normoxic DA contraction and hypoxic fetoplacental and PA vasoconstriction share some mechanistic features. Activation of neutral sphingomyelinase (nSMase) and subsequent ceramide production has been involved in hypoxic pulmonary vasoconstriction (HPV). Herein we aimed to study the possible role of nSMase-derived ceramide as a common factor in the acute oxygen-sensing function of specialized vascular tissues. Results: The nSMase inhibitor GW4869 and an anticeramide antibody reduced the hypoxic vasoconstriction in chicken PA and chorioallantoic arteries (CA) and the normoxic contraction of chicken DA. Incubation with interference RNA targeted to SMPD3 also inhibited HPV. Moreover, ceramide and reactive oxygen species production were increased by hypoxia in PA and by normoxia in DA. Either bacterial sphingomyelinase or ceramide mimicked the contractile responses of hypoxia in PA and CA and those of normoxia in the DA. Furthermore, ceramide inhibited voltage-gated potassium currents present in smooth muscle cells from PA and DA. Finally, the role of nSMase in acute oxygen sensing was also observed in human PA and DA. Innovation: These data provide evidence for the proposal that nSMase-derived ceramide is a critical player in acute oxygen-sensing in specialized vascular tissues. Conclusion: Our results indicate that an increase in ceramide generation is involved in the vasoconstrictor responses induced by two opposite stimuli, such as hypoxia (in PA and CA) and normoxia (in DA). Antioxid. Redox Signal. 20, 1–14. PMID:23725018

  17. Synthesis, characterization, photophysical and oxygen-sensing properties of a novel europium(III) complex

    NASA Astrophysics Data System (ADS)

    Feng, Nan; Xie, Jing; Zhang, Dawei

    2010-09-01

    In this paper, we report the synthesis, characterization, crystal structure, and photophysical properties of a novel Eu 3+ complex of Eu(DBM) 3IPD, where DBM = 1,3-diphenyl-propane-1,3-dione and IPD = 4-(1H-imidazo[4,5-f][1,10]phenanthrolin-2-yl)-N,N-diphenylaniline. Its elementary application for oxygen-sensing application is also investigated by doping it into a silica matrix of MCM-41. Experimental data suggest that the 20 mg/g doped Eu(DBM) 3IPD/MCM-41 system exhibits a high sensitivity of 3.6 towards molecular oxygen with a good linear relationship of R2 = 0.9987. In addition, the 20 mg/g doped Eu(DBM) 3IPD/MCM-41 system owns a quick response of 8 s towards oxygen, along with its excellent atmosphere insensitivity and photobleaching resistance. All these results suggest that both Eu(DBM) 3IPD and Eu(DBM) 3IPD/MCM-41 systems are promising candidates for oxygen-sensing optical sensors.

  18. Two–Photon Oxygen Sensing with Quantum Dot–Porphyrin Conjugates

    PubMed Central

    Lemon, Christopher M.; Karnas, Elizabeth; Bawendi, Moungi G.; Nocera, Daniel G.

    2013-01-01

    Supramolecular assemblies of a quantum dot (QD) associated to palladium(II) porphyrins have been developed to detect oxygen (pO2) in organic solvents. Palladium porphyrins are sensitive in the 0–160 torr range, making them ideal phosphors for in vivo biological oxygen quantification. Porphyrins with meso pyridyl substituents bind to the surface of the QD to produce self–assembled nanosensors. Appreciable overlap between QD emission and porphyrin absorption features results in efficient Förster resonance energy transfer (FRET) for signal transduction in these sensors. The QD serves as a photon antenna, enhancing porphyrin emission under both one– and two–photon excitation, demonstrating that QD–palladium porphyrin conjugates may be used for oxygen sensing over physiological oxygen ranges. PMID:23978247

  19. Interaction of Hydrogen Sulfide with Oxygen Sensing under Hypoxia

    PubMed Central

    Wu, Bo; Teng, Huajian; Zhang, Li; Li, Hong; Li, Jing; Wang, Lina; Li, Hongzhu

    2015-01-01

    Based on the discovery of endogenous H2S production, many in depth studies show this gasotransmitter with a variety of physiological and pathological functions. Three enzymes, cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (MST), are involved in enzymatic production of H2S. Emerging evidence has elucidated an important protective role of H2S in hypoxic conditions in many mammalian systems. However, the mechanisms by which H2S senses and responses to hypoxia are largely elusive. Hypoxia-inducible factors (HIFs) function as key regulators of oxygen sensing, activating target genes expression under hypoxia. Recent studies have shown that exogenous H2S regulates HIF action in different patterns. The activation of carotid bodies is a sensitive and prompt response to hypoxia, rapidly enhancing general O2 supply. H2S has been identified as an excitatory mediator of hypoxic sensing in the carotid bodies. This paper presents a brief review of the roles of these two pathways which contribute to hypoxic sensing of H2S. PMID:26078818

  20. Melatonin and the von Hippel-Lindau/HIF-1 oxygen sensing mechanism: A review.

    PubMed

    Vriend, Jerry; Reiter, Russel J

    2016-04-01

    There are numerous reports that melatonin inhibits the hypoxia-inducible factor, HIF-1α, and the HIF-1α-inducible gene, VEGF, both in vivo and in vitro. Through the inhibition of the HIF-1-VEGF pathway, melatonin reduces hypoxia-induced angiogenesis. Herein we discuss the interaction of melatonin with HIF-1α and HIF-1α-inducible genes in terms of what is currently known concerning the HIF-1α hypoxia response element (HIF-1α-HRE) pathway. The von Hippel-Lindau protein (VHL), also known as the VHL tumor suppressor, functions as part of a ubiquitin ligase complex which recognizes HIF-1α as a substrate. As such, VHL is part of the oxygen sensing mechanism of the cell. Under conditions of hypoxia, HIF-1α stimulates the transcription of numerous HIF-1α-induced genes, including EPO, VEGF, and PFKFB3; the latter is an enzyme which regulates glycolysis. Data from several studies show that ROS generated in mitochondria under conditions of hypoxia stimulate HIF-1α. Since melatonin acts as an antioxidant and reduces ROS, these data suggest that the antioxidant action of melatonin could account for reduced HIF-1, less VEGF, and reduced glycolysis in cancer cells (Warburg effect). A direct or indirect inhibitory action (via the reduction in ROS) of melatonin on proteasome activity would account for much of the published data.

  1. Oxygen Sensing for Industrial Safety — Evolution and New Approaches

    PubMed Central

    Willett, Martin

    2014-01-01

    The requirement for the detection of oxygen in industrial safety applications has historically been met by electrochemical technologies based on the consumption of metal anodes. Products using this approach have been technically and commercially successful for more than three decades. However, a combination of new requirements is driving the development of alternative approaches offering fresh opportunities and challenges. This paper reviews some key aspects in the evolution of consumable anode products and highlights recent developments in alternative technologies aimed at meeting current and anticipated future needs in this important application. PMID:24681673

  2. Spatially monitoring oxygen level in 3D microfabricated cell culture systems using optical oxygen sensing beads.

    PubMed

    Wang, Lin; Acosta, Miguel A; Leach, Jennie B; Carrier, Rebecca L

    2013-04-21

    Capability of measuring and monitoring local oxygen concentration at the single cell level (tens of microns scale) is often desirable but difficult to achieve in cell culture. In this study, biocompatible oxygen sensing beads were prepared and tested for their potential for real-time monitoring and mapping of local oxygen concentration in 3D micro-patterned cell culture systems. Each oxygen sensing bead is composed of a silica core loaded with both an oxygen sensitive Ru(Ph2phen3)Cl2 dye and oxygen insensitive Nile blue reference dye, and a poly-dimethylsiloxane (PDMS) shell rendering biocompatibility. Human intestinal epithelial Caco-2 cells were cultivated on a series of PDMS and type I collagen based substrates patterned with micro-well arrays for 3 or 7 days, and then brought into contact with oxygen sensing beads. Using an image analysis algorithm to convert florescence intensity of beads to partial oxygen pressure in the culture system, tens of microns-size oxygen sensing beads enabled the spatial measurement of local oxygen concentration in the microfabricated system. Results generally indicated lower oxygen level inside wells than on top of wells, and local oxygen level dependence on structural features of cell culture surfaces. Interestingly, chemical composition of cell culture substrates also appeared to affect oxygen level, with type-I collagen based cell culture systems having lower oxygen concentration compared to PDMS based cell culture systems. In general, results suggest that oxygen sensing beads can be utilized to achieve real-time and local monitoring of micro-environment oxygen level in 3D microfabricated cell culture systems.

  3. Biomedical Applications of Sodium MRI In Vivo

    PubMed Central

    Madelin, Guillaume; Regatte, Ravinder R.

    2013-01-01

    In this article, we present an up-to-date overview of the potential biomedical applications of sodium MRI in vivo. Sodium MRI is a subject of increasing interest in translational imaging research as it can give some direct and quantitative biochemical information on the tissue viability, cell integrity and function, and therefore not only help the diagnosis but also the prognosis of diseases and treatment outcomes. It has already been applied in vivo in most of human tissues, such as brain for stroke or tumor detection and therapeutic response, in breast cancer, in articular cartilage, in muscle and in kidney, and it was shown in some studies that it could provide very useful new information not available through standard proton MRI. However, this technique is still very challenging due to the low detectable sodium signal in biological tissue with MRI and hardware/software limitations of the clinical scanners. The article is divided in three parts: (1) the role of sodium in biological tissues, (2) a short review on sodium magnetic resonance, and (3) a review of some studies on sodium MRI on different organs/diseases to date. PMID:23722972

  4. n-Propyl gallate activates hypoxia-inducible factor 1 by modulating intracellular oxygen-sensing systems.

    PubMed

    Kimura, Motohide; Takabuchi, Satoshi; Tanaka, Tomoharu; Murata, Miyahiko; Nishi, Kenichiro; Oda, Seiko; Oda, Tomoyuki; Kanai, Michiyuki; Fukuda, Kazuhiko; Kizaka-Kondoh, Shinae; Adachi, Takehiko; Takabayashi, Arimichi; Semenza, Gregg L; Hirota, Kiichi

    2008-04-01

    HIF-1 (hypoxia-inducible factor 1) is a master regulator of cellular adaptive responses to hypoxia. The expression and transcriptional activity of the HIF-1alpha subunit is stringently controlled by intracellular oxygen tension through the action of prolyl and asparaginyl hydroxylases. In the present study we demonstrate that PG (n-propyl gallate) activates HIF-1 and expression of its downstream target genes under normoxic conditions in cultured cells and in mice. The stability and transcriptional activity of HIF-1alpha are increased by PG. PG treatment inhibits the interaction between HIF-1alpha and VHL (von Hippel-Lindau protein) and promotes the interaction between HIF-1alpha and p300, indicating that PG inhibits the activity of both prolyl and asparaginyl HIF-1alpha hydroxylases. We conclude that PG activates HIF-1 and enhances the resultant gene expression by directly affecting the intracellular oxygen sensing system in vitro and in vivo and that PG represents a lead compound for the development of a non-toxic activator of HIF-1.

  5. Phosphorescent Platinum(II) and Palladium(II) Complexes with Azatetrabenzoporphyrins—New Red Laser Diode-Compatible Indicators for Optical Oxygen Sensing

    PubMed Central

    2010-01-01

    A new class of oxygen indicators is described. Platinum(II) and palladium(II) complexes of azatetrabenzoporphyrins occupy an intermediate position between tetrabenzoporphyrins and phthalocyanines and combine features of both. The new dyes are excitable in the red part of the spectrum and possess strong room-temperature NIR phosphorescence. Other features include excellent spectral compatibility with the red laser diodes and 632.8 nm line of He−Ne laser, excellent photostability, and significantly shorter decay times than for the respective meso-tetraphenyltetrabenzoporphyrins. Applicability of the complexes for optical oxygen sensing is demonstrated. PMID:20186289

  6. Exceptional Oxygen Sensing Properties of New Blue Light-Excitable Highly Luminescent Europium(III) and Gadolinium(III) Complexes

    PubMed Central

    Borisov, Sergey M.; Fischer, Roland; Saf, Robert; Klimant, Ingo

    2016-01-01

    New europium(III) and gadolinium(III) complexes bearing 8-hydroxyphenalenone antenna combine efficient absorption in the blue part of the spectrum and strong emission in polymers at room temperature. The Eu(III) complexes show characteristic red luminescence whereas the Gd(III) dyes are strongly phosphorescent. The luminescence quantum yields are about 20% for the Eu(III) complexes and 50% for the Gd(III) dyes. In contrast to most state-of-the-art Eu(III) complexes the new dyes are quenched very efficiently by molecular oxygen. The luminescence decay times of the Gd(III) complexes exceed 1 ms which ensures exceptional sensitivity even in polymers of moderate oxygen permeability. These sensors are particularly suitable for trace oxygen sensing and may be good substitutes for Pd(II) porphyrins. The photophysical and sensing properties can be tuned by varying the nature of the fourth ligand. The narrow-band emission of the Eu(III) allows efficient elimination of the background light and autofluorescence and is also very attractive for use e.g. in multi-analyte sensors. The highly photostable indicators incorporated in nanoparticles are promising for imaging applications. Due to the straightforward preparation and low cost of starting materials the new dyes represent a promising alternative to the state-of-the-art oxygen indicators particularly for such applications as e.g. food packaging. PMID:27158252

  7. Aspartate 141 Is the Fourth Ligand of the Oxygen-sensing [4Fe-4S]2+ Cluster of Bacillus subtilis Transcriptional Regulator Fnr*

    PubMed Central

    Gruner, Ines; Frädrich, Claudia; Böttger, Lars H.; Trautwein, Alfred X.; Jahn, Dieter; Härtig, Elisabeth

    2011-01-01

    The Bacillus subtilis redox regulator Fnr controls genes of the anaerobic metabolism in response to low oxygen tension. An unusual structure for the oxygen-sensing [4Fe-4S]2+ cluster was detected by a combination of genetic experiments with UV-visible and Mössbauer spectroscopy. Asp-141 was identified as the fourth iron-sulfur cluster ligand besides three Cys residues. Exchange of Asp-141 with Ala abolished functional in vivo complementation of an fnr knock-out strain by the mutagenized fnr gene and in vitro DNA binding of the recombinant regulator FnrD141A. In contrast, substitution of Asp-141 with Cys preserved [4Fe-4S]2+ structure and regulator function. PMID:21068385

  8. Silicon-on-glass pore network micromodels with oxygen-sensing fluorophore films for chemical imaging and defined spatial structure.

    PubMed

    Grate, Jay W; Kelly, Ryan T; Suter, Jonathan; Anheier, Norm C

    2012-11-21

    Pore network microfluidic models were fabricated by a silicon-on-glass technique that provides the precision advantage of dry etched silicon while creating a structure that is transparent across all microfluidic channels and pores, and can be imaged from either side. A silicon layer is bonded to an underlying borosilicate glass substrate and thinned to the desired height of the microfluidic channels and pores. The silicon is then patterned and through-etched by deep reactive ion etching (DRIE), with the underlying glass serving as an etch stop. After bonding on a transparent glass cover plate, one obtains a micromodel in oxygen impermeable materials with water-wet surfaces where the microfluidic channels are transparent and structural elements such as the pillars creating the pore network are opaque. The advantageous features of this approach in a chemical imaging application are demonstrated by incorporating a Pt porphyrin fluorophore in a PDMS film serving as the oxygen-sensing layer and a bonding surface, or in a polystyrene film coated with a PDMS layer for bonding. The sensing of a dissolved oxygen gradient was demonstrated using fluorescence lifetime imaging, and it is shown that different matrix polymers lead to optimal use in different ranges of oxygen concentration. Imaging with the opaque pillars in between the observation direction and the continuous fluorophore film yields images that retain defined spatial structure in the sensor image.

  9. Silicon-on-glass pore network micromodels with oxygen-sensing fluorophore films for chemical imaging and defined spatial structure

    SciTech Connect

    Grate, Jay W.; Kelly, Ryan T.; Suter, Jonathan D.; Anheier, Norman C.

    2012-11-21

    Pore network microfluidic models were fabricated by a silicon-on-glass technique that provides the precision advantage of dry etched silicon while creating a structure that is transparent across all microfluidic channels and pores, and can be imaged from either side. A silicon layer is bonded to an underlying borosilicate glass substrate and thinned to the desired height of the microfluidic channels and pores. The silicon is then patterned and through-etched by deep reactive ion etching (DRIE), with the underlying glass serving as an etch stop. After bonding on a transparent glass cover plate, one obtains a micromodel in oxygen impermeable materials with water wet surfaces where the microfluidic channels are transparent and structural elements such as the pillars creating the pore network are opaque. The micromodel can be imaged from either side. The advantageous features of this approach in a chemical imaging application are demonstrated by incorporating a Pt porphyrin fluorophore in a PDMS film serving as the oxygen sensing layer and a bonding surface, or in a polystyrene film coated with a PDMS layer for bonding. The sensing of a dissolved oxygen gradient was demonstrated using fluorescence lifetime imaging, and it is shown that different matrix polymers lead to optimal use in different ranges dissolved oxygen concentration. Imaging with the opaque pillars in between the observation direction and the continuous fluorophore film yields images that retain spatial information in the sensor image.

  10. Oxygen-sensing mechanisms and the regulation of redox-responsive transcription factors in development and pathophysiology

    PubMed Central

    Haddad, John J

    2002-01-01

    How do organisms sense the amount of oxygen in the environment and respond appropriately when the level of oxygen decreases? Oxygen sensing and the molecular stratagems underlying the process have been the focus of an endless number of investigations trying to find an answer to the question: "What is the identity of the oxygen sensor?" Dynamic changes in pO2 constitute a potential signaling mechanism for the regulation of the expression and activation of reduction-oxidation (redox)-sensitive and oxygen-responsive transcription factors, apoptosis-signaling molecules and inflammatory cytokines. The transition from placental to lung-based respiration causes a relatively hyperoxic shift or oxidative stress, which the perinatal, developing lung experiences during birth. This variation in ΔpO2, in particular, differentially regulates the compartmentalization and functioning of the transcription factors hypoxia-inducible factor-1α (HIF-1α) and nuclear factor-κB (NF-κB). In addition, oxygen-evoked regulation of HIF-1α and NF-κB is closely coupled with the intracellular redox state, such that modulating redox equilibrium affects their responsiveness at the molecular level (expression/transactivation). The differential regulation of HIF-1α and NF-κB in vitro is paralleled by oxygen-sensitive and redox-dependent pathways governing the regulation of these factors during the transition from placental to lung-based respiration ex utero. The birth transition period in vivo and ex utero also regulates apoptosis signaling pathways in a redox-dependent manner, consistent with NF-κB being transcriptionally regulated in order to play an anti-apoptotic function. An association is established between oxidative stress conditions and the augmentation of an inflammatory state in pathophysiology, regulated by the oxygen- and redox-sensitive pleiotropic cytokines. PMID:12537605

  11. Are rare-earth nanoparticles suitable for in vivo applications?

    PubMed

    Liu, Chunyan; Hou, Yi; Gao, Mingyuan

    2014-10-29

    Rare earth (RE) nanoparticles have attracted considerable attention due to their unique optical and magnetic properties associated with f-electrons. The recent accomplishments in RE nanoparticle synthesis have aroused great interest of scientists to further explore their biomedical applications. This Research News summarizes recent achievements in controlled synthesis of magnetic and luminescent RE nanoparticles, surface modification, and toxicity studies of RE nanomaterials, and highlights state-of-the-art in in vivo applications of RE nanoparticles.

  12. Molecular evolution of the metazoan PHD-HIF oxygen-sensing system.

    PubMed

    Rytkönen, Kalle T; Williams, Tom A; Renshaw, Gillian M; Primmer, Craig R; Nikinmaa, Mikko

    2011-06-01

    Metazoans rely on aerobic energy production, which requires an adequate oxygen supply. During reduced oxygen supply (hypoxia), the most profound changes in gene expression are mediated by transcription factors known as hypoxia-inducible factors (HIFs). HIF alpha proteins are commonly posttranslationally regulated by prolyl-4-hydroxylase (PHD) enzymes, which are direct "sensors" of cellular oxygen levels. We examined the molecular evolution of the metazoan PHD-HIF oxygen-sensing system by constructing complete phylogenies for PHD and HIF alpha genes and used computational tools to characterize the molecular changes underlying the functional divergence of PHD and HIF alpha duplicates. The presence of PHDs in metazoan genomes predates the emergence of HIF alphas. Our analysis revealed an unexpected diversity of PHD genes and HIF alpha sequence characteristics in invertebrates, suggesting that the simple oxygen-sensing systems of Caenorhabditis and Drosophila may not be typical of other invertebrate bilaterians. We studied the early vertebrate evolution of the system by sequencing these genes in early-diverging cartilaginous fishes, elasmobranchs. Cartilaginous fishes appear to have three paralogs of both PHD and HIF alpha. The novel sequences were used as outgroups for a detailed molecular analysis of PHD and HIF alpha duplicates in a major air-breathing vertebrate lineage, the mammals, and a major water-breathing vertebrate lineage, the teleosts. In PHDs, functionally divergent amino acid sites were detected near the HIF alpha-binding channel and beta2beta3 loop that defines its substrate specificity. In HIF alphas, more functional divergence was found in teleosts than in mammals, especially in the HIF-1 alpha PAS domain and HIF-2 alpha oxygen-dependent degradation (ODD) domains, which interact with PHDs. Overall, in the vertebrates, elevated substitution rates in the HIF-2 alpha N-terminal ODD domain, together with a functional divergence associated with the known

  13. Application of in vivo laser scanning microscope in dermatology

    NASA Astrophysics Data System (ADS)

    Lademann, Juergen; Richter, H.; Otberg, N.; Lawrenz, F.; Blume-Peytavi, U.; Sterry, W.

    2003-10-01

    The state of the art of in-vivo and in-vitro penetration measurements of topically applied substances is described. Only optical techniques represent online measuring methods based on the absorption or scattering properties of the topically applied substances. Laser scanning microscopy (LSM) has become a promising method for investigations in dermatology and skin physiology, after it was possible to analyze the skin surface on any body side in-vivo. In the present paper the application of a dermatological laser scanning microscope for penetration and distribution measurements of topically applied substances is described. The intercellular and follicular penetration pathways were studied.

  14. 3D bioprinting and its in vivo applications.

    PubMed

    Hong, Nhayoung; Yang, Gi-Hoon; Lee, JaeHwan; Kim, GeunHyung

    2017-01-20

    The purpose of 3D bioprinting technology is to design and create functional 3D tissues or organs in situ for in vivo applications. 3D cell-printing, or additive biomanufacturing, allows the selection of biomaterials and cells (bioink), and the fabrication of cell-laden structures in high resolution. 3D cell-printed structures have also been used for applications such as research models, drug delivery and discovery, and toxicology. Recently, numerous attempts have been made to fabricate tissues and organs by using various 3D printing techniques. However, challenges such as vascularization are yet to be solved. This article reviews the most commonly used 3D cell-printing techniques with their advantages and drawbacks. Furthermore, up-to-date achievements of 3D bioprinting in in vivo applications are introduced, and prospects for the future of 3D cell-printing technology are discussed. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2017.

  15. Cellular Oxygen Sensing: Crystal Structure of Hypoxia-Inducible Factor Prolyl Hydroxylase (PHD2)

    SciTech Connect

    McDonough,M.; Li, V.; Flashman, E.; Chowdhury, R.; Mohr, C.; Lienard, B.; Zondlo, J.; Oldham, N.; Clifton, I.; et al.

    2006-01-01

    Cellular and physiological responses to changes in dioxygen levels in metazoans are mediated via the posttranslational oxidation of hypoxia-inducible transcription factor (HIF). Hydroxylation of conserved prolyl residues in the HIF-{alpha} subunit, catalyzed by HIF prolyl-hydroxylases (PHDs), signals for its proteasomal degradation. The requirement of the PHDs for dioxygen links changes in dioxygen levels with the transcriptional regulation of the gene array that enables the cellular response to chronic hypoxia; the PHDs thus act as an oxygen-sensing component of the HIF system, and their inhibition mimics the hypoxic response. We describe crystal structures of the catalytic domain of human PHD2, an important prolyl-4-hydroxylase in the human hypoxic response in normal cells, in complex with Fe(II) and an inhibitor to 1.7 Angstroms resolution. PHD2 crystallizes as a homotrimer and contains a double-stranded {beta}-helix core fold common to the Fe(II) and 2-oxoglutarate-dependant dioxygenase family, the residues of which are well conserved in the three human PHD enzymes (PHD 1-3). The structure provides insights into the hypoxic response, helps to rationalize a clinically observed mutation leading to familial erythrocytosis, and will aid in the design of PHD selective inhibitors for the treatment of anemia and ischemic disease.

  16. Diversity of Magneto-Aerotactic Behaviors and Oxygen Sensing Mechanisms in Cultured Magnetotactic Bacteria

    PubMed Central

    Lefèvre, Christopher T.; Bennet, Mathieu; Landau, Livnat; Vach, Peter; Pignol, David; Bazylinski, Dennis A.; Frankel, Richard B.; Klumpp, Stefan; Faivre, Damien

    2014-01-01

    Microorganisms living in gradient environments affect large-scale processes, including the cycling of elements such as carbon, nitrogen or sulfur, the rates and fate of primary production, and the generation of climatically active gases. Aerotaxis is a common adaptation in organisms living in the oxygen gradients of stratified environments. Magnetotactic bacteria are such gradient-inhabiting organisms that have a specific type of aerotaxis that allows them to compete at the oxic-anoxic interface. They biomineralize magnetosomes, intracellular membrane-coated magnetic nanoparticles, that comprise a permanent magnetic dipole that causes the cells to align along magnetic field lines. The magnetic alignment enables them to efficiently migrate toward an optimal oxygen concentration in microaerobic niches. This phenomenon is known as magneto-aerotaxis. Magneto-aerotaxis has only been characterized in a limited number of available cultured strains. In this work, we characterize the magneto-aerotactic behavior of 12 magnetotactic bacteria with various morphologies, phylogenies, physiologies, and flagellar apparatus. We report six different magneto-aerotactic behaviors that can be described as a combination of three distinct mechanisms, including the reported (di-)polar, axial, and a previously undescribed mechanism we named unipolar. We implement a model suggesting that the three magneto-aerotactic mechanisms are related to distinct oxygen sensing mechanisms that regulate the direction of cells’ motility in an oxygen gradient. PMID:25028894

  17. A green-emitting Cu complex for oxygen-sensing purpose: Synthesis, characterization and photophysical features

    NASA Astrophysics Data System (ADS)

    Hui, Han; Wei, Li; Zhentao, Liu; Xiangen, Han

    2015-05-01

    In the present work, a green-emitting Cu(I) complex [Cu(BT-Et)(POP)]BF4 was synthesized and fully characterized, where BT-Et = 4-(1-ethyl-1H-benzo[d]imidazol-2-yl)thiazole, POP = bis(2-(diphenylphosphanyl)phenyl) ether, respectively. An ethyl group was connected onto the diamine ligand to breach π-π attraction within solid [Cu(BT-Et)(POP)]BF4, favoring O2 molecule attack and sensitivity improvement. Its molecular identity was confirmed by single crystal analysis and theoretical calculation. [Cu(BT-Et)(POP)]BF4 emitted long-lived green emission peaking at 521 nm upon photoexcitation which was vulnerable towards O2 molecule, making itself a potential oxygen sensing material. [Cu(BT-Et)(POP)]BF4 was then doped into a silica supporting matrix MCM-41. The resulting composite samples showed sensing behavior towards O2 molecule, with short response time of 10 s and sensitivity of 5.56.

  18. Thickness Dependency of Thin Film Samaria Doped Ceria for Oxygen Sensing

    SciTech Connect

    Sanghavi, Rahul P.; Nandasiri, Manjula I.; Kuchibhatla, Satyanarayana V N T; Jiang, Weilin; Varga, Tamas; Nachimuthu, Ponnusamy; Engelhard, Mark H.; Shutthanandan, V.; Thevuthasan, Suntharampillai; Kayani, Asghar N.; Prasad, Shalini

    2011-01-01

    High temperature oxygen sensors are widely used for exhaust gas monitoring in automobiles. This particular study explores the use of thin film single crystalline samaria doped ceria as the oxygen sensing material. Desired signal to noise ratio can be achieved in a material system with high conductivity. From previous studies it is established that 6 atomic percent samarium doping is the optimum concentration for thin film samaria doped ceria to achieve high ionic conductivity. In this study, the conductivity of the 6 atomic percent samaria doped ceria thin film is measured as a function of the sensing film thickness. Hysteresis and dynamic response of this sensing platform is tested for a range of oxygen pressures from 0.001 Torr to 100 Torr for temperatures above 673 K. An attempt has been made to understand the physics behind the thickness dependent conductivity behavior of this sensing platform by developing a hypothetical operating model and through COMSOL simulations. This study can be used to identify the parameters required to construct a fast, reliable and compact high temperature oxygen sensor.

  19. Diversity of magneto-aerotactic behaviors and oxygen sensing mechanisms in cultured magnetotactic bacteria.

    PubMed

    Lefèvre, Christopher T; Bennet, Mathieu; Landau, Livnat; Vach, Peter; Pignol, David; Bazylinski, Dennis A; Frankel, Richard B; Klumpp, Stefan; Faivre, Damien

    2014-07-15

    Microorganisms living in gradient environments affect large-scale processes, including the cycling of elements such as carbon, nitrogen or sulfur, the rates and fate of primary production, and the generation of climatically active gases. Aerotaxis is a common adaptation in organisms living in the oxygen gradients of stratified environments. Magnetotactic bacteria are such gradient-inhabiting organisms that have a specific type of aerotaxis that allows them to compete at the oxic-anoxic interface. They biomineralize magnetosomes, intracellular membrane-coated magnetic nanoparticles, that comprise a permanent magnetic dipole that causes the cells to align along magnetic field lines. The magnetic alignment enables them to efficiently migrate toward an optimal oxygen concentration in microaerobic niches. This phenomenon is known as magneto-aerotaxis. Magneto-aerotaxis has only been characterized in a limited number of available cultured strains. In this work, we characterize the magneto-aerotactic behavior of 12 magnetotactic bacteria with various morphologies, phylogenies, physiologies, and flagellar apparatus. We report six different magneto-aerotactic behaviors that can be described as a combination of three distinct mechanisms, including the reported (di-)polar, axial, and a previously undescribed mechanism we named unipolar. We implement a model suggesting that the three magneto-aerotactic mechanisms are related to distinct oxygen sensing mechanisms that regulate the direction of cells' motility in an oxygen gradient.

  20. Proteomic analysis reveals diverse proline hydroxylation-mediated oxygen-sensing cellular pathways in cancer cells

    PubMed Central

    Liu, Bing; Gao, Yankun; Ruan, Hai-Bin; Chen, Yue

    2016-01-01

    Proline hydroxylation is a critical cellular mechanism regulating oxygen-response pathways in tumor initiation and progression. Yet, its substrate diversity and functions remain largely unknown. Here, we report a system-wide analysis to characterize proline hydroxylation substrates in cancer cells using an immunoaffinity-purification assisted proteomics strategy. We identified 562 sites from 272 proteins in HeLa cells. Bioinformatic analysis revealed that proline hydroxylation substrates are significantly enriched with mRNA processing and stress-response cellular pathways with canonical and diverse flanking sequence motifs. Structural analysis indicates a significant enrichment of proline hydroxylation participating in the secondary structure of substrate proteins. Our study identified and validated Brd4, a key transcription factor, as a novel proline hydroxylation substrate. Functional analysis showed that the inhibition of proline hydroxylation pathway significantly reduced the proline hydroxylation abundance on Brd4 and affected Brd4-mediated transcriptional activity as well as cell proliferation in AML leukemia cells. Taken together, our study identified a broad regulatory role of proline hydroxylation in cellular oxygen-sensing pathways and revealed potentially new targets that dynamically respond to hypoxia microenvironment in tumor cells. PMID:27764789

  1. Rational design of fluorophores for in vivo applications.

    PubMed

    Ptaszek, Marcin

    2013-01-01

    Several classes of small organic molecules exhibit properties that make them suitable for fluorescence in vivo imaging. The most promising candidates are cyanines, squaraines, boron dipyrromethenes, porphyrin derivatives, hydroporphyrins, and phthalocyanines. The recent designing and synthetic efforts have been dedicated to improving their optical properties (shift the absorption and emission maxima toward longer wavelengths and increase the brightness) as well as increasing their stability and water solubility. The most notable advances include development of encapsulated cyanine dyes with increased stability and water solubility, squaraine rotaxanes with increased stability, long-wavelength-absorbing boron dipyrromethenes, long-wavelength-absorbing porphyrin and hydroporphyrin derivatives, and water-soluble phthalocyanines. Recent advances in luminescence and bioluminescence have made self-illuminating fluorophores available for in vivo applications. Development of new types of hydroporphyrin energy-transfer dyads gives the promise for further advances in in vivo multicolor imaging.

  2. Oxygen Sensing Difluoroboron β-Diketonate Polylactide Materials with Tunable Dynamic Ranges for Wound Imaging.

    PubMed

    DeRosa, Christopher A; Seaman, Scott A; Mathew, Alexander S; Gorick, Catherine M; Fan, Ziyi; Demas, James N; Peirce, Shayn M; Fraser, Cassandra L

    2016-11-23

    Difluoroboron β-diketonate poly(lactic acid) materials exhibit both fluorescence (F) and oxygen sensitive room-temperature phosphorescence (RTP). Introduction of halide heavy atoms (Br and I) is an effective strategy to control the oxygen sensitivity in these materials. A series of naphthyl-phenyl (nbm) dye derivatives with hydrogen, bromide and iodide substituents were prepared for comparison. As nanoparticles, the hydrogen derivative was hypersensitive to oxygen (0-0.3%), while the bromide analogue was suited for hypoxia detection (0-3% O2). The iodo derivative, BF2nbm(I)PLA, showed excellent F to RTP peak separation and an 0-100% oxygen sensitivity range unprecedented for metal-free RTP emitting materials. Due to the dual emission and unconventionally long RTP lifetimes of these O2 sensing materials, a portable, cost-effective camera was used to quantify oxygen levels via lifetime and red/green/blue (RGB) ratiometry. The hypersensitive H dye was well matched to lifetime detection, simultaneous lifetime and ratiometric imaging was possible for the bromide analogue, whereas the iodide material, with intense RTP emission and a shorter lifetime, was suited for RGB ratiometry. To demonstrate the prospects of this camera/material design combination for bioimaging, iodide boron dye-PLA nanoparticles were applied to a murine wound model to detect oxygen levels. Surprisingly, wound oxygen imaging was achieved without covering (i.e. without isolating from ambient conditions, air). Additionally, would healing was monitored via wound size reduction and associated oxygen recovery, from hypoxic to normoxic. These single-component materials provide a simple tunable platform for biological oxygen sensing that can be deployed to spatially resolve oxygen in a variety of environments.

  3. Fluorescence-lifetime-based sensors: oxygen sensing and other biomedical applications

    NASA Astrophysics Data System (ADS)

    Randers-Eichhorn, Lisa; Bartlett, Roscoe A.; Sipior, Jeffrey; Frey, Douglas D.; Carter, Gary M.; Lakowicz, Joseph R.; Rao, Govind

    1996-05-01

    Murine hybridomas were cultivated in tissue culture flasks. Dissolved oxygen tensions in the gas and liquid phases during cell growth were measured non-invasively by an optical oxygen sensor. Readings were made with caps both cracked open and completely closed. During cell growth, gas phase oxygen concentrations remained near atmospheric levels, while the oxygen tension at the bottom of the flasks eventually reached zero. These results suggest that the widespread practice of cracking open tissue culture flask caps during cell growth with a view to supplying adequate oxygen to cells is ineffective and unnecessary. The mass transfer characteristics of the tissue culture flask indicate the dominant resistance to oxygen mass transfer to the cells was the liquid media. The mass transfer rates through the liquid layer under standard laboratory conditions were found to be greater than those predicted by diffusion alone, suggesting microscale mixing. Volumetric and specific oxygen consumption rates were calculated from the sensor data, and were comparable to published values. A recently developed single fiber optic oxygen sensor is described. This new sensor will provide oxygen concentrations at various levels in the tissue culture flasks, allowing more accurate modeling of oxygen diffusion.

  4. Amphiphilic Fluorinated Polymer Nanoparticle Film Formation and Dissolved Oxygen Sensing Application

    NASA Astrophysics Data System (ADS)

    Gao, Yu; Zhu, Huie; Yamamoto, Shunsuke; Miyashita, Tokuji; Mitsuishi, Masaya

    2016-04-01

    Fluorinated polymer nanoparticle films were prepared by dissolving amphiphilic fluorinated polymer, poly (N-1H, 1H-pentadecafluorooctylmethacrylamide) (pC7F15MAA) in two miscible solvents (AK-225 and acetic acid). A superhydrophobic and porous film was obtained by dropcasting the solution on substrates. With higher ratios of AK-225 to acetic acid, pC7F15MAA was densified around acetic acid droplets, leading to the formation of pC7F15MAA nanoparticles. The condition of the nanoparticle film preparation was investigated by varying the mixing ratio or total concentration. A highly sensitive dissolved oxygen sensor system was successfully prepared utilizing a smart surface of superhydrophobic and porous pC7F15MAA nanoparticle film. The sensitivity showed I0/I40 = 126 in the range of dissolved oxygen concentration of 0 ~ 40 mg L-1. The oxygen sensitivity was compared with that of previous reports.

  5. Clinical applications of in vivo fluorescence confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Oh, Chilhwan; Park, Sangyong; Kim, Junhyung; Ha, Seunghan; Park, Gyuman; Lee, Gunwoo; Lee, Onseok; Chun, Byungseon; Gweon, Daegab

    2008-02-01

    Living skin for basic and clinical research can be evaluated by Confocal Laser Scanning Microscope (CLSM) non-invasively. CLSM imaging system can achieve skin image its native state either "in vivo" or "fresh biopsy (ex vivo)" without fixation, sectioning and staining that is necessary for routine histology. This study examines the potential fluorescent CLSM with a various exogenous fluorescent contrast agent, to provide with more resolution images in skin. In addition, in vivo fluorescent CLSM researchers will be extended a range of potential clinical application. The prototype of our CLSM system has been developed by Prof. Gweon's group. The operating parameters are composed of some units, such as illuminated wavelength 488 nm, argon illumination power up to 20mW on the skin, objective lens, 0.9NA oil immersion, axial resolution 1.0μm, field of view 200μm x 100μm (lateral resolution , 0.3μm). In human volunteer, fluorescein sodium was administrated topically and intradermally. Animal studies were done in GFP transgenic mouse, IRC mouse and pig skin. For imaging of animal skin, fluorescein sodium, acridine orange, and curcumine were used for fluorescein contrast agent. We also used the GFP transgenic mouse for fluorescein CLSM imaging. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. Curcumin is a yellow food dye that has similar fluorescent properties to fluorescein sodium. Acridin Orange can be highlight nuclei in viable keratinocyte. In vivo CLSM of transgenic GFP mouse enable on in vivo, high resolution view of GFP expressing skin tissue. GFP signals are brightest in corneocyte, kertinocyte, hair and eccrine gland. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. In

  6. Applications of nuclear technologies for in-vivo elemental analysis

    SciTech Connect

    Cohn, S.H.; Ellis, K.J.; Vartsky, D.; Wielopolski, L.

    1982-01-01

    Measurement facilities developed, to date, include a unique whole-body-counter, (WBC); a total-body neutron-activation facility (TBNAA); and a partial-body activation facility (PBNAA). A variation of the prompt-gamma neutron-activation technique for measuring total-body nitrogen was developed to study body composition of cancer patients and the effect of nutritional regimens on the composition. These new techniques provide data in numerous clinical studies not previously amenable to investigation. The development and perfection of these techniques provide unique applications of radiation and radioisotopes to the early diagnosis of certain diseases and the evaluation of therapeutic programs. The PBNAA technique has been developed and calibrated for in-vivo measurement of metals. Development has gone forward on prompt-gamma neutron activation for the measurement of cadmium, x-ray fluorescence (XRF) for measurement of iron. Other techniques are being investigated for in-vivo measurement of metals such as silicon and beryllium.

  7. Therapeutic targeting of oxygen-sensing prolyl hydroxylases abrogates ATF4-dependent neuronal death and improves outcomes after brain hemorrhage in several rodent models

    PubMed Central

    Karuppagounder, Saravanan S.; Alim, Ishraq; Khim, Soah J.; Bourassa, Megan W.; Sleiman, Sama F.; John, Roseleen; Thinnes, Cyrille C.; Yeh, Tzu-Lan; Demetriades, Marina; Neitemeier, Sandra; Cruz, Dana; Gazaryan, Irina; Killilea, David W.; Morgenstern, Lewis; Xi, Guohua; Keep, Richard F.; Schallert, Timothy; Tappero, Ryan V.; Zhong, Jian; Cho, Sunghee; Maxfield, Frederick R.; Holman, Theodore R.; Culmsee, Carsten; Fong, Guo-Hua; Su, Yijing; Ming, Guo-li; Song, Hongjun; Cave, John W.; Schofield, Christopher J.; Colbourne, Frederick; Coppola, Giovanni; Ratan, Rajiv R.

    2017-01-01

    Disability or death due to intracerebral hemorrhage (ICH) is attributed to blood lysis, liberation of iron, and consequent oxidative stress. Iron chelators bind to free iron and prevent neuronal death induced by oxidative stress and disability due to ICH, but the mechanisms for this effect remain unclear. We show that the hypoxia-inducible factor prolyl hydroxylase domain (HIF-PHD) family of iron-dependent, oxygen-sensing enzymes are effectors of iron chelation. Molecular reduction of the three HIF-PHD enzyme isoforms in the mouse striatum improved functional recovery after ICH. A low-molecular-weight hydroxyquinoline inhibitor of the HIF-PHD enzymes, adaptaquin, reduced neuronal death and behavioral deficits after ICH in several rodent models without affecting total iron or zinc distribution in the brain. Unexpectedly, protection from oxidative death in vitro or from ICH in vivo by adaptaquin was associated with suppression of activity of the prodeath factor ATF4 rather than activation of an HIF-dependent prosurvival pathway. Together, these findings demonstrate that brain-specific inactivation of the HIF-PHD metalloenzymes with the blood-brain barrier-permeable inhibitor adaptaquin can improve functional outcomes after ICH in several rodent models. PMID:26936506

  8. Therapeutic targeting of oxygen-sensing prolyl hydroxylases abrogates ATF4-dependent neuronal death and improves outcomes after brain hemorrhage in several rodent models

    DOE PAGES

    Karuppagounder, Saravanan S.; Alim, Ishraq; Khim, Soah J.; ...

    2016-03-02

    Disability or death due to intracerebral hemorrhage (ICH) is attributed to blood lysis, liberation of iron and consequent oxidative stress. Iron chelators bind to free iron and prevent neuronal death induced by oxidative stress and disability due to ICH, but the mechanisms for this effect remain unclear. Here we show that the hypoxia-inducible factor prolyl-hydroxylase (HIF- PHD) family of iron-dependent oxygen sensing enzymes are effectors of iron chelation. Molecular reduction of the three HIF-PHD enzyme isoforms in mouse striatum improved functional recovery following ICH. A low molecular weight hydroxyquinoline inhibitor of the HIF-PHDs, adaptaquin, reduced neuronal death and behavioral deficitsmore » following ICH in several rodent models without affecting total iron or zinc distribution in the brain. Unexpectedly, protection from oxidative death in vitro or from ICH in vivo by adaptaquin was associated with suppression of expression of the prodeath factor ATF4 rather than activation of a HIF-dependent prosurvival pathway. In conclusion, together these findings demonstrate that brain-specific inactivation of the HIF-PHD metalloenzymes with the blood-brain barrier permeable inhibitor adaptaquin can improve functional outcomes following ICH in multiple rodent species.« less

  9. Therapeutic targeting of oxygen-sensing prolyl hydroxylases abrogates ATF4-dependent neuronal death and improves outcomes after brain hemorrhage in several rodent models

    SciTech Connect

    Karuppagounder, Saravanan S.; Alim, Ishraq; Khim, Soah J.; Bourassa, Megan W.; Sleiman, Sama F.; John, Roseleen; Thinnes, Cyrille C.; Yeh, Tzu-Lan; Demetriades, Marina; Neitemeier, Sandra; Cruz, Dana; Gazaryan, Irina; Killilea, David W.; Morgenstern, Lewis; Xi, Guohu; Keep, Richard F.; Schallert, Timothy; Tappero, Ryan V.; Zhong, Jian; Cho, Sunghee; Maxfield, Frederick R.; Holman, T. R.; Culmsee, Carsten; Fong, Guo-Hua -H.; Su, Yijing; Ming, Guo-li; Song, Hongjun; Cave, John W.; Schofield, Christopher J.; Colbourne, Frederick; Coppola, Giovanni; Ratan, Rajiv R.

    2016-03-02

    Disability or death due to intracerebral hemorrhage (ICH) is attributed to blood lysis, liberation of iron and consequent oxidative stress. Iron chelators bind to free iron and prevent neuronal death induced by oxidative stress and disability due to ICH, but the mechanisms for this effect remain unclear. Here we show that the hypoxia-inducible factor prolyl-hydroxylase (HIF- PHD) family of iron-dependent oxygen sensing enzymes are effectors of iron chelation. Molecular reduction of the three HIF-PHD enzyme isoforms in mouse striatum improved functional recovery following ICH. A low molecular weight hydroxyquinoline inhibitor of the HIF-PHDs, adaptaquin, reduced neuronal death and behavioral deficits following ICH in several rodent models without affecting total iron or zinc distribution in the brain. Unexpectedly, protection from oxidative death in vitro or from ICH in vivo by adaptaquin was associated with suppression of expression of the prodeath factor ATF4 rather than activation of a HIF-dependent prosurvival pathway. In conclusion, together these findings demonstrate that brain-specific inactivation of the HIF-PHD metalloenzymes with the blood-brain barrier permeable inhibitor adaptaquin can improve functional outcomes following ICH in multiple rodent species.

  10. Therapeutic targeting of oxygen-sensing prolyl hydroxylases abrogates ATF4-dependent neuronal death and improves outcomes after brain hemorrhage in several rodent models.

    PubMed

    Karuppagounder, Saravanan S; Alim, Ishraq; Khim, Soah J; Bourassa, Megan W; Sleiman, Sama F; John, Roseleen; Thinnes, Cyrille C; Yeh, Tzu-Lan; Demetriades, Marina; Neitemeier, Sandra; Cruz, Dana; Gazaryan, Irina; Killilea, David W; Morgenstern, Lewis; Xi, Guohua; Keep, Richard F; Schallert, Timothy; Tappero, Ryan V; Zhong, Jian; Cho, Sunghee; Maxfield, Frederick R; Holman, Theodore R; Culmsee, Carsten; Fong, Guo-Hua; Su, Yijing; Ming, Guo-li; Song, Hongjun; Cave, John W; Schofield, Christopher J; Colbourne, Frederick; Coppola, Giovanni; Ratan, Rajiv R

    2016-03-02

    Disability or death due to intracerebral hemorrhage (ICH) is attributed to blood lysis, liberation of iron, and consequent oxidative stress. Iron chelators bind to free iron and prevent neuronal death induced by oxidative stress and disability due to ICH, but the mechanisms for this effect remain unclear. We show that the hypoxia-inducible factor prolyl hydroxylase domain (HIF-PHD) family of iron-dependent, oxygen-sensing enzymes are effectors of iron chelation. Molecular reduction of the three HIF-PHD enzyme isoforms in the mouse striatum improved functional recovery after ICH. A low-molecular-weight hydroxyquinoline inhibitor of the HIF-PHD enzymes, adaptaquin, reduced neuronal death and behavioral deficits after ICH in several rodent models without affecting total iron or zinc distribution in the brain. Unexpectedly, protection from oxidative death in vitro or from ICH in vivo by adaptaquin was associated with suppression of activity of the prodeath factor ATF4 rather than activation of an HIF-dependent prosurvival pathway. Together, these findings demonstrate that brain-specific inactivation of the HIF-PHD metalloenzymes with the blood-brain barrier-permeable inhibitor adaptaquin can improve functional outcomes after ICH in several rodent models.

  11. In vivo Coherent Raman Imaging for Neuroscience Applications

    NASA Astrophysics Data System (ADS)

    Cote, Daniel

    2010-08-01

    The use of coherent Raman imaging is described for applications in neuroscience. Myelin imaging of the spinal cord can be performed with Raman imaging through the use of the vibration in carbon-hydrogen bonds, dominant in lipids. First, we demonstrate in vivo histomorphometry in live animal for characterization of myelin-related nervous system pathologies. This is used to characterize spinal cord health during multiple sclerosis. Second, Raman spectroscopy of tissue is discussed. We discuss the challenges that live animal imaging brings, together with important aspects of coherent Raman imaging in tissue.

  12. In vitro - in vivo correlation: from theory to applications.

    PubMed

    Emami, Jaber

    2006-01-01

    A key goal in pharmaceutical development of dosage forms is a good understanding of the in vitro and in vivo performance of the dosage forms. One of the challenges of biopharmaceutics research is correlating in vitro drug release information of various drug formulations to the in vivo drug profiles (IVIVC). Thus the need for a tool to reliably correlate in vitro and in vivo drug release data has exceedingly increased. Such a tool shortens the drug development period, economizes the resources and leads to improved product quality. Increased activity in developing IVIVCs indicates the value of IVIVCs to the pharmaceutical industry. IVIVC can be used in the development of new pharmaceuticals to reduce the number of human studies during the formulation development as the main objective of an IVIVC is to serve as a surrogate for in vivo bioavailability and to support biowaivers. It supports and/or validates the use of dissolution methods and specification settings. This is because the IVIVC includes in vivo relevance to in vitro dissolution specifications. It can also assist in quality control for certain scale-up and post-approval changes (SUPAC). With the proliferation of modified-release products, it becomes necessary to examine the concept of IVIVC in greater depth. Investigations of IVIVC are increasingly becoming an integral part of extended release drug development. There must be some in vitro means of assuring that each batch of the same product will perform identically in vivo. This review article represents the FDA guidance, development, evaluation, and validation of an IVIVC to grant biowaivers, and to set dissolution specifications for oral dosage forms, biopharmaceutics classification systems (BCS), BCS biowaivers, application of BCS in IVIVC development and concept of mapping. The importance of dissolution media and methodology and pharmacokinetic studies in the context of IVIVC has been highlighted. The review also covers the literature examples of IVIVCs

  13. Biological oxygen sensing via two-photon absorption by an Ir(III) complex using a femtosecond fiber laser

    NASA Astrophysics Data System (ADS)

    Moritomo, Hiroki; Fujii, Akinari; Suzuki, Yasutaka; Yoshihara, Toshitada; Tobita, Seiji; Kawamata, Jun

    2016-09-01

    Near-infrared two-photon absorption of the phosphorescent Ir(III) complex (2,4-pentanedionato-κO 2,κO 4)bis[2-(6-phenanthridinyl-κN)benzo[b]thien-3-yl-κC]iridium (BTPHSA) was characterized. It exhibited a 800-1200 nm two-photon absorption band, and thus could be electronically excited by 1030-nm femtosecond Ti:sapphire and Yb-doped fiber lasers. By using BTPHSA, oxygen concentrations in human embryonic kidney 293 (HEK293) cells were imaged. These results demonstrate two-photon oxygen sensing of live tissues via easily operable excitation sources.

  14. Microfabricated, amperometric, enzyme-based biosensors for in vivo applications.

    PubMed

    Weltin, Andreas; Kieninger, Jochen; Urban, Gerald A

    2016-07-01

    Miniaturized electrochemical in vivo biosensors allow the measurement of fast extracellular dynamics of neurotransmitter and energy metabolism directly in the tissue. Enzyme-based amperometric biosensing is characterized by high specificity and precision as well as high spatial and temporal resolution. Aside from glucose monitoring, many systems have been introduced mainly for application in the central nervous system in animal models. We compare the microsensor principle with other methods applied in biomedical research to show advantages and drawbacks. Electrochemical sensor systems are easily miniaturized and fabricated by microtechnology processes. We review different microfabrication approaches for in vivo sensor platforms, ranging from simple modified wires and fibres to fully microfabricated systems on silicon, ceramic or polymer substrates. The various immobilization methods for the enzyme such as chemical cross-linking and entrapment in polymer membranes are discussed. The resulting sensor performance is compared in detail. We also examine different concepts to reject interfering substances by additional membranes, aspects of instrumentation and biocompatibility. Practical considerations are elaborated, and conclusions for future developments are presented. Graphical Abstract ᅟ.

  15. Ex vivo laser lipolysis assisted with radially diffusing optical applicator

    NASA Astrophysics Data System (ADS)

    Hwang, Jieun; Hau, Nguyen Trung; Park, Sung Yeon; Rhee, Yun-Hee; Ahn, Jin-Chul; Kang, Hyun Wook

    2016-05-01

    Laser-assisted lipolysis has been implemented to reduce body fat in light of thermal interactions with adipose tissue. However, using a flat fiber with high irradiance often needs rapid cannula movements and even undesirable thermal injury due to direct tissue contact. The aim of the current study was to explore the feasibility of a radially diffusing optical applicator to liquefy the adipose tissue for effective laser lipolysis. The proposed diffuser was evaluated with a flat fiber in terms of temperature elevation and tissue liquefaction after laser lipolysis with a 980-nm wavelength. Given the same power (20 W), the diffusing applicator generated a 30% slower temperature increase with a 25% lower maximum temperature (84±3.2°C in 1 min p<0.001) in the tissue, compared with the flat fiber. Under the equivalent temperature development, the diffuser induced up to fivefold larger area of the adipose liquefaction due to radial light emission than the flat fiber. Ex vivo tissue tests for 5-min irradiation demonstrated that the diffuser (1.24±0.15 g) liquefied 66% more adipose tissue than the flat fiber (0.75±0.05 g). The proposed diffusing applicator can be a feasible therapeutic device for laser lipolysis due to low temperature development and wide coverage of thermal treatment.

  16. Mutual antagonism between hypoxia-inducible factors 1α and 2α regulates oxygen sensing and cardio-respiratory homeostasis.

    PubMed

    Yuan, Guoxiang; Peng, Ying-Jie; Reddy, Vaddi Damodara; Makarenko, Vladislav V; Nanduri, Jayasri; Khan, Shakil A; Garcia, Joseph A; Kumar, Ganesh K; Semenza, Gregg L; Prabhakar, Nanduri R

    2013-05-07

    Breathing and blood pressure are under constant homeostatic regulation to maintain optimal oxygen delivery to the tissues. Chemosensory reflexes initiated by the carotid body and catecholamine secretion from the adrenal medulla are the principal mechanisms for maintaining respiratory and cardiovascular homeostasis; however, the underlying molecular mechanisms are not known. Here, we report that balanced activity of hypoxia-inducible factor-1 (HIF-1) and HIF-2 is critical for oxygen sensing by the carotid body and adrenal medulla, and for their control of cardio-respiratory function. In Hif2α(+/-) mice, partial HIF-2α deficiency increased levels of HIF-1α and NADPH oxidase 2, leading to an oxidized intracellular redox state, exaggerated hypoxic sensitivity, and cardio-respiratory abnormalities, which were reversed by treatment with a HIF-1α inhibitor or a superoxide anion scavenger. Conversely, in Hif1α(+/-) mice, partial HIF-1α deficiency increased levels of HIF-2α and superoxide dismutase 2, leading to a reduced intracellular redox state, blunted oxygen sensing, and impaired carotid body and ventilatory responses to chronic hypoxia, which were corrected by treatment with a HIF-2α inhibitor. None of the abnormalities observed in Hif1α(+/-) mice or Hif2α(+/-) mice were observed in Hif1α(+/-);Hif2α(+/-) mice. These observations demonstrate that redox balance, which is determined by mutual antagonism between HIF-α isoforms, establishes the set point for hypoxic sensing by the carotid body and adrenal medulla, and is required for maintenance of cardio-respiratory homeostasis.

  17. FIBER OPTICAL MICRO-DETECTORS FOR OXYGEN SENSING IN POWER PLANTS

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III

    2004-10-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. High temperature measurements of the emission of clusters in sol gel films show that the luminescence intensity from the films follow a 1/T relationship from room temperature to 150 C, and then declines at a slower rate at higher temperatures. The large number of photons available at 230 C is consistent with simple low cost optics for fiber optic probes based on the emission from clusters in sol gel films.

  18. An optical biopsy system with miniaturized Raman and spectral imaging probes; in vivo animal and ex vivo clinical application studies

    NASA Astrophysics Data System (ADS)

    Sato, Hidetoshi; Suzuki, Toshiaki; Andriana, Bibin B.; Morita, Shin'ichi; Maruyama, Atsushi; Shinzawa, Hideyuki; Komachi, Yuichi; Kanai, Gen'ichi; Ura, Nobuo; Masutani, Koji; Matsuura, Yuji; Toi, Masakazu; Shimosegawa, Toru; Ozaki, Yukihiro

    2009-02-01

    An optical biopsy system which equips miniaturized Raman probes, a miniaturized endoscope and a fluorescent image probe has been developed for in vivo studies of live experimental animals. The present report describes basic optical properties of the system and its application studies for in vivo cancer model animals and ex vivo human cancer tissues. It was developed two types of miniaturized Raman probes, micro Raman probe (MRP) made of optical fibers and ball lens hollow optical fiber Raman probe (BHRP) made of single hollow optical fiber (HOF) with a ball lens. The former has rather large working distance (WD), up to one millimeter. The latter has small WD (~300μm) which depends on the focal length of the ball lens. Use of multiple probes with different WD allows one to obtain detailed information of subsurface tissues in the totally noninvasive manner. The probe is enough narrow to be inserted into a biopsy needle (~19G), for observations of the lesion at deeper inside bodies. The miniaturized endoscope has been applied to observe progression of a stomach cancer in the same rat lesion. It was succeeded to visualize structure of non-stained cancer tissue in live model animals by the fluorescent image technique. The system was also applied to ex vivo studies of human breast and stomach cancers.

  19. FIBER OPTICAL MICRO-DETECTORS FOR OXYGEN SENSING IN POWER PLANTS

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III

    2004-07-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Alkali salts of Mo{sub 6}Cl{sub 12} were synthesized and heated to 280 C for one hour in air. Optical measurements of the thermally treated material confirm the potential of the salts as lumophores in high temperature fiber optic sensors. In addition sol-gel films containing Mo{sub 6}Cl{sub 12} were dip coated on quartz substrates and heated at 200 C for one hour. Conditions were developed for successfully immobilizing monomeric complexes that are compatible with sol-gel processing.

  20. Fiber Optical Micro-detectors for Oxygen Sensing in Power Plants

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D. J. Osborn; Po Zhang

    2006-09-30

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Our approach towards immobilizing the potassium salt of the molybdenum cluster, K{sub 2}Mo{sub 6}Cl{sub 14}, at the far end of an optical fiber is to embed the cluster in a thermally cured sol-gel matrix particle. Due to the improved mechanical properties of this approach high temperature sensor measurements were performed up to 100 C. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  1. Fiber Optical Micro-detectors for Oxygen Sensing in Power Plants

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn; Po Zhang

    2006-06-30

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Our approach towards immobilizing the potassium salt of the molybdenum cluster, K{sub 2}Mo{sub 6}Cl{sub 14}, at the far end of an optical fiber is to embed the cluster in a thermally cured sol-gel matrix particle. This particle-in-binder approach affords fibers with greatly improved mechanical properties, as compared to previous approaches. The sensor was characterized in 2-21% gas phase oxygen at 40, 70 and 100 C. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  2. Fiber Optical Micro-detectors for Oxygen Sensing in Power Plants

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D. J. Osborn; Po Zhang

    2006-09-30

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications has been developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. We report on a fiber optic technique for detection of gas phase oxygen up to 100 C based on the {sup 3}O{sub 2} quenching of the luminescence from molybdenum chloride clusters, K{sub 2}Mo{sub 6}Cl{sub 14}. The inorganic sensing film is a composite of sol-gel particles embedded in a thin, oxygen permeable sol-gel binder. The particles are comprised of thermally stable, luminescent K{sub 2}Mo{sub 6}Cl{sub 14} clusters dispersed in a fully equilibrated sol-gel matrix. From 40 to 100 C, the fiber sensor switches {approx}6x in intensity in response to alternating pulses of <0.001% O2 and 21% O{sub 2} between two well defined levels with a response time of 10 s. The sensor signal is a few nW for an input pump power of 250 {micro}W. The normalized sensor signal is linear with molar oxygen concentration and fits the theoretical Stern-Volmer relationship. Although the sensitivity decreases with temperature, sensitivity at 100 C is 160 [O{sub 2}]{sup -1}. These parameters are well suited for in-situ, real-time monitoring of oxygen for industrial process control applications.

  3. Linear oxygen-sensing response from a rhenium complex induced by heavy atom: synthesis, characterization, photophysical study and sensing performance.

    PubMed

    Wan, Pu; Zhao, Lun; Wang, Lisha; Xu, Guangyang

    2013-08-01

    In this paper, we synthesized a Br-containing ligand of 2-(4-bromophenyl)-5-(pyridin-2-yl)-1,3,4-oxadiazole and its corresponding Re(I) complex. Their synthesis, characterization, single crystal structure, electronic transitions and photophysical property were presented and discussed in detail. This Re(I) complex was found to be a yellow emitter with slim π→π* radiative decay contribution, and its emission was also found to be sensitive towards O2. By doping this Re(I) complex into a polymer matrix, the oxygen-sensing performance of the resulted composite nanofibers was also investigated. Owing to the porous structure of the supporting matrix, the optimal sample gave the highest sensitivity of 3.91 with short response time of only 9 s. In addition, the linearity of the Stern-Volmer plots was greatly improved due to the highly pure emissive center triggered by heavy-atom turbulence effect from Br atom, as indicted by theoretical calculation result.

  4. Linear oxygen-sensing response from a rhenium complex induced by heavy atom: Synthesis, characterization, photophysical study and sensing performance

    NASA Astrophysics Data System (ADS)

    Pu, Wan; Lun, Zhao; Lisha, Wang; Guangyang, Xu

    2013-08-01

    In this paper, we synthesized a Br-containing ligand of 2-(4-bromophenyl)-5-(pyridin-2-yl)-1,3,4-oxadiazole and its corresponding Re(I) complex. Their synthesis, characterization, single crystal structure, electronic transitions and photophysical property were presented and discussed in detail. This Re(I) complex was found to be a yellow emitter with slim π → π* radiative decay contribution, and its emission was also found to be sensitive towards O2. By doping this Re(I) complex into a polymer matrix, the oxygen-sensing performance of the resulted composite nanofibers was also investigated. Owing to the porous structure of the supporting matrix, the optimal sample gave the highest sensitivity of 3.91 with short response time of only 9 s. In addition, the linearity of the Stern-Volmer plots was greatly improved due to the highly pure emissive center triggered by heavy-atom turbulence effect from Br atom, as indicted by theoretical calculation result.

  5. FIBER OPTICAL MICRO-DETECTORS FOR OXYGEN SENSING IN POWER PLANTS

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III; Po Zhang

    2005-07-01

    A reflection mode fiber optic oxygen sensor is being developed that can operate at high temperatures for power plant applications. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Two critical materials issues are the cluster's ability to withstand high temperatures when immobilized in a porous the sol-gel support, and whether after heating to high temperatures, the sol-gel matrix maintains a high and constant permeability to oxygen to support rapid quenching of luminescence. We used a composite materials approach to prepare stable sensing layers on optical fibers. We dispersed 60 w/w% of a pre-cured sol-gel composite containing the potassium salt of molybdenum clusters (K{sub 2}Mo{sub 6}Cl{sub 14}) into a sol-gel binder solution, and established the conditions necessary for deposition of sol-gel films on optical fibers and planar substrates. The fiber sensor has an output signal of 5 nW when pumped with an inexpensive commercial 365 nm ultraviolet light emitting diode (LED). Quenching of the sensor signal by oxygen was observed up to a gas temperature of 175 C with no degradation of the oxygen permeability of the composite after high temperature cycling. On planar substrates the cluster containing composite responds within <1 second to a gas exchange from nitrogen to oxygen, indicating the feasibility of real-time oxygen detection.

  6. Fiber Optical Micro-detectors for Oxygen Sensing in Power Plants

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III; Po Zhang

    2006-05-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Previously we described a particle-in-binder approach to immobilizing the potassium salt of the molybdenum cluster, K{sub 2}Mo{sub 6}Cl{sub 14}, at the tips of optical fibers. Compared to previous methods, the particle-in-binder approach affords fibers with greatly improved mechanical properties. The response of the sensor to oxygen at 40, 70 and 100 C was measured in 2-21% gas phase oxygen. The normalized sensor signal is linear with molar oxygen concentration and fits the theoretical Stern-Volmer relationship. Although the sensitivity decreases with temperature, at 100 C the sensitivity is 160 [O{sub 2}]{sup -1}. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  7. FIBER OPTICAL MICRO-DETECTORS FOR OXYGEN SENSING IN POWER PLANTS

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III; Po Zhang

    2005-04-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. One of the critical materials issues is to demonstrate that the luminescent cluster immobilized in the sol-gel porous support can withstand high temperature. At the same time the sol-gel matrix must have a high permeability to oxygen. Using a potassium salt of the molybdenum clusters, K{sub 2}Mo{sub 6}Cl{sub 14}, we have established the conditions necessary for deposition of optical quality sol-gel films. From spectroscopic measurements of the film we have shown that the cluster luminescence is stable following heat cycling of 54 hours at 200 C. Quenching of a factor of 1.5X between pure nitrogen and 21% oxygen was observed from in-situ measurements of films heated directly at 200 C. An automated system for characterizing fiber optic oxygen sensors up to 220 C with a temporal resolution better than 10 s is under construction. We estimate a signal of 6 x 10{sup 8} photons/s after complete quenching in 21% oxygen. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  8. FIBER OPTICAL MICRO-DETECTORS FOR OXYGEN SENSING IN POWER PLANTS

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III

    2004-04-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. The luminescence of Mo{sub 6}Cl{sub 12} immobilized in a sol-gel matrix was measured as a function of heater temperature up to 200 C, in an inert environment. While the luminescence decreased with temperature, the integrated intensity at 200 C should be sufficient to enable detection of the luminescence in a fiber geometry. Previously we found that aging Mo{sub 6}Cl{sub 12} at temperatures above 250 C converts the canary yellow Mo{sub 6}Cl{sub 12} to a non-luminescent gray solid. Optical and thermal aging experiments show that the alkali metal salts of Mo{sub 6}Cl{sub 12} have higher thermal stabilities and remain luminescent after aging at 280 C.

  9. FIBER OPTICAL MICRO-DETECTORS FOR OXYGEN SENSING IN POWER PLANTS

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III; Po Zhang

    2005-01-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. One of the critical materials issues is to demonstrate that the luminescent cluster immobilized in the sol-gel porous support can withstand high temperature. At the same time the sol-gel matrix must have a high permeability to oxygen. Using a potassium salt of the molybdenum clusters, K{sub 2}Mo{sub 6}Cl{sub 14}, we have established the conditions necessary for deposition of optical quality sol-gel films. From spectroscopic measurements of the film we have shown that the cluster luminescence is stable following heat cycling of 1 hour at 250 C. Quenching of a factor of 4X between pure nitrogen and 21% oxygen was observed for films cured directly at 200 C. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  10. Fiber Optical Micro-detectors for Oxygen Sensing in Power Plants

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III; Po Zhang

    2006-01-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Previously we described a particle-in-binder approach to immobilizing the potassium salt of a molybdenum cluster, K{sub 2}Mo{sub 6}Cl{sub 14}, at the tips of optical fibers. Compared to previous methods, the particle-in-binder approach affords fibers with greatly improved mechanical properties. We have extensively characterized two fiber sensors at high temperature. We obtain quenching ratios between pure nitrogen and 21% oxygen as high as 3.9 x at 70 C. For the first sensor at 60 C we obtained a {+-} 1% variation in the quenching ratio over 6 cycles of measurement, and monitored the device performance over 23 days. We were able to operate the second sensor continuously for 14 hours at 70 C, and the sensor quenching ratio was stable to 5% over that time period. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  11. Ex vivo culture of the intestinal epithelium: strategies and applications.

    PubMed

    Leushacke, Marc; Barker, Nick

    2014-08-01

    Limited pools of resident adult stem cells are critical effectors of epithelial renewal in the intestine throughout life. Recently, significant progress has been made regarding the isolation and in vitro propagation of fetal and adult intestinal stem cells in mammals. It is now possible to generate ever-expanding, three-dimensional epithelial structures in culture that closely parallel the in vivo epithelium of the intestine. Growing such organotypic epithelium ex vivo facilitates a detailed description of endogenous niche factors or stem-cell characteristics, as they can be monitored in real time. Accordingly, this technology has already greatly contributed to our understanding of intestinal adult stem-cell renewal and differentiation. Transplanted organoids have also been proven to readily integrate into, and effect the long-term repair of, mouse colonic epithelia in vivo, establishing the organoid culture as a promising tool for adult stem cell/gene therapy. In another exciting development, novel genome-editing techniques have been successfully employed to functionally repair disease loci in cultured intestinal stem cells from human patients with a hereditary defect. It is anticipated that this technology will be instrumental in exploiting the regenerative medicine potential of human intestinal stem cells for treating human disorders in the intestinal tract and for creating near-physiological ex vivo models of human gastrointestinal disease.

  12. Fiber Optical Micro-detectors for Oxygen Sensing in Power Plants

    SciTech Connect

    Gregory L. Baker; Ruby N. Ghosh; D.J. Osborn III; Po Zhang

    2005-10-01

    A reflection mode fiber optic oxygen sensor that can operate at high temperatures for power plant applications is being developed. The sensor is based on the {sup 3}O{sub 2} quenching of the red emission from hexanuclear molybdenum chloride clusters. Previously we immobilized the potassium salt of a molybdenum cluster, K{sub 2}M{sub 6}Cl{sub 14}, in a sol-gel matrix and showed that the luminescence is stable after 54 hours at 200 C, but the quenching ratios were low and the films delaminated after thermal cycling due to densification of the matrix. Three new approaches to solve decreased quenching over time and delamination of films off fiber tips were investigated. In the first approach K{sub 2}Mo{sub 6}Cl{sub 14} embedded in cured sol-gel particles were incorporated into a TEOS based sol-gel. These gave enhanced quenching (6x), but delaminated. Our second approach was to use a commercial cyanoacrylate glue to immobilize the particles onto the tip of an optical fiber. This gave better adhesion and good quenching initially, but eventually the glue degraded upon heating. Our third approach was to use a 55% OtMOS/ TEOS sol-gel binder. Films based on this new sol-gel binder show high quenching ({approx}6x) and superior mechanical stability even after thermal cycling. Sensor measurements on an optical fiber containing K{sub 2}Mo{sub 6}Cl{sub 14} embedded in cured sol-gel particles were obtained from 100 to 25 C. The signal intensity in nitrogen was stable at 2.8 {+-} 0.2 nW, and the quenching ratio (ratio of signal in N{sub 2} vs. 21 % O{sub 2}) varied from 4.4 to 6.9X. These are promising results for a high temperature fiber optical oxygen sensor based on molybdenum chloride clusters.

  13. Molybdenum chloride incorporated sol-gel materials for oxygen sensing above room temperature

    NASA Astrophysics Data System (ADS)

    Osborn, D. J., III

    Maximizing the efficiency of the combustion process requires the ability to sense oxygen levels over a broad range of concentrations with fast response times under rapidly varying conditions of pressure and temperature to maintain the correct fuel/oxygen ratio in real-time. Quenching of the luminescence from organometallic compounds by oxygen has been used to develop a number of fiber-based sensors. A major drawback of these organometallic indicators for combustion applications is that the chromophores degrade with time, have a limited operational temperature range, typically room temperature +/-25°C, and lack long-term reliability. This work investigates luminescent molybdenum clusters based on Mo6Cl12 were as replacements for organometallic indicators. A study of the high temperature stability of Mo6Cl 12 in air revealed irreversible changes in the optical absorption spectrum at T >250°C and a loss of the red luminescence characteristic of the pristine clusters. Thermal aging experiments run in air and under nitrogen point to oxidation of the clusters as the cause of the change in optical properties. X-ray powder diffraction measurements on samples annealed at 300°C under controlled conditions are consistent with oxidation of Mo6Cl 12 to form MoO3. Optical and thermal aging experiments show that K2Mo6Cl14•1H2O, the alkali metal salt of Mo6Cl12, has higher thermal stability and remains luminescent after long-term aging in air at 280°C. Methods were developed for depositing K2Mo6Cl14•1H 2O-incorporated sol--gel films on planar and optical fiber substrates by dip coating and spray coating. The mechanical properties of the films depended on the film thickness; thin films were stable, but cracks often formed in the thicker films needed for sensors. This problem was addressed using two strategies: altering the components of the sol--gel solutions used to embed the clusters and by devising a composite approach to sensing layers where a slurry of fully cured sol

  14. Full-field OCT: ex vivo and in vivo biological imaging applications

    NASA Astrophysics Data System (ADS)

    Grieve, Katharine; Dubois, Arnaud; Moneron, Gael; Guyot, Elvire; Boccara, Albert C.

    2005-04-01

    We present results of studies in embryology and ophthalmology performed using our ultrahigh-resolution full-field OCT system. We also discuss recent developments to our ultrashort acquisition time full-field optical coherence tomography system designed to allow in vivo biological imaging. Preliminary results of high-speed imaging in biological samples are presented. The core of the experimental setup is the Linnik interferometer, illuminated by a white light source. En face tomographic images are obtained in real-time without scanning by computing the difference of two phase-opposed interferometric images recorded by high-resolution CCD cameras. An isotropic spatial resolution of ~1 μm is achieved thanks to the short source coherence length and the use of high numerical aperture microscope objectives. A detection sensitivity of ~90 dB is obtained by means of image averaging and pixel binning. In ophthalmology, reconstructed xz images from rat ocular tissue are presented, where cellular-level structures in the retina are revealed, demonstrating the unprecedented resolution of our instrument. Three-dimensional reconstructions of the mouse embryo allowing the study of the establishment of the anterior-posterior axis are shown. Finally we present the first results of embryonic imaging using the new rapid acquisition full-field OCT system, which offers an acquisition time of 10 μs per frame.

  15. In Vivo Application of Optogenetics for Neural Circuit Analysis

    PubMed Central

    2012-01-01

    Optogenetics combines optical and genetic methods to rapidly and reversibly control neural activities or other cellular functions. Using genetic methods, specific cells or anatomical pathways can be sensitized to light through exogenous expression of microbial light activated opsin proteins. Using optical methods, opsin expressing cells can be rapidly and reversibly controlled by pulses of light of specific wavelength. With the high spatial temporal precision, optogenetic tools have enabled new ways to probe the causal role of specific cells in neural computation and behavior. Here, we overview the current state of the technology, and provide a brief introduction to the practical considerations in applying optogenetics in vivo to analyze neural circuit functions. PMID:22896801

  16. Progress connecting multi-disciplinary geoscience communities through the VIVO semantic web application

    NASA Astrophysics Data System (ADS)

    Gross, M. B.; Mayernik, M. S.; Rowan, L. R.; Khan, H.; Boler, F. M.; Maull, K. E.; Stott, D.; Williams, S.; Corson-Rikert, J.; Johns, E. M.; Daniels, M. D.; Krafft, D. B.

    2015-12-01

    UNAVCO, UCAR, and Cornell University are working together to leverage semantic web technologies to enable discovery of people, datasets, publications and other research products, as well as the connections between them. The EarthCollab project, an EarthCube Building Block, is enhancing an existing open-source semantic web application, VIVO, to address connectivity gaps across distributed networks of researchers and resources related to the following two geoscience-based communities: (1) the Bering Sea Project, an interdisciplinary field program whose data archive is hosted by NCAR's Earth Observing Laboratory (EOL), and (2) UNAVCO, a geodetic facility and consortium that supports diverse research projects informed by geodesy. People, publications, datasets and grant information have been mapped to an extended version of the VIVO-ISF ontology and ingested into VIVO's database. Data is ingested using a custom set of scripts that include the ability to perform basic automated and curated disambiguation. VIVO can display a page for every object ingested, including connections to other objects in the VIVO database. A dataset page, for example, includes the dataset type, time interval, DOI, related publications, and authors. The dataset type field provides a connection to all other datasets of the same type. The author's page will show, among other information, related datasets and co-authors. Information previously spread across several unconnected databases is now stored in a single location. In addition to VIVO's default display, the new database can also be queried using SPARQL, a query language for semantic data. EarthCollab will also extend the VIVO web application. One such extension is the ability to cross-link separate VIVO instances across institutions, allowing local display of externally curated information. For example, Cornell's VIVO faculty pages will display UNAVCO's dataset information and UNAVCO's VIVO will display Cornell faculty member contact and

  17. Application of an improved in vivo counting program to Korean nuclear power plants.

    PubMed

    Kong, Tae Young; Kim, Hee Geun

    2009-01-01

    During the in vivo counting of individuals using a whole body counter at nuclear power plants (NPPs), external skin contamination is occasionally mistaken for internal radioactive contamination. This not only confuses the degree of external contamination and internal contamination, but can also result in the excessively conservative estimation of radioactive contamination. In this paper, previous experiments to improve in vivo counting at NPPs are introduced briefly and the practical application of techniques presented in those experiments to Korean NPPs is demonstrated in detail.

  18. Clinical applications of in vivo neutron-activation analysis

    SciTech Connect

    Cohn, S.H.

    1982-01-01

    In vivo neutron activation has opened a new era of both clinical diagnosis and therapy evaluation, and investigation into and modelling of body composition. The techniques are new, but it is already clear that considerable strides can be made in increasing accuracy and precision, increasing the number of elements susceptible to measurement, enhancing uniformity, and reducing the dose required for the measurement. The work presently underway will yield significant data on a variety of environmental contaminants such as Cd. Compositional studies are determining the level of vital constituents such as nitrogen and potassium in both normal subjects and in patients with a variety of metabolic disorders. Therapeutic programs can be assessed while in progress.

  19. In-vivo neutron activation analysis: principles and clinical applications

    SciTech Connect

    Cohn, S.H.

    1982-01-01

    In vivo neutron activation has opened a new era of both clinical diagnosis and therapy evaluation, and investigation into and modelling of body composition. The techniques are new, but it is already clear that considerable strides can be made in increasing accuracy and precision, increasing the number of elements susceptible to measurement, enhancing uniformity, and reducing the dose required for the measurement. The work presently underway will yield significant data on a variety of environmental contaminants such as Cd. Compositional studies are determining the level of vital constituents such as nitrogen and potassium in both normal subjects and in patients with a variety of metabolic disorders. Therapeutic programs can be assessed while in progress. It seems likely that by the end of this century there will have been significant progress with this research tool, and exciting insights obtained into the nature and dynamics of human body composition.

  20. Carbon nanotubes from synthesis to in vivo biomedical applications.

    PubMed

    Sajid, Muhammad Imran; Jamshaid, Usama; Jamshaid, Talha; Zafar, Nadiah; Fessi, H; Elaissari, Abdelhamid

    2016-03-30

    Owing to their unique and interesting properties, extensive research round the globe has been carried out on carbon nanotubes and carbon nanotubes based systems to investigate their practical usefulness in biomedical applications. The results from these studies demonstrate a great promise in their use in targeted drug delivery systems, diagnostic techniques and in bio-analytical applications. Although, carbon nanotubes possess quite interesting properties, which make them potential candidates in the biomedical science, but they also have some inherent properties which arise great concern regarding their biosafety. In this comprehensive review, we have discussed different aspects of carbon nanotubes and carbon nanotube based systems related to biomedical applications. In the beginning, a short historical account of these tiny yet powerful particles is given followed by discussion regarding their types, properties, methods of synthesis, large scale production method, purification techniques and characterization aspects of carbon nanotubes. In the second part of the review, the functionalization of carbon nanotubes is reviewed in detail, which is not only important to make them biocompatible and stable in biological systems but also render them a great property of loading various biomolecules, diagnostic and therapeutic moieties resulting in diversified applications. In the final part of the review, emphasis is given on the pharmacokinetic aspects of carbon nanotubes including administration routes, absorption mechanisms, distribution and elimination of carbon nanotubes based systems. Lastly, a comprehensive account about the potential biomedical applications has been given followed by insights into the future.

  1. A Telemedicine Application to Schedule Temperature in an In Vivo Sensor Network for Cancer Treatment

    PubMed Central

    Kamal, Rossi; Lee, Seok-Geun

    2012-01-01

    Abstract Wireless communication has played a significant role in modern healthcare systems. However, the death toll from chronic diseases, such as cancer, continues to increase. Hyperthermia combined with radiotherapy and/or chemotherapy is a promising strategy for cancer treatment, and temperature control is critical for the success of this intervention. In vivo sensors are an emerging technology in healthcare. Thermal awareness has also received attention in in vivo sensor research. In this context, we have been motivated to use in vivo sensors to regulate the temperature changes in cancer cells during combined treatment. Limitations in existing in vivo thermal-aware routing algorithms motivated us to use the in vivo “lightweight rendezvous routing” approach. However, smartphone-driven telemedicine applications are proliferating to provide remote healthcare and collaborative consultation, required in combined therapies. In this context, we have proposed a telemedicine application where a smartphone not only regulates temperature scheduling in in vivo sensors, but also communicates with local or remote clinicians to maintain collaborative efforts for combined therapies against cancer. PMID:23234425

  2. Histotripsy for Pediatric Cardiac Applications: In Vivo Neonatal Pig Model

    NASA Astrophysics Data System (ADS)

    Miller, Ryan M.; Owens, Gabe; Ensing, Gregory; Ludomirsky, Achiau; Cain, Charles; Xu, Zhen

    2010-03-01

    This study investigated the in vivo feasibility of using histotripsy to non-invasively create a flow channel between the ventricles by generating a perforation of the ventricular septum, clinically referred to as a ventricular septum defect (VSD). The overall goal is to develop a non-invasive procedure to aid in the treatment of neonatal patients with complex congenital heart diseases such as Hypoplastic Left Heart Syndrome (HLHS). Histotripsy is a therapeutic ultrasound technique that produces mechanical fractionation of soft tissue through controlled cavitation. The study was conducted in a live and intact neonatal pig model. The ventricular septum in the neonatal pig heart was treated with histotripsy delivered by a spherically focused 1 MHz transducer positioned outside the chest wall. Histotripsy treatment was applied using 5-cycle ultrasound pulses at 1 kHz pulse repetition frequency with 12-18 MPa peak negative pressure. The treatment was guided and monitored with ultrasound imaging. In all nine subjects treated, a bubble cloud was generated on the ventricular septum using histotripsy, and visualized with ultrasound imaging. Within 20 seconds to 4 minutes following the initiation of a bubble cloud, a VSD was created in all nine pigs and confirmed by the detection of blood flow through the ventricular septum with color Doppler ultrasound. Gross morphology and histology on all hearts showed a demarcated perforation in the ventricular septum. This study shows that a VSD can be created in an intact neonatal animal using extracorporeal histotripsy under real-time ultrasound guidance.

  3. Deep tissue fluorescence imaging and in vivo biological applications

    NASA Astrophysics Data System (ADS)

    Crosignani, Viera; Dvornikov, Alexander; Aguilar, Jose S.; Stringari, Chiara; Edwards, Robert; Mantulin, William W.; Gratton, Enrico

    2012-11-01

    We describe a novel technical approach with enhanced fluorescence detection capabilities in two-photon microscopy that achieves deep tissue imaging, while maintaining micron resolution. Compared to conventional two-photon microscopy, greater imaging depth is achieved by more efficient harvesting of fluorescence photons propagating in multiple-scattering media. The system maintains the conventional two-photon microscopy scheme for excitation. However, for fluorescence collection the detection system harvests fluorescence photons directly from a wide area of the turbid sample. The detection scheme relies on a wide area detector, minimal optical components and an emission path bathed in a refractive-index-matching fluid that minimizes emission photon losses. This detection scheme proved to be very efficient, allowing us to obtain high resolution images at depths up to 3 mm. This technique was applied to in vivo imaging of the murine small intestine (SI) and colon. The challenge is to image normal and diseased tissue in the whole live animal, while maintaining high resolution imaging at millimeter depth. In Lgr5-GFP mice, we have been successful in imaging Lgr5-eGFP positive stem cells, present in SI and colon crypt bases.

  4. Optogenetic tools for in vivo applications in neonatal mice

    NASA Astrophysics Data System (ADS)

    Zhang, Yue; Qin, Nan; Diao, Yupu; Guan, Yangtai; Fan, Lu; Crair, Michael C.; Zhang, Jiayi

    2012-10-01

    Spontaneous neural activities exist early in development and their spatiotemporal patterns play important roles in the development of sensory maps such as maps of retinotopy in the visual system. We summarized different optogenetic tools, including transgenic mouse lines, viral-mediated transfection and electroporation methods to enable the expression of light-gated channelrhodopsin (ChR2) in retinal ganglion cells (RGCs) before the onset of vision. Patch-clamp and extracellular recording experiments verified that activities of ChR2-expressing cells were precisely manipulated by the patterns of optical stimuli. In chronic stimulation experiments, light-emitting diodes controlled the activity patterns of ChR2-expressing RGCs in vivo. Changes in the retinotopic map in Superior Colliculus (SC) were examined by quantifying the relative sizes of fluorescently labeled target zones. Our results revealed that various optogenetic and optical tools can manipulate retinal activities with precise temporal patterns. These techniques can be readily used in studying the development of the central nervous system of neonatal rodents.

  5. Comparison of in vivo and ex vivo laser scanning microscopy and multiphoton tomography application for human and porcine skin imaging

    NASA Astrophysics Data System (ADS)

    Darvin, M. E.; Richter, H.; Zhu, Y. J.; Meinke, M. C.; Knorr, F.; Gonchukov, S. A.; Koenig, K.; Lademann, J.

    2014-07-01

    Two state-of-the-art microscopic optical methods, namely, confocal laser scanning microscopy in the fluorescence and reflectance regimes and multiphoton tomography in the autofluorescence and second harmonic generation regimes, are compared for porcine skin ex vivo and healthy human skin in vivo. All skin layers such as stratum corneum (SC), stratum spinosum (SS), stratum basale (SB), papillary dermis (PD) and reticular dermis (RD) as well as transition zones between these skin layers are measured noninvasively at a high resolution, using the above mentioned microscopic methods. In the case of confocal laser scanning microscopy (CLSM), measurements in the fluorescence regime were performed by using a fluorescent dye whose topical application on the surface is well suited for the investigation of superficial SC and characterisation of the skin barrier function. For investigations of deeply located skin layers, such as SS, SB and PD, the fluorescent dye must be injected into the skin, which markedly limits fluorescence measurements using CLSM. In the case of reflection CLSM measurements, the obtained results can be compared to the results of multiphoton tomography (MPT) for all skin layers excluding RD. CLSM cannot distinguish between dermal collagen and elastin measuring their superposition in the RD. By using MPT, it is possible to analyse the collagen and elastin structures separately, which is important for the investigation of anti-aging processes. The resolution of MPT is superior to CLSM. The advantages and limitations of both methods are discussed and the differences and similarities between human and porcine skin are highlighted.

  6. Comparison of in vivo and ex vivo laser scanning microscopy and multiphoton tomography application for human and porcine skin imaging

    SciTech Connect

    Darvin, M E; Richter, H; Zhu, Y J; Meinke, M C; Knorr, F; Lademann, J; Gonchukov, S A; Koenig, K

    2014-07-31

    Two state-of-the-art microscopic optical methods, namely, confocal laser scanning microscopy in the fluorescence and reflectance regimes and multiphoton tomography in the autofluorescence and second harmonic generation regimes, are compared for porcine skin ex vivo and healthy human skin in vivo. All skin layers such as stratum corneum (SC), stratum spinosum (SS), stratum basale (SB), papillary dermis (PD) and reticular dermis (RD) as well as transition zones between these skin layers are measured noninvasively at a high resolution, using the above mentioned microscopic methods. In the case of confocal laser scanning microscopy (CLSM), measurements in the fluorescence regime were performed by using a fluorescent dye whose topical application on the surface is well suited for the investigation of superficial SC and characterisation of the skin barrier function. For investigations of deeply located skin layers, such as SS, SB and PD, the fluorescent dye must be injected into the skin, which markedly limits fluorescence measurements using CLSM. In the case of reflection CLSM measurements, the obtained results can be compared to the results of multiphoton tomography (MPT) for all skin layers excluding RD. CLSM cannot distinguish between dermal collagen and elastin measuring their superposition in the RD. By using MPT, it is possible to analyse the collagen and elastin structures separately, which is important for the investigation of anti-aging processes. The resolution of MPT is superior to CLSM. The advantages and limitations of both methods are discussed and the differences and similarities between human and porcine skin are highlighted. (laser biophotonics)

  7. In vivo confocal microscopy in dermatology: from research to clinical application

    NASA Astrophysics Data System (ADS)

    Ulrich, Martina; Lange-Asschenfeldt, Susanne

    2013-06-01

    Confocal laser scanning microscopy (CLSM) represents an emerging technique for the noninvasive histomorphological analysis of skin in vivo and has shown its applicability for dermatological research as well as its value as an adjunct tool in the clinical management of skin cancer patients. Herein, we aim to give an overview on the current clinical indications for CLSM in dermatology and also highlight the diverse applications of CLSM in dermatological research.

  8. Red-Shifted Aequorin Variants Incorporating Non-Canonical Amino Acids: Applications in In Vivo Imaging

    PubMed Central

    Grinstead, Kristen M.; Rowe, Laura; Ensor, Charles M.; Joel, Smita; Daftarian, Pirouz; Dikici, Emre; Zingg, Jean-Marc; Daunert, Sylvia

    2016-01-01

    The increased importance of in vivo diagnostics has posed new demands for imaging technologies. In that regard, there is a need for imaging molecules capable of expanding the applications of current state-of-the-art imaging in vivo diagnostics. To that end, there is a desire for new reporter molecules capable of providing strong signals, are non-toxic, and can be tailored to diagnose or monitor the progression of a number of diseases. Aequorin is a non-toxic photoprotein that can be used as a sensitive marker for bioluminescence in vivo imaging. The sensitivity of aequorin is due to the fact that bioluminescence is a rare phenomenon in nature and, therefore, it does not suffer from autofluorescence, which contributes to background emission. Emission of bioluminescence in the blue-region of the spectrum by aequorin only occurs when calcium, and its luciferin coelenterazine, are bound to the protein and trigger a biochemical reaction that results in light generation. It is this reaction that endows aequorin with unique characteristics, making it ideally suited for a number of applications in bioanalysis and imaging. Herein we report the site-specific incorporation of non-canonical or non-natural amino acids and several coelenterazine analogues, resulting in a catalog of 72 cysteine-free, aequorin variants which expand the potential applications of these photoproteins by providing several red-shifted mutants better suited to use in vivo. In vivo studies in mouse models using the transparent tissue of the eye confirmed the activity of the aequorin variants incorporating L-4-iodophehylalanine and L-4-methoxyphenylalanine after injection into the eye and topical addition of coelenterazine. The signal also remained localized within the eye. This is the first time that aequorin variants incorporating non-canonical amino acids have shown to be active in vivo and useful as reporters in bioluminescence imaging. PMID:27367859

  9. In Vivo Application and Localization of Transcranial Focused Ultrasound Using Dual-Mode Ultrasound Arrays

    PubMed Central

    Haritonova, Alyona; Liu, Dalong; Ebbini, Emad S.

    2015-01-01

    Focused ultrasound (FUS) has been proposed for a variety of transcranial applications, including neuromodulation, tumor ablation, and blood brain barrier opening. A flurry of activity in recent years has generated encouraging results demonstrating its feasibility in these and other applications. To date, monitoring of FUS beams have been primarily accomplished using MR guidance, where both MR thermography and elastography have been used. The recent introduction of real-time dual-mode ultrasound array (DMUA) systems offers a new paradigm in transcranial focusing. In this paper, we present first experimental results of ultrasound-guided transcranial FUS (tFUS) application in a rodent brain, both ex vivo and in vivo. DMUA imaging is used for visualization of the treatment region for placement of the focal spot within the brain. This includes the detection and localization of pulsating blood vessels at or near the target point(s). In addition, DMUA imaging is used to monitor and localize the FUS-tissue interactions in real-time. In particular, a concave (40-mm radius of curvature), 32-element, 3.5 MHz DMUA prototype was used for imaging and tFUS application in ex vivo and in vivo rat model. The ex vivo experiments were used to evaluate the point spread function (psf) of the transcranial DMUA imaging at various points within the brain. In addition, DMUA-based transcranial ultrasound thermography measurements were compared with thermocouple measurements of subtherapeutic tFUS heating in rat brain ex vivo. The ex vivo setting was also used to demonstrate the DMUA capability to produce localized thermal lesions. The in vivo experiments were designed to demonstrate the ability of the DMUA to apply, monitor, and localize subtherapeutic tFUS patterns that could be beneficial in transient blood brain barrier opening. The results show that, while the DMUA focus is degraded due to the propagation through the skull, it still produces localized heating effects within sub

  10. Nanodiamonds for Medical Applications: Interaction with Blood in Vitro and in Vivo.

    PubMed

    Tsai, Lin-Wei; Lin, Yu-Chung; Perevedentseva, Elena; Lugovtsov, Andrei; Priezzhev, Alexander; Cheng, Chia-Liang

    2016-07-12

    Nanodiamonds (ND) have emerged to be a widely-discussed nanomaterial for their applications in biological studies and for medical diagnostics and treatment. The potentials have been successfully demonstrated in cellular and tissue models in vitro. For medical applications, further in vivo studies on various applications become important. One of the most challenging possibilities of ND biomedical application is controllable drug delivery and tracing. That usually assumes ND interaction with the blood system. In this work, we study ND interaction with rat blood and analyze how the ND surface modification and coating can optimize the ND interaction with the blood. It was found that adsorption of a low concentration of ND does not affect the oxygenation state of red blood cells (RBC). The obtained in vivo results are compared to the results of in vitro studies of nanodiamond interaction with rat and human blood and blood components, such as red blood cells and blood plasma. An in vivo animal model shows ND injected in blood attach to the RBC membrane and circulate with blood for more than 30 min; and ND do not stimulate an immune response by measurement of proinflammatory cytokine TNF-α with ND injected into mice via the caudal vein. The results further confirm nanodiamonds' safety in organisms, as well as the possibility of their application without complicating the blood's physiological conditions.

  11. Nanodiamonds for Medical Applications: Interaction with Blood in Vitro and in Vivo

    PubMed Central

    Tsai, Lin-Wei; Lin, Yu-Chung; Perevedentseva, Elena; Lugovtsov, Andrei; Priezzhev, Alexander; Cheng, Chia-Liang

    2016-01-01

    Nanodiamonds (ND) have emerged to be a widely-discussed nanomaterial for their applications in biological studies and for medical diagnostics and treatment. The potentials have been successfully demonstrated in cellular and tissue models in vitro. For medical applications, further in vivo studies on various applications become important. One of the most challenging possibilities of ND biomedical application is controllable drug delivery and tracing. That usually assumes ND interaction with the blood system. In this work, we study ND interaction with rat blood and analyze how the ND surface modification and coating can optimize the ND interaction with the blood. It was found that adsorption of a low concentration of ND does not affect the oxygenation state of red blood cells (RBC). The obtained in vivo results are compared to the results of in vitro studies of nanodiamond interaction with rat and human blood and blood components, such as red blood cells and blood plasma. An in vivo animal model shows ND injected in blood attach to the RBC membrane and circulate with blood for more than 30 min; and ND do not stimulate an immune response by measurement of proinflammatory cytokine TNF-α with ND injected into mice via the caudal vein. The results further confirm nanodiamonds’ safety in organisms, as well as the possibility of their application without complicating the blood’s physiological conditions. PMID:27420044

  12. [Application of numerical convolution in in vivo/in vitro correlation research].

    PubMed

    Yue, Peng

    2009-01-01

    This paper introduced the conception and principle of in vivo/in vitro correlation (IVIVC) and convolution/deconvolution methods, and elucidated in details the convolution strategy and method for calculating the in vivo absorption performance of the pharmaceutics according to the their pharmacokinetic data in Excel, then put the results forward to IVIVC research. Firstly, the pharmacokinetic data ware fitted by mathematical software to make up the lost points. Secondly, the parameters of the optimal fitted input function were defined by trail-and-error method according to the convolution principle in Excel under the hypothesis that all the input functions fit the Weibull functions. Finally, the IVIVC between in vivo input function and the in vitro dissolution was studied. In the examples, not only the application of this method was demonstrated in details but also its simplicity and effectiveness were proved by comparing with the compartment model method and deconvolution method. It showed to be a powerful tool for IVIVC research.

  13. Application of in vivo measurements for the management of cyanobacteria breakthrough into drinking water treatment plants.

    PubMed

    Zamyadi, Arash; Dorner, Sarah; Ndong, Mouhamed; Ellis, Donald; Bolduc, Anouka; Bastien, Christian; Prévost, Michèle

    2014-02-01

    The increasing presence of potentially toxic cyanobacterial blooms in drinking water sources and within drinking water treatment plants (DWTPs) has been reported worldwide. The objectives of this study are to validate the application of in vivo probes for the detection and management of cyanobacteria breakthrough inside DWTPs, and to verify the possibility of treatment adjustment based on intensive real-time monitoring. In vivo phycocyanin YSI probes were used to monitor the fate of cyanobacteria in raw water, clarified water, filtered water, and chlorinated water in a full scale DWTP. Simultaneous samples were also taken for microscopic enumeration. The in vivo probe was successfully used to detect the incoming densities of high cyanobacterial cell number into the clarification process and their breakthrough into the filtered water. In vivo probes were used to trace the increase in floating cells over the clarifier, a robust sign of malfunction of the coagulation-sedimentation process. Pre-emptive treatment adjustments, based on in vivo probe monitoring, resulted in successful removal of cyanobacterial cells. The field results on validation of the probes with cyanobacterial bloom samples showed that the probe responses are highly linear and can be used to trigger alerts to take action.

  14. Applications of the direct photon absorption technique for measuring bone mineral content in vivo. Determination of body composition in vivo

    NASA Technical Reports Server (NTRS)

    Cameron, J. R.

    1972-01-01

    The bone mineral content, BMC, determined by monoenergetic photon absorption technique, of 29 different locations on the long bones and vertebral columns of 24 skeletons was measured. Compressive tests were made on bone from these locations in which the maximum load and maximum stress were measured. Also the ultimate strain, modulus of elasticity and energy absorbed to failure were determined for compact bone from the femoral diaphysis and cancellous bone from the eighth through eleventh thoracic vertebrae. Correlations and predictive relationships between these parameters were examined to investigate the applicability of using the BMC at sites normally measured in vivo, i.e. radius and ulna in estimating the BMC and/or strength of the spine or femoral neck. It was found that the BMC at sites on the same bone were highly correlated r = 0.95 or better; the BMC at sites on different bones were also highly interrelated, r = 0.85. The BMC at various sites on the long bones could be estimated to between 10 and 15 per cent from the BMC of sites on the radius or ulna.

  15. Hepatic ablation with multiple interstitial ultrasound applicators: initial ex vivo and computational studies

    NASA Astrophysics Data System (ADS)

    Prakash, Punit; Salgaonkar, Vasant A.; Burdette, E. Clif; Diederich, Chris J.

    2011-03-01

    Radiofrequency (RF) ablation has emerged as an effective method for treating liver tumors under 3 cm in diameter. Multiple applicator devices and techniques - using RF, microwave and other modalities - are under development for thermal ablation of large and irregularly-shaped liver tumors. Interstitial ultrasound (IUS) applicators, comprised of linear arrays of independently powered tubular transducers, enable 3D control of the spatial power deposition profile and simultaneous ablation with multiple applicators. We evaluated IUS applicator configurations (parallel, converging and diverging implants) suitable for percutaneous and laparascopic placement with experiments in ex vivo bovine tissue and computational models. Ex vivo ablation zones measured 4.6+/-0.5 x 4.2+/-0.5 × 3.3+/-0.5 cm3 and 5.6+/-0.5 × 4.9+/-0.5 x 2.8+/-0.3 cm3 using three parallel applicators spaced 2 and 3 cm apart, respectively, and 4.0+/-0.3 × 3.2+/-0.4 × 2.9+/-0.2 cm3 using two parallel applicators spaced 2 cm apart. Computational models indicate in vivo ablation zones up to 4.5 × 4.4 × 5.5 cm3 and 5.7 × 4.8 × 5.2 cm3, using three applicators spaced 2 and 3 cm apart, respectively. Converging and diverging implant patterns can also be employed for conformal ablation of irregularly-shaped tumor margins by tailoring power levels along each device. Simultaneously powered interstitial ultrasound devices can create tailored ablation zones comparable to currently available RF devices and similarly sized microwave antennas.

  16. Metered Cryospray™: a novel uniform, controlled, and consistent in vivo application of liquid nitrogen cryogenic spray

    PubMed Central

    Mulcahey, Thomas I; Coad, James E; Fan, Wei Li; Grasso, Daniel J; Hanley, Brian M; Hawkes, Heather V; McDermott, Sean A; O’Connor, John P; Sheets, Ellen E; Vadala, Charles J

    2017-01-01

    In this article, a novel cryotherapy approach using a uniform, controlled, and consistent in vivo application of liquid nitrogen (LN2) spray as a Metered Cryospray™ (MCS) process is described. Although MCS may be used for many potential clinical applications, this paper focuses on the development that led to the controlled and consistent delivery of radial LN2 cryogen spray in order to generate a uniform circumferential effect and how the amount of MCS can be adapted to specifically ablate targeted diseases within a patient’s lumen such as an airway or esophagus. PMID:28255257

  17. In vivo toxicity of enoxaparin encapsulated in mucoadhesive nanoparticles: Topical application in a wound healing model

    NASA Astrophysics Data System (ADS)

    Huber, S. C.; Marcato, P. D.; Barbosa, R. M.; Duran, N.; Annichino-Bizzacchi, J. M.

    2013-04-01

    Wound healing comprises four distinct phases and involves many cell events and biologic markers. The use of nanoparticles for topical application has gaining attention due to its deeper penetration in the skin and the retention capacity of the drug in the site of application. In this study the effect and toxicity of mucoadhesive polymeric nanoparticles loaded with enoxaparin was evaluated in in vivo model of skin ulcer. Our results showed an interesting formulation based on mucoadhesive nanoparticles with enoxaparin that improved wound healing without cytotoxicity in vitro in all endpoint evaluated. Then, this semi-solid formulation is a promising option for skin ulcer treatment.

  18. Quantitative in vivo receptor binding. I. Theory and application to the muscarinic cholinergic receptor

    SciTech Connect

    Frey, K.A.; Ehrenkaufer, R.L.; Beaucage, S.; Agranoff, B.W.

    1985-02-01

    A novel approach to in vivo receptor binding experiments is presented which allows direct quantitation of binding site densities. The method is based on an equilibrium model of tracer uptake and is designed to produce a static distribution proportional to receptor density and to minimize possible confounding influences of regional blood flow, blood-brain barrier permeability, and nonspecific binding. This technique was applied to the measurement of regional muscarinic cholinergic receptor densities in rat brain using (/sup 3/H)scopolamine. Specific in vivo binding of scopolamine demonstrated saturability, a pharmacologic profile, and regional densities which are consistent with interaction of the tracer with the muscarinic receptor. Estimates of receptor density obtained with the in vivo method and in vitro measurements in homogenates were highly correlated. Furthermore, reduction in striatal muscarinic receptors following ibotenic acid lesions resulted in a significant decrease in tracer uptake in vivo, indicating that the correlation between scopolamine distribution and receptor density may be used to demonstrate pathologic conditions. We propose that the general method presented here is directly applicable to investigation of high affinity binding sites for a variety of radioligands.

  19. Optical brain imaging in vivo: techniques and applications from animal to man

    PubMed Central

    Hillman, Elizabeth M. C.

    2008-01-01

    Optical brain imaging has seen 30 years of intense development, and has grown into a rich and diverse field. In-vivo imaging using light provides unprecedented sensitivity to functional changes through intrinsic contrast, and is rapidly exploiting the growing availability of exogenous optical contrast agents. Light can be used to image microscopic structure and function in vivo in exposed animal brain, while also allowing noninvasive imaging of hemodynamics and metabolism in a clinical setting. This work presents an overview of the wide range of approaches currently being applied to in-vivo optical brain imaging, from animal to man. Techniques include multispectral optical imaging, voltage sensitive dye imaging and speckle-flow imaging of exposed cortex, in-vivo two-photon microscopy of the living brain, and the broad range of noninvasive topography and tomography approaches to near-infrared imaging of the human brain. The basic principles of each technique are described, followed by examples of current applications to cutting-edge neuroscience research. In summary, it is shown that optical brain imaging continues to grow and evolve, embracing new technologies and advancing to address ever more complex and important neuroscience questions. PMID:17994863

  20. Techniques and Applications of in vivo Diffusion Imaging of Articular Cartilage

    PubMed Central

    Raya, José G.

    2014-01-01

    Early in the process of osteoarthritis (OA) the composition (water, proteoglycan [PG], and collagen) and structure of articular cartilage is altered leading to changes in its mechanical properties. A technique that can assess the composition and structure of the cartilage in vivo can provide insight in the mechanical integrity of articular cartilage and become a powerful tool for the early diagnosis of OA. Diffusion tensor imaging (DTI) has been proposed as a biomarker for cartilage composition and structure. DTI is sensitive to the PG content through the mean diffusivity (MD) and to the collagen architecture through the fractional anisotropy (FA). However, the acquisition of DTI of articular cartilage in vivo is challenging due to the short T2 of articular cartilage (~40 ms at 3 T) and the high resolution needed (0.5–0.7 mm in plane) to depict the cartilage anatomy. We describe the pulse sequences used for in vivo DTI of articular cartilage and discus general strategies for protocol optimization. We provide a comprehensive review of measurements of DTI of articular cartilage from ex vivo validation experiments to its recent clinical applications. PMID:25865215

  1. Optical brain imaging in vivo: techniques and applications from animal to man.

    PubMed

    Hillman, Elizabeth M C

    2007-01-01

    Optical brain imaging has seen 30 years of intense development, and has grown into a rich and diverse field. In-vivo imaging using light provides unprecedented sensitivity to functional changes through intrinsic contrast, and is rapidly exploiting the growing availability of exogenous optical contrast agents. Light can be used to image microscopic structure and function in vivo in exposed animal brain, while also allowing noninvasive imaging of hemodynamics and metabolism in a clinical setting. This work presents an overview of the wide range of approaches currently being applied to in-vivo optical brain imaging, from animal to man. Techniques include multispectral optical imaging, voltage sensitive dye imaging and speckle-flow imaging of exposed cortex, in-vivo two-photon microscopy of the living brain, and the broad range of noninvasive topography and tomography approaches to near-infrared imaging of the human brain. The basic principles of each technique are described, followed by examples of current applications to cutting-edge neuroscience research. In summary, it is shown that optical brain imaging continues to grow and evolve, embracing new technologies and advancing to address ever more complex and important neuroscience questions.

  2. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell...

  3. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell...

  4. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell...

  5. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell...

  6. 21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Tissue culture media for human ex vivo tissue and cell culture processing applications. 876.5885 Section 876.5885 Food and Drugs FOOD AND DRUG... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell...

  7. Principles and application of an in vivo swine assay for the determination of arsenic bioavailability in contaminated matrices.

    PubMed

    Rees, Matthew; Sansom, Lloyd; Rofe, Allan; Juhasz, Albert L; Smith, Euan; Weber, John; Naidu, Ravi; Kuchel, Tim

    2009-04-01

    The assessment of arsenic (As) bioavailability from contaminated matrices is a crucial parameter for reducing the uncertainty when estimating exposure for human health risk assessment. In vivo assessment of As utilising swine is considered an appropriate model for human health risk assessment applications as swine are remarkably similar to humans in terms of physiology and As metabolism. While limited in vivo As bioavailability data is available in the literature, few details have been provided regarding technical considerations for performing in vivo assays. This paper describes, with examples, surgical, experimental design and analytical issues associated with performing chronic and acute in vivo swine assays to determine As bioavailability in contaminated soil and food.

  8. Oxygen Sensing via the Ethylene Response Transcription Factor RAP2.12 Affects Plant Metabolism and Performance under Both Normoxia and Hypoxia1[OPEN

    PubMed Central

    Paul, Melanie Verena; Iyer, Srignanakshi; Lehmann, Martin

    2016-01-01

    Subgroup-VII-ethylene-response-factor (ERF-VII) transcription factors are involved in the regulation of hypoxic gene expression and regulated by proteasome-mediated proteolysis via the oxygen-dependent branch of the N-end-rule pathway. While research into ERF-VII mainly focused on their role to regulate anoxic gene expression, little is known on the impact of this oxygen-sensing system in regulating plant metabolism and growth. By comparing Arabidopsis (Arabidopsis thaliana) plants overexpressing N-end-rule-sensitive and insensitive forms of the ERF-VII-factor RAP2.12, we provide evidence that oxygen-dependent RAP2.12 stability regulates central metabolic processes to sustain growth, development, and anoxic resistance of plants. (1) Under normoxia, overexpression of N-end-rule-insensitive Δ13RAP2.12 led to increased activities of fermentative enzymes and increased accumulation of fermentation products, which were accompanied by decreased adenylate energy states and starch levels, and impaired plant growth and development, indicating a role of oxygen-regulated RAP2.12 degradation to prevent aerobic fermentation. (2) In Δ13RAP2.12-overexpressing plants, decreased carbohydrate reserves also led to a decrease in anoxic resistance, which was prevented by external Suc supply. (3) Overexpression of Δ13RAP2.12 led to decreased respiration rates, changes in the levels of tricarboxylic acid cycle intermediates, and accumulation of a large number of amino acids, including Ala and γ-amino butyric acid, indicating a role of oxygen-regulated RAP2.12 abundance in controlling the flux-modus of the tricarboxylic acid cycle. (4) The increase in amino acids was accompanied by increased levels of immune-regulatory metabolites. These results show that oxygen-sensing, mediating RAP2.12 degradation is indispensable to optimize metabolic performance, plant growth, and development under both normoxic and hypoxic conditions. PMID:27372243

  9. The application of dermal papillary rings in dermatology by in vivo confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Xiang, W. Z.; Xu, A. E.; Xu, J.; Bi, Z. G.; Shang, Y. B.; Ren, Q. S.

    2010-08-01

    Confocal laser scanning microscopy (CLSM) allows noninvasive visualization of human skin in vivo, without needing to fix or section the tissue. Melanocytes and pigmented keratinocytes at the level of the basal layer form bright dermal papillary rings which are readily amenable to identify in confocal images. Our purpose was to explore the role of dermal papillary rings in assessment of lesion location, the diagnosis, differential diagnosis of lesions and assessment of therapeutic efficacy by in vivo CLSM. Seventy-one patients were imaged with the VivaScope 1500 reflectance confocal microscope provided by Lucid, Inc. The results indicate that dermal papillary rings can assess the location of lesion; the application of dermal papillary rings can provide diagnostic support and differential diagnosis for vitiligo, nevus depigmentosus, tinea versicolor, halo nevus, common nevi, and assess the therapeutic efficacy of NBUVB phototherapy plus topical 0.1 percent tacrolimus ointment for vitiligo. In conclusion, our findings indicate that the dermal papillary rings play an important role in the assessment the location of lesion, diagnosis, differential diagnosis of lesions and assessment of therapeutic efficacy by in vivo CLSM. CLSM may be a promising tool for noninvasive examination in dermatology. However, larger studies are needed to expand the application of dermal papillary rings in dermatology.

  10. In vivo studies of polyacrylate nanoparticle emulsions for topical and systemic applications.

    PubMed

    Greenhalgh, Kerriann; Turos, Edward

    2009-03-01

    We have recently reported on a new nanomedicine containing antibiotic-conjugated polyacrylate nanoparticles, which has shown activity against methicillin-resistant Staphylococcus aureus (MRSA) in vitro and no cytotoxicity toward human dermal cells. The water-based nanoparticle emulsion is capable of solubilizing lipophilic antibiotics for systemic administration, and the nanoparticle drug delivery vehicle has shown protective properties for antibiotics from hydrolytic cleavage by bacterial penicillinases, thus rejuvenating the drug's activity against resistant microbes such as MRSA. Here we report the first in vivo study of this penicillin-conjugated nanoparticle emulsion in determining toxicological responses initiated upon systemic and topical application in a murine model. Favorable results were observed in vivo upon both routes of administration and, when topically applied to a dermal abrasion model, the emulsion enhanced wound healing by an average of 3 to 5 days. This study suggests that polyacrylate nanoparticle-containing emulsions may afford promising opportunities for treating both skin and systemic infections.

  11. Real-time co-registration using novel ultrasound technology: ex vivo validation and in vivo applications

    PubMed Central

    Yang, Eric Y.; Polsani, Venkateshwar R.; Washburn, Michael J.; Zang, William; Hall, Anne L.; Virani, Salim S.; Hodge, Megan S.; Parker, Dan; Kerwin, William S.; Lawrie, Gerald M.; Garami, Zsolt; Ballantyne, Christie M.; Morrisett, Joel D.; Nambi, Vijay

    2011-01-01

    OBJECTIVES To evaluate whether a novel GPS-like position-sensing technology will enable accurate co-registration of images between imaging modalities. BACKGROUND Co-registration of images obtained by different imaging modalities will allow for comparison and fusion between imaging modalities, and therefore has significant clinical and research implications. We compared US and MR images of carotid endarterectomy (CEA) specimens using a novel position-sensing technology that uses an electromagnetic (EM) transmitter and sensors mounted on a US transducer. We then evaluated in vivo US-US and US-MRI co-registration. METHODS Thirteen CEA specimens underwent 3.0 Tesla MRI, after which images were uploaded to a LOGIQ E9 3D (GE Healthcare) US system and registered by identifying 2–3 common points. A similar method was used to evaluate US-MRI co-registration in patients with carotid atherosclerosis. For carotid intima-media thickness (C-IMT) measurements, ten volunteers underwent bilateral carotid US scans co-registered to 3D US maps created on the initial visit, with a repeat scan 2 days later. RESULTS For the CEA specimens, there was a mean 20 (standard error [SE] 2.0) frames per MRI slice. The mean frame difference, over 33 registration markers, between MR and US images for Readers 1 and 2 was −2.82 ± 19.32 (mean ± 95% confidence interval [CI]) frames and 2.09 ± 14.68 (mean ± 95% CI) frames, respectively. The US-MRI intraclass correlation coefficients (ICC) for the first and second readers were 0.995 and 0.997, respectively. For patients with carotid atherosclerosis, the mean US frames per MRI slice (9 [SE 2.3]) was within range of that observed with CEA specimens. Inter-visit, intra-reader, and inter-reader reproducibility of C-IMT measurements were consistently high (side-averaged ICC >0.9). CONCLUSION Accurate co-registration between US and other modalities is feasible with a GPS-like technology, which has significant clinical and research applicability. PMID

  12. In vivo evaluation of drug delivery after ultrasound application: A new use for the photoacoustic technique

    NASA Astrophysics Data System (ADS)

    Barja, P. R.; Acosta-Avalos, D.; Rompe, P. C. B.; Dos Anjos, F. H.; Marciano, F. R.; da Silva, M. D.

    2005-06-01

    Ultrasound application is a therapeutical resource widely employed in physiotherapy. One of its applications is the phonophoresis, a technique in which the ultrasound radiation is utilized to deliver drugs through the skin to soft tissues. The proposal of our study was to employ the Photoacoustic Technique to evaluate the efficacy of such treatment, analyzing if phonophoresis could enhance drug delivery through skin when compared to the more traditional method of manual massage. The configuration of the system employed was such that it was possible to perform in vivo measurements, which is a pre-requisite for this kind of study. The changes observed in the photoacoustic signal amplitude after each form of drug application were attributed to changes in the thermal effusivity of the system, due to penetration of the drug. The technique was able to detect differences in drug delivery between the specified physiotherapy treatments, indicating that phonophoresis enhances drug absorption by tissue.

  13. Fabricated micro-nano devices for in vivo and in vitro biomedical applications.

    PubMed

    Barkam, Swetha; Saraf, Shashank; Seal, Sudipta

    2013-01-01

    In recent years, the innovative use of microelectromechanical systems (MEMSs) and nanoelectromechanical systems (NEMSs) in biomedical applications has opened wide opportunities for precise and accurate human diagnostics and therapeutics. The introduction of nanotechnology in biomedical applications has facilitated the exact control and regulation of biological environments. This ability is derived from the small size of the devices and their multifunctional capabilities to operate at specific sites for selected durations of time. Researchers have developed wide varieties of unique and multifunctional MEMS/NEMS devices with micro and nano features for biomedical applications (BioMEMS/NEMS) using the state of the art microfabrication techniques and biocompatible materials. However, the integration of devices with the biological milieu is still a fundamental issue to be addressed. Devices often fail to operate due to loss of functionality, or generate adverse toxic effects inside the body. The in vitro and in vivo performance of implantable BioMEMS such as biosensors, smart stents, drug delivery systems, and actuation systems are researched extensively to understand the interaction of the BioMEMS devices with physiological environments. BioMEMS developed for drug delivery applications include microneedles, microreservoirs, and micropumps to achieve targeted drug delivery. The biocompatibility of BioMEMS is further enhanced through the application of tissue and smart surface engineering. This involves the application of nanotechnology, which includes the modification of surfaces with polymers or the self-assembly of monolayers of molecules. Thereby, the adverse effects of biofouling can be reduced and the performance of devices can be improved in in vivo and in vitro conditions.

  14. Application of locked nucleic acids to improve aptamer in vivo stability and targeting function

    PubMed Central

    Schmidt, Kathrin S.; Borkowski, Sandra; Kurreck, Jens; Stephens, Andrew W.; Bald, Rolf; Hecht, Maren; Friebe, Matthias; Dinkelborg, Ludger; Erdmann, Volker A.

    2004-01-01

    Aptamers are powerful candidates for molecular imaging applications due to a number of attractive features, including rapid blood clearance and tumor penetration. We carried out structure–activity relationship (SAR) studies with the Tenascin-C binding aptamer TTA1, which is a promising candidate for application in tumor imaging with radioisotopes. The aim was to improve its in vivo stability and target binding. We investigated the effect of thermal stabilization of the presumed non-binding double-stranded stem region on binding affinity and resistance against nucleolytic degradation. To achieve maximal thermal stem stabilization melting experiments with model hexanucleotide duplexes consisting of unmodified RNA, 2′-O-methyl RNA (2′-OMe), 2′-Fluoro RNA (2′-F) or Locked Nucleic Acids (LNAs) were initially carried out. Extremely high melting temperatures have been found for an LNA/LNA duplex. TTA1 derivatives with LNA and 2′-OMe modifications within the non-binding stem have subsequently been synthesized. Especially, the LNA-modified TTA1 derivative exhibited significant stem stabilization and markedly improved plasma stability while maintaining its binding affinity to the target. In addition, higher tumor uptake and longer blood retention was found in tumor-bearing nude mice. Thus, our strategy to introduce LNA modifications after the selection procedure is likely to be generally applicable to improve the in vivo stability of aptamers without compromising their binding properties. PMID:15509871

  15. Applications of nuclear techniques for in vivo body composition studies at Brookhaven National Laboratory

    SciTech Connect

    Cohn, S.H.; Ellis, K.J.; Vartsky, D.; Vaswani, A.N.; Wielopolski, L.

    1981-01-01

    A series of technical developments and their clinical applications in various nuclear technologies at Brookhaven National Laboratory is described. These include the development of a portable neutron activation facility for measuring cadmium in vivo in kidney and liver, a technique for the measurement of body iron utilizing nuclear resonant scattering of gamma rays, a non-invasive measure of the skeletal levels of lead by an x-ray fluorescence technique, and the development of a pulsed Van de Graaff generator as a source of pulsed neutrons for the measurement of lung silicon. (ACR)

  16. Phosphorescent nanoparticles for quantitative measurements of oxygen profiles in vitro and in vivo

    PubMed Central

    Choi, Nak Won; Verbridge, Scott S.; Williams, Rebecca M.; Chen, Jin; Kim, Ju-Young; Schmehl, Russel; Farnum, Cornelia E.; Zipfel, Warren R.; Fischbach, Claudia; Stroock, Abraham D.

    2012-01-01

    We present the development and characterization of nanoparticles loaded with a custom phosphor; we exploit these nanoparticles to perform quantitative measurements of the concentration of oxygen within three-dimensional (3-D) tissue cultures in vitro and blood vessels in vivo. We synthesized a customized ruthenium (Ru)-phosphor and incorporated it into polymeric nanoparticles via self-assembly. We demonstrate that the encapsulated phosphor is non-toxic with and without illumination. We evaluated two distinct modes of employing the phosphorescent nanoparticles for the measurement of concentrations of oxygen: 1) in vitro, in a 3-D microfluidic tumor model via ratiometric measurements of intensity with an oxygen-insensitive fluorophore as a reference, and 2) in vivo, in mouse vasculature using measurements of phosphorescence lifetime. With both methods, we demonstrated micrometer-scale resolution and absolute calibration to the dissolved oxygen concentration. Based on the ease and customizability of the synthesis of the nanoparticles and the flexibility of their application, these oxygen-sensing polymeric nanoparticles will find a natural home in a range of biological applications, benefiting studies of physiological as well as pathological processes in which oxygen availability and concentration play a critical role. PMID:22240511

  17. Oxygen sensing by primary cardiac fibroblasts: a key role of p21(Waf1/Cip1/Sdi1).

    PubMed

    Roy, Sashwati; Khanna, Savita; Bickerstaff, Alice A; Subramanian, Sukanya V; Atalay, Mustafa; Bierl, Michael; Pendyala, Srikanth; Levy, Dana; Sharma, Nidhi; Venojarvi, Mika; Strauch, Arthur; Orosz, Charles G; Sen, Chandan K

    2003-02-21

    In mammalian organs under normoxic conditions, O2 concentration ranges from 12% to <0.5%, with O2 approximately 14% in arterial blood and <10% in the myocardium. During mild hypoxia, myocardial O2 drops to approximately 1% to 3% or lower. In response to chronic moderate hypoxia, cells adjust their normoxia set point such that reoxygenation-dependent relative elevation of PO2 results in perceived hyperoxia. We hypothesized that O2, even in marginal relative excess of the PO2 to which cardiac cells are adjusted, results in activation of specific signal transduction pathways that alter the phenotype and function of these cells. To test this hypothesis, cardiac fibroblasts (CFs) isolated from adult murine ventricle were cultured in 10% or 21% O2 (hyperoxia relative to the PO2 to which cells are adjusted in vivo) and were compared with those cultured in 3% O2 (mild hypoxia). Compared with cells cultured in 3% O2, cells that were cultured in 10% or 21% O2 demonstrated remarkable reversible G2/M arrest and a phenotype indicative of differentiation to myofibroblasts. These effects were independent of NADPH oxidase function. CFs exposed to high O2 exhibited higher levels of reactive oxygen species production. The molecular signature response to perceived hyperoxia included (1) induction of p21, cyclin D1, cyclin D2, cyclin G1, Fos-related antigen-2, and transforming growth factor-beta1, (2) lowered telomerase activity, and (3) activation of transforming growth factor-beta1 and p38 mitogen-activated protein kinase. CFs deficient in p21 were resistant to such O2 sensitivity. This study raises the vital broad-based issue of controlling ambient O2 during the culture of primary cells isolated from organs.

  18. Oxygen Sensing in Drosophila: Multiple Isoforms of the Prolyl Hydroxylase Fatiga Have Different Capacity to Regulate HIFα/Sima

    PubMed Central

    Dekanty, Andrés; Wappner, Pablo

    2010-01-01

    Background The Hypoxia Inducible Factor (HIF) mediates cellular adaptations to low oxygen. Prolyl-4-hydroxylases are oxygen sensors that hydroxylate the HIF alpha-subunit, promoting its proteasomal degradation in normoxia. Three HIF-prolyl hydroxylases, encoded by independent genes, PHD1, PHD2, and PHD3, occur in mammals. PHD2, the longest PHD isoform includes a MYND domain, whose biochemical function is unclear. PHD2 and PHD3 genes are induced in hypoxia to shut down HIF dependent transcription upon reoxygenation, while expression of PHD1 is oxygen-independent. The physiologic significance of the diversity of the PHD oxygen sensors is intriguing. Methodology and Principal Findings We have analyzed the Drosophila PHD locus, fatiga, which encodes 3 isoforms, FgaA, FgaB and FgaC that are originated through a combination of alternative initiation of transcription and alternative splicing. FgaA includes a MYND domain and is homologous to PHD2, while FgaB and FgaC are shorter isoforms most similar to PHD3. Through a combination of genetic experiments in vivo and molecular analyses in cell culture, we show that fgaB but not fgaA is induced in hypoxia, in a Sima-dependent manner, through a HIF-Responsive Element localized in the first intron of fgaA. The regulatory capacity of FgaB is stronger than that of FgaA, as complete reversion of fga loss-of-function phenotypes is observed upon transgenic expression of the former, and only partial rescue occurs after expression of the latter. Conclusions and Significance Diversity of PHD isoforms is a conserved feature in evolution. As in mammals, there are hypoxia-inducible and non-inducible Drosophila PHDs, and a fly isoform including a MYND domain co-exists with isoforms lacking this domain. Our results suggest that the isoform devoid of a MYND domain has stronger regulatory capacity than that including this domain. PMID:20811646

  19. [Advances in ex vivo expansion and immunotherapy application of regulatory T cells].

    PubMed

    Yan, Li; Shao, Zong-Hong

    2015-04-01

    CD4+ CD25+ regulatory T cells (Treg) play a fundamental role in the establishment and maintenance of immune tolerance. In a some of experimental models, it was found that Tregs can quench autoimmune diseases, maintain allogeneic transplants, and prevent allergic diseases. A major obstacle to their clinical application is related to their definitive phenotype and very limited number of these cells in peripheral circulation, no more than 5%-10% of total CD4+ T cells. Recent progress of technologies for Treg sorting with multicolor flow cytometry and immuno-absorbing columns has overcome these obstacles, and opened the doors to the clinical application of Treg. This review highlight the characteristics of Treg, describe the current information of cell sorting and ex vivo expansion techniques, and outline the adoptive transfer experiments and clinical trials of immunotherapy that have been developed in recent years. It is foreseeable that Treg adoptive transfusion will be a promising immunosuppressive therapy.

  20. In vivo 783-channel diffuse reflectance imaging system and its application

    NASA Astrophysics Data System (ADS)

    Yang, Joon-Mo; Han, Yong-Hui; Yoon, Gilwon; Ahn, Byung Soo; Lee, Byung-Cheon; Soh, Kwang-Sup

    2007-08-01

    A fiber-based reflectance imaging system was constructed to produce in vivo absorption spectroscopic images of biological tissues with diffuse light in the cw domain. The principal part of this system is the 783-channel fiber probe, composed of 253 illumination fibers and 530 detection fibers distributed in a 20×20 mm square region. During illumination with the 253 illumination fibers, diffuse reflected lights are collected by the 530 detection fibers and recorded simultaneously as an image with an electron multiplying CCD camera for fast data acquisition. After signal acquisition, a diffuse reflectance image was reconstructed by applying the spectral normalization method we devised. To test the applicability of the spectral normalization, we conducted two phantom experiments with chicken breast tissue and white Delrin resin by using animal blood as an optical inhomogeneity. In the Delrin phantom experiment, we present images produced by two methods, spectral normalization and reference signal normalization, along with a comparison of the two. To show the feasibility of our system for biomedical applications, we took images of a human vein in vivo with the spectral normalization method.

  1. The application of graphene for in vitro and in vivo electrochemical biosensing.

    PubMed

    Janegitz, Bruno Campos; Silva, Tiago Almeida; Wong, Ademar; Ribovski, Laís; Vicentini, Fernando Campanhã; Taboada Sotomayor, Maria Del Pilar; Fatibello-Filho, Orlando

    2017-03-15

    Advances in analysis are required for rapid and reliable clinical diagnosis. Graphene is a 2D material that has been extensively used in the development of devices for the medical proposes due to properties such as an elevated surface area and excellent electrical conductivity. On the other hand, architectures have been designed with the incorporation of different biological recognition elements such as antibodies/antigens and DNA probes for the proposition of immunosensors and genosensors. This field presents a great progress in the last few years, which have opened up a wide range of applications. Here, we highlight a rather comprehensive overview of the interesting properties of graphene for in vitro, in vivo, and point-of-care electrochemical biosensing. In the course of the paper, we first introduce graphene, electroanalytical methods (potentiometry, voltammetry, amperometry and electrochemical impedance spectroscopy) followed by an overview of the prospects and possible applications of this material in electrochemical biosensors. In this context, we discuss some relevant trends including the monitoring of multiple biomarkers for cancer diagnostic, implantable devices for in vivo sensing and, development of point-of-care devices to real-time diagnostics.

  2. Multi-factorial in vivo stable isotope fractionation: causes, correlations, consequences and applications.

    PubMed

    Schmidt, Hanns-Ludwig; Robins, Richard J; Werner, Roland A

    2015-01-01

    Many physical and chemical processes in living systems are accompanied by isotope fractionation on H, C, N, O and S. Although kinetic or thermodynamic isotope effects are always the basis, their in vivo manifestation is often modulated by secondary influences. These include metabolic branching events or metabolite channeling, metabolite pool sizes, reaction mechanisms, anatomical properties and compartmentation of plants and animals, and climatological or environmental conditions. In the present contribution, the fundamentals of isotope effects and their manifestation under in vivo conditions are outlined. The knowledge about and the understanding of these interferences provide a potent tool for the reconstruction of physiological events in plants and animals, their geographical origin, the history of bulk biomass and the biosynthesis of defined representatives. It allows the use of isotope characteristics of biomass for the elucidation of biochemical pathways and reaction mechanisms and for the reconstruction of climatic, physiological, ecological and environmental conditions during biosynthesis. Thus, it can be used for the origin and authenticity control of food, the study of ecosystems and animal physiology, the reconstruction of present and prehistoric nutrition chains and paleaoclimatological conditions. This is demonstrated by the outline of fundamental and application-orientated examples for all bio-elements. The aim of the review is to inform (advanced) students from various disciplines about the whole potential and the scope of stable isotope characteristics and fractionations and to provide them with a comprehensive introduction to the literature on fundamental aspects and applications.

  3. Non invasive in vivo investigation of hepatobiliary structure and function in STII medaka (Oryzias latipes): methodology and applications

    PubMed Central

    Hardman, Ron C; Kullman, Seth W; Hinton, David E

    2008-01-01

    Background A novel transparent stock of medaka (Oryzias latipes; STII), recessive for all pigments found in chromatophores, permits transcutaneous imaging of internal organs and tissues in living individuals. Findings presented describe the development of methodologies for non invasive in vivo investigation in STII medaka, and the successful application of these methodologies to in vivo study of hepatobiliary structure, function, and xenobiotic response, in both 2 and 3 dimensions. Results Using brightfield, and widefield and confocal fluorescence microscopy, coupled with the in vivo application of fluorescent probes, structural and functional features of the hepatobiliary system, and xenobiotic induced toxicity, were imaged at the cellular level, with high resolution (< 1 μm), in living individuals. The findings presented demonstrate; (1) phenotypic response to xenobiotic exposure can be investigated/imaged in vivo with high resolution (< 1 μm), (2) hepatobiliary transport of solutes from blood to bile can be qualitatively and quantitatively studied/imaged in vivo, (3) hepatobiliary architecture in this lower vertebrate liver can be studied in 3 dimensions, and (4) non invasive in vivo imaging/description of hepatobiliary development in this model can be investigated. Conclusion The non-invasive in vivo methodologies described are a unique means by which to investigate biological structure, function and xenobiotic response with high resolution in STII medaka. In vivo methodologies also provide the future opportunity to integrate molecular mechanisms (e.g., genomic, proteomic) of disease and toxicity with phenotypic changes at the cellular and system levels of biological organization. While our focus has been the hepatobiliary system, other organ systems are equally amenable to in vivo study, and we consider the potential for discovery, within the context of in vivo investigation in STII medaka, as significant. PMID:18838008

  4. Sol-gel synthesized Sr4Al14O25:Eu2+/Dy3+ blue-green phosphorous as oxygen sensing materials

    NASA Astrophysics Data System (ADS)

    Aydin, Ilkyaz; Ertekin, Kadriye; Demirci, Selim; Gultekin, Serdar; Celik, Erdal

    2016-12-01

    In this study, we utilized newly synthesized Sr4Al14O25:Eu2+/Dy3+ blue-green phosphors along with silver nanoparticles (AgNPs) for fabrication of oxygen sensitive materials. To the best of our knowledge oxygen sensing mechanism of the offered design is totally different from the previously published works. One-component silicone: poly (1-trimethylsilyl-1-propyne), two component phenyl bearing silicone, plasticized polymethylmethacrylate, and ethylcellulose (EC) were tested as matrix materials. Electrospun fibers, porous and smooth thin films were produced by electrospinning or knife coating technique. Oxygen induced luminescence of the phosphors at 544 nm was followed as the analytical signal. Utilization of silver nanoparticles in silicone along with phosphors resulted with a 7.14 fold enhancement in the signal intensity and significant spectral response towards oxygen competing with the signals of the oxygen sensors utilizing metalloporphyrins or ruthenium complexes. We observed high sensitivity and stability, increased surface area and an enhancement in all sensor dynamics. Linearity of the calibration plots was superior for the pO2 range of 0.0-20.0% with respect to the previously reported ones. When stored at the ambient air of the laboratory there was no significant drift in signal intensity after 12 months. Our sensitivity and stability tests are still in progress.

  5. A feasibility study of in vivo applications of single beam acoustic tweezers

    SciTech Connect

    Li, Ying Lee, Changyang; Chen, Ruimin; Zhou, Qifa; Shung, K. Kirk

    2014-10-27

    Tools that are capable of manipulating micro-sized objects have been widely used in such fields as physics, chemistry, biology, and medicine. Several devices, including optical tweezers, atomic force microscope, micro-pipette aspirator, and standing surface wave type acoustic tweezers have been studied to satisfy this need. However, none of them has been demonstrated to be suitable for in vivo and clinical studies. Single beam acoustic tweezers (SBAT) is a technology that uses highly focused acoustic beam to trap particles toward the beam focus. Its feasibility was first theoretically and experimentally demonstrated by Lee and Shung several years ago. Since then, much effort has been devoted to improving this technology. At present, the tool is capable of trapping a microparticle as small as 1 μm, as well as a single red blood cell. Although in comparing to other microparticles manipulating technologies, SBAT has advantages of providing stronger trapping force and deeper penetration depth in tissues, and producing less tissue damage, its potential for in vivo applications has yet been explored. It is worth noting that ultrasound has been used as a diagnostic tool for over 50 years and no known major adverse effects have been observed at the diagnostic energy level. This paper reports the results of an initial attempt to assess the feasibility of single beam acoustic tweezers to trap microparticles in vivo inside of a blood vessel. The acoustic intensity of SBAT under the trapping conditions that were utilized was measured. The mechanical index and thermal index at the focus of acoustic beam were found to be 0.48 and 0.044, respectively, which meet the standard of commercial diagnostic ultrasound system.

  6. Application of Gold Nanorods for Photothermal Therapy in Ex Vivo Human Oesophagogastric Adenocarcinoma.

    PubMed

    Singh, Mohan; Harris-Birtill, David C C; Zhou, Yu; Gallina, Maria E; Cass, Anthony E G; Hanna, George B; Elson, Daniel S

    2016-03-01

    Gold nanoparticles are chemically fabricated and tuned to strongly absorb near infrared (NIR) light, enabling deep optical penetration and therapy within human tissues, where sufficient heating induces tumour necrosis. In our studies we aim to establish the optimal gold nanorod (GNR) concentration and laser power for inducing hyperthermic effects in tissues and test this photothermal effect on ex vivo human oesophagogastric adenocarcinoma. The ideal GNR concentration and NIR laser power that would elicit sufficient hyperthermia for tumour necrosis was pre-determined on porcine oesophageal tissues. Human ex vivo oesophageal and gastric adenocarcinoma tissues were incubated with GNR solutions and a GNR-free control solution with corresponding healthy tissues for comparison, then irradiated with NIR light for 10 minutes. Temperature rise was found to vary linearly with both the concentration of GNRs and the laser power. Human ex vivo oesophageal and gastric tissues consistently demonstrated a significant temperature rise when incubated in an optimally concentrated GNR solution (3 x 10(10) GNRs/ml) prior to NIR irradiation delivered at an optimal power (2 W/cm2). A mean temperature rise of 27 degrees C was observed in tissues incubated with GNRs, whereas only a modest 2 degrees C rise in tissues not exposed to any GNRs. This study evaluates the photothermal effects of GNRs on oesophagogastric tissue examines their application in the minimally invasive therapeutics of oesophageal and gastric adenocarcinomas. This could potentially be an effective method of clinically inducing irreversible oesophagogastric tumour photodestruction, with minimal collateral damage expected in (healthy) tissues free from GNRs.

  7. Application of Ulex europaeus agglutinin I-modified liposomes for oral vaccine: Ex Vivo bioadhesion and in Vivo immunity.

    PubMed

    Li, KeXin; Zhao, Xiuli; Xu, Shiyi; Pang, DaHai; Yang, ChunRong; Chen, DaWei

    2011-01-01

    The conjugation of Ulex europaeus agglutinin I (UEAI) onto surface of liposomes has been demonstrated to effectively improve the intestinal absorption of antigen, subsequently induced strong mucosal and systemic immune responses. In this context, we prepared bovine serum albumin (BSA)-encapsulating UEAI-modified liposomes (UEAI-LIP) and unmodified ones (LIP). The specific bioadhesion on mice gastro-intestinal mucosa was studied ex vivo. An important increase of interaction between UEAI-conjugated liposomes and the intestinal segments with Peyer's Patches (PPs) was observed compared with the unconjugated one (p<0.01). However, under the presence of α-L-fucose, which is the reported specific sugar for UEAI, specifically inhibited the activity of these conjugates. The immune-stimulating activity in vivo was studied by measuring immunoglobulin G (IgG) levels in serum and immunoglobulin A (IgA) levels in intestinal mucosal secretions following oral administration of BSA solution, LIP and UEAI-LIP in mice. Results indicate that antigen encapsulated in liposomes, especially the UEAI-modified ones, was favorable for inducing immune response. At 42 d after the first immunization, the highest IgG and IgA antibody levels produced by UEAI-LIP occurred, respectively showing 4.4-fold and 5-fold higher levels compared to those of the groups receiving BSA alone. This data demonstrated high potential of UEAI-modified liposomes for their use as carrier for oral vaccines.

  8. Orthogonal spin labeling using click chemistry for in vitro and in vivo applications.

    PubMed

    Kucher, Svetlana; Korneev, Sergei; Tyagi, Swati; Apfelbaum, Ronja; Grohmann, Dina; Lemke, Edward A; Klare, Johann P; Steinhoff, Heinz-Jürgen; Klose, Daniel

    2017-02-01

    Site-directed spin labeling for EPR- and NMR spectroscopy has mainly been achieved exploiting the specific reactivity of cysteines. For proteins with native cysteines or for in vivo applications, an alternative coupling strategy is required. In these cases click chemistry offers major benefits by providing a fast and highly selective, biocompatible reaction between azide and alkyne groups. Here, we establish click chemistry as a tool to target unnatural amino acids in vitro and in vivo using azide- and alkyne-functionalized spin labels. The approach is compatible with a variety of labels including reduction-sensitive nitroxides. Comparing spin labeling efficiencies from the copper-free with the strongly reducing copper(I)-catalyzed azide-alkyne click reaction, we find that the faster kinetics for the catalyzed reaction outrun reduction of the labile nitroxide spin labels and allow quantitative labeling yields within short reaction times. Inter-spin distance measurements demonstrate that the novel side chain is suitable for paramagnetic NMR- or EPR-based conformational studies of macromolecular complexes.

  9. An Alumina Toughened Zirconia Composite for Dental Implant Application: In Vivo Animal Results

    PubMed Central

    Schierano, Gianmario; Faga, Maria Giulia; Menicucci, Giulio; Sabione, Cristian; Genova, Tullio; von Degerfeld, Mitzy Mauthe; Peirone, Bruno; Cassenti, Adele; Cassoni, Paola; Carossa, Stefano

    2015-01-01

    Ceramic materials are widely used for biomedical applications because of their remarkable biological and mechanical properties. Composites made of alumina and zirconia are particularly interesting owing to their higher toughness with respect to the monolithic materials. On this basis, the present study is focused on the in vivo behavior of alumina toughened zirconia (ATZ) dental implants treated with a hydrothermal process. A minipig model was implemented to assess the bone healing through histology and mRNA expression at different time points (8, 14, 28, and 56 days). The novel ATZ implant was compared to a titanium clinical standard. The implants were analyzed in terms of microstructure and surface roughness before in vivo tests. The most interesting result deals with a statistically significant higher digital histology index for ATZ implants with respect to titanium standard at 56 days, which is an unprecedented finding, to the authors' knowledge. Even if further investigations are needed before proposing the clinical use in humans, the tested material proved to be a promising candidate among the possible ceramic dental implants. PMID:25945324

  10. Orthogonal spin labeling using click chemistry for in vitro and in vivo applications

    NASA Astrophysics Data System (ADS)

    Kucher, Svetlana; Korneev, Sergei; Tyagi, Swati; Apfelbaum, Ronja; Grohmann, Dina; Lemke, Edward A.; Klare, Johann P.; Steinhoff, Heinz-Jürgen; Klose, Daniel

    2017-02-01

    Site-directed spin labeling for EPR- and NMR spectroscopy has mainly been achieved exploiting the specific reactivity of cysteines. For proteins with native cysteines or for in vivo applications, an alternative coupling strategy is required. In these cases click chemistry offers major benefits by providing a fast and highly selective, biocompatible reaction between azide and alkyne groups. Here, we establish click chemistry as a tool to target unnatural amino acids in vitro and in vivo using azide- and alkyne-functionalized spin labels. The approach is compatible with a variety of labels including reduction-sensitive nitroxides. Comparing spin labeling efficiencies from the copper-free with the strongly reducing copper(I)-catalyzed azide-alkyne click reaction, we find that the faster kinetics for the catalyzed reaction outrun reduction of the labile nitroxide spin labels and allow quantitative labeling yields within short reaction times. Inter-spin distance measurements demonstrate that the novel side chain is suitable for paramagnetic NMR- or EPR-based conformational studies of macromolecular complexes.

  11. Ex vivo evaluation of a microneedle array device for transdermal application.

    PubMed

    Indermun, Sunaina; Choonara, Yahya E; Kumar, Pradeep; du Toit, Lisa C; Modi, Girish; van Vuuren, Sandy; Luttge, Regina; Pillay, Viness

    2015-12-30

    A new approach of transdermal drug delivery is the use of microneedles. This promising technique offers the potential to be broadly used for drug administration as it enables the dramatic increase in permeation of medicaments across the stratum corneum. The potential of microneedles has evolved to spawn a plethora of potential transdermal applications. In order to advance the microneedle capabilities and possibly revolutionize advanced drug delivery, this study introduces a novel transdermal electro-modulated hydrogel-microneedle array (EMH-MNA) device composed of a nano-porous, embeddable ceramic microneedle array as well as an optimized EMH for the electro-responsive delivery of indomethacin through the skin. The ex vivo permeation as well as drug release experiments were performed on porcine skin tissue to ascertain the electro-responsive capabilities of the device. In addition, the microbial permeation ability of the microneedles across the viable epidermis in both microneedle-punctured skin as well as hypodermic needle-punctured skin was determined. Ex vivo evaluation of the EMH-MNA device across porcine skin demonstrated that without electro-stimulation, significantly less drug release was obtained (±0.4540mg) as compared to electro-stimulation (±2.93mg).

  12. In vivo study of nanostructured akermanite/PEO coating on biodegradable magnesium alloy for biomedical applications.

    PubMed

    Razavi, Mehdi; Fathi, Mohammadhossein; Savabi, Omid; Vashaee, Daryoosh; Tayebi, Lobat

    2015-05-01

    The major issue for biodegradable magnesium alloys is the fast degradation and release of hydrogen gas. In this article, we aim to overcome these disadvantages by using a surface modified magnesium implant. We have recently coated AZ91 magnesium implants by akermanite (Ca2 MgSi2 O7 ) through the combined electrophoretic deposition (EPD) and plasma electrolytic oxidation (PEO) methods. In this work, we performed the in vitro and in vivo examinations of these coated implants using L-929 cell line and rabbit animal model. The in vitro study confirmed the higher cytocompatibility of the coated implants compare to the uncoated ones. For the in vivo experiment, the rod samples were implanted into the greater trochanter of rabbits and monitored for two months. The results indicated a noticeable biocompatibility improvement of the coated implants which includes slower implant weight loss, reduction in Mg ion released from the coated samples in the blood plasma, lower release of hydrogen bubbles, increase in the amount of bone formation and ultimately lower bone inflammation after the surgery according to the histological images. Our data exemplifies that the proper surface treatment of the magnesium implants can improve their biocompatibility under physiological conditions to make them applicable in clinical uses. © 2014 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103A: 1798-1808, 2015.

  13. In vivo neurochemistry with emission tomography and magnetic resonance spectroscopy: clinical applications.

    PubMed

    Del Sole, Angelo; Gambini, Anna; Falini, Andrea; Lecchi, Michela; Lucignani, Giovanni

    2002-10-01

    The assessment of neurochemical processes in vivo has received much attention in the past decade as techniques such as positron or single photon emission tomography (PET and SPET), and magnetic resonance spectroscopy (MRS) have become more available. With PET and SPET, basic processes, such as blood flow and oxygen or glucose metabolism, can be regionally assessed, along with more specific functions such as the production, release, and reuptake of neurotransmitters and their occupancy of specific receptors. At the same time, MRS can reveal changes in concentration of several hydrogenate compounds in the brain. All these methods have been extensively applied for research in neurology, and some applications have reached the clinical level, namely for the study of degenerative diseases, motor-neuron diseases, movement disorders, cerebrovascular diseases, and epilepsy. This article focuses on the most relevant information that can be obtained with these complementary techniques to help clinicians in the assessment of neurological diseases.

  14. In vivo application of a small molecular weight antifungal protein of Penicillium chrysogenum (PAF)

    SciTech Connect

    Palicz, Zoltán; Jenes, Ágnes; Gáll, Tamás; Miszti-Blasius, Kornél; Kollár, Sándor; Kovács, Ilona; Emri, Miklós; Márián, Teréz; Leiter, Éva; Pócsi, István; Csősz, Éva; Kalló, Gergő; Hegedűs, Csaba; Virág, László; Csernoch, László; Szentesi, Péter

    2013-05-15

    The antifungal protein of Penicillium chrysogenum (PAF) inhibits the growth of important pathogenic filamentous fungi, including members of the Aspergillus family and some dermatophytes. Furthermore, PAF was proven to have no toxic effects on mammalian cells in vitro. To prove that PAF could be safely used in therapy, experiments were carried out to investigate its in vivo effects. Adult mice were inoculated with PAF intranasally in different concentrations, up to 2700 μg·kg{sup −1} daily, for 2 weeks. Even at the highest concentration – a concentration highly toxic in vitro for all affected molds – used, animals neither died due to the treatment nor were any side effects observed. Histological examinations did not find pathological reactions in the liver, in the kidney, and in the lungs. Mass spectrometry confirmed that a measurable amount of PAF was accumulated in the lungs after the treatment. Lung tissue extracts from PAF treated mice exerted significant antifungal activity. Small-animal positron emission tomography revealed that neither the application of physiological saline nor that of PAF induced any inflammation while the positive control lipopolysaccharide did. The effect of the drug on the skin was examined in an irritative dermatitis model where the change in the thickness of the ears following PAF application was found to be the same as in control and significantly less than when treated with phorbol-12-myristate-13-acetate used as positive control. Since no toxic effects of PAF were found in intranasal application, our result is the first step for introducing PAF as potential antifungal drug in therapy. - Highlights: • PAF, the antifungal protein of Penicillium chrysogenum, was not toxic in mice. • Its intranasal application didn't induce pathological reactions in the lung. • PAF retained its antifungal activity in lung extracts. • Its application on the skin did not cause inflammation.

  15. A computational atlas of the hippocampal formation using ex vivo, ultra-high resolution MRI: Application to adaptive segmentation of in vivo MRI.

    PubMed

    Iglesias, Juan Eugenio; Augustinack, Jean C; Nguyen, Khoa; Player, Christopher M; Player, Allison; Wright, Michelle; Roy, Nicole; Frosch, Matthew P; McKee, Ann C; Wald, Lawrence L; Fischl, Bruce; Van Leemput, Koen

    2015-07-15

    Automated analysis of MRI data of the subregions of the hippocampus requires computational atlases built at a higher resolution than those that are typically used in current neuroimaging studies. Here we describe the construction of a statistical atlas of the hippocampal formation at the subregion level using ultra-high resolution, ex vivo MRI. Fifteen autopsy samples were scanned at 0.13 mm isotropic resolution (on average) using customized hardware. The images were manually segmented into 13 different hippocampal substructures using a protocol specifically designed for this study; precise delineations were made possible by the extraordinary resolution of the scans. In addition to the subregions, manual annotations for neighboring structures (e.g., amygdala, cortex) were obtained from a separate dataset of in vivo, T1-weighted MRI scans of the whole brain (1mm resolution). The manual labels from the in vivo and ex vivo data were combined into a single computational atlas of the hippocampal formation with a novel atlas building algorithm based on Bayesian inference. The resulting atlas can be used to automatically segment the hippocampal subregions in structural MRI images, using an algorithm that can analyze multimodal data and adapt to variations in MRI contrast due to differences in acquisition hardware or pulse sequences. The applicability of the atlas, which we are releasing as part of FreeSurfer (version 6.0), is demonstrated with experiments on three different publicly available datasets with different types of MRI contrast. The results show that the atlas and companion segmentation method: 1) can segment T1 and T2 images, as well as their combination, 2) replicate findings on mild cognitive impairment based on high-resolution T2 data, and 3) can discriminate between Alzheimer's disease subjects and elderly controls with 88% accuracy in standard resolution (1mm) T1 data, significantly outperforming the atlas in FreeSurfer version 5.3 (86% accuracy) and

  16. Fluorescence spectroscopy of gastrointestinal tumors: in vitro studies and in vivo clinical applications

    NASA Astrophysics Data System (ADS)

    Angelova, L.; Borisova, E.; Zhelyazkova, Al.; Keremedchiev, M.; Vladimirov, B.; Avramov, L.

    2013-11-01

    The limitations of standard endoscopy for detection and evaluation of cancerous changes in the gastrointestinal tract (GIT) are significant challenges and initiate development of new diagnostic modalities. Therefore many spectral and optical techniques are applied recently into the clinical practice for obtaining qualitatively and quantitatively new data from gastrointestinal neoplasia with different levels of clinical applicability and diagnostic success. Fluorescence imaging has been one of the most promising technologies in this area. The technique is very topical with its practical application in intra-operative, image-guided resection of tumors, because it permits minimal surgery intervention and friendly therapeutic conditions. The investigations presented here are based on in vitro measurements of excitation-emission matrices (EEM) for GIT neoplasia and in vivo measurements in the frames of initial clinical trial for tumor fluorescence spectra detection, applied for introduction of spectroscopic diagnostic system for optical biopsy of GIT tumors in the daily clinical practice of the University Hospital "Queen Jiovanna - ISUL"- Sofia. Autofluorescence and exogenous fluorescence signals are detected from normal mucosa, inflammation, dysphasia and carcinoma and main spectral features are evaluated. The systems and methods developed for diagnosis and monitoring could open new dimensions in diagnostic and real-time tumor resection. This will make the entire procedure more personal, patient friendly and effective and will help for further understanding of the tumor nature.

  17. Topical application of solubilized Reseda luteola extract reduces ultraviolet B-induced inflammation in vivo.

    PubMed

    Casetti, F; Jung, W; Wölfle, U; Reuter, J; Neumann, K; Gilb, B; Wähling, A; Wagner, S; Merfort, I; Schempp, C M

    2009-09-04

    We investigated the skin tolerance and anti-inflammatory potential of a nanoparticular solubilisate of a luteolin-rich Reseda extract (s-RE) in two independent studies in vivo. Reseda luteola extract containing 40% flavonoids was solubilized with polysorbate, resulting in product micelles with a diameter of 10 (+/-1.5)nm. Standardized inflammation was induced by irradiating test areas on the back of healthy volunteers with defined doses of ultraviolet B (UVB). In the first study different concentrations of s-RE were tested in 10 volunteers to evaluate dose-dependency of anti-inflammatory effects of s-RE. In the second randomized, double-blind, placebo-controlled study a defined concentration of s-RE (2.5%w/w) was tested in 40 volunteers in comparison to the vehicle (glycerol) and hydrocortisone (1%w/w). s-RE dose-dependently reduced UVB-induced erythema when applied 30 min before irradiation. To a lesser extent, topical application of s-RE after irradiation also reduced UVB-induced erythema. s-RE was as effective as hydrocortisone, whereas the vehicle had no effect. Occlusive application of s-RE on non-irradiated test sites did not cause any skin irritation. Due to excellent skin tolerance combined with potent anti-inflammatory properties s-RE bears potential especially for the prevention but also for the treatment of inflammatory skin conditions such as UV-induced erythema.

  18. Application of FRET Technology to the In Vivo Evaluation of Therapeutic Nucleic Acids (ANTs)

    NASA Astrophysics Data System (ADS)

    Benítez-Hess, María Luisa; Alvarez-Salas, Luis Marat

    2007-02-01

    Developing applications for therapeutic nucleic acids (TNAs) (i.e. ribozymes, antisense oligodeoxynucleotides (AS-ODNs), siRNA and aptamers) requires a reporter system designed to rapidly evaluate their in vivo effect. To this end we designed a reporter system based on the fluorescence resonance energy transfer (FRET) engineered to release the FRET effect produced by two green fluorescent protein (GFP) variants linked by a TNA target site. Because the FRET effect occurs instantaneously when two fluorophores are very close to each other (>100nm) stimulating emission of the acceptor fluorophore by the excitation of the donor fluorophore it has been widely use to reveal interactions between molecules. The present system (FRET2) correlates the FRET effect with the in vivo activity of distinct types of TNAs based on a model consisting of RNA from human papillomavirus type 16 (HPV-16) previously shown accessible to TNAs. HPV-16 is the most common papillomavirus associated with cervical cancer, the leading cause of death by cancer in México. The FRET2 system was first tested in vitro and then used in bacteria in which transcription is linked to translation allowing controlled expression and rapid evaluation of the FRET2 protein. To assure accessibility of the target mRNA to TNAs, the FRET2 mRNA was probed by RNaseH assays prior FRET testing. The fluorescence features of the FRET2 system was tested with different FRET-producing GFP donor-acceptor pairs leading to selection of green (donor) and yellow (acceptor) variants of GFP as the most efficient. Modifications in aminoacid composition and linker length of the target sequence did not affect FRET efficiency. In vivo AS-ODN-mediated destruction of the chimerical FRET2 reporter mRNA resulted in the recovery of GFP fluorescent spectrum in a concentration and time dependent manner. Reported anti-HPV ribozymes were also tested with similar results. Therefore, we conclude that the FRET effect can be a useful tool in the

  19. In vivo applications of electrospun tissue-engineered vascular grafts: a review.

    PubMed

    Rocco, Kevin A; Maxfield, Mark W; Best, Cameron A; Dean, Ethan W; Breuer, Christopher K

    2014-12-01

    There is great clinical demand for synthetic vascular grafts with improved long-term efficacy. The ideal vascular conduit is easily implanted, nonthrombogenic, biocompatible, resists aneurysmal dilatation, and ultimately degrades or is assimilated as the patient remodels the graft into tissue resembling native vessel. The field of vascular tissue engineering offers an opportunity to design the ideal synthetic graft, and researchers have evaluated a variety of methods and materials for use in graft construction. Electrospinning is one method that has received considerable attention within tissue engineering for constructing so-called tissue scaffolds. Tissue scaffolds are temporary, porous structures which are commonly composed of bioresorbable polymers that promote native tissue ingrowth and have degradation kinetics compatible with a patient's rate of extracellular matrix production in order to successfully transit from synthetic conduits into neovessels. In this review, we summarize the history of tissue-engineered vascular grafts (TEVG), focusing on scaffolds generated by the electrospinning process, and discuss in vivo applications. We review the materials commonly employed in this approach and the preliminary results after implantation in animal models in order to gauge clinical viability of the electrospinning process for TEVG construction. Scientists have studied electrospinning technology for decades, but only recently has it been orthotopically evaluated in animal models such as TEVG. Advantages of electrospun TEVG include ease of construction, favorable cellular interactions, control of scaffold features such as fiber diameter and pore size, and the ability to choose from a variety of polymers possessing a range of mechanical and chemical properties and degradation kinetics. Given its advantages, electrospinning technology merits investigation for use in TEVG, but an emphasis on long-term in vivo evaluation is required before its role in clinical vascular

  20. Novel biomaterial for transdermal application: in vitro and in vivo characterization.

    PubMed

    Mundada, A S; Avari, J G

    2011-08-01

    The objective of the present study was to evaluate a novel film forming biomaterial for its potential application in the preparation of unilaminate transdermal adhesive matrix systems. The biomaterial, Damar Batu (DB), was tried alone and in combination with Eudragit RL100 as a matrixing agent in the preparation of transdermal patches. Developed transdermal patches of Diltiazem hydrochloride (DH) were evaluated for thickness uniformity, weight uniformity, folding endurance and drug content. USP dissolution apparatus V was used for in vitro drug release studies. Modified Franz diffusion cell used for permeation study using excised human cadaver skin. Total 6 formulations were developed and on the basis of in vitro drug release and in vitro skin permeation profile F5 composed of DB: Eudragit RL100 (60:40) and carrying 20 %w/w DH was selected as an optimized formulation for in vivo study. The in vivo study results showed that F5 achieved the Cmax of about 269.76 ± 1.52 ng/mL in 6 h and sustained the release of the drug till 24 h. The skin irritation study results proved that the novel biomaterial is non-sensitizing and non-irritating. Drug-polymer interaction study carried out to check the compatibility of drug and polymer showed the intactness of the drug in the formulation proving the compatibility of the polymer. It can be proposed from the outcome of the present study that by applying suitable adhesive layer and backing membrane, DB: Eudragit RL100 (60:40) transdermal patches can be of potential therapeutic use.

  1. Application of NIR fluorescent markers to quantify expression level of HER2 receptors in carcinomas in vivo

    NASA Astrophysics Data System (ADS)

    Chernomordik, Victor; Hassan, Moinuddin; Lee, Sang Bong; Zielinski, Rafal; Capala, Jacek; Gandjbakhche, Amir

    2010-02-01

    HER2 overexpression has been associated with a poor prognosis and resistance to therapy in breast cancer patients. However, quantitative estimates of this important characteristic have been limited to ex vivo ELISA essays of tissue biopsies and/or PET. We develop a novel approach in optical imaging, involving specific probes, not interfering with the binding of the therapeutic agents, thus, excluding competition between therapy and imaging. Affibody-based molecular probes seem to be ideal for in vivo analysis of HER2 receptors using near-infrared optical imaging. Fluorescence intensity distributions, originating from specific markers in the tumor area, can reveal the corresponding fluorophore concentration. We use temporal changes of the signal from a contrast agent, conjugated with HER2-specific Affibody as a signature to monitor in vivo the receptors status in mice with different HER2 over-expressed tumor models. Kinetic model, incorporating saturation of the bound ligands in the tumor area due to HER2 receptor concentration, is suggested to analyze relationship between tumor cell characteristics, i.e., HER2 overexpression, obtained by traditional ("golden standard") ex vivo methods (ELISA), and parameters, estimated from the series of images in vivo. Observed correlation between these parameters and HER2 overexpression substantiates application of our approach to quantify HER2 concentration in vivo.

  2. Synthesis and surface modification of magnetic nanoparticles for in vivo biomedical applications

    NASA Astrophysics Data System (ADS)

    Sun, Conroy Ghin Chee

    enhancement both in vitro and in vivo in MRI experiments. The successful application of such smart molecular imaging probes will have a significant clinical impact on improved diagnosis and treatment of malignant tumors.

  3. Multiphoton fluorescence recovery after photobleaching: Advancements for novel in vivo applications

    NASA Astrophysics Data System (ADS)

    Sullivan, Kelley Diane

    Multiphoton fluorescence recovery after photobleaching (MP-FRAP) is a laser microscopy technique used to probe the transport properties of macromolecules in biological systems. MP-FRAP utilizes two-photon fluorescence and photobleaching to produce a three-dimensionally resolved diffusion coefficient for an ensemble of molecules in the region of the two-photon focal volume. This thesis describes two fundamental improvements to the MP-FRAP technique, which are vital steps to enable MP-FRAP to be applied to the complex in vivo environment. In Chapter 1, we lay the groundwork for our discussion of these advancements by introducing the MP-FRAP technique and the physics upon which it is based. We begin with a description of fluorescence and diffusion and discuss their importance in biomedical research. Next, we describe how two-photon fluorescence and photobleaching are applied to a diffusing system to measure the diffusion coefficient via fluorescence recovery after photobleaching (FRAP). Then, we take the reader through the evolution of FRAP, which leads to the application of two- photon fluorescence and photobleaching to produce MP-FRAP. Along the way, we highlight applications and advancements of the FRAP techniques, and introduce fluorescence correlation spectroscopy, a popular complement to FRAP. In Chapter 2, we collect the experimental methods for the studies presented in Chapters 3 and 4. We begin with an in-depth discussion of our work to build and troubleshoot our MP-FRAP apparatus, followed by a detailed description of our data analysis protocol. Next, we delve into the specific methods for producing computer generated data and fits, as well as in vitro and in vivo experimental data, for our work in Chap. 3 on improving MP-FRAP to measure diffusion in the presence of convective flow. We end with a description of the Monte Carlo algorithm we developed for our work in Chap. 4 to model diffusion and multiphoton fluorescence recovery after photobleaching in the

  4. In vivo selection to identify bacterial strains with enhanced ecological performance in synbiotic applications.

    PubMed

    Krumbeck, Janina A; Maldonado-Gomez, María X; Martínez, Inés; Frese, Steven A; Burkey, Thomas E; Rasineni, Karuna; Ramer-Tait, Amanda E; Harris, Edward N; Hutkins, Robert W; Walter, Jens

    2015-04-01

    One strategy for enhancing the establishment of probiotic bacteria in the human intestinal tract is via the parallel administration of a prebiotic, which is referred to as a synbiotic. Here we present a novel method that allows a rational selection of putative probiotic strains to be used in synbiotic applications: in vivo selection (IVS). This method consists of isolating candidate probiotic strains from fecal samples following enrichment with the respective prebiotic. To test the potential of IVS, we isolated bifidobacteria from human subjects who consumed increasing doses of galactooligosaccharides (GOS) for 9 weeks. A retrospective analysis of the fecal microbiota of one subject revealed an 8-fold enrichment in Bifidobacterium adolescentis strain IVS-1 during GOS administration. The functionality of GOS to support the establishment of IVS-1 in the gastrointestinal tract was then evaluated in rats administered the bacterial strain alone, the prebiotic alone, or the synbiotic combination. Strain-specific quantitative real-time PCR showed that the addition of GOS increased B. adolescentis IVS-1 abundance in the distal intestine by nearly 2 logs compared to rats receiving only the probiotic. Illumina 16S rRNA sequencing not only confirmed the increased establishment of IVS-1 in the intestine but also revealed that the strain was able to outcompete the resident Bifidobacterium population when provided with GOS. In conclusion, this study demonstrated that IVS can be used to successfully formulate a synergistic synbiotic that can substantially enhance the establishment and competitiveness of a putative probiotic strain in the gastrointestinal tract.

  5. In Vivo Selection To Identify Bacterial Strains with Enhanced Ecological Performance in Synbiotic Applications

    PubMed Central

    Krumbeck, Janina A.; Maldonado-Gomez, María X.; Martínez, Inés; Frese, Steven A.; Burkey, Thomas E.; Rasineni, Karuna; Ramer-Tait, Amanda E.; Harris, Edward N.; Hutkins, Robert W.

    2015-01-01

    One strategy for enhancing the establishment of probiotic bacteria in the human intestinal tract is via the parallel administration of a prebiotic, which is referred to as a synbiotic. Here we present a novel method that allows a rational selection of putative probiotic strains to be used in synbiotic applications: in vivo selection (IVS). This method consists of isolating candidate probiotic strains from fecal samples following enrichment with the respective prebiotic. To test the potential of IVS, we isolated bifidobacteria from human subjects who consumed increasing doses of galactooligosaccharides (GOS) for 9 weeks. A retrospective analysis of the fecal microbiota of one subject revealed an 8-fold enrichment in Bifidobacterium adolescentis strain IVS-1 during GOS administration. The functionality of GOS to support the establishment of IVS-1 in the gastrointestinal tract was then evaluated in rats administered the bacterial strain alone, the prebiotic alone, or the synbiotic combination. Strain-specific quantitative real-time PCR showed that the addition of GOS increased B. adolescentis IVS-1 abundance in the distal intestine by nearly 2 logs compared to rats receiving only the probiotic. Illumina 16S rRNA sequencing not only confirmed the increased establishment of IVS-1 in the intestine but also revealed that the strain was able to outcompete the resident Bifidobacterium population when provided with GOS. In conclusion, this study demonstrated that IVS can be used to successfully formulate a synergistic synbiotic that can substantially enhance the establishment and competitiveness of a putative probiotic strain in the gastrointestinal tract. PMID:25616794

  6. Experimental model to measure the increase of dental pulp temperature in vivo during laser application

    NASA Astrophysics Data System (ADS)

    Nicola, Ester M. D.; Junqueira, Silvio L. M.; Busato, Mara S.

    1994-09-01

    Carbon dioxide laser has been used in dental surgery. The existence of healthy teeth, which have pulp vitality needing to be preserved, is observed in a great number of cases. In this work we describe an experimental model which provides the measurement of temperature in pulp chamber `in vivo,' during oral surgeries in which the CO2 laser beam is applied to gingival tissue. The problems met during the search for the best way to place the thermal probe regarding the diameter and depth of pulp chamber and the thickness of the tissue layer formed by gum and maxillary bone are discussed. We use a thermocouple placed in the pulp chamber of superior canine teeth in dogs. After that, the probe was also placed between gum and dental root. Since the temperature at gingival surface was known, it was easy to determine the rise in temperature at pulp chamber and also to observe the thermal gradient from gum to tissue to bone, thus avoiding pulp damage during laser applications.

  7. Segmented surface coil resonator for in vivo EPR applications at 1.1 GHz

    NASA Astrophysics Data System (ADS)

    Petryakov, Sergey; Samouilov, Alexandre; Chzhan-Roytenberg, Michael; Kesselring, Eric; Sun, Ziqi; Zweier, Jay L.

    2009-05-01

    A four-loop segmented surface coil resonator (SSCR) with electronic frequency and coupling adjustments was constructed with 18 mm aperture and loading capability suitable for in vivo Electron Paramagnetic Resonance (EPR) spectroscopy and imaging applications at L-band. Increased sample volume and loading capability were achieved by employing a multi-loop three-dimensional surface coil structure. Symmetrical design of the resonator with coupling to each loop resulted in high homogeneity of RF magnetic field. Parallel loops were coupled to the feeder cable via balancing circuitry containing varactor diodes for electronic coupling and tuning over a wide range of loading conditions. Manually adjusted high Q trimmer capacitors were used for initial tuning with subsequent tuning electronically controlled using varactor diodes. This design provides transparency and homogeneity of magnetic field modulation in the sample volume, while matching components are shielded to minimize interference with modulation and ambient RF fields. It can accommodate lossy samples up to 90% of its aperture with high homogeneity of RF and modulation magnetic fields and can function as a surface loop or a slice volume resonator. Along with an outer coaxial NMR surface coil, the SSCR enabled EPR/NMR co-imaging of paramagnetic probes in living rats to a depth of 20 mm.

  8. Retinal electrophysiology for toxicology studies: applications and limits of ERG in animals and ex vivo recordings.

    PubMed

    Rosolen, Serge Georges; Kolomiets, Bogdan; Varela, Oscar; Picaud, Serge

    2008-06-01

    Assessing retinal drug toxicity is becoming increasingly important as different molecules are now developed for the treatment of neurodegenerative diseases and vascular disorders. In pharmacology and toxicology, the electroretinogram (ERG) and the multielectrode array (MEA) recording techniques can be used to quantify the possible side effects of retino-active xenobiotics. Toxicity testing requires the use of rodent as well as non-rodent models for extrapolation to the human model when determining risk and safety. Animal species differ in their retinal anatomo-physiology: most rodents used in toxicology studies are essentially nocturnal species, whereas the non-rodent laboratory species normally used (e.g. dogs, pigs and monkeys) are diurnal. The ratio between the photoreceptor populations which varies from species to species, should be considered when designing the experiment protocol and the interpretation. The described ERG procedures are designed to comply with all applicable good laboratory practice standards. Use of these procedures should yield an acceptable level of intra- and inter-subject variability for compiling a historical database, and for detecting possible retinal toxicity in animal studies. They could therefore be used as specific and standardized tools for screening of potential retinotoxic molecules in drug discovery and development in order to compare methods and results with those obtained in human electrophysiological assessments. Recording of ganglion cell light responses on ex vivo retina with the MEA technique can further demonstrate how retino-active xenobiotics affect retinal visual information processing by eliminating potential obstacles related to bioavailability and blood barrier permeability.

  9. Skin imaged by femtosecond laser irradiation: a risk assessment for in vivo applications

    NASA Astrophysics Data System (ADS)

    Fischer, F.; Volkmer, B.; Puschmann, S.; Greinert, R.; Breitbart, W.; Kiefer, J.; Wepf, R.

    2006-04-01

    During the last couple of years new imaging techniques using femtosecond lasers (fs-lasers) in the near infrared spectral range evolved for a variety of in vitro applications. We wanted to know, whether fs-lasers have a non-invasive imaging potential for in vivo applications for human skin. So far, little is known about possible risks of this irradiation type. To estimate the risk of irradiation damage in human skin we used a "biological dosimeter" in this investigation. We irradiated fresh human skin samples with both an fs-laser and a solar simulator (UV-source) for comparison. DNA damage introduced in the epidermis was evaluated using fluorescent antibodies against cyclobutane-pyrimidin-dimers (CPDs) in combination with immuno-fluorescence image analysis. Various fs-irradiation regimes were evaluated differing in laser power and step width of horizontal irradiation scans. When using 15 mW or 30 mW fs-laser power combined with horizontal irradiation scans applied every 5 mm in depth around the epidermal-dermal junction no induction of CPDs was found. However, induction of CPDs could be seen using 60 mW laser power and 5 μm step width. Narrowing the step width to 1 mm and using increasing laser power (up to 35 mW) from the surface of the skin to the epidermis led to CPD formation, too. Quantitative comparison of CPD production at various laser regimes with CPD production using a solar simulator was done. We could show that the number of CPDs formed by the 60 mW laser irradiation regime is comparable to an UV-irradiation giving 0.6 MED (minimal erythemal dose). The smaller step width laser irradiation regime (1 μm step width and up to 35 mW) was comparable to a UV-irradiation regime resulting in 0.45 MED.

  10. Combining whispering gallery mode lasers and microstructured optical fibers for in-vivo biosensing applications

    NASA Astrophysics Data System (ADS)

    François, A.; Rowland, K. J.; Reynolds, T.; Nicholls, S. J.; Monro, T. M.

    2013-10-01

    Whispering Gallery Modes (WGMs) have been widely studied for the past 20 years for various applications, including biological sensing. While the different WGM-based sensing approaches reported in the literature enable useful sensor characteristics, at present this technology is not yet mature, mainly for practical reasons. Our work has been focused on developing a simple, yet efficient, WGM-based sensing platform capable of being used as a dip sensor for in-vivo biosensing applications. We recently demonstrated that a dye-doped polymer microresonator, supporting WGMs, positioned onto the tip of a suspended core Microstructured Optical Fiber can be used as a dip sensor. In this architecture, the resonator is located on an air hole next to the fiber core at the fiber's tip, enabling a significant portion of the sphere to overlap with the guided light emerging from the fiber tip. This architecture offers significant benefits that have never been reported in the literature in terms of radiation efficiency, compared to the standard freestanding resonators, which arise from breaking the symmetry of the resonator. In addition to providing the remote excitation and collection of the WGMs' signal, the fiber also allows easy manipulation of the microresonator and the use this sensor in a dip sensing architecture, alleviating the need for a complex microfluidic interface. Here, we present our recent results on the microstructured fiber tip WGM-based sensor, including its lasing behavior and enhancement of the radiation efficiency as a function of the position of the resonator on the fiber tip. We also show that this platform can be used for clinical diagnostics and applying this technology to the detection of Troponin T, an acute myocardial infarction biomarker, down to a concentration of 7.4 pg/mL.

  11. PVP- coated naringenin nanoparticles for biomedical applications - In vivo toxicological evaluations.

    PubMed

    Kumar, R Pradeep; Abraham, Annie

    2016-09-25

    Naringenin (NAR) is one of the naturally occurring flavonoids found in citrus fruits and exerts a wide variety of pharmacological activities. The clinical relevance of naringenin is limited by its low solubility and minimal bioavailability, owing to its largely hydrophobic ring structure. The aim of the present study is to develop a novel naringenin nanoparticle system (NAR NP) using simple nanoprecipitation technique with polyvinylpyrrolidone (PVP) as the hydrophilic carrier. The synthesized nanoparticles were characterized using XRD, FTIR, SEM and EDX. The characterization study revealed the nanoscale properties and the interactions between NAR and PVP. In vivo toxicological evaluations were carried out at various doses (1, 5, 10 & 50 mg/kg body wt) in male Sprague-Dawley rats in comparison with silver nanoparticle (AgNP) at toxic concentration (50 mg/kg body wt). The altered hepatotoxicity markers, hematology parameters and antioxidant defense system were observed in AgNP- treated rats. But NAR NP - treated rats did not show any biochemical alterations and improved the antioxidant defense indices when compared to control group, by virtue of the pharmacological properties exerted by NAR. The modulatory effect of NAR NP over inflammatory and stress signaling cascades were confirmed by the normalized mRNA expressions of NF-κB, TNF-α and IL-6. The histopathological analysis of liver, kidney and heart reinforce our findings. These studies provide preliminary answers to some of the key biological issues raised over the use and safety of nanoparticles for diagnostic and therapeutic applications. Consequently, we suggest that the safe NAR NP can be used to reduce the dosage of NAR, improve its bioavailability and merits further investigation for therapeutic applications.

  12. In vivo pH monitoring using boron doped diamond microelectrode and silver needles: application to stomach disorder diagnosis.

    PubMed

    Fierro, Stéphane; Seishima, Ryo; Nagano, Osamu; Saya, Hideyuki; Einaga, Yasuaki

    2013-11-19

    This study presents the in vivo electrochemical monitoring of pH using boron doped diamond (BDD) microelectrode and silver needles for potential application in medical diagnosis. Accurate calibration curve for pH determination were obtained through in vitro electrochemical measurements. The increase induced in stomach pH by treatment with pantoprazole was used to demonstrate that it is possible to monitor the pH in vivo using the simple and noninvasive system proposed herein. Using the results of the in vivo and in vitro experiments, a quantitative analysis of the increase in stomach pH is also presented. It is proposed that the catheter-free pH monitoring system presented in this study could be potentially employed in any biological environment.

  13. Development of HiLo Microscope and its use in In-Vivo Applications

    NASA Astrophysics Data System (ADS)

    Patel, Shreyas J.

    The functionality of achieving optical sectioning in biomedical research is invaluable as it allows for visualization of a biological sample at different depths while being free of background scattering. Most current microscopy techniques that offer optical sectioning, unfortunately, require complex instrumentation and thus are generally costly. HiLo microscopy, on the other hand, offers the same functionality and advantage at a relatively low cost. Hence, the work described in this thesis involves the design, build, and application of a HiLo microscope. More specifically, a standalone HiLo microscope was built in addition to implementing HiLo microscopy on a standard fluorescence microscope. In HiLo microscopy, optical sectioning is achieved by acquiring two different types of images per focal plane. One image is acquired under uniform illumination and the other is acquired under speckle illumination. These images are processed using an algorithm that extracts in-focus information and removes features and glare that occur as a result of background fluorescence. To show the benefits of the HiLo microscopy, several imaging experiments on various samples were performed under a HiLo microscope and compared against a traditional fluorescence microscope and a confocal microscope, which is considered the gold standard in optical imaging. In-vitro and ex-vivo imaging was performed on a set of pollen grains, and optically cleared mouse brain and heart slices. Each of these experiments showed great reduction in background scattering at different depths under HiLo microscopy. More importantly, HiLo imaging of optically cleared heart slice demonstrated emergence of different vasculature at different depths. Reduction of out-of-focus light increased the spatial resolution and allowed better visualization of capillary vessels. Furthermore, HiLo imaging was tested in an in-vivo model of a rodent dorsal window chamber model. When imaging the same sample under confocal microscope

  14. Three-photon luminescence of gold nanorods and its applications for high contrast tissue and deep in vivo brain imaging.

    PubMed

    Wang, Shaowei; Xi, Wang; Cai, Fuhong; Zhao, Xinyuan; Xu, Zhengping; Qian, Jun; He, Sailing

    2015-01-01

    Gold nanoparticles can be used as contrast agents for bio-imaging applications. Here we studied multi-photon luminescence (MPL) of gold nanorods (GNRs), under the excitation of femtosecond (fs) lasers. GNRs functionalized with polyethylene glycol (PEG) molecules have high chemical and optical stability, and can be used as multi-photon luminescent nanoprobes for deep in vivo imaging of live animals. We have found that the depth of in vivo imaging is dependent upon the transmission and focal capability of the excitation light interacting with the GNRs. Our study focused on the comparison of MPL from GNRs with two different aspect ratios, as well as their ex vivo and in vivo imaging effects under 760 nm and 1000 nm excitation, respectively. Both of these wavelengths were located at an optically transparent window of biological tissue (700-1000 nm). PEGylated GNRs, which were intravenously injected into mice via the tail vein and accumulated in major organs and tumor tissue, showed high image contrast due to distinct three-photon luminescence (3PL) signals upon irradiation of a 1000 nm fs laser. Concerning in vivo mouse brain imaging, the 3PL imaging depth of GNRs under 1000 nm fs excitation could reach 600 μm, which was approximately 170 μm deeper than the two-photon luminescence (2PL) imaging depth of GNRs with a fs excitation of 760 nm.

  15. Three-Photon Luminescence of Gold Nanorods and Its Applications for High Contrast Tissue and Deep In Vivo Brain Imaging

    PubMed Central

    Wang, Shaowei; Xi, Wang; Cai, Fuhong; Zhao, Xinyuan; Xu, Zhengping; Qian, Jun; He, Sailing

    2015-01-01

    Gold nanoparticles can be used as contrast agents for bio-imaging applications. Here we studied multi-photon luminescence (MPL) of gold nanorods (GNRs), under the excitation of femtosecond (fs) lasers. GNRs functionalized with polyethylene glycol (PEG) molecules have high chemical and optical stability, and can be used as multi-photon luminescent nanoprobes for deep in vivo imaging of live animals. We have found that the depth of in vivo imaging is dependent upon the transmission and focal capability of the excitation light interacting with the GNRs. Our study focused on the comparison of MPL from GNRs with two different aspect ratios, as well as their ex vivo and in vivo imaging effects under 760 nm and 1000 nm excitation, respectively. Both of these wavelengths were located at an optically transparent window of biological tissue (700-1000 nm). PEGylated GNRs, which were intravenously injected into mice via the tail vein and accumulated in major organs and tumor tissue, showed high image contrast due to distinct three-photon luminescence (3PL) signals upon irradiation of a 1000 nm fs laser. Concerning in vivo mouse brain imaging, the 3PL imaging depth of GNRs under 1000 nm fs excitation could reach 600 μm, which was approximately 170 μm deeper than the two-photon luminescence (2PL) imaging depth of GNRs with a fs excitation of 760 nm. PMID:25553113

  16. Selective perturbation of in vivo linear energy transfer using high- Z vaginal applicators for Cf-252 brachytherapy

    NASA Astrophysics Data System (ADS)

    Rivard, M. J.; Evans, K. E.; Leal, L. C.; Kirk, B. L.

    2004-01-01

    Californium-252 ( 252Cf) brachytherapy sources emit both neutrons and photons, and have the potential to vastly improve the current standard-of-practice for brachytherapy. While hydrogenous materials readily attenuate the 252Cf fission energy neutrons, high- Z materials are utilized to attenuate the 252Cf gamma-rays. These differences in shielding materials may be exploited when treating with a vaginal applicator to possibly improve patient survival through perturbation of the in vivo linear energy transfer radiation.

  17. Biosensors based on inorganic nanoparticles with biomimetic properties: Biomedical applications and in vivo cytotoxicity measurements

    NASA Astrophysics Data System (ADS)

    Ispas, Cristina R.

    . This work introduces a new generic approach of improving the sensitivity of oxidase-based enzymatic assays and indicates that ceria and its mixture with other metal oxide nanoparticles could be used to minimize the problems associated with variations of the oxygen. These materials have great potential in bioanalytical and biotechnological applications and offer great opportunities for development of implantable sensing devices for in vivo and in vitro monitoring of analytes of clinical relevance. Additionally, this thesis evaluates the toxicity of different metal and metal oxide nanoparticles by using zebrafish embryos as a toxicological target. Because of their similarities with other vertebrates, rapid development and low cost, zebrafish embryos are ideal animal models for probing toxicological effects of engineered nanomaterials. Among the nanomaterials tested, nickel nanoparticles were characterized by high toxicity and induced delayed development and morphological malformations, while metal oxides nanoparticles (i.e. ceria nanoparticles) had no toxic effects.

  18. Sustained Growth of the Ex Vivo Ablation Zones' Critical Short Axis Using Gas-cooled Radiofrequency Applicators

    SciTech Connect

    Rempp, Hansjoerg; Scharpf, Marcus; Voigtlaender, Matthias; Schraml, Christina; Schmidt, Diethard; Fend, Falko; Claussen, Claus D.; Enderle, Markus D.; Pereira, Philippe L.; Clasen, Stephan

    2011-02-15

    Purpose: To evaluate the ablation zones created with a gas-cooled bipolar radiofrequency applicator performed on ex vivo bovine liver tissue. Materials and Methods: A total of 320 ablations with an internally gas-cooled bipolar radiofrequency applicator were performed on fresh ex vivo bovine liver tissue, varying the ablation time (5, 10, 15, and 20 min), power (20, 30, 40, and 50 W), and gas pressure of the CO{sub 2} used for cooling (585, 600, 615, 630, 645 psi), leading to a total of 80 different parameter combinations. Size and shape of the white coagulation zone were assessed. Results: The largest complete ablation zone was achieved after 20 min of implementing 50 W and 645 psi, resulting in a short axis of mean 46 {+-} 1 mm and a long axis of 56 {+-} 2 mm (mean {+-} standard deviation). Short-axis diameters increased between 5 and 20 min of ablation time at 585 psi (increase of the short axis was 45% at 30 W, 29% at 40 W, and 39% at 50 W). This increase was larger at 645 psi (113% at 30 W, 67% at 40 W, and 70% at 50 W). Macroscopic assessment and NADH (nicotinamide adenine dinucleotide) staining revealed incompletely ablated tissue along the needle track in 18 parameter combinations including low-power settings (20 and 30 W) and different cooling levels and ablation times. Conclusion: Gas-cooled radiofrequency applicators increase the short-axis diameter of coagulation in an ex vivo setting if appropriate parameters are selected.

  19. Windows on the human body--in vivo high-field magnetic resonance research and applications in medicine and psychology.

    PubMed

    Moser, Ewald; Meyerspeer, Martin; Fischmeister, Florian Ph S; Grabner, Günther; Bauer, Herbert; Trattnig, Siegfried

    2010-01-01

    Analogous to the evolution of biological sensor-systems, the progress in "medical sensor-systems", i.e., diagnostic procedures, is paradigmatically described. Outstanding highlights of this progress are magnetic resonance imaging (MRI) and spectroscopy (MRS), which enable non-invasive, in vivo acquisition of morphological, functional, and metabolic information from the human body with unsurpassed quality. Recent achievements in high and ultra-high field MR (at 3 and 7 Tesla) are described, and representative research applications in Medicine and Psychology in Austria are discussed. Finally, an overview of current and prospective research in multi-modal imaging, potential clinical applications, as well as current limitations and challenges is given.

  20. Application of a practical method for the isocenter point in vivo dosimetry by a transit signal

    NASA Astrophysics Data System (ADS)

    Piermattei, Angelo; Fidanzio, Andrea; Azario, Luigi; Grimaldi, Luca; D'Onofrio, Guido; Cilla, Savino; Stimato, Gerardina; Gaudino, Diego; Ramella, Sara; D'Angelillo, Rolando; Cellini, Francesco; Trodella, Lucio; Russo, Aniello; Iadanza, Luciano; Zucca, Sergio; Fusco, Vincenzo; Di Napoli, Nicola; Gambacorta, Maria Antonietta; Balducci, Mario; Cellini, Numa; Deodato, Francesco; Macchia, Gabriella; Morganti, Alessio G.

    2007-08-01

    This work reports the results of the application of a practical method to determine the in vivo dose at the isocenter point, Diso, of brain thorax and pelvic treatments using a transit signal St. The use of a stable detector for the measurement of the signal St (obtained by the x-ray beam transmitted through the patient) reduces many of the disadvantages associated with the use of solid-state detectors positioned on the patient as their periodic recalibration, and their positioning is time consuming. The method makes use of a set of correlation functions, obtained by the ratio between St and the mid-plane dose value, Dm, in standard water-equivalent phantoms, both determined along the beam central axis. The in vivo measurement of Diso required the determination of the water-equivalent thickness of the patient along the beam central axis by the treatment planning system that uses the electron densities supplied by calibrated Hounsfield numbers of the computed tomography scanner. This way it is, therefore, possible to compare Diso with the stated doses, Diso,TPS, generally used by the treatment planning system for the determination of the monitor units. The method was applied in five Italian centers that used beams of 6 MV, 10 MV, 15 MV x-rays and 60Co γ-rays. In particular, in four centers small ion-chambers were positioned below the patient and used for the St measurement. In only one center, the St signals were obtained directly by the central pixels of an EPID (electronic portal imaging device) equipped with commercial software that enabled its use as a stable detector. In the four centers where an ion-chamber was positioned on the EPID, 60 pelvic treatments were followed for two fields, an anterior-posterior or a posterior-anterior irradiation and a lateral-lateral irradiation. Moreover, ten brain tumors were checked for a lateral-lateral irradiation, and five lung tumors carried out with three irradiations with different gantry angles were followed. One center

  1. Applications of phosphorescent materials for in-vivo imaging of brain structure and function

    NASA Astrophysics Data System (ADS)

    Boverman, Gregory; Shi, Xiaolei; Cotero, Victoria E.; Filkins, Robert J.; Srivastava, Alok M.; Lorraine, Peter W.; Neculaes, Vasile B.; Ishaque, A. N.

    2016-03-01

    A number of approaches have been developed for in-vivo imaging of neural function at the time scale of action potentials and at the spatial resolution of individual neurons. Remarkable results have been obtained with optogenetics, although the need for genetic modification is an important limitation of these approaches. Similarly, voltage and ion-sensitive dyes allow for optical imaging of action potentials but toxicity remains a problem. Additionally, optical techniques are often only able to be used up to a limited depth. Our preliminary work has shown that nanoparticles of common phosphorescent materials, believed to be generally non-toxic, specifically lutetium oxide and strontium aluminate, can be utilized for cellular imaging, for tomographic imaging, and that the particles can be designed to adhere to neurons. Additionally, lutetium oxide has been shown to be highly X-ray luminescent, potentially allowing for imaging deep within the brain, if the particles can be targeted properly. In ex vivo experiments, we have shown that the phosphorescence of strontium aluminate particles is significantly affected by electric fields similar in strength to those found in the vicinity of the cellular membrane of a neuron. This phenomenon is consistent with early published reports in the electroluminescence literature, namely the Gudden-Pohl effect. We will show results of the ex vivo imaging and dynamic electrical stimulation experiments. We will also show some preliminary ex vivo cell culture results, and will describe plans for future research, focusing on potential in both cell cultures and in vivo for animal models.

  2. Application of electrical stimulation for functional tissue engineering in vitro and in vivo

    NASA Technical Reports Server (NTRS)

    Radisic, Milica (Inventor); Park, Hyoungshin (Inventor); Langer, Robert (Inventor); Freed, Lisa (Inventor); Vunjak-Novakovic, Gordana (Inventor)

    2013-01-01

    The present invention provides new methods for the in vitro preparation of bioartificial tissue equivalents and their enhanced integration after implantation in vivo. These methods include submitting a tissue construct to a biomimetic electrical stimulation during cultivation in vitro to improve its structural and functional properties, and/or in vivo, after implantation of the construct, to enhance its integration with host tissue and increase cell survival and functionality. The inventive methods are particularly useful for the production of bioartificial equivalents and/or the repair and replacement of native tissues that contain electrically excitable cells and are subject to electrical stimulation in vivo, such as, for example, cardiac muscle tissue, striated skeletal muscle tissue, smooth muscle tissue, bone, vasculature, and nerve tissue.

  3. High molecular weight DNA assembly in vivo for synthetic biology applications.

    PubMed

    Juhas, Mario; Ajioka, James W

    2017-05-01

    DNA assembly is the key technology of the emerging interdisciplinary field of synthetic biology. While the assembly of smaller DNA fragments is usually performed in vitro, high molecular weight DNA molecules are assembled in vivo via homologous recombination in the host cell. Escherichia coli, Bacillus subtilis and Saccharomyces cerevisiae are the main hosts used for DNA assembly in vivo. Progress in DNA assembly over the last few years has paved the way for the construction of whole genomes. This review provides an update on recent synthetic biology advances with particular emphasis on high molecular weight DNA assembly in vivo in E. coli, B. subtilis and S. cerevisiae. Special attention is paid to the assembly of whole genomes, such as those of the first synthetic cell, synthetic yeast and minimal genomes.

  4. Informatics approach using metabolic reactivity classifiers to link in vitro to in vivo data in application to the ToxCast Phase I dataset

    EPA Science Inventory

    Strategic combinations and tiered application of alternative testing methods to replace or minimize the use of animal models is attracting much attention. With the advancement of high throughput screening (HTS) assays and legacy databases providing in vivo testing results, suffic...

  5. Application of the front detection photopiroelectric configuration to the study of in vivo human skin

    NASA Astrophysics Data System (ADS)

    Gutierrez-Juarez, G.; Pichardo-Molina, J. L.; Rocha-Osornio, L. N.; Huerta-Franco, R.; Ivanov, R.; Huerta-Franco, B.; Cordova-Fraga, T.; Vargas-Luna, M.

    2005-06-01

    We report a novel method for measurements in vivo of the penetration of topically applied substances by inverse photopyroelectric configuration. This configuration was used to obtain the thermal effusivity, as a function of time, of in vivo human skin with ointments. This thermal magnitude was employed to characterize the penetration on the anterior-face of the volunteers forearm. This thermal effusivity was fitted with an exponential function in order to obtain a parameter (characteristic time) for the penetration. The substances used were a sunscreen and Vick Vaporub ointment. We found that the sunscreen have a characteristic time bigger that the Vick Vaporub ointment. The feasibility of skin hydration studies are discussed.

  6. Applications of stable, nonradioactive isotope tracers in in vivo human metabolic research

    PubMed Central

    Kim, Il-Young; Suh, Sang-Hoon; Lee, In-Kyu; Wolfe, Robert R

    2016-01-01

    The human body is in a constant state of turnover, that is, being synthesized, broken down and/or converted to different compounds. The dynamic nature of in vivo kinetics of human metabolism at rest and in stressed conditions such as exercise and pathophysiological conditions such as diabetes and cancer can be quantitatively assessed with stable, nonradioactive isotope tracers in conjunction with gas or liquid chromatography mass spectrometry and modeling. Although measurements of metabolite concentrations have been useful as general indicators of one's health status, critical information on in vivo kinetics of metabolites such as rates of production, appearance or disappearance of metabolites are not provided. Over the past decades, stable, nonradioactive isotope tracers have been used to provide information on dynamics of specific metabolites. Stable isotope tracers can be used in conjunction with molecular and cellular biology tools, thereby providing an in-depth dynamic assessment of metabolic changes, as well as simultaneous investigation of the molecular basis for the observed kinetic responses. In this review, we will introduce basic principles of stable isotope methodology for tracing in vivo kinetics of human or animal metabolism with examples of quantifying certain aspects of in vivo kinetics of carbohydrate, lipid and protein metabolism. PMID:26795236

  7. Applications of stable, nonradioactive isotope tracers in in vivo human metabolic research.

    PubMed

    Kim, Il-Young; Suh, Sang-Hoon; Lee, In-Kyu; Wolfe, Robert R

    2016-01-15

    The human body is in a constant state of turnover, that is, being synthesized, broken down and/or converted to different compounds. The dynamic nature of in vivo kinetics of human metabolism at rest and in stressed conditions such as exercise and pathophysiological conditions such as diabetes and cancer can be quantitatively assessed with stable, nonradioactive isotope tracers in conjunction with gas or liquid chromatography mass spectrometry and modeling. Although measurements of metabolite concentrations have been useful as general indicators of one's health status, critical information on in vivo kinetics of metabolites such as rates of production, appearance or disappearance of metabolites are not provided. Over the past decades, stable, nonradioactive isotope tracers have been used to provide information on dynamics of specific metabolites. Stable isotope tracers can be used in conjunction with molecular and cellular biology tools, thereby providing an in-depth dynamic assessment of metabolic changes, as well as simultaneous investigation of the molecular basis for the observed kinetic responses. In this review, we will introduce basic principles of stable isotope methodology for tracing in vivo kinetics of human or animal metabolism with examples of quantifying certain aspects of in vivo kinetics of carbohydrate, lipid and protein metabolism.

  8. Development of in vivo Constitutive Models for Liver: Application to Surgical Simulation

    PubMed Central

    Lister, Kevin; Gao, Zhan; Desai, Jaydev P.

    2011-01-01

    Advancements in real-time surgical simulation techniques have provided the ability to utilize more complex nonlinear constitutive models for biological tissues which result in increased haptic and graphic accuracy. When developing such a model, verification is necessary to determine the accuracy of the force response as well as the magnitude of tissue deformation for tool-tissue interactions. In this study, we present an experimental device which provides the ability to obtain force-displacement information as well as surface deformation of porcine liver for in vivo probing tasks. In addition, the system is capable of accurately determining the geometry of the liver specimen. These combined attributes provide the context required to simulate the experiment with accurate boundary conditions, whereby the only variable in the analysis is the material properties of the liver specimen. During the simulation, effects of settling due to gravity have been taken into account by a technique which incorporates the proper internal stress conditions in the model without altering the geometry. Initially, an Ogden model developed from ex vivo tension and compression experimentation is run through the simulation to determine the efficacy of utilizing an ex vivo model for simulation of in vivo probing tasks on porcine liver. Subsequently, a method for improving upon the ex vivo model was developed using different hyperelastic models such that increased accuracy could be achieved for the force characteristics compared to the displacement characteristics, since changes in the force variation would be more perceptible to a user in the simulation environment, while maintaining a high correlation with the surface displacement data. Furthermore, this study also presents the probing simulation which includes the capsule surrounding the liver. PMID:21161684

  9. Development of in vivo constitutive models for liver: application to surgical simulation.

    PubMed

    Lister, Kevin; Gao, Zhan; Desai, Jaydev P

    2011-03-01

    Advancements in real-time surgical simulation techniques have provided the ability to utilize more complex nonlinear constitutive models for biological tissues which result in increased haptic and graphic accuracy. When developing such a model, verification is necessary to determine the accuracy of the force response as well as the magnitude of tissue deformation for tool-tissue interactions. In this study, we present an experimental device which provides the ability to obtain force-displacement information as well as surface deformation of porcine liver for in vivo probing tasks. In addition, the system is capable of accurately determining the geometry of the liver specimen. These combined attributes provide the context required to simulate the experiment with accurate boundary conditions, whereby the only variable in the analysis is the material properties of the liver specimen. During the simulation, effects of settling due to gravity have been taken into account by a technique which incorporates the proper internal stress conditions in the model without altering the geometry. Initially, an Ogden model developed from ex vivo tension and compression experimentation is run through the simulation to determine the efficacy of utilizing an ex vivo model for simulation of in vivo probing tasks on porcine liver. Subsequently, a method for improving upon the ex vivo model was developed using different hyperelastic models such that increased accuracy could be achieved for the force characteristics compared to the displacement characteristics, since changes in the force variation would be more perceptible to a user in the simulation environment, while maintaining a high correlation with the surface displacement data. Furthermore, this study also presents the probing simulation which includes the capsule surrounding the liver.

  10. A new strategy for in vivo spectral editing. Application to GABA editing using selective homonuclear polarization transfer spectroscopy

    NASA Astrophysics Data System (ADS)

    Shen, Jun; Yang, Jehoon; Choi, In-Young; Li, Shizhe Steve; Chen, Zhengguang

    2004-10-01

    A novel single-shot in vivo spectral editing method is proposed in which the signal to be detected, is regenerated anew from the thermal equilibrium magnetization of a source to which it is J-coupled. The thermal equilibrium magnetization of the signal to be detected together with those of overlapping signals are suppressed by single-shot gradient dephasing prior to the signal regeneration process. Application of this new strategy to in vivo GABA editing using selective homonuclear polarization transfer allows complete suppression of overlapping creatine and glutathione while detecting the GABA-4 methylene resonance at 3.02 ppm with an editing yield similar to that of conventional editing methods. The NAA methyl group at 2.02 ppm was simultaneously detected and can be used as an internal navigator echo for correcting the zero order phase and frequency shifts and as an internal reference for concentration. This new method has been demonstrated for robust in vivo GABA editing in the rat brain and for study of GABA synthesis after acute vigabatrin administration.

  11. Illuminating the Undergraduate Behavioral Neuroscience Laboratory: A Guide for the in vivo Application of Optogenetics in Mammalian Model Organisms.

    PubMed

    Roberts, Bradley M; Jarrin, Sarah E; Mathur, Brian N; Bailey, Aileen M

    2016-01-01

    Optogenetics is a technology that is growing rapidly in neuroscience, establishing itself as a fundamental investigative tool. As this tool is increasingly utilized across the neuroscience community and is one of the primary research techniques being presented at neuroscience conferences and in journals, we believe that it is important that this technology is introduced into the undergraduate neuroscience research laboratory. While there has been a significant body of work concentrated to deploy optogenetics in invertebrate model organisms, little to no work has focused on brining this technology to mammalian model organisms in undergraduate neuroscience laboratories. The establishment of in vivo optogenetics could provide for high-impact independent research projects for upper-level undergraduate students. Here we review the considerations for establishing in vivo optogenetics with the use of rodents in an undergraduate laboratory setting and provide some cost-saving guidelines to assist in making optogenetic technologies financially accessible. We discuss opsin selection, cell-specific opsin expression strategies, species selection, experimental design, selection of light delivery systems, and the construction of implantable optical fibers for the application of in vivo optogenetics in rodents.

  12. Synthesis, characterization, and application of reversible PDLLA-PEG-PDLLA copolymer thermogels in vitro and in vivo

    PubMed Central

    Shi, Kun; Wang, Ya-Li; Qu, Ying; Liao, Jin-Feng; Chu, Bing-Yang; Zhang, Hua-Ping; Luo, Feng; Qian, Zhi-Yong

    2016-01-01

    In this study, a series of injectable thermoreversible and thermogelling PDLLA-PEG-PDLLA copolymers were developed and a systematic evaluation of the thermogelling system both in vitro and in vivo was performed. The aqueous PDLLA-PEG-PDLLA solutions above a critical gel concentration could transform into hydrogel spontaneously within 2 minutes around the body temperature in vitro or in vivo. Modulating the molecular weight, block length and polymer concentration could adjust the sol-gel transition behavior and the mechanical properties of the hydrogels. The gelation was thermally reversible due to the physical interaction of copolymer micelles and no crystallization formed during the gelation. Little cytotoxicity and hemolysis of this polymer was found, and the inflammatory response after injecting the hydrogel to small-animal was acceptable. In vitro and in vivo degradation experiments illustrated that the physical hydrogel could retain its integrity as long as several weeks and eventually be degraded by hydrolysis. A rat model of sidewall defect-bowel abrasion was employed, and a significant reduction of post-operative adhesion has been found in the group of PDLLA-PEG-PDLLA hydrogel-treated, compared with untreated control group and commercial hyaluronic acid (HA) anti-adhesion hydrogel group. As such, this PDLLA-PEG-PDLLA hydrogel might be a promising candidate of injectable biomaterial for medical applications. PMID:26752008

  13. Illuminating the Undergraduate Behavioral Neuroscience Laboratory: A Guide for the in vivo Application of Optogenetics in Mammalian Model Organisms

    PubMed Central

    Roberts, Bradley M.; Jarrin, Sarah E.; Mathur, Brian N.; Bailey, Aileen M.

    2016-01-01

    Optogenetics is a technology that is growing rapidly in neuroscience, establishing itself as a fundamental investigative tool. As this tool is increasingly utilized across the neuroscience community and is one of the primary research techniques being presented at neuroscience conferences and in journals, we believe that it is important that this technology is introduced into the undergraduate neuroscience research laboratory. While there has been a significant body of work concentrated to deploy optogenetics in invertebrate model organisms, little to no work has focused on brining this technology to mammalian model organisms in undergraduate neuroscience laboratories. The establishment of in vivo optogenetics could provide for high-impact independent research projects for upper-level undergraduate students. Here we review the considerations for establishing in vivo optogenetics with the use of rodents in an undergraduate laboratory setting and provide some cost-saving guidelines to assist in making optogenetic technologies financially accessible. We discuss opsin selection, cell-specific opsin expression strategies, species selection, experimental design, selection of light delivery systems, and the construction of implantable optical fibers for the application of in vivo optogenetics in rodents. PMID:27385919

  14. Tracking stem cells for cardiovascular applications in vivo: focus on imaging techniques

    PubMed Central

    Fu, Yingli; Azene, Nicole; Xu, Yi; Kraitchman, Dara L

    2011-01-01

    Despite rapid translation of stem cell therapy into clinical practice, the treatment of cardiovascular disease using embryonic stem cells, adult stem and progenitor cells or induced pluripotent stem cells has not yielded satisfactory results to date. Noninvasive stem cell imaging techniques could provide greater insight into not only the therapeutic benefit, but also the fundamental mechanisms underlying stem cell fate, migration, survival and engraftment in vivo. This information could also assist in the appropriate choice of stem cell type(s), delivery routes and dosing regimes in clinical cardiovascular stem cell trials. Multiple imaging modalities, such as MRI, PET, SPECT and CT, have emerged, offering the ability to localize, monitor and track stem cells in vivo. This article discusses stem cell labeling approaches and highlights the latest cardiac stem cell imaging techniques that may help clinicians, research scientists or other healthcare professionals select the best cellular therapeutics for cardiovascular disease management. PMID:22287982

  15. Luminescent magnetic quantum dots for in vitro/in vivo imaging and applications in therapeutics.

    PubMed

    Acharya, Amitabha

    2013-06-01

    The quest for design of newer/advanced methods for medical diagnosis and targeted therapeutics are of utmost interest and challenging too, because of its importance in clinical diagnosis. Currently available diagnosis methodologies have their own disadvantages. These shortcomings can be overcome by using multimodal imaging systems where two or more imaging modalities may be coupled. Nanoparticles being widely studied for targeted drug delivery and as biological contrasting agents, might play a decisive role in such findings. This review is focused towards the ongoing research in the area of hybrid nanocomposites which can be used for both as MRI contrasting agent (magnetic nanoparticles) and molecular imaging studies (using fluorescent quantum dots) at in vitro and in vivo level. Though several reports are available in literature for such bimodal imaging systems, their clinical trials are very restricted, possibly because of the lack of communication between the in vitro and in vivo studies. This review is expected to bridge the gap between such studies.

  16. The application of PEDRI to the study of free radicals in vivo

    NASA Astrophysics Data System (ADS)

    Foster, M. A.; Seimenis, I.; Lurie, D. J.

    1998-07-01

    Proton-electron double-resonance imaging (PEDRI) has considerable value for study of the distribution and elimination pathways of nitroxide free radicals (NFRs). This has been illustrated by its use in studies of kidney function in the living rat in which the NFR proxyl carboxylic acid (PCA) has been employed as a `tracer'. The technique, at its present stage of development, can demonstrate location of PCA in enough detail to observe the passage through kidney cortex and medulla differentially, and to see the NFR within the major abdominal blood vessels. These studies are helping towards an understanding of the metabolic fate of PCA, as well as providing information about kidney performance after challenge with a nephrotoxin. In addition, nitric oxide complexes, formed in vivo by providing rats with a nitrite-rich diet, have been observed ex vivo using PEDRI and field-cycled DNP.

  17. Miniature Uncooled Infrared Sensitive Detectors for in Vivo Biomedical Imaging Applications

    SciTech Connect

    Datskos, P. G.; Demos, S. G.; Rajic, S.

    1998-06-01

    Broadband infrared (OR) radiation detectors have been developed using miniature, inexpensive, mass produced microcantilevers capable of detecting temperature differences as small as lea(-6) K. Microcantilevers made out of semiconductor materials can be used either as uncurled photon or thermal detectors. Mounted on a probe mm in diameter a number of microcantilevers can be accommodated in the working channel of existing endoscopes for in vivo proximity focus measurements inside the human body.

  18. Application of XRF to measure strontium in human bone in vivo

    SciTech Connect

    Wielopolski, L.; Vartsky, D.; Yasumura, S.; Cohn, S.H.

    1982-01-01

    As a basis for better understanding the role that Sr fulfills in human body, it is desirable to measure directly the main Sr store in human body. Although strontium is omnipresent in human tissues, 99% is stored inthe mineral portion of the bone. In the present study x-ray fluorescence (XRF) was applied to measure the strontium content of the tibial shaft in vivo. The feasibility studies showed that normal levels of stable strontium in the bone can be measured successfully.

  19. In vivo elemental analysis by counting neutron-induced gamma rays for medical and biological applications

    NASA Astrophysics Data System (ADS)

    Kehayias, Joseph J.; Ma, Ruimei; Zhuang, Hong; Moore, Robert; Dowling, Lisa

    1995-03-01

    Non-invasive in vivo elemental analysis is a technique used to assess human body composition which is indicative of nutritional status and health condition. The in vivo measurement of the body's major elements is used for a variety of medical studies requiring the determination of the body's compartments (protein, fat, water, bone). Whole body gamma-ray counters, consisting of Nal(Tl) crystal detectors in a shielded room, are used for measuring in vivo the body's Ca, Cl, Na and P by delayed neutron activation analysis. Thermal neutrons from a moderated 238Pu-Be source are used for the measurement of total body nitrogen (and thus protein) and chlorine at low radiation exposure (0.80 mSv). The resulting high energy prompt gamma-rays from nitrogen (10.83 MeV) and chlorine (6.11 MeV) are detected simultaneously with the irradiation. Body fat (the main energy store) and fat distribution (which relates to risk for cardiovascular disease) are measured by detecting C and O in vivo through fast neutron inelastic scattering. A small sealed D-T neutron generator is used for the pulsed (4 - 8 KHz) production of fast neutrons. Carbon and oxygen are detected by counting the 4.44 and 6.13 MeV gamma-rays resulting from the inelastic scattering of the fast neutrons from the 12C and 16O nuclei, respectively. One use of this method is the systematic study of the mechanisms driving the age-associated depletion of the metabolizing, oxygen-consuming cellular compartment of the body. The understanding of this catabolism may suggest ways to maintain lean tissue and thus to preserve quality of life for the very old.

  20. In vivo Real-Time Mass Spectrometry for Guided Surgery Application

    NASA Astrophysics Data System (ADS)

    Fatou, Benoit; Saudemont, Philippe; Leblanc, Eric; Vinatier, Denis; Mesdag, Violette; Wisztorski, Maxence; Focsa, Cristian; Salzet, Michel; Ziskind, Michael; Fournier, Isabelle

    2016-05-01

    Here we describe a new instrument (SpiderMass) designed for in vivo and real-time analysis. In this instrument ion production is performed remotely from the MS instrument and the generated ions are transported in real-time to the MS analyzer. Ion production is promoted by Resonant Infrared Laser Ablation (RIR-LA) based on the highly effective excitation of O-H bonds in water molecules naturally present in most biological samples. The retrieved molecular patterns are specific to the cell phenotypes and benign versus cancer regions of patient biopsies can be easily differentiated. We also demonstrate by analysis of human skin that SpiderMass can be used under in vivo conditions with minimal damage and pain. Furthermore SpiderMass can also be used for real-time drug metabolism and pharmacokinetic (DMPK) analysis or food safety topics. SpiderMass is thus the first MS based system designed for in vivo real-time analysis under minimally invasive conditions.

  1. In vivo Real-Time Mass Spectrometry for Guided Surgery Application

    PubMed Central

    Fatou, Benoit; Saudemont, Philippe; Leblanc, Eric; Vinatier, Denis; Mesdag, Violette; Wisztorski, Maxence; Focsa, Cristian; Salzet, Michel; Ziskind, Michael; Fournier, Isabelle

    2016-01-01

    Here we describe a new instrument (SpiderMass) designed for in vivo and real-time analysis. In this instrument ion production is performed remotely from the MS instrument and the generated ions are transported in real-time to the MS analyzer. Ion production is promoted by Resonant Infrared Laser Ablation (RIR-LA) based on the highly effective excitation of O-H bonds in water molecules naturally present in most biological samples. The retrieved molecular patterns are specific to the cell phenotypes and benign versus cancer regions of patient biopsies can be easily differentiated. We also demonstrate by analysis of human skin that SpiderMass can be used under in vivo conditions with minimal damage and pain. Furthermore SpiderMass can also be used for real-time drug metabolism and pharmacokinetic (DMPK) analysis or food safety topics. SpiderMass is thus the first MS based system designed for in vivo real-time analysis under minimally invasive conditions. PMID:27189490

  2. Measurement of passive skeletal muscle mechanical properties in vivo: recent progress, clinical applications, and remaining challenges.

    PubMed

    Bilston, Lynne E; Tan, Kristy

    2015-02-01

    The ability to measure and quantify the properties of skeletal muscle in vivo as a method for understanding its complex physiological and pathophysiological behavior is important in numerous clinical settings, including rehabilitation. However, this remains a challenge to date due to the lack of a "gold standard" technique. Instead, there are a myriad of measuring techniques each with its own set of pros and cons. This review discusses the current state-of-the-art in elastography imaging techniques, i.e., ultrasound and magnetic resonance elastography, as applied to skeletal muscle, and briefly reviews other methods of measuring muscle mechanical behavior in vivo. While in vivo muscle viscoelastic properties can be measured, these techniques are largely limited to static or quasistatic measurements. Emerging elastography techniques are able to quantify muscle anisotropy and large deformation effects on stiffness, but, validation and optimization of these newer techniques is required. The development of reliable values for the mechanical properties of muscle across the population using these techniques are required to enable them to become more useful in rehabilitation and other clinical settings.

  3. A biocompatible in vivo ligation reaction and its application for noninvasive bioluminescent imaging of protease activity in living mice.

    PubMed

    Godinat, Aurélien; Park, Hyo Min; Miller, Stephen C; Cheng, Ke; Hanahan, Douglas; Sanman, Laura E; Bogyo, Matthew; Yu, Allen; Nikitin, Gennady F; Stahl, Andreas; Dubikovskaya, Elena A

    2013-05-17

    The discovery of biocompatible reactions had a tremendous impact on chemical biology, allowing the study of numerous biological processes directly in complex systems. However, despite the fact that multiple biocompatible reactions have been developed in the past decade, very few work well in living mice. Here we report that D-cysteine and 2-cyanobenzothiazoles can selectively react with each other in vivo to generate a luciferin substrate for firefly luciferase. The success of this "split luciferin" ligation reaction has important implications for both in vivo imaging and biocompatible labeling strategies. First, the production of a luciferin substrate can be visualized in a live mouse by bioluminescence imaging (BLI) and furthermore allows interrogation of targeted tissues using a "caged" luciferin approach. We therefore applied this reaction to the real-time noninvasive imaging of apoptosis associated with caspase 3/7. Caspase-dependent release of free D-cysteine from the caspase 3/7 peptide substrate Asp-Glu-Val-Asp-D-Cys (DEVD-(D-Cys)) allowed selective reaction with 6-amino-2-cyanobenzothiazole (NH(2)-CBT) in vivo to form 6-amino-D-luciferin with subsequent light emission from luciferase. Importantly, this strategy was found to be superior to the commercially available DEVD-aminoluciferin substrate for imaging of caspase 3/7 activity. Moreover, the split luciferin approach enables the modular construction of bioluminogenic sensors, where either or both reaction partners could be caged to report on multiple biological events. Lastly, the luciferin ligation reaction is 3 orders of magnitude faster than Staudinger ligation, suggesting further applications for both bioluminescence and specific molecular targeting in vivo.

  4. Phenotype and functional evaluation of ex vivo generated antigen-specific immune effector cells with potential for therapeutic applications.

    PubMed

    Han, Shuhong; Huang, Yuju; Liang, Yin; Ho, Yuchin; Wang, Yichen; Chang, Lung-Ji

    2009-08-06

    Ex vivo activation and expansion of lymphocytes for adoptive cell therapy has demonstrated great success. To improve safety and therapeutic efficacy, increased antigen specificity and reduced non-specific response of the ex vivo generated immune cells are necessary. Here, using a complete protein-spanning pool of pentadecapeptides of the latent membrane protein 2A (LMP2A) of Epstein-Barr virus (EBV), a weak viral antigen which is associated with EBV lymphoproliferative diseases, we investigated the phenotype and function of immune effector cells generated based on IFN-gamma or CD137 activation marker selection and dendritic cell (DC) activation. These ex vivo prepared immune cells exhibited a donor- and antigen-dependent T cell response; the IFN-gamma-selected immune cells displayed a donor-related CD4- or CD8-dominant T cell phenotype; however, the CD137-enriched cells showed an increased ratio of CD4 T cells. Importantly, the pentadecapeptide antigens accessed both class II and class I MHC antigen processing machineries and effectively activated EBV-specific CD4 and CD8 T cells. Phenotype and kinetic analyses revealed that the IFN-gamma and the CD137 selections enriched more central memory T (Tcm) cells than did the DC-activation approach, and after expansion, the IFN-gamma-selected effector cells showed the highest level of antigen-specificity and effector activities. While all three approaches generated immune cells with comparable antigen-specific activities, the IFN-gamma selection followed by ex vivo expansion produced high quality and quantity of antigen-specific effector cells. Our studies presented the optimal approach for generating therapeutic immune cells with potential for emergency and routine clinical applications.

  5. Clinical applicability of in vivo fluorescence confocal microscopy for noninvasive diagnosis and therapeutic monitoring of nonmelanoma skin cancer.

    PubMed

    Astner, Susanne; Dietterle, Susanne; Otberg, Nina; Röwert-Huber, Hans-Joachim; Stockfleth, Eggert; Lademann, Jürgen

    2008-01-01

    Excisional biopsies and routine histology remains the gold standard for the histomorphologic evaluation of normal and diseased skin. However, there is increasing interest in the development of noninvasive optical technologies for evaluation, diagnosis, and monitoring of skin disease in vivo. Fluorescent confocal microscopy is an innovative optical technology that has previously been used for morphologic evaluation of live human tissue. We evaluate the clinical applicability of a fluorescent confocal laser scanning microscope (FLSM) for a systematic evaluation of normal and diseased skin in vivo and in correlation with routine histology. A total of 40 patients were recruited to participate in the study. Skin sites of 10 participants with no prior history of skin disease served as controls and to evaluate topographic variations of normal skin in vivo. Thirty patients with a suspected diagnosis of nonmelanoma skin cancer were evaluated, whereby FLSM features of actinic keratoses (AK) and basal cell carcinoma (BCC) were recorded in an observational analysis. Selected BCCs were monitored for their skin response to topical therapy using Imiquimod as an immune-response modifier. A commercially available fluorescence microscope (OptiScan Ltd., Melbourne, Australia) was used to carry out all FLSM evaluations. Common FLSM features to AK and BCC included nuclear pleomorphism at the level of the granular and spinous layer and increased vascularity in the superficial dermal compartment. Even though the presence of superficial disruption and mere atypia of epidermal keratinocytes was more indicative of AK, the nesting of atypical basal cells, increased blood vessel tortuosity, and nuclear polarization were more typical for BCC. All diagnoses were confirmed by histology. FLSM allowed a monitoring of the local immune response following therapy with Imiquimod and demonstrated a continuous normalization of diseased skin on repeated evaluations over time. This study illustrates

  6. Simple and effective exercise design for assessing in vivo mitochondrial function in clinical applications using 31P magnetic resonance spectroscopy

    PubMed Central

    Sleigh, Alison; Lupson, Victoria; Thankamony, Ajay; Dunger, David B.; Savage, David B.; Carpenter, T. Adrian; Kemp, Graham J.

    2016-01-01

    The growing recognition of diseases associated with dysfunction of mitochondria poses an urgent need for simple measures of mitochondrial function. Assessment of the kinetics of replenishment of the phosphocreatine pool after exercise using 31P magnetic resonance spectroscopy can provide an in vivo measure of mitochondrial function; however, the wider application of this technique appears limited by complex or expensive MR-compatible exercise equipment and protocols not easily tolerated by frail participants or those with reduced mental capacity. Here we describe a novel in-scanner exercise method which is patient-focused, inexpensive, remarkably simple and highly portable. The device exploits an MR-compatible high-density material (BaSO4) to form a weight which is attached directly to the ankle, and a one-minute dynamic knee extension protocol produced highly reproducible measurements of post-exercise PCr recovery kinetics in both healthy subjects and patients. As sophisticated exercise equipment is unnecessary for this measurement, our extremely simple design provides an effective and easy-to-implement apparatus that is readily translatable across sites. Its design, being tailored to the needs of the patient, makes it particularly well suited to clinical applications, and we argue the potential of this method for investigating in vivo mitochondrial function in new cohorts of growing clinical interest. PMID:26751849

  7. PEGylated gold nanorods as optical trackers for biomedical applications: an in vivo and in vitro comparative study

    NASA Astrophysics Data System (ADS)

    Abdelrasoul, Gaser N.; Magrassi, Raffaella; Dante, Silvia; d'Amora, Marta; Scotto d'Abbusco, Marco; Pellegrino, Teresa; Diaspro, Alberto

    2016-06-01

    Gold nanorods (AuNRs) are eligible for a variety of biological applications including cell imaging, sensing, and photothermal therapy thanks to their optical properties. The aim of this work is to show how AuNRs could be employed as non-photobleachable optical contrast agents for biomedical applications. In order to demonstrate the feasibility of their use as optical trackers, we employed two-photon emission confocal microscopy on cells incubated with PEGylated AuNRs. Remarkably, AuNRs were localized mostly in the perinuclear zone and microscopy characterization showed the presence of a considerable number of rods inside cell nuclei. Furthermore, we estimated the toxicity and the efficiency of cellular uptake of the PEGylated AuNRs as a function of administered dose on HeLa/3T3 cell lines and on zebrafish during development, employed as an in vivo model. Eventually, we observed good agreement between in vivo and in vitro experiments. The employed AuNRs were prepared through a photochemical protocol here improved by tuning the amount of the cationic surfactant cetyltrimethylammonium bromide for the achievement of AuNRs at two different aspect ratios. Furthermore we also investigated if the AuNR aspect ratio influenced the toxicity and the efficiency of cellular uptake of the PEGylated AuNRs in HeLa/3T3 cell lines and in zebrafish embryos.

  8. In-vitro Optimization of Nanoparticle-Cell Labeling Protocols for In-vivo Cell Tracking Applications.

    PubMed

    Betzer, Oshra; Meir, Rinat; Dreifuss, Tamar; Shamalov, Katerina; Motiei, Menachem; Shwartz, Amit; Baranes, Koby; Cohen, Cyrille J; Shraga-Heled, Niva; Ofir, Racheli; Yadid, Gal; Popovtzer, Rachela

    2015-10-28

    Recent advances in theranostic nanomedicine can promote stem cell and immune cell-based therapy. Gold nanoparticles (GNPs) have been shown to be promising agents for in-vivo cell-tracking in cell-based therapy applications. Yet a crucial challenge is to develop a reliable protocol for cell upload with, on the one hand, sufficient nanoparticles to achieve maximum visibility of cells, while on the other hand, assuring minimal effect of particles on cell function and viability. Previous studies have demonstrated that the physicochemical parameters of GNPs have a critical impact on their efficient uptake by cells. In the current study we have examined possible variations in GNP uptake, resulting from different incubation period and concentrations in different cell-lines. We have found that GNPs effectively labeled three different cell-lines - stem, immune and cancer cells, with minimal impairment to cell viability and functionality. We further found that uptake efficiency of GNPs into cells stabilized after a short period of time, while GNP concentration had a significant impact on cellular uptake, revealing cell-dependent differences. Our results suggest that while heeding the slight variations within cell lines, modifying the loading time and concentration of GNPs, can promote cell visibility in various nanoparticle-dependent in-vivo cell tracking and imaging applications.

  9. In-vitro Optimization of Nanoparticle-Cell Labeling Protocols for In-vivo Cell Tracking Applications

    PubMed Central

    Betzer, Oshra; Meir, Rinat; Dreifuss, Tamar; Shamalov, Katerina; Motiei, Menachem; Shwartz, Amit; Baranes, Koby; Cohen, Cyrille J.; Shraga-Heled, Niva; Ofir, Racheli; Yadid, Gal; Popovtzer, Rachela

    2015-01-01

    Recent advances in theranostic nanomedicine can promote stem cell and immune cell-based therapy. Gold nanoparticles (GNPs) have been shown to be promising agents for in-vivo cell-tracking in cell-based therapy applications. Yet a crucial challenge is to develop a reliable protocol for cell upload with, on the one hand, sufficient nanoparticles to achieve maximum visibility of cells, while on the other hand, assuring minimal effect of particles on cell function and viability. Previous studies have demonstrated that the physicochemical parameters of GNPs have a critical impact on their efficient uptake by cells. In the current study we have examined possible variations in GNP uptake, resulting from different incubation period and concentrations in different cell-lines. We have found that GNPs effectively labeled three different cell-lines - stem, immune and cancer cells, with minimal impairment to cell viability and functionality. We further found that uptake efficiency of GNPs into cells stabilized after a short period of time, while GNP concentration had a significant impact on cellular uptake, revealing cell-dependent differences. Our results suggest that while heeding the slight variations within cell lines, modifying the loading time and concentration of GNPs, can promote cell visibility in various nanoparticle-dependent in-vivo cell tracking and imaging applications. PMID:26507853

  10. Quercetin-loaded PLGA nanoparticles: a highly effective antibacterial agent in vitro and anti-infection application in vivo

    NASA Astrophysics Data System (ADS)

    Sun, Dongdong; Li, Nuan; Zhang, Weiwei; Yang, Endong; Mou, Zhipeng; Zhao, Zhiwei; Liu, Haiping; Wang, Weiyun

    2016-01-01

    Nanotechnology-based approaches have tremendous potential for enhancing efficacy against infectious diseases. PLGA-based nanoparticles as drug delivery carrier have shown promising potential, owing to their sizes and related unique properties. This article aims to develop nanosized poly ( d, l-lactide-co-glycolide) PLGA nanoparticle formulation loaded with quercetin (QT). QT is an antioxidant and antibacterial compound isolated from Chinese traditional medicine with low skin permeability and extreme water insolubility. The quercetin-loaded PLGA nanoparticles (PQTs) were synthesized by emulsion-solvent evaporation method and stabilized by coating with poly (vinyl alcohol). The characteristics of PQTs were analyzed by Fourier transform infrared spectroscopy, Ultraviolet-Visible spectroscopy, scanning electron microscope, transmission electron microscopy, and atomic force microscopy, respectively. The PQTs showed a spherical shape with an average size of 100-150 nm. We compared the antibacterial effects of PQTs against Escherichia coli ( E. coli) and Micrococcus tetragenus ( M. tetragenus).The PQTs produced stronger antibacterial activity to E. coli than that to M. tetragenus through disrupting bacterial cell wall integrity. The antibacterial ratio was increased with the increasing dosages and incubation time. Next, we tested the in vivo antibacterial activity in mice. No noticeable organ damage was captured from H&E-staining organ slices, suggesting the promise of using PQTs for in vivo applications. The results of this study demonstrated the interaction between bacteria and PLGA-based nanoparticles, providing encouragement for conducting further investigations on properties and antimicrobial activity of the PQTs in clinical application.

  11. In Vivo Assessment of Neurotransmitters and Modulators with Magnetic Resonance Spectroscopy: Application to Schizophrenia

    PubMed Central

    Wijtenburg, S. Andrea; Yang, Shaolin; Fischer, Bernard A.; Rowland, Laura M.

    2015-01-01

    In vivo measurement of neurotransmitters and modulators is now feasible with advanced proton magnetic resonance spectroscopy (1H-MRS) techniques. This review provides a basic tutorial of MRS, describes the methods available to measure brain glutamate, glutamine, γ-aminobutyric acid, glutathione, N-acetylaspartylglutamate, glycine, and serine at magnetic field strengths of 3Tesla or higher, and summarizes the neurochemical findings in schizophrenia. Overall, 1H-MRS holds great promise for producing biomarkers that can serve as treatment targets, prediction of disease onset, or illness exacerbation in schizophrenia and other brain diseases. PMID:25614132

  12. In vitro and in vivo evaluation of an alumina-zirconia composite for arthroplasty applications.

    PubMed

    Roualdes, Olivier; Duclos, Marie-Eve; Gutknecht, Dan; Frappart, Lucien; Chevalier, Jérôme; Hartmann, Daniel J

    2010-03-01

    In order to improve the reliability and the mechanical properties of orthopaedic hip prosthesis, new ceramic composites starting with nanosized powders of alumina and zirconia have been recently developed. The aim of the present study was to investigate the biological tolerance of one of these sintered ceramics and of its alumina and zirconia constitutive nanosized powders with both in vitro and in vivo approaches. At first, osteoblasts and fibroblasts were cultured either upon sintered ceramic discs with polished or rough surfaces or in the presence of the corresponding alumina or zirconia powders at various concentrations. Thereafter, we chronically injected these powders in the knee articulation of rats. In vitro, the materials showed no deleterious effect on cell proliferation, extra-cellular matrix production (human type I collagen and fibronectin) or on cell morphology. In vivo, the histological examination showed only a very moderate and non-specific granulomatous response of the synovial membrane but no major inflammation as clinically described with metals or polyethylene wear debris. Besides its improved physical properties, this recently developed alumina-zirconia composite showed satisfactory biocompatibility.

  13. In-vivo high resolution corneal imaging and analysis on animal models for clinical applications

    NASA Astrophysics Data System (ADS)

    Hong, Jesmond; Shinoj, V. K.; Murukeshan, V. M.; Baskaran, M.; Aung, Tin

    2015-07-01

    A simple and low cost optical probe system for the high resolution imaging of the cornea is proposed, based on a Gaussian beam epi-illumination configuration. Corneal topography is obtained by moving the scanning spot across the eye in a raster fashion whereas pachymetry data is achieved by reconstructing the images obtained at different depths. The proposed prototype has been successfully tested on porcine eye samples ex vivo and subsequently on laboratory animals, such as the New Zealand White Rabbit, in vivo. This proposed system and methodology pave the way for realizing a simple and inexpensive optical configuration for pachymetry and keratometry readings, with achievable resolution up to the cellular level. This novel and non-contact high resolution imaging modality demonstrates high intraobserver reproducibility and repeatability. Together with its sophisticated data analysis strategies and safety profile, it is believed to complement existing imaging modalities in the assessment and evaluation of corneal diseases, which enable a decrease in morbidity and improvement in the effectiveness of subsequent treatment.

  14. Development of a small-scale bioreactor: application to in vivo NMR measurement.

    PubMed

    Gmati, Dorra; Chen, Jingkui; Jolicoeur, Mario

    2005-01-20

    A perfused bioreactor allowing in vivo NMR measurement was developed and validated for Eschscholtzia californica cells. The bioreactor was made of a 10-mm NMR tube. NMR measurement of the signal-to-noise ratio was optimized using a sedimented compact bed of cells that were retained in the bioreactor by a supporting filter. Liquid medium flow through the cell bed was characterized from a mass balance on oxygen and a dispersive hydrodynamic model. Cell bed oxygen demand for 4 h perfusion required a minimal medium flow rate of 0.8 mL/min. Residence time distribution assays at 0.8-2.6 mL/min suggest that the cells are subjected to a uniform nutrient environment along the cell bed. Cell integrity was maintained for all culture conditions since the release of intracellular esterases was not significant even after 4 h of perfusion. In vivo NMR was performed for (31)P NMR and the spectrum can be recorded after only 10 min of spectral accumulation (500 scans) with peaks identified as G-6P, F-6P, cytoplasmic Pi, vacuolar Pi, ATP(gamma) and ADP(beta), ATP(alpha) and ADP(alpha), NADP and NDPG, NDPG and ATP(beta). Cell viability was shown to be maintained as (31)P chemical shifts were constant with time for all the identified nuclei, thus suggesting constant intracellular pH.

  15. Application of laser scan microscopy in vivo for wound healing characterization

    NASA Astrophysics Data System (ADS)

    Czaika, V.; Alborova, A.; Sterry, W.; Lademann, J.; Koch, S.

    2010-09-01

    Considering the advancing age of the population, wound healing disturbances are becoming increasingly important in clinical routine. The development of wound healing creams and lotions as well as therapy control require an objective evaluation of the wound healing process, which represents the destruction of the barrier. Therefore, transepidermal water loss measurements are often carried out. These measurements have the disadvantage that they are disturbed by the interstitial fluid, which is located on the surface of chronic wounds and also by water components of the creams and lotions. Additionally, the TEWL measurements are very sensitive to temperature changes and to the anxiety of the volunteers. In the present study, in vivo laser scanning microscopy was used to analyze the reepithelialization and barrier recovery of standardized wounds produced by the suction blister technique. It was demonstrated that this non-invasive, on-line spectroscopic method allows the evaluation of the wound healing process, without any disturbances. It was found that the wound healing starts not only from the edges of the wound, but also out of the hair follicles. The in vivo laser scanning microscopy is well suited to evaluate the efficacy of wound healing creams and for therapy control.

  16. A new in vivo Raman probe for enhanced applicability to the body.

    PubMed

    Pudney, Paul D A; Bonnist, Eleanor Y M; Caspers, Peter J; Gorce, Jean-Philippe; Marriot, Chris; Puppels, Gerwin J; Singleton, Scott; van der Wolf, Martin J G

    2012-08-01

    This paper describes a new in vivo Raman probe that allows investigation of areas of the body that are otherwise difficult to access. It is coupled to a previously described commercially available in vivo Raman spectrometer that samples the skin through an optical flat. In the work presented here, the laser light emerges from a smaller pen-shaped probe. It thus works on the same principles as the original spectrometer, while its relative performance in terms of signal-to-noise ratio of the spectra and obtained spatial resolution is only slightly diminished. It allows the window to be placed against the subject in more curved and recessed areas of subject's body and also for them to be more comfortable while the measurements take place. Results from three areas of the body that have previously been very difficult to study are described, the mouth, axilla, and scalp. Results from the scalp and axilla strata cornea (SC) show significant differences from the "normal" SC of the volar forearm. For instance, the scalp is observed to have lower amounts of natural moisturizing factors (NMF) compared to the volar forearm within the same subjects. Also for both the axilla and scalp the lipids show a change in order as compared to the lipids in the volar forearm and also differences from each other. The potential significance of these observations is discussed. Further, we show how we can probe the mouth, in this case observing the presence of the astringent tea polyphenol epigallocatechin gallate within the oral mucosa.

  17. Application of a new high-speed magnetic deformable mirror for in-vivo retinal imaging

    NASA Astrophysics Data System (ADS)

    Balderas-Mata, Sandra E.; Jones, Steven M.; Zawadzki, Robert J.; Werner, John S.

    2011-08-01

    Nowadays in ophthalmologic practice several commercial instruments are available to image patient retinas in vivo. Many modern fundus cameras and confocal scanning laser ophthalmoscopes allow acquisition of two dimensional en face images of the retina with both back reflected as well as fluorescent light. Additionally, optical coherence tomography systems allow non-invasive probing of three-dimensional retinal morphology. For all of these instruments the available lateral resolution is limited by optical quality of the human eye used as the imaging objective. To improve lateral resolution and achieve diffraction-limited imaging, adaptive optics (AO) can be implemented with any of these imaging systems to correct both static and dynamic aberrations inherent in human eyes. Most of the wavefront correctors used previously in AO systems have limited dynamic range and an insufficient number of actuators to achieve diffraction-limited correction of most human eyes. Thus, additional corrections were necessary, either by trial lenses or additional deformable mirrors (DMs). The UC Davis AO flood-illuminated fundus camera system described in this paper has been previously used to acquire in vivo images of the photoreceptor mosaic and for psychophysical studies on normal and diseased retinas. These results were acquired using a DM manufactured by Litton ITEK (DM109), which has 109 actuators arranged in a hexagonal array below a continuous front-surface mirror. It has an approximate surface actuator stroke of +/-2μm. Here we present results with a new hi-speed magnetic DM manufactured by ALPAO (DM97, voice coil technology), which has 97 actuators and similar inter-actuator stroke (>3μm, mirror surface) but much higher low-order aberration correction (defocus stroke of at least +/-30μm) than the previous one. In this paper we report results of testing performance of the ALPAO DM for the correction of human eye aberrations. Additionally changes made to our AO flood

  18. Double tuned 23Na 1H nuclear magnetic resonance birdcage for application on mice in vivo

    NASA Astrophysics Data System (ADS)

    Lanz, Titus; Ruff, Jan; Weisser, Alexander; Haase, Axel

    2001-05-01

    The design and the characterization of a double tuned nuclear magnetic birdcage resonator is presented. It abandons quadrature drive and uses the two orthogonal modes of the birdcage for two different frequencies. In order to tune the birdcage to frequencies that are far apart, the number of legs is reduced to only four. This limits the homogeneity of the rf field, but enables the birdcage to be tuned to very different frequencies without further B1 field distortions. Following a brief explanation of the theory of the coil design, a 23Na 1H four leg birdcage for in vivo measurements on mice is presented. The performance of the coil is demonstrated in experiments on both a phantom and a mouse.

  19. Intelligent spectral signature bio-imaging in vivo for surgical applications

    NASA Astrophysics Data System (ADS)

    Jeong, Jihoon; Frykman, Philip K.; Gaon, Mark; Chung, Alice P.; Lindsley, Erik H.; Hwang, Jae Y.; Farkas, Daniel L.

    2007-02-01

    Multi-spectral imaging provides digital images of a scene or object at a large, usually sequential number of wavelengths, generating precise optical spectra at every pixel. We use the term "spectral signature" for a quantitative plot of optical property variations as a function of wavelengths. We present here intelligent spectral signature bio-imaging methods we developed, including automatic signature selection based on machine learning algorithms and database search-based automatic color allocations, and selected visualization schemes matching these approaches. Using this intelligent spectral signature bio-imaging method, we could discriminate normal and aganglionic colon tissue of the Hirschsprung's disease mouse model with over 95% sensitivity and specificity in various similarity measure methods and various anatomic organs such as parathyroid gland, thyroid gland and pre-tracheal fat in dissected neck of the rat in vivo.

  20. Regulatory Experience with In Vivo In Vitro Correlations (IVIVC) in New Drug Applications.

    PubMed

    Suarez-Sharp, Sandra; Li, Min; Duan, John; Shah, Heta; Seo, Paul

    2016-11-01

    In the past two decades, in vitro in vivo correlation (IVIVC) has been considered an important tool for supporting biowaivers, setting dissolution acceptance criteria, and more recently in the Quality by Design (QbD) framework promoting the establishment of clinically meaningful drug product specifications using dissolution as the endpoint. Based on our review experience at the FDA, for the purposes of this article, we analyzed the current state of regulatory submissions containing IVIVC approaches and discussed the successes and failures from the perspectives of study design to methodology. In the past decade, the overall acceptance rate of the IVIVC submissions is about 40%. Moreover, the number of IVIVC studies seen in the submissions per year is not increasing. Establishing clinically meaningful drug product specifications through the linkages between the identified critical quality attributes and in vivo performance is key for developing a quality drug product. To achieve this goal, there is an imminent need for addressing the issues behind a low success rate in IVIVC development. The results from the current analysis revealed that special considerations should be taken in areas such as (1) selection of appropriate number/kind of formulations for IVIVC development/validation, (2) construction of exploratory plots to guide model building and selection, (3) investigation of the reasons of inconclusive predictability, (4) improvement on the quality and richness of the data, and (5) avoidance of over parameterization. The development and incorporation of biopredictive dissolution methods and the use of non-conventional approaches, including mechanistic/physiologically based approaches, should be explored to increase the likelihood of IVIVC success.

  1. Application Of Micro-Highspeed Flow Visualization In Study Of Blood Cells Rheology In Vivo

    NASA Astrophysics Data System (ADS)

    Gui-shah, Li; Ni, Liang; Yu-ju, Lin; Jian, Zhang; Qiang, Wang

    1990-01-01

    A new experimental method has been developed in study of rheological behaviour of single red blood cell (RBC) in passing through the capillaries in vivo, using the technique of micro-highspeed cinecamera and micro-highspeed video system. It is one of the most important topics in the study of microcirculatory theories that fur-ther understand the deformability of RBC, flow states, velocities and dynamic mechanimi. A micro-highspeed flow visualization system consisted of essential elements: a biological microscope, a highspeed cinecmera with 35 mm film, a highspeed motion analysis system SP2000 (Kodak U.S.A) and a cold-light source etc. We have investigated the rheological parameters of single RBC in vivo in single capillaries which are about 3.3 to 6.9 um in diameters. The RBCs velocities are 0.1 to 0.25 mm/sec, and maximum shear stress on the outside surface of RBC is 13.8 dyn/cml, and maximum extension of RBC is 10.3 um. In aforementioned experiment, the highspeed flow visualization system frequency at 530 frames/sec and 200 frames/sec were used respectively. In addition, the vasomotion of precapillary sphincters have been measured and a complicated coupling phenomena between the RBC and sphincter have also been recorded and analysed. The experiment were performed with intravital hamsters and frogs. The results obtained by this system shown that the method designed by us are an effective tool in the study of rheological behaviour of single RBC in passing through the blood capillaries in vivoz.

  2. In vivo determination of the diclofenac skin reservoir: comparison between passive, occlusive, and iontophoretic application

    PubMed Central

    Clijsen, Ron; Baeyens, Jean Pierre; Barel, André Odilon; Clarys, Peter

    2015-01-01

    Aim There is scarce information concerning the pharmacodynamic behavior of topical substances used in the physiotherapy setting. The aim of the present study was to estimate the formation and emptying of the diclofenac (DF) skin reservoir after passive, semiocclusive, and electrically assisted applications of DF. Subjects and methods Five different groups of healthy volunteers (ntotal=60, 23 male and 37 female), participated in this study. A 1% DF (Voltaren Emulgel) formulation (12 mg) was applied on the volar forearms on randomized defined circular skin areas of 7 cm2. DF was applied for 20 minutes under three different conditions at the same time. The presence of DF in the skin results in a reduction of the methyl nicotinate (MN) response. To estimate the bioavailability of DF in the skin, MN responses at different times following initial DF application (1.5, 6, 24, 32, 48, 72, 96, and 120 hours) were analyzed. Results At 1.5 hours after the initial DF application, a significant decrease in MN response was detected for the occluded and iontophoretic delivery. Passive application resulted in a decrease of the MN response from 6 hours post-DF application. The inhibition remained up to 32 hours post-DF application for the iontophoretic delivery, 48 hours for the occluded application, and 72 hours for the passive delivery. At 96 and 120 hours post-DF application none of the MN responses was inhibited. Conclusion The formation and emptying of a DF skin reservoir was found to be dependent on the DF-application mode. Penetration-enhanced delivery resulted in a faster emptying of the reservoir. PMID:25709408

  3. Medical applications of in vivo neutron inelastic scattering and neutron activation analysis: Technical similarities to detection of explosives and contraband

    NASA Astrophysics Data System (ADS)

    Kehayias, J. J.

    2001-07-01

    Nutritional status of patients can be evaluated by monitoring changes in elemental body composition. Fast neutron activation (for N and P) and neutron inelastic scattering (for C and O) are used in vivo to assess elements characteristic of specific body compartments. There are similarities between the body composition techniques and the detection of hidden explosives and narcotics. All samples have to be examined in depth and the ratio of elements provides a "signature" of the chemical of interest. The N/H and C/O ratios measure protein and fat content in the body. Similarly, a high C/O ratio is characteristic of narcotics and a low C/O together with a strong presence of N is a signature of some explosives. The available time for medical applications is about 20 min—compared to a few seconds for the detection of explosives—but the permitted radiation exposure is limited. In vivo neutron analysis is used to measure H, O, C, N, P, Na, Cl, and Ca for the study of the mechanisms of lean tissue depletion with aging and wasting diseases, and to investigate methods of preserving function and quality of life in the elderly.

  4. Optically deviated focusing method based high-speed SD-OCT for in vivo retinal clinical applications

    NASA Astrophysics Data System (ADS)

    Wijesinghe, Ruchire Eranga; Park, Kibeom; Kim, Pilun; Oh, Jaeryung; Kim, Seong-Woo; Kim, Kwangtae; Kim, Beop-Min; Jeon, Mansik; Kim, Jeehyun

    2016-04-01

    The aim of this study is to provide accurately focused, high-resolution in vivo human retinal depth images using an optically deviated focusing method with spectral-domain optical coherence tomography (SD-OCT) system. The proposed method was applied to increase the retinal diagnosing speed of patients with various values of retinal distances (i.e., the distance between the crystalline eye lens and the retina). The increased diagnosing speed was facilitated through an optical modification in the OCT sample arm configuration. Moreover, the optical path length matching process was compensated using the proposed optically deviated focusing method. The developed system was mounted on a bench-top cradle to overcome the motion artifacts. Further, we demonstrated the capability of the system by carrying out in vivo retinal imaging experiments. The clinical trials confirmed that the system was effective in diagnosing normal and abnormal retinal layers as several retinal abnormalities were identified using non-averaged single-shot OCT images, which demonstrate the feasibility of the method for clinical applications.

  5. Topical application of ex vivo expanded endothelial progenitor cells promotes vascularisation and wound healing in diabetic mice.

    PubMed

    Asai, Jun; Takenaka, Hideya; Ii, Masaaki; Asahi, Michio; Kishimoto, Saburo; Katoh, Norito; Losordo, Douglas W

    2013-10-01

    Impaired wound healing leading to skin ulceration is a serious complication of diabetes and may be caused by defective angiogenesis. Endothelial progenitor cells (EPCs) can augment neovascularisation in the ischaemic tissue. Experiments were performed to test the hypothesis that locally administered EPCs can promote wound healing in diabetes. Full-thickness skin wounds were created on the dorsum of diabetic mice. EPCs were obtained from bone marrow mononuclear cells (BMMNCs) and applied topically to the wound immediately after surgery. Vehicle and non-selective BMMNCs were used as controls. Wound size was measured on days 5, 10 and 14 after treatment, followed by resection, histological analysis and quantification of vascularity. Topical application of EPCs significantly promoted wound healing, as assessed by closure rate and wound vascularity. Immunostaining revealed that transplanted EPCs induced increased expression of vascular endothelial growth factor and basic fibroblast growth factor. Few EPCs were observed in the neovasculature based on in vivo staining of the functional vasculature. Ex vivo expanded EPCs promote wound healing in diabetic mice via mechanisms involving increased local cytokine expression and enhanced neovascularisation of the wound. This strategy exploiting the therapeutic capacity of autologously derived EPCs may be a novel approach to skin repair in diabetes.

  6. Application of the moving-actuator type pump as a ventricular assist device: in vitro and in vivo studies.

    PubMed

    Lee, H S; Rho, Y R; Park, C Y; Hwang, C M; Kim, W G; Sun, K; Choi, M J; Lee, K K; Cheong, J T; Shim, E B; Min, B G

    2002-06-01

    A moving actuator type pump has been developed as a multifunctional Korean artificial heart (AnyHeart). The pump consists of a moving actuator as an energy converter, right and left sacs, polymer (or mechanical) valves, and a rigid polyurethane housing. The actuator containing a brushless DC motor moves back and forth on an epicyclical gear train to produce a pendular motion, which compresses both sacs alternately. Of its versatile functions of ventricular assist device and total artificial heart use, we have evaluated the system performance as a single or biventricular assist device through in vitro and in vivo experiments. Pump performance and anatomical feasibility were tested using various animals of different sizes. In the case of single ventricular assist device (VAD) use, one of the sacs remained empty and a mini-compliance chamber was attached to either an outflow or inflow port of the unused sac. The in vitro and in vivo studies show acceptable performance and pump behavior. Further extensive study is required to proceed to human application.

  7. Potential application of silver nanoparticles to control the infectivity of Rift Valley fever virus in vitro and in vivo.

    PubMed

    Borrego, Belén; Lorenzo, Gema; Mota-Morales, Josué D; Almanza-Reyes, Horacio; Mateos, Francisco; López-Gil, Elena; de la Losa, Nuria; Burmistrov, Vasily A; Pestryakov, Alexey N; Brun, Alejandro; Bogdanchikova, Nina

    2016-07-01

    In this work we have tested the potential antiviral activity of silver nanoparticles formulated as Argovit™ against Rift Valley fever virus (RVFV). The antiviral activity of Argovit was tested on Vero cell cultures and in type-I interferon receptor deficient mice (IFNAR (-/-) mice) by two different approaches: (i) different dilutions of Argovit were added to previously infected cells or administrated to animals infected with a lethal dose of virus; (ii) virus was pre-incubated with different dilutions of Argovit before inoculation in mice or cells. Though the ability of silver nanoparticles to control an ongoing RVFV infection in the conditions tested was limited, the incubation of virus with Argovit before the infection led to a reduction of the infectivity titers both in vitro and in vivo. These results reveal the potential application of silver nanoparticles to control the infectivity of RVFV, which is an important zoonotic pathogen.

  8. Enhanced in vivo visualization of the microcirculation by topical application of fructose solution confirmed with correlation mapping optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Enfield, Joey; McGrath, James; Daly, Susan M.; Leahy, Martin

    2016-08-01

    Changes within the microcirculation can provide an early indication of the onset of a plethora of ailments. Various techniques have thus been developed that enable the study of microcirculatory irregularities. Correlation mapping optical coherence tomography (cmOCT) is a recently proposed technique, which enables mapping of vasculature networks at the capillary level in a noninvasive and noncontact manner. This technique is an extension of conventional optical coherence tomography (OCT) and is therefore likewise limited in the penetration depth of ballistic photons in biological media. Optical clearing has previously been demonstrated to enhance the penetration depth and the imaging capabilities of OCT. In order to enhance the achievable maximum imaging depth, we propose the use of optical clearing in conjunction with the cmOCT technique. We demonstrate in vivo a 13% increase in OCT penetration depth by topical application of a high-concentration fructose solution, thereby enabling the visualization of vessel features at deeper depths within the tissue.

  9. The antioxidant capacity of milk--the application of different methods in vitro and in vivo.

    PubMed

    Cloetens, L; Panee, J; Åkesson, B

    2013-11-03

    Milk contains a wide array of compounds with established or putative pro- or anti-oxidant function. The functions of these compounds have been intensively studied. This review focusses on some important aspects in this wide field namely the methodology for measurement of the total antioxidant capacity (TAC), the content of TAC and some related compounds in human and animal milks and infant formulas, and the effect of milk intake on antioxidant status in the body and on the activity of dietary flavonoids as studied in vitro and in vivo. Regarding methodology TAC in milk can be measured by spectrophotometric and electrochemical methods and some of their characteristics are reviewed. Milk, whey, high-molecular-weight and low-molecular-weight (LMW) fractions of whey have all been found to have antioxidant capacity using these techniques. The major antioxidant in the LMW fraction has been identified as urate. An extensive literature survey was made regarding data on the antioxidant capacity and related variables of milk obtained from different sources (human milk, infant formulas and animal milk) and subjected to different treatments. Differences in TAC between milks from different sources have been observed but due to the variety of techniques used no clear pattern is evident at present. Another important aspect is the putative effects of the intake of milk products on the antioxidant status of the consumer. A few studies performed in adults and premature infants are reviewed and it is stated that too little information is available to make any firm conclusions in this regard. Finally, a high interest has been devoted to the possible interference of milk with the antioxidant properties of flavonoid-rich food like tea. Most in vitro studies show an inhibition by milk on tea flavonoid activity whereas the results from the corresponding in vivo studies are equivocal. Our general conclusion is that several compounds in various milk fractions contribute to the antioxidant

  10. Programmable oligonucleotide probes design and applications for in situ and in vivo RNA imaging in cells

    NASA Astrophysics Data System (ADS)

    Cheglakov, Zoya

    Unequal spreading of mRNA is a frequent experience observed in varied cell lines. The study of cellular processes dynamics and precise localization of mRNAs offers a vital toolbox to target specific proteins in precise cytoplasmic areas and provides a convenient instrument to uncover their mechanisms and functions. Latest methodological innovations have allowed imaging of a single mRNA molecule in situ and in vivo. Today, Fluorescent In Situ Hybridization (FISH) methods allow the studying of mRNA expression and offer a vital toolbox for accurate biological models. Studies enable analysis of the dynamics of an individual mRNA, have uncovered the multiplex RNA transport systems. With all current approaches, a single mRNA tracking in the mammalian cells is still challenging. This thesis describes mRNA detection methods based on programmable fluorophore-labeled DNA structures complimentary to native targets providing an accurate mRNA imaging in mammalian cells. First method represents beta-actin (ACTB) transcripts in situ detection in human cells, the technique strategy is based on programmable DNA probes, amplified by rolling circle amplification (RCA). The method reports precise localization of molecule of interest with an accuracy of a single-cell. Visualization and localization of specific endogenous mRNA molecules in real-time in vivo has the promising to innovate cellular biology studies, medical analysis and to provide a vital toolbox in drugs invention area. Second method described in this thesis represents miR-21 miRNA detection within a single live-cell resolution. The method using fluorophore-labeled short synthetic DNAs probes forming a stem-loop shape and generating Fluorescent Resonance Energy Transfer (FRET) as a result of target-probes hybridization. Catalytic nucleic acid (DNAzymes) probes are cooperative tool for precise detection of different mRNA targets. With assistance of a complementary fluorophore-quencher labeled substrate, the DNAzymes provide

  11. Near-IR-Absorbing Gold Nanoframes with Enhanced Physiological Stability and Improved Biocompatibility for In Vivo Biomedical Applications.

    PubMed

    Wang, Liying; Chen, Yunching; Lin, Hsin Yao; Hou, Yung-Te; Yang, Ling-Chu; Sun, Aileen Y; Liu, Jia-Yu; Chang, Chien-Wen; Wan, Dehui

    2017-02-01

    This paper describes the synthesis of near-infrared (NIR)-absorbing gold nanoframes (GNFs) and a systematic study comparing their physiological stability and biocompatibility with those of hollow Au-Ag nanoshells (GNSs), which have been used widely as photothermal agents in biomedical applications because of their localized surface plasmon resonance (LSPR) in the NIR region. The GNFs were synthesized in three steps: galvanic replacement, Au deposition, and Ag dealloying, using silver nanospheres (SNP) as the starting material. The morphology and optical properties of the GNFs were dependent on the thickness of the Au coating layer and the degree of Ag dealloying. The optimal GNF exhibited a robust spherical skeleton composed of a few thick rims, but preserved the distinctive LSPR absorbance in the NIR region-even when the Ag content within the skeleton was only 10 wt %, 4-fold lower than that of the GNSs. These GNFs displayed an attractive photothermal conversion ability and great photothermal stability, and could efficiently kill 4T1 cancer cells through light-induced heating. Moreover, the GNFs preserved their morphology and optical properties after incubation in biological media (e.g., saline, serum), whereas the GNSs were unstable under the same conditions because of rapid dissolution of the considerable silver content with the shell. Furthermore, the GNFs had good biocompatibility with normal cells (e.g., NIH-3T3 and hepatocytes; cell viability for both cells: >90%), whereas the GNSs exhibited significant dose-dependent cytotoxicity (e.g., cell viability for hepatocytes at 1.14 nM: ca. 11%), accompanied by the induction of reactive oxygen species. Finally, the GNFs displayed good biocompatibility and biosafety in an in vivo mouse model; in contrast, the accumulation of GNSs caused liver injury and inflammation. Our results suggest that GNFs have great potential to serve as stable, biocompatible NIR-light absorbers for in vivo applications, including cancer

  12. In vivo experiment leading to clinical application of an electrohydraulic ventricular assist device with magnetic coupling.

    PubMed

    Kim, W G; Choi, J S; Won, Y S; Jo, Y H; Park, S K; Chung, C I; Kim, J; Min, B G; Ahn, H; Rho, J R

    1999-01-01

    We developed an electrohydraulic ventricular assist device with magnetic coupling. The integrated system consists of a blood pump, a water conduit for pressure transmission, a bellows type pumping sac, an actuator for transforming the circular motion of a motor to the linear motion of a pusher plate attached to the pumping sac with magnetic coupling, and a controller. The purpose of the coupling was to prevent excessive sucking against the atrial wall. Number 21 Medtronic Hall (Irvine, CA) mechanical valves were used in the inflow and outflow ports of the blood pump. Maximum dynamic stroke volume was 48 ml, and against a mean afterload of 100 mm Hg, maximum pump output was 7 L/min. Chronic in vivo experiments were performed in three sheep, and during these evaluations the system showed no noticeable problems related to mechanical or electronic devices. When left atrial pressure decreased below 0 mm Hg, the magnetic coupling system decoupled the pumping sac and pusher plate with satisfactory reliability. The device was clinically applied in a postoperative patient with chronic dilating cardiomyopathy, and no significant device related problems ensued. These results prove that the electrohydraulic ventricular assist system with magnetic coupling is a suitable ventricular assist device.

  13. In vivo biochemistry: Applications for small molecule biosensors in plant biology

    PubMed Central

    Jones, Alexander; Grossmann, Guido; Danielson, Jonas Å.H.; Sosso, Davide; Chen, Li-Qing; Ho, Cheng Hsun; Frommer, Wolf B.

    2013-01-01

    Summary Revolutionary new technologies, namely in the areas of DNA sequencing and molecular imaging, continue to impact new discoveries in plant science and beyond. For decades we have been able to determine properties of enzymes, receptors and transporters in vitro or in heterologous systems, and more recently been able to analyze their regulation at the transcriptional level, use GFP reporters to obtain insights into cellular and subcellular localization, and measure ion and metabolite levels with unprecedented precision using mass spectrometry. However, we lack key information on location and dynamics of the substrates of enzymes, receptors and transporters, and on the regulation of these proteins in their cellular environment. Such information can now be obtained by transitioning from in vitro to in vivo biochemistry using biosensors. Genetically encoded fluorescent protein-based sensors for ion and metabolite dynamics provide highly resolved spatial and temporal information, and are complemented by sensors for pH, redox, voltage, and tension. They serve as powerful tools for identifying missing processes (e.g. glucose transport across ER membranes), components (e.g. SWEET sugar transporters for cellular sugar efflux), and signaling networks (e.g. from systematic screening of mutants that affect sugar transport or cytosolic and vacuolar pH). Combined with the knowledge of properties of enzymes and transporters and their interactions with the regulatory machinery, biosensors promise to be key diagnostic tools for systems and synthetic biology. PMID:23587939

  14. Simultaneous Determination of 11 Illicit Phenethylamines in Hair by LC-MS-MS: In Vivo Application.

    PubMed

    Nieddu, Maria; Burrai, Lucia; Demontis, Maria Piera; Varoni, Maria Vittoria; Baralla, Elena; Trignano, Claudia; Boatto, Gianpiero

    2015-09-01

    Existing phenethylamines are a class of synthetic compounds that differ from each other only in small changes to a largely conserved chemical structure. The recreational and illicit use of phenethylamines is a widespread problem. A simple procedure for the simultaneous quantitative determination in hair of 11 phenethylamines that are officially recognized as illicit by Italian legislation (p-methoxyamphetamine; p-methoxymethamphetamine; 3,4,5-trimethoxyamphetamine; 2,5-dimethoxyamphetamine; 2,5-dimethoxy-4-methylamphetamine; 2,5-dimethoxy-4-ethylamphetamine; 2,5-dimethoxy-4-bromoamphetamine; 2,5-dimethoxy-4-bromophenethylamine; 2,5-dimethoxy-4-iodophenethylamine; 2,5-dimethoxy-4-ethylthiophenethylamine and 2,5-dimethoxy-4-n-propylthiophenethylamine) has been developed and validated. Extraction from the matrix was performed after incubation in methanolic HCl and filtered reconstituted extracts were injected into a liquid chromatography/tandem mass spectrometry system (LC-MS-MS) without any further purification steps. This validated LC-MS-MS method has been used to determine the in vivo accumulation/retention of the above target analytes in hair after repeat oral administration to rats. This experiment further permitted investigation of the effect of pigmentation on the uptake of these phenethylamines by hair and the effect of hair pigmentation. The developed method could potentially be used for forensic and toxicological purposes, in the detection and quantitation of these illicit substances in human hair in workplace drug testing; drug-facilitated crime investigation; driver re-licensing; determining drug abuse history and postmortem toxicology.

  15. Endosomolytic anionic polymer for the cytoplasmic delivery of siRNAs in localized in vivo applications

    PubMed Central

    Khormaee, Sariah; Choi, Yong; Shen, Michael J.; Xu, Biying; Wu, Haitao; Griffiths, Gary L.; Chen, Rongjun; Slater, Nigel K. H.; Park, John K.

    2013-01-01

    The use of small interfering RNAs (siRNAs) to down-regulate the expression of disease-associated proteins carries significant promise for the treatment of a variety of clinical disorders. One of the main barriers to the widespread clinical use of siRNAs, however, is their entrapment and degradation within the endolysosomal pathway of target cells. Here we report the trafficking and function of PP75, a non-toxic, biodegradable, lipid membrane disruptive anionic polymer composed of phenylalanine derivatized poly(L-lysine iso-phthalamide). PP75 is readily endocytosed by cells, safely permeabilizes endolysosomes in a pH dependent manner and facilitates the transfer of co-endocytosed materials directly into the cytoplasm. The covalent attachment of siRNAs to PP75 using disulfide linkages generates conjugates that effectively traffic siRNAs to the cytoplasm of target cells both in vitro and in vivo. In a subcutaneous malignant glioma tumor model, a locally delivered PP75-stathmin siRNA conjugate decreases stathmin expression in tumor cells and, in combination with the nitrosourea chemotherapy carmustine, is highly effective at inhibiting tumor growth. PP75 may be clinically useful for the local delivery of siRNAs, in particular for the treatment of solid tumors. PMID:24273480

  16. Propagation of normal human epithelial cell populations using an in vivo culture system. Description and applications.

    PubMed Central

    Klein-Szanto, A. J.; Terzaghi, M.; Mirkin, L. D.; Martin, D.; Shiba, M.

    1982-01-01

    A new model using xenotransplanted human epithelia was developed for the study of toxic and carcinogenic effects of chemicals. Epithelial cells from the respiratory tract of 4 male and 3 female premature and fullterm fetuses were enzymatically removed and inoculated into deepithelialized rat tracheas. These were sealed at both ends and transplanted subcutaneously into nude mice. After 3-4 weeks, a normal mucociliary epithelium covered the tracheal lumen. At this stage the epithelial cells could be isolated again and transplanted into new denuded rat tracheas. This passaging could be repeated up to six times, each permitting an amplification factor of approximately 3. Tracheal transplants containing cells of human origin (in vivo Passages 2-4) were treated with 7,12-dimethylbenz(a)anthracene. Hyperplasias, squamous metaplasias, and dysplasias were seen 1-8 weeks after initiation of treatment, indicating that the responses of human and rodent epithelial cells to polycyclic aromatic hydrocarbons are similar. Initial experiments with skin and esophageal epithelia suggest that other covering epithelia could also be used in this fashion for evaluation of toxicants and carcinogens that are likely to come into contact with these tissues. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:6821529

  17. Raman spectroscopy and the spectral correlation index for predicting wound healing outcome: towards in vivo application

    NASA Astrophysics Data System (ADS)

    Berger, Adam G.; Crane, Nicole J.; Elster, Eric A.

    2016-03-01

    Combat wounds are sometimes confounded by healing complications that are not as prevalent in civilian wounds due to their high energy etiology. One complication of wound healing is dehiscence, where a surgically closed wound reopens after closure. This complication can have serious consequences for the patient, but knowledge about the molecular composition of the wound bed beyond what is readily visible may help clinicians mitigate these complications. It is necessary to develop techniques that can be used in vivo to assess and predict wound healing pointof- care so that care-takers can decide the best way to make informed clinical decisions regarding their patient's healing. Raman spectroscopy is a perfect candidate for predicting wound healing due to its ability to provide a detailed molecular fingerprint of the wound bed noninvasively. Here, we study the spectral correlation index, a measure of orthogonality, with ten reference tissue components to stratify wounds based on how they heal. We analyze these indexes over time to show the modulation of these tissue components over the wound healing process. Results show that qualitative observation of the spectra cannot reveal major differences between the dehisced and normal healing wounds, but the spectral correlation index can. Analysis of the spectral correlations across the wound healing process demonstrates the changes throughout the wound healing process, showing that early differences in tissue components may portend wound healing. Furthermore, Raman spectroscopy coupled with the spectral correlation index presents as a possible point-of-care tool for enabling discrimination of wounds with impaired healing.

  18. In Vivo Fate of Carbon Nanotubes with Different Physicochemical Properties for Gene Delivery Applications.

    PubMed

    Cifuentes-Rius, Anna; Boase, Nathan R B; Font, Ines; Coronas, Nuria; Ramos-Perez, Victor; Thurecht, Kristofer J; Borrós, Salvador

    2017-04-05

    Gene therapy has arisen as a pioneering technique to treat diseases by direct employment of nucleic acids as medicine. The major historical problem is to develop efficient and safe systems for the delivery of therapeutic genes into the target cells. Carbon nanotubes (CNTs) have demonstrated considerable promise as delivery vectors due to their (i) high aspect ratio and (ii) capacity to translocate through plasma membranes, known as the nanoneedle effect. To leverage these advantages, close attention needs to be paid to the physicochemical characteristics of the CNTs used. CNTs with different diameters (thinner and thicker) were treated by chemical oxidation to produce shorter fragments. Rigid (thick) and flexible (thin) CNTs, and their shortened versions, were coated with polyallylamine (ppAA) by plasma-enhanced chemical vapor deposition. The ppAA coating leads to a positively charged CNT surface that is able to electrostatically bind the green fluorescent protein plasmid reporter. This study shows how rigidity and length can affect their (i) behavior in biological media, (ii) ability to transfect in vitro, and (iii) biodistribution in vivo. This study also generates a set of basic design rules for the development of more efficient CNT-based gene-delivery vectors.

  19. In vivo application of short-lag spatial coherence imaging in human liver.

    PubMed

    Jakovljevic, Marko; Trahey, Gregg E; Nelson, Rendon C; Dahl, Jeremy J

    2013-03-01

    We present the results of a patient study conducted to assess the performance of two novel imaging methods, namely short-lag spatial coherence (SLSC) and harmonic spatial coherence imaging (HSCI), in an in vivo liver environment. Similar in appearance to the B-mode images, SLSC and HSCI images are based solely on the spatial coherence of fundamental and harmonic echo data, respectively, and do not depend on the echo magnitude. SLSC and HSCI suppress incoherent echo signals and thus tend to reduce clutter. The SLSC and HSCI images of 17 patients demonstrated sharper delineation of blood vessel walls, suppressed clutter inside the vessel lumen, and showed reduced speckle in surrounding tissue compared to matched B-modes. Target contrast and contrast-to-noise ratio (CNR) show statistically significant improvements between fundamental B-mode and SLSC imaging and between harmonic B-mode and HSCI imaging (in all cases p < 0.001). The magnitude of improvement in contrast and CNR increases as the overall quality of B-mode images decreases. Poor-quality fundamental B-mode images (where image quality classification is based on both contrast and CNR) exhibit the highest improvements in both contrast and CNR (288% improvement in contrast and 533% improvement in CNR).

  20. Nanomiemgel - A Novel Drug Delivery System for Topical Application - In Vitro and In Vivo Evaluation

    PubMed Central

    Somagoni, Jaganmohan; Boakye, Cedar H. A.; Godugu, Chandraiah; Patel, Apurva R.; Mendonca Faria, Henrique Antonio; Zucolotto, Valtencir; Singh, Mandip

    2014-01-01

    Aim The objective of this study was to formulate and evaluate a unique matrix mixture (nanomiemgel) of nanomicelle and nanoemulsion containing aceclofenac and capsaicin using in vitro and in vivo analyses and to compare it to a marketed formulation (Aceproxyvon). Methods Nanomicelles were prepared using Vitamin E TPGS by solvent evaporation method and nanoemulsion was prepared by high-pressure homogenization method. In vitro drug release and human skin permeation studies were performed and analyzed using HPLC. The efficiency of nanomiemgel as a delivery system was investigated using an imiquimod-induced psoriatic like plaque model developed in C57BL/6 mice. Results Atomic Force Microscopy images of the samples exhibited a globular morphology with an average diameter of 200, 250 and 220 nm for NMI, NEM and NMG, respectively. Nanomiemgel demonstrated a controlled release drug pattern and induced 2.02 and 1.97-fold more permeation of aceclofenac and capsaicin, respectively than Aceproxyvon through dermatomed human skin. Nanomiemgel also showed 2.94 and 2.09-fold greater Cmax of aceclofenac and capsaicin, respectively than Aceproxyvon in skin microdialysis study in rats. The PASI score, ear thickness and spleen weight of the imiquimod-induced psoriatic-like plaque model were significantly (p<0.05) reduced in NMG treated mice compared to free drug, NEM, NMI & Aceproxyvon. Conclusion Using a new combination of two different drug delivery systems (NEM+NMI), the absorption of the combined system (NMG) was found to be better than either of the individual drug delivery systems due to the utilization of the maximum possible paths of absorption available for that particular drug. PMID:25546392

  1. In vivo efficacy and bioavailability of lumefantrine: Evaluating the application of Pheroid technology.

    PubMed

    du Plessis, Lissinda H; Govender, Katya; Denti, Paolo; Wiesner, Lubbe

    2015-11-01

    The oral absorption of compounds with low aqueous solubility, such as lumefantrine, is typically limited by the dissolution rate in the gastro-intestinal tract, resulting in erratic absorption and highly variable bioavailability. In previous studies we reported on the ability of Pheroid vesicles to improve the bioavailability of poorly soluble drugs. In the present study a Pro-Pheroid formulation, a modification of the previous formulation, was applied to improve the solubility of lumefantrine after oral administration and compared to lumefantrine in DMSO:water (1:9 v/v) solution (reference solution). A bioavailability study of lumefantrine was conducted in a mouse model in fed and fasted states. When using the reference solution, the bioavailability of the lumefantrine heavily depended on food intake, resulting in a 2.7 times higher bioavailability in the fed state when compared to the fasted state. It also showed large between-subject variability. When formulated using Pro-Pheroid, the bioavailability of lumefantrine was 3.5 times higher as compared to lumefantrine in the reference solution and fasting state. Pro-Pheroid also dramatically reduced the effects of food intake and the between-subject variability for bioavailability observed with the reference. In vivo antimalarial efficacy was also evaluated with lumefantrine formulated using Pro-Pheroid technology compared to the reference solution. The results indicated that lumefantrine in Pro-Pheroid formulation exhibited improved antimalarial activity in vitro by 46.8%, when compared to the reference. The results of the Peters' 4-day suppressive test indicated no significant difference in the efficacy or mean survival time of the mice in the Pro-Pheroid formulation and reference test groups when compared to the positive control, chloroquine. These findings suggest that using the Pro-Pheroid formulation improves the bioavailability of lumefantrine, eliminates the food effect associated with lumefantrine as well

  2. In vivo/ex vivo targeting of Langerhans cells after topical application of the immune response modifier TMX-202: confocal Raman microscopy and histology analysis

    NASA Astrophysics Data System (ADS)

    Darvin, Maxim E.; Thiede, Gisela; Ascencio, Saul Mujica; Schanzer, Sabine; Richter, Heike; Vinzón, Sabrina E.; Hasche, Daniel; Rösl, Frank; May, Roberto; Hazot, Yohan; Tamarkin, Dov; Lademann, Juergen

    2016-05-01

    The increased ability of TMX-202 (derivative of imiquimod) to penetrate the intact stratum corneum (SC) and the follicular orifices of porcine ear skin was shown ex vivo using confocal Raman microscopy and laser scanning microscopy. Moreover, to assess whether TMX-202 is able to reach the immune cells, Langerhans cells extracted from pretreated human skin were investigated ex vivo using confocal Raman microscopy combined with multivariate statistical methods. Tracking the Raman peak of dimethyl sulfoxide centered at 690 cm-1, the absorption of TMX-202 containing formulation by Langerhans cells was shown. To answer the question whether the TMX-202 active ingredient is able to reach Langerhans cells, the attraction of immune cells to TMX-202 containing formulation treated skin was measured in the in vivo rodent model Mastomys coucha. The results show that TMX-202 active ingredient is able to reach Langerhans cells after penetrating through the intact skin and subsequently attract immune cells. Both the intercellular/transcellular as well as the follicular pathways allow the penetration through the intact barrier of the SC.

  3. Noninvasive fluorescence excitation spectroscopy during application of 5-aminolevulinic acid in vivo.

    PubMed

    Juzenas, Petras; Juzeniene, Asta; Kaalhus, Olav; Iani, Vladimir; Moan, Johan

    2002-10-01

    The fluorescence of PpIX induced by topical application of 5-aminolevulinic acid (ALA) in normal mouse skin was studied noninvasively by means of a fibre optic probe. The fluorescence excitation spectrum of PpIX exhibits five distinct peaks at around 408. 510, 543, 583 and 633 nm under fluorescence monitoring at the second emission peak of PpIX (705 nm). The transmission of the excitation light is wavelength dependent: the long wavelength light (>600 nm) penetrates deeper into the tissues by a factor of 6 compared with the short wavelength light (<590 nm). Thus, the fluorescence excitation spectrum of PpIX measured on the surface of the skin can be used to estimate the depth of the penetration of topically applied ALA. The fluorescence excitation spectra calculated for the depth 1.1 mm obtained the best fit with the experimentally measured spectra after topical application of ALA.

  4. Surface coatings shape the protein corona of SPIONs with relevance to their application in vivo.

    PubMed

    Jedlovszky-Hajdú, Angéla; Bombelli, Francesca Baldelli; Monopoli, Marco P; Tombácz, Etelka; Dawson, Kenneth A

    2012-10-23

    Superparamagnetic iron oxide nanoparticles (SPIONs) have proved their use in many biomedical applications, such as drug delivery, hyperthermia, and MRI (magnetic resonance imaging) contrast agents. Due to their instability in fluids, several surface coatings have been used to both stabilize and tune the properties of these nanoparticles (NPs) according to their applications. These coatings will strongly modify their surface properties and influence their interaction with the environment proteins in a relevant biological medium with a clear impact on their function. It is well-accepted that a protein corona is immediately formed when nanoparticles come in contact with a biological milieu, and the emergent bionano interface represents the biological identity of the particles. Here, we investigate how a different coating on the same magnetic core can influence the protein corona composition and structure with clear relevance to application of these NPs in medicine. In particular, we have studied the structure and composition of the protein corona-SPION complexes of magnetite nanoparticles stabilized with citric acid, poly(acrylic acid), or double layer oleic acid by a range of approaches, including dynamic light scattering, nanoparticle tracking analysis, differential centrifugal sedimentation, infrared spectroscopy, 1-D SDS gel electrophoresis, and mass spectroscopy.

  5. PEGylation of superparamagnetic iron oxide nanoparticle for drug delivery applications with decreased toxicity: an in vivo study

    NASA Astrophysics Data System (ADS)

    Prabhu, Suma; Mutalik, Srinivas; Rai, Sharada; Udupa, Nayanabhirama; Rao, Bola Sadashiva Satish

    2015-10-01

    Superparamagnetic iron oxide nanoparticles (SPIONs) are evolving as a mainstay across various applications in the field of Science and Technology. SPIONs have enticed attention on the grounds of their unique physicochemical properties as well as potential applications in magnetic hyperthermia, immunoassays, as a contrast agent in magnetic resonance imaging and targeted drug delivery among others. Toward this goal, we synthesized SPIONs by chemical co-precipitation and PEGylated it. PEGylated SPIONs (PS) were studied for its detailed in vivo toxicity profile, in view of further surface engineering for its clinical applications. The intravenous LD50(14) of the PS was ascertained as 508.16 ± 41.52 mg/kg b wt. Histopathology of the vital organs of the animals injected with acute toxic doses showed pathological changes in spleen, lung, liver, and kidney. Accumulation of SPION was found in the aforementioned organs as confirmed by Prussian blue staining. Further, 1/10th dose of LD50(14) of PS and the Bare SPION (BS) was used to analyze a detailed toxicity profile, including genotoxicity (micronuclei formation and chromosomal aberration assays), organ-specific toxicity (a detailed serum biochemical analysis), and also determination of oxidative stress. The results of toxicity profile indicated no significant toxicity due to systemic exposure of PS. Atomic absorption spectroscopy (AAS) analysis confirmed the accumulation of SPION majorly in lungs, liver spleen, and kidneys. The present study thus indicated an optimal dose of PS which could be used for surface modification for targeted drug delivery applications with least toxicity.

  6. Modulating Gold Nanoparticle in vivo Delivery for Photothermal Therapy Applications Using a T Cell Delivery System

    NASA Astrophysics Data System (ADS)

    Kennedy, Laura Carpin

    This thesis reports new gold nanoparticle-based methods to treat chemotherapy-resistant and metastatic tumors that frequently evade conventional cancer therapies. Gold nanoparticles represent an innovative generation of diagnostic and treatment agents due to the ease with which they can be tuned to scatter or absorb a chosen wavelength of light. One area of intensive investigation in recent years is gold nanoparticle photothermal therapy (PTT), in which gold nanoparticles are used to heat and destroy cancer. This work demonstrates the utility of gold nanoparticle PTT against two categories of cancer that are currently a clinical challenge: trastuzumab-resistant breast cancer and metastatic cancer. In addition, this thesis presents a new method of gold nanoparticle delivery using T cells that increases gold nanoparticle tumor accumulation efficiency, a current challenge in the field of PTT. I ablated trastuzumab-resistant breast cancer in vitro for the first time using anti-HER2 labeled silica-gold nanoshells, demonstrating the potential utility of PTT against chemotherapy-resistant cancers. I next established for the first time the use of T cells as gold nanoparticle vehicles in vivo. When incubated with gold nanoparticles in culture, T cells can internalize up to 15000 nanoparticles per cell with no detrimental effects to T cell viability or function (e.g. migration and cytokine secretion). These AuNP-T cells can be systemically administered to tumor-bearing mice and deliver gold nanoparticles four times more efficiently than by injecting free nanoparticles. In addition, the biodistribution of AuNP-T cells correlates with the normal biodistribution of T cell carrier, suggesting the gold nanoparticle biodistribution can be modulated through the choice of nanoparticle vehicle. Finally, I apply gold nanoparticle PTT as an adjuvant treatment for T cell adoptive transfer immunotherapy (Hyperthermia-Enhanced Immunotherapy or HIT) of distant tumors in a melanoma mouse

  7. Chemical modifications of artificial restriction DNA cutter (ARCUT) to promote its in vivo and in vitro applications

    PubMed Central

    Komiyama, Makoto

    2014-01-01

    ABSTRACT Recently, completely chemistry-based tools for site-selective scission of DNA (ARCUT) have been prepared by combining 2 strands of pseudo-complementary PNA (pcPNA: site-selective activator) and a Ce(IV)-EDTA complex (molecular scissors). Its site-specificity is sufficient to cut the whole human genome at one predetermined site. In this first-generation ARCUT, however, there still remain several problems to be solved for wider applications. This review presents recent approaches to solve these problems. They are divided into (i) covalent modification of pcPNA with other functional groups and (ii) new strategies using conventional PNA, in place of pcPNA, as site-selective activator. Among various chemical modifications, conjugation with positively-charged nuclear localization signal peptide is especially effective. Furthermore, unimolecular activators, a single strand of which successfully activates the target site in DNA for site-selective scission, have been also developed. As the result of these modifications, the site-selective scission by Ce(IV)-EDTA was achieved promptly even under high salt conditions which are otherwise unfavourable for double-duplex invasion. Furthermore, it has been shown that “molecular crowding effect,” which characterizes the inside of living cells, enormously promotes the invasion, and thus the invasion seems to proceed effectively and spontaneously in the cells. Strong potential of pcPNA for further applications in vivo and in vitro has been confirmed. PMID:26744220

  8. Scalable wide-field optical coherence tomography-based angiography for in vivo imaging applications

    PubMed Central

    Xu, Jingjiang; Wei, Wei; Song, Shaozhen; Qi, Xiaoli; Wang, Ruikang K.

    2016-01-01

    Recent advances in optical coherence tomography (OCT)-based angiography have demonstrated a variety of biomedical applications in the diagnosis and therapeutic monitoring of diseases with vascular involvement. While promising, its imaging field of view (FOV) is however still limited (typically less than 9 mm2), which somehow slows down its clinical acceptance. In this paper, we report a high-speed spectral-domain OCT operating at 1310 nm to enable wide FOV up to 750 mm2. Using optical microangiography (OMAG) algorithm, we are able to map vascular networks within living biological tissues. Thanks to 2,048 pixel-array line scan InGaAs camera operating at 147 kHz scan rate, the system delivers a ranging depth of ~7.5 mm and provides wide-field OCT-based angiography at a single data acquisition. We implement two imaging modes (i.e., wide-field mode and high-resolution mode) in the OCT system, which gives highly scalable FOV with flexible lateral resolution. We demonstrate scalable wide-field vascular imaging for multiple finger nail beds in human and whole brain in mice with skull left intact at a single 3D scan, promising new opportunities for wide-field OCT-based angiography for many clinical applications. PMID:27231630

  9. Stabilization of Collagen-Model, Triple-Helical Peptides for In Vitro and In Vivo Applications

    PubMed Central

    Bhowmick, Manishabrata; Fields, Gregg B.

    2014-01-01

    The triple-helical structure of collagen has been accurately reproduced in numerous chemical and recombinant model systems. Triple-helical peptides and proteins have found application for dissecting collagen-stabilizing forces, isolating receptor- and protein-binding sites in collagen, mechanistic examination of collagenolytic proteases, and development of novel biomaterials. Introduction of native-like sequences into triple-helical constructs can reduce the thermal stability of the triple-helix to below that of the physiological environment. In turn, incorporation of nonnative amino acids and/or templates can enhance triple-helix stability. We presently describe approaches by which triple-helical structure can be modulated for use under physiological or near-physiological conditions. PMID:24014440

  10. New fluorescence imaging probe with high spatial resolution for in vivo applications.

    PubMed

    Bonnans, V; Gharbi, T; Pieralli, C; Wacogne, B; Humbert, Ph

    2004-01-01

    A new fiberized fluorescence imaging probe is presented. This device can potentially be used for a wide range of biological or medical applications. By exploiting the chromatic aberrations of gradient index lenses, the excitation blue or near-UV excitation light is focused on the sample surface, while the red fluorescence signal is efficiently launched back to collecting fibers. The excitation fiber is single mode at the working wavelength so that a resolution of 5 microm is obtained over a scanning area of several square millimeters. Experimental fluorescence images are presented. They concern either self-fabricated fluorescent microsamples or views of leaves that constitute an example of biological tissues analysis. The probe can also be adapted for spectroscopic investigations.

  11. Stabilization of collagen-model, triple-helical peptides for in vitro and in vivo applications.

    PubMed

    Bhowmick, Manishabrata; Fields, Gregg B

    2013-01-01

    The triple-helical structure of collagen has been accurately reproduced in numerous chemical and recombinant model systems. Triple-helical peptides and proteins have found application for dissecting collagen-stabilizing forces, isolating receptor- and protein-binding sites in collagen, mechanistic examination of collagenolytic proteases, and development of novel biomaterials. Introduction of native-like sequences into triple-helical constructs can reduce the thermal stability of the triple-helix to below that of the physiological environment. In turn, incorporation of nonnative amino acids and/or templates can enhance triple-helix stability. We presently describe approaches by which triple-helical structure can be modulated for use under physiological or near-physiological conditions.

  12. Transurethral ultrasound applicators with directional heating patterns for prostate thermal therapy: in vivo evaluation using magnetic resonance thermometry.

    PubMed

    Diederich, C J; Stafford, R J; Nau, W H; Burdette, E C; Price, R E; Hazle, J D

    2004-02-01

    A catheter-based transurethral ultrasound applicator with angularly directional heating patterns has been designed for prostate thermal therapy and evaluated in canine prostate in vivo using MRI to monitor and assess performance. The ultrasound transducer array (3.5 mm diameter tubular transducers, 180 degrees active sectors, approximately 7.5 MHz) was integrated to a flexible delivery catheter (4 mm OD), and encapsulated within an expandable balloon (35 mm x 10 mm OD, 80 ml min(-1) ambient water) for coupling and cooling of the prostatic urethra. These devices were used to thermally coagulate targeted portions of the canine prostate (n = 2) while using MR thermal imaging (MRTI) to monitor the therapy. MRI was also used for target definition, positioning of the applicator, and evaluation of target viability post-therapy. MRTI was based upon the complex phase-difference mapping technique using an interleaved gradient echo-planar imaging sequence with lipid suppression. MRTI derived temperature distributions, thermal dose exposures, T1-contrast enhanced MR images, and histology of sectioned prostates were used to define destroyed tissue zones and characterize the three-dimensional heating patterns. The ultrasound applicators produced approximately 180 degrees directed zones of thermal coagulation within targeted tissue which extended 15-20 mm radially to the outer boundary of the prostate within 15 min. Transducer activation lengths of 17 mm and 24 mm produced contiguous zones of coagulation extending axially approximately 18 mm and approximately 25 mm from base to apex, respectively. Peak temperatures around 90 degrees C were measured, with approximately 50 degrees C-52 degrees C corresponding to outer boundary t43 = 240 min at approximately 15 min treatment time. These devices are MRI compatible, and when coupled with multiplanar MRTI provide a means for selectively controlling the length and sector angle of therapeutic thermal treatment in the prostate.

  13. A new cortical thickness mapping method with application to an in vivo finite element model.

    PubMed

    Kim, Young Ho; Kim, Jong-Eun; Eberhardt, Alan W

    2014-01-01

    Finite element modelling of musculoskeletal systems, with geometrical structures constructed from computed tomography (CT) scans, is a useful and powerful tool for biomechanical studies. The use of CT scans from living human subjects, however, is still limited. Accurate reconstruction of thin cortical bone structures from CT scans of living human subjects is especially problematic, due to low CT resolution that results from mandatory low radiation doses and/or involuntary movements of the subject. In this study, a new method for mapping cortical thickness is described. Using the method, cortical thickness measurements of a coxal (pelvis) bone obtained from CT scans of a cadaver were mapped to the coxal geometry as obtained through CT scans of a live human subject, resulting in accurate cortical thickness while maintaining geometric fidelity of the live subject. The mapping procedure includes shape-preserving parameterisation, mesh movement and interpolation of thickness using a search algorithm. The methodology is applicable to modelling of other bones where accurate cortical thickness is needed and for which such data exist.

  14. A USPL functional system with articulated mirror arm for in-vivo applications in dentistry

    NASA Astrophysics Data System (ADS)

    Schelle, Florian; Meister, Jörg; Dehn, Claudia; Oehme, Bernd; Bourauel, Christoph; Frentzen, Mathias

    Ultra-short pulsed laser (USPL) systems for dental application have overcome many of their initial disadvantages. However, a problem that has not yet been addressed and solved is the beam delivery into the oral cavity. The functional system that is introduced in this study includes an articulated mirror arm, a scanning system as well as a handpiece, allowing for freehand preparations with ultra-short laser pulses. As laser source an Nd:YVO4 laser is employed, emitting pulses with a duration of tp < 10 ps at a repetition rate of up to 500 kHz. The centre wavelength is at 1064 nm and the average output power can be tuned up to 9 W. The delivery system consists of an articulated mirror arm, to which a scanning system and a custom made handpiece are connected, including a 75 mm focussing lens. The whole functional system is compact in size and moveable. General characteristics like optical losses and ablation rate are determined and compared to results employing a fixed setup on an optical table. Furthermore classical treatment procedures like cavity preparation are being demonstrated on mammoth ivory. This study indicates that freehand preparation employing an USPL system is possible but challenging, and accompanied by a variety of side-effects. The ablation rate with fixed handpiece is about 10 mm3/min. Factors like defocussing and blinding affect treatment efficiency. Laser sources with higher average output powers might be needed in order to reach sufficient preparation speeds.

  15. Cestode parasites: application of in vivo and in vitro models for studies on the host-parasite relationship.

    PubMed

    Siles-Lucas, Mar; Hemphill, Andrew

    2002-01-01

    Cestode worms, commonly also known as 'flat' worms or tapeworms, are an important class of endoparasitic organisms. In order to complete their life cycle, they infect intermediate and definitive hosts in succession, through oral ingestion of eggs or larvae, respectively. Serious disease in humans or other mammalian hosts is mostly caused by the larval stages. Echinococcus spp. and Taenia spp. have been extensively investigated in the laboratory due to the fact that they represent important veterinary medical challenges and also cause grave diseases in humans. In contrast, Hymenolepis spp. and Mesocestoides spp. infections are relatively rare in humans, but these parasites have been extensively studied because their life cycle stages can be easily cultured in vitro, and can also be conveniently maintained in laboratory animal hosts. Thus they are more easily experimentally accessible, and represent important models for investigating the various aspects of cestode biology. This review will focus on in vitro and in vivo models which have been developed for studies on the host-parasite relationship during infection with Echinococcus, Taenia, Hymenolepis, Mesocestoides and Spirometra, and will cover the use of these models to investigate the morphology and ultrastructure of respective genera, the immunological relationship with the host and the development of vaccination approaches, as well as applications of these models for studies on parasite metabolism, physiology and gene expression. In addition, the use of these models in the development of chemotherapeutic measures against cestode infections is reviewed.

  16. Temperature dependence of the optoacoustic transformation efficiency in ex vivo tissues for application in monitoring thermal therapies.

    PubMed

    Nikitin, Sergey M; Khokhlova, Tatiana D; Pelivanov, Ivan M

    2012-06-01

    The calibration dependencies of the optoacoustic (OA) transformation efficiency on tissue temperature are obtained for the application in OA temperature monitoring during thermal therapies. Accurate measurement of the OA signal amplitude versus temperature is performed in different ex vivo tissues in the temperature range 25°C to 80°C. The investigated tissues were selected to represent different structural components: chicken breast (skeletal muscle), porcine lard (fatty tissue), and porcine liver (richly perfused tissue). Backward mode of the OA signal detection and a narrow probe laser beam were used in the experiments to avoid the influence of changes in light scattering with tissue coagulation on the OA signal amplitude. Measurements were performed in heating and cooling regimes. Characteristic behavior of the OA signal amplitude temperature dependences in different temperature ranges were described in terms of changes in different structural components of the tissue samples. The accuracy of temperature reconstruction from the obtained calibration dependencies for the investigated tissue types is evaluated.

  17. Applications of in vivo electron paramagnetic resonance (EPR) spectroscopy: measurements of pO2 and NO in endotoxin shock.

    PubMed

    Jackson, S K; Madhani, M; Thomas, M; Timmins, G S; James, P E

    2001-03-31

    Recent developments of EPR instrumentation that allow the use of large tissue samples or whole animals and the ability to image spatially resolved EPR signals has led to novel applications of EPR spectroscopy in vivo. Utilising a 1 GHz EPR spectrometer with a 3.4-cm birdcage resonator, it was possible to detect and measure nitric oxide and oxygen in the livers of mice with lipopolysaccharide (LPS)-induced septic shock. Nitric oxide was detected as the nitric oxide (NO) complex of Fe-diethyldithiocarbamic acid (Fe-DETC) while pO2 was measured from the EPR linewidth of the oxygen-sensitive coal material 'gloxy'. LPS treatment stimulated the production of nitric oxide in the liver and the general circulation and the oxygenation of liver tissue was decreased. Selective placement of the EPR probes allowed images of nitric oxide and oxygen to be obtained in the liver. The spectral and spatial information obtained with this technique will allow improved understanding of the pathophysiology of such diseases.

  18. In vitro and in vivo degradation evaluation of novel iron-bioceramic composites for bone implant applications.

    PubMed

    Ulum, M F; Arafat, A; Noviana, D; Yusop, A H; Nasution, A K; Abdul Kadir, M R; Hermawan, H

    2014-03-01

    Biodegradable metals such as magnesium, iron and their alloys have been known as potential materials for temporary medical implants. However, most of the studies on biodegradable metals have been focusing on optimizing their mechanical properties and degradation behavior with no emphasis on improving their bioactivity behavior. We therefore investigated the possibility of improving iron biodegradation rate and bioactivity by incorporating various bioactive bioceramics. The iron-based bioceramic (hydroxyapatite, tricalcium phosphate and biphasic calcium phosphate) composites were prepared by mechanical mixing and sintering process. Degradation studies indicated that the addition of bioceramics lowered the corrosion potential of the composites and slightly increased their corrosion rate compared to that of pure iron. In vitro cytotoxicity results showed an increase of cellular activity when rat smooth muscle cells interacted with the degrading composites compared to pure iron. X-ray radiogram analysis showed a consistent degradation progress with that found in vivo and positive tissue response up to 70 days implantation in sheep animal model. Therefore, the iron-based bioceramic composites have the potential to be used for biodegradable bone implant applications.

  19. Construction of Chinese adult male phantom library and its application in the virtual calibration of in vivo measurement.

    PubMed

    Chen, Yizheng; Qiu, Rui; Li, Chunyan; Wu, Zhen; Li, Junli

    2016-03-07

    In vivo measurement is a main method of internal contamination evaluation, particularly for large numbers of people after a nuclear accident. Before the practical application, it is necessary to obtain the counting efficiency of the detector by calibration. The virtual calibration based on Monte Carlo simulation usually uses the reference human computational phantom, and the morphological difference between the monitored personnel with the calibrated phantom may lead to the deviation of the counting efficiency. Therefore, a phantom library containing a wide range of heights and total body masses is needed. In this study, a Chinese reference adult male polygon surface (CRAM_S) phantom was constructed based on the CRAM voxel phantom, with the organ models adjusted to match the Chinese reference data. CRAM_S phantom was then transformed to sitting posture for convenience in practical monitoring. Referring to the mass and height distribution of the Chinese adult male, a phantom library containing 84 phantoms was constructed by deforming the reference surface phantom. Phantoms in the library have 7 different heights ranging from 155 cm to 185 cm, and there are 12 phantoms with different total body masses in each height. As an example of application, organ specific and total counting efficiencies of Ba-133 were calculated using the MCNPX code, with two series of phantoms selected from the library. The influence of morphological variation on the counting efficiency was analyzed. The results show only using the reference phantom in virtual calibration may lead to an error of 68.9% for total counting efficiency. Thus the influence of morphological difference on virtual calibration can be greatly reduced using the phantom library with a wide range of masses and heights instead of a single reference phantom.

  20. Construction of Chinese adult male phantom library and its application in the virtual calibration of in vivo measurement

    NASA Astrophysics Data System (ADS)

    Chen, Yizheng; Qiu, Rui; Li, Chunyan; Wu, Zhen; Li, Junli

    2016-03-01

    In vivo measurement is a main method of internal contamination evaluation, particularly for large numbers of people after a nuclear accident. Before the practical application, it is necessary to obtain the counting efficiency of the detector by calibration. The virtual calibration based on Monte Carlo simulation usually uses the reference human computational phantom, and the morphological difference between the monitored personnel with the calibrated phantom may lead to the deviation of the counting efficiency. Therefore, a phantom library containing a wide range of heights and total body masses is needed. In this study, a Chinese reference adult male polygon surface (CRAM_S) phantom was constructed based on the CRAM voxel phantom, with the organ models adjusted to match the Chinese reference data. CRAMS phantom was then transformed to sitting posture for convenience in practical monitoring. Referring to the mass and height distribution of the Chinese adult male, a phantom library containing 84 phantoms was constructed by deforming the reference surface phantom. Phantoms in the library have 7 different heights ranging from 155 cm to 185 cm, and there are 12 phantoms with different total body masses in each height. As an example of application, organ specific and total counting efficiencies of Ba-133 were calculated using the MCNPX code, with two series of phantoms selected from the library. The influence of morphological variation on the counting efficiency was analyzed. The results show only using the reference phantom in virtual calibration may lead to an error of 68.9% for total counting efficiency. Thus the influence of morphological difference on virtual calibration can be greatly reduced using the phantom library with a wide range of masses and heights instead of a single reference phantom.

  1. Gellan gum injectable hydrogels for cartilage tissue engineering applications: in vitro studies and preliminary in vivo evaluation.

    PubMed

    Oliveira, João T; Santos, Tírcia C; Martins, Luís; Picciochi, Ricardo; Marques, Alexandra P; Castro, António G; Neves, Nuno M; Mano, João F; Reis, Rui L

    2010-01-01

    Gellan gum is a polysaccharide that we have previously proposed for applications in the cartilage tissue engineering field. In this work, gellan gum hydrogels were tested for their ability to be used as injectable systems using simple processing methods, able to deliver and maintain chondrocytes by in situ gelation, and support cell viability and production of extracellular matrix (ECM). Rheological measurements determined that the sol-gel transition occurred near the body temperature at 39 degrees C, upon temperature decrease, in approximately 20 s. Gellan gum discs shows a storage compression modulus of around 80 kPa at a frequency of 1 Hz by dynamic mechanical analysis. Human articular chondrocytes were encapsulated in the gels, cultured in vitro for total periods of 56 days, and analyzed for cell viability and ECM production. Calcein AM staining showed that cell kept viable after 14 days and the histological analysis and real-time quantitative polymerase chain reaction revealed that hyaline-like cartilage ECM was synthesized. Finally, the in vivo performance of the gellan gum hydrogels, in terms of induced inflammatory reaction and integration into the host tissue, was evaluated by subcutaneous implantation in Balb/c mice for 21 days. Histological analysis showed a residual fibrotic capsule at the end of the experiments. Dynamic mechanical analysis revealed that the gels were stable throughout the experiments while evidencing a tendency for decreasing mechanical properties, which was consistent with weight measurements. Altogether, the results demonstrate the adequacy of gellan gum hydrogels processed by simple methods for noninvasive injectable applications toward the formation of a functional cartilage tissue-engineered construct and originally report the preliminary response of a living organism to the subcutaneous implantation of the gellan gum hydrogels. These are the two novel features of this work.

  2. Probe depth matters in dermal microdialysis sampling of benzoic acid after topical application: an ex vivo study in human skin.

    PubMed

    Holmgaard, R; Benfeldt, E; Bangsgaard, N; Sorensen, J A; Brosen, K; Nielsen, F; Nielsen, J B

    2012-01-01

    Microdialysis (MD) in the skin - dermal microdialysis (DMD) - is a unique technique for sampling of topically as well as systemically administered drugs at the site of action, e.g. sampling of dermatological drug concentrations in the dermis. Debate has concerned the existence of a correlation between the depth of the sampling device - the probe - in the dermis and the amount of drug sampled following topical drug administration. This study evaluates the relation between probe depth and drug sampling using dermal DMD sampling ex vivo in human skin. We used superficial (<1 mm), intermediate (1-2 mm) and deep (>2 mm) positioning of the linear MD probe in the dermis of human abdominal skin, followed by topical application of 4 mg/ml of benzoic acid (BA) in skin chambers overlying the probes. Dialysate was sampled every hour for 12 h and analysed for BA content by high-performance liquid chromatography. Probe depth was measured by 20-MHz ultrasound scanning. The area under the time-versus-concentration curve (AUC) describes the drug exposure in the tissue during the experiment and is a relevant parameter to compare for the 3 dermal probe depths investigated. The AUC(0-12) were: superficial probes: 3,335 ± 1,094 μg·h/ml (mean ± SD); intermediate probes: 2,178 ± 1,068 μg·h/ml, and deep probes: 1,159 ± 306 μg·h/ml. AUC(0-12) sampled by the superficial probes was significantly higher than that of samples from the intermediate and deeply positioned probes (p value <0.05). There was a significant inverse correlation between probe depth and AUC(0-12) sampled by the same probe (p value <0.001, r(2) value = 0.5). The mean extrapolated lag-times (±SD) for the superficial probes were 0.8 ± 0.1 h, for the intermediate probes 1.7 ± 0.5 h, and for the deep probes 2.7 ± 0.5 h, which were all significantly different from each other (p value <0.05). In conclusion, this paper demonstrates that there is an inverse relationship between the depth of the probe in the dermis

  3. In vitro and in vivo corrosion properties of new iron-manganese alloys designed for cardiovascular applications.

    PubMed

    Drynda, Andreas; Hassel, Thomas; Bach, Friedrich Wilhelm; Peuster, Matthias

    2015-04-01

    The principle of biodegradation for the production of temporary implant materials (e.g. stents) plays an important role in the treatment of congenital heart defects. In the last decade several attempts have been made with different alloy materials-mainly based on iron and magnesium. None of the currently available materials in this field have demonstrated satisfying results and have therefore not found entry into broad clinical practice. While magnesium or magnesium alloy systems corrode too fast, the corrosion rate of pure iron-stents is too slow for cardiovascular applications. In the last years FeMn alloy systems were developed with the idea that galvanic effects, caused by different electrochemical properties of Fe and Mn, would increase the corrosion rate. In vitro tests with alloys containing up to 30% Mn showed promising results in terms of biocompatibility. This study deals with the development of new FeMn alloy systems with lower Mn concentrations (FeMn 0.5 wt %, FeMn 2.7 wt %, FeMn 6.9 wt %) to avoid Mn toxicity. Our results show, that these alloys exhibit good mechanical features as well as suitable in vitro biocompatibility and corrosion properties. In contrast, the evaluation of these alloys in a mouse model led to unexpected results-even after 9 months no significant corrosion was detectable. Preliminary SEM investigations showed that passivation layers (FeMn phosphates) might be the reason for corrosion resistance. If this can be proved in further experiments, strategies to prevent or dissolve those layers need to be developed to expedite the in vivo corrosion of FeMn alloys.

  4. A volume birdcage coil with an adjustable sliding tuner ring for neuroimaging in high field vertical magnets: ex and in vivo applications at 21.1 T

    PubMed Central

    Qian, Chunqi; Masad, Ihssan S.; Rosenberg, Jens T.; Elumalai, Malathy; Brey, William W.; Grant, Samuel C.; Gor’kov, Peter L.

    2012-01-01

    A tunable 900 MHz transmit/receive volume coil was constructed for 1H MR imaging of biological samples in a 21.1 T vertical bore magnet. To accommodate a diverse range of specimen and RF loads at such a high frequency, a sliding-ring adaptation of a low-pass birdcage was implemented through simultaneous alteration of distributed capacitance. To make efficient use of the constrained space inside the vertical bore, a modular probe design was implemented with a bottom-adjustable tuning and matching apparatus. The sliding ring coil displays good homogeneity and sufficient tuning range for different samples of various dimensions representing large span of RF loads. High resolution in vivo and ex vivo images of large rats (up to 350 g), mice and human postmortem tissues were obtained to demonstrate coil functionality and to provide examples of potential applications at 21.1 T. PMID:22750638

  5. Characteristics and Applications of the ToxRefDB In Vivo Datasets from Chronic, Reproductive and Developmental Assays

    EPA Science Inventory

    ToxRefDB was developed to store data from in vivo animal toxicity studies. The initial focus was populating ToxRefDB with pesticide registration toxicity data that has been historically stored as hard-copy and scanned documents by the Office of Pesticide Programs. A significant p...

  6. Application of a partial-thickness human ex vivo skin culture model in cutaneous wound healing study.

    PubMed

    Xu, Wei; Jong Hong, Seok; Jia, Shengxian; Zhao, Yanan; Galiano, Robert D; Mustoe, Thomas A

    2012-04-01

    A number of in vivo and ex vivo skin models have been applied to human wound healing studies. A reliable skin model, which recapitulates the features of human wound repair, is essential for the clinical and mechanical investigation of human cutaneous wound healing. Full-skin ex vivo culture systems have been used in wound healing studies. However, important structures of the skin, such as the differentiation of keratinocytes and epidermis-dermis junction, are poorly characterized in this model. This study aims to develop an optimized partial-thickness human ex vivo skin culture (HESC) model to maintain human skin characteristics in vitro. During our culture, the basal layer, suprabasal layer, and stratum granulosum layer of epidermis were preserved until day 8. Analyses of hemidesmosome proteins, bullous pemphigoid antigen 1 (BP180) and 2 (BP230), showed that the integrity of the basement membrane of the epidermis was well preserved in the HESC model. In contrast, an organotypic culture with human keratinocytes and fibroblasts failed to show an integrated basement membrane. Maintenance of skin structure by histological analysis and proliferation of epidermal keratinocytes by Ki67 staining were observed in our model for 12 days. Complete re-epithelialization of the wounding area was observed at day 6 post wounding when a superficial incisional wound was created. The expression of Ki-67 and keratin 6, indicators of activated keratinocytes in epidermis, was significantly upregulated and new collagen synthesis was found in the dermis during the wound healing process. As control, we also used organotypic culture in studying the differentiation of the keratinocyte layers and incisional wound repair. It turned out that our model has advantage in these study fields. The results suggest that our HESC model retains important elements of in vivo skin and has significant advantages for the wound healing studies in vitro.

  7. Development of a protocol for selection of genes fit for the in vivo knockdown method and its application to insulin receptor substrate genes in mice.

    PubMed

    Saito, Mikako; Kakutani, Yukari; Kaburagi, Misako; Funabashi, Hisakage; Matsuoka, Hideaki

    2013-01-01

    Prediabetes model mice in which more than one gene associated with diabetes is knocked down simultaneously are potentially useful for pharmaceutical and medical studies of diabetes. However, the effective conditions for sufficient knockdown in vivo are dependent on the intrinsic properties of the target genes. It is necessary to investigate which genes are applicable or not to the in vivo knockdown method. In this study, insulin receptor substrate 1 and 2 (Irs-1, Irs-2) were selected as target genes. Effective siRNAs against the respective genes were designed, and their efficacy was confirmed by cell-based experiments. Based on the results of siRNAs, shRNA expression vectors against Irs-1 and Irs-2 were constructed, respectively. Their efficacy was also confirmed by cell-based experiments. A hydrodynamic method was applied to the delivery of the vectors to mice. This method was found to be effective for predominant delivery to the liver by demonstrative delivery of an EGFP expression vector and successive histochemical analysis. Fifty micrograms of the shRNA expression vector was injected into the tail vein. After 24 h, the liver, pancreas, and muscle were isolated, and the expression levels of Irs-1 and Irs-2 were analyzed by quantitative RT-PCR. In the liver, Irs-2 was effectively knocked down to 60% of the control level, but Irs-1 was not influenced even under the same conditions. The protocol developed here is feasible for the selection of genes fit for in vivo knockdown method.

  8. Near-infrared luminescent cubic silicon carbide nanocrystals for in vivo biomarker applications: an ab initio study.

    PubMed

    Somogyi, Bálint; Zólyomi, Viktor; Gali, Adam

    2012-12-21

    Molecule-sized fluorescent emitters are much sought-after to probe biomolecules in living cells. We demonstrate here by time-dependent density functional calculations that the experimentally achievable 1-2 nm sized silicon carbide nanocrystals can emit light in the near-infrared region after introducing appropriate color centers in them. These near-infrared luminescent silicon carbide nanocrystals may act as ideal fluorophores for in vivo bioimaging.

  9. Application of Fluorescent Protein Expressing Strains to Evaluation of Anti-Tuberculosis Therapeutic Efficacy In Vitro and In Vivo

    PubMed Central

    Kong, Ying; Yang, Dong; Cirillo, Suat L. G.; Li, Shaoji; Akin, Ali; Francis, Kevin P.; Maloney, Taylor; Cirillo, Jeffrey D.

    2016-01-01

    The slow growth of Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), hinders development of new diagnostics, therapeutics and vaccines. Using non-invasive real-time imaging technologies to monitor the disease process in live animals would facilitate TB research in all areas. We developed fluorescent protein (FP) expressing Mycobacterium bovis BCG strains for in vivo imaging, which can be used to track bacterial location, and to quantify bacterial load in live animals. We selected an optimal FP for in vivo imaging, by first cloning six FPs: tdTomato, mCherry, mPlum, mKate, Katushka and mKeima, into mycobacteria under either a mycobacterial Hsp60 or L5 promoter, and compared their fluorescent signals in vitro and in vivo. Fluorescence from each FP-expressing strain was measured with a multimode reader using the optimal excitation and emission wavelengths for the FP. After normalizing bacterial numbers with optical density, the strain expressing L5-tdTomato displayed the highest fluorescence. We used the tdTomato-labeled M. bovis BCG to obtain real-time images of pulmonary infections in living mice and rapidly determined the number of bacteria present. Further comparison between L5-tdTomato and Hsp60-tdTomato revealed that L5-tdTomato carried four-fold more tdTomato gene copies than Hsp60-tdTomato, which eventually led to higher protein expression of tdTomato. Evaluating anti-TB efficacy of rifampicin and isoniazid therapy in vitro and in vivo using the L5-tdTomato strain demonstrated that this strain can be used to identify anti-TB therapeutic efficacy as quickly as 24 h post-treatment. These M. bovis BCG reporter strains represent a valuable new tool for evaluation of therapeutics, vaccines and virulence. PMID:26934495

  10. Application of twyman-green interferometer for evaluation of in vivo breakup characteristic of the human tear film.

    PubMed

    Licznerski, T J; Kasprzak, H T; Kowalik, W

    1999-01-01

    The paper presents an interferometric method of assessing the in vivo stability of the precorneal tear film. To observe dynamic effects on a human cornea the Twyman-Green interferometer with television frame speed digital registration synchronized with a laser flash was used. The instrument was applied to the human cornea in vivo. The results of the experiment, both tear film distribution and its dynamics, are presented. The proposed interferometric setup can be used to evaluate the breakup characteristics of the tear film, its distribution, and to examine its dynamic changes. The breakup profiles and their cross sections calculated from the interferogram analysis are presented. The depth of recorded breakup, calculated on the basis of interferogram analysis, amounts to about 1.5 μm. The proposed method has the advantage of being noncontact and applies only a low-energy laser beam to the eye. This provides noninvasive viewing of human cornea in vivo and makes it possible to observe the kinetics of its tear film deterioration. © 1999 Society of Photo-Optical Instrumentation Engineers.

  11. Comparison of the sensitivities of common in vitro and in vivo assays of estrogenic activity: application of chemical toxicity distributions.

    PubMed

    Dobbins, Laura L; Brain, Richard A; Brooks, Bryan W

    2008-12-01

    A number of contaminants in municipal effluent discharges are estrogen agonists to fish. Whereas several in vitro and in vivo techniques have been developed to assess the estrogenic activity of these compounds or ambient environmental samples, previous comparisons of the relative sensitivities of these approaches remain inconclusive. We employed a probabilistic hazard assessment approach using chemical toxicity distributions (CTDs) to perform a novel evaluation of relative sensitivities of six common in vitro and in vivo assays. We predicted that there was an 8.3% (human breast ademocarcinoma cell line, MCF-7, assay), 6.3% (yeast estrogen screen assay), or 1.9% (fish hepatocyte vitellogenin, VTG, assay) probability of detecting a compound in aquatic systems that will elicit an estrogenic response at concentrations at or below 0.1 microg/L, suggesting that the MCF-7 assay was the most sensitive in vitro assay evaluated in this study. The probabilities of eliciting the estrogenic response of VTG induction at a concentration less than 0.1 microg/L in rainbow trout, fathead minnow, and Japanese medaka were determined at 29.9, 26.2, and 18.8%, respectively. Thus, rainbow trout VTG induction was the most sensitive in vivo assay assessed. Subsequently, CTDs may provide a useful technique for hazard assessment of chemical classes for which exposure data are limited and for chemicals with common toxicological mechanisms and modes of action.

  12. Use-dependent effects of lidocaine on conduction in canine myocardium: application of the modulated receptor hypothesis in vivo.

    PubMed

    Davis, J; Matsubara, T; Scheinman, M M; Katzung, B; Hondeghem, L H

    1986-07-01

    Lidocaine is a commonly used antiarrhythmic drug that causes use-dependent blockade of sodium channels in vitro and reduces conduction velocity in vitro and in vivo. According to the modulated receptor hypothesis of antiarrhythmic drug action, lidocaine has a low affinity for rested sodium channels but a high affinity for open and inactivated channels. In the present experiments, we characterized use-dependent conduction slowing and recovery from slowing by lidocaine in anesthetized dogs. The His-to-ventricular conduction interval was used as the indicator of conduction velocity. We found that prolongation of conduction time was greater as the stimulation frequency was increased. Moreover, on abruptly changing the stimulation frequency, a new steady-state conduction time was approached in two to three depolarizations. On discontinuation of stimulation, the conduction time of progressively less premature extrastimuli shortened exponentially with a terminal phase time constant of 152 +/- 115 msec. These effects by lidocaine were enhanced during acidosis and enhancement was reversed by correction of the acidosis. It is concluded that the effects in vivo of lidocaine on conduction under several conditions of rate, rhythm, and pH are similar to its effects on the maximum upstroke velocity of the action potential in vitro. Although these experiments were not designed to validate the modulated receptor hypothesis, it appears that the modulated receptor hypothesis can predict the effects of lidocaine on conduction in vivo.

  13. αTCP ceramic doped with dicalcium silicate for bone regeneration applications prepared by powder metallurgy method: in vitro and in vivo studies.

    PubMed

    Velasquez, Pablo; Luklinska, Zofia B; Meseguer-Olmo, Luis; Mate-Sanchez de Val, Jose E; Delgado-Ruiz, Rafael A; Calvo-Guirado, Jose L; Ramirez-Fernandez, Ma P; de Aza, Piedad N

    2013-07-01

    This study reports on the in vitro and in vivo behavior of α-tricalcium phosphate (αTCP) and also αTCP doped with either 1.5 or 3.0 wt % of dicalcium silicate (C2 S). The ceramics were successfully prepared by powder metallurgy method combined with homogenization and heat treatment procedures. All materials were composed of a single-phase, αTCP in the case of a pure material, or solid solution of C2 S in αTCP for the doped αTCP, which were stable at room temperature. The ceramics were tested for bioactivity in simulated body fluid, cell culture medium containing adult mesenchymal stem cells of human origin, and in animals. Analytical scanning electron microscopy combined with chemical elemental analysis was used and Fourier transform infrared and conventional histology methods. The in vivo behavior of the ceramics matched the in vitro results, independently of the C2 S content in αTCP. Carbonated hydroxyapatite (CHA) layer was formed on the surface and within the inner parts of the specimens in all cases. A fully mineralized new bone growing in direct contact with the implants was found under the in vivo conditions. The bioactivity and biocompatibility of the implants increased with the C2 S content in αTCP. The C2 S doped ceramics also favoured a phase transformation of αTCP into CHA, important for full implant integration during the natural bone healing processes. αTCP ceramic doped with 3.0 wt % C2 S showed the best bioactive in vitro and in vivo properties of all the compositions and hence could be of interest in specific applications for bone restorative purposes.

  14. Development of a disposable magnetically levitated centrifugal blood pump (MedTech Dispo) intended for bridge-to-bridge applications--two-week in vivo evaluation.

    PubMed

    Nagaoka, Eiki; Someya, Takeshi; Kitao, Takashi; Kimura, Taro; Ushiyama, Tomohiro; Hijikata, Wataru; Shinshi, Tadahiko; Arai, Hirokuni; Takatani, Setsuo

    2010-09-01

    Last year, we reported in vitro pump performance, low hemolytic characteristics, and initial in vivo evaluation of a disposable, magnetically levitated centrifugal blood pump, MedTech Dispo. As the first phase of the two-stage in vivo studies, in this study we have carried out a 2-week in vivo evaluation in calves. Male Holstein calves with body weight of 62.4–92.2 kg were used. Under general anesthesia, a left heart bypass with a MedTech Dispo pump was instituted between the left atrium and the descending aorta via left thoracotomy. Blood-contacting surface of the pump was coated with a 2-methacryloyloxyethyl phosphorylcholine polymer. Post-operatively, with activated clotting time controlled at 180–220 s using heparin and bypass flow rate maintained at 50 mL/kg/min, plasma-free hemoglobin (Hb), coagulation, and major organ functions were analyzed for evaluation of biocompatibility. The animals were electively sacrificed at the completion of the 2-week study to evaluate presence of thrombus inside the pump,together with an examination of major organs. To date, we have done 13 MedTech Dispo implantations, of which three went successfully for a 2-week duration. In these three cases, the pump produced a fairly constant flow of 50 mL/Kg/min. Neurological disorders and any symptoms of thromboembolism were not seen. Levels of plasma-free Hb were maintained very low. Major organ functions remained within normal ranges. Autopsy results revealed no thrombus formation inside the pump. In the last six cases, calves suffered from severe pneumonia and they were excluded from the analysis. The MedTech Dispo pump demonstrated sufficient pump performance and biocompatibility to meet requirements for 1-week circulatory support. The second phase (2-month in vivo study) is under way to prove the safety and efficacy of MedTech Dispo for 1-month applications.

  15. A Biocompatible In Vivo Ligation Reaction and its Application for Non-Invasive Bioluminescent Imaging of Protease Activity in Living Mice

    PubMed Central

    Godinat, Aurélien; Park, Hyo Min; Miller, Stephen C.; Cheng, Ke; Hanahan, Douglas; Sanman, Laura E.; Bogyo, Matthew; Yu, Allen; Nikitin, Gennady F.; Stahl, Andreas; Dubikovskaya, Elena A.

    2013-01-01

    The discovery of biocompatible reactions has had a tremendous impact on chemical biology, allowing the study of numerous biological processes directly in complex systems. However, despite the fact that multiple biocompatible reactions have been developed in the past decade, very few work well in living mice. Here we report that D-cysteine and 2-cyanobenzothiazoles can selectively react with each other in vivo to generate a luciferin substrate for firefly luciferase. The success of this “split luciferin” ligation reaction has important implications for both in vivo imaging and biocompatible labeling strategies. First, the production of a luciferin substrate can be visualized in a live mouse by bioluminescence imaging (BLI), and furthermore allows interrogation of targeted tissues using a “caged” luciferin approach. We therefore applied this reaction to the real-time non-invasive imaging of apoptosis associated with caspase 3/7. Caspase-dependent release of free D-cysteine from the caspase 3/7 peptide substrate Asp-Glu-Val-Asp-D-Cys (DEVD-(D-Cys)) allowed selective reaction with 6-amino-2-cyanobenzothiazole (NH2-CBT) in vivo to form 6-amino-D-luciferin with subsequent light emission from luciferase. Importantly, this strategy was found to be superior to the commercially-available DEVD-aminoluciferin substrate for imaging of caspase 3/7 activity. Moreover, the split luciferin approach enables the modular construction of bioluminogenic sensors, where either or both reaction partners could be caged to report on multiple biological events. Lastly, the luciferin ligation reaction is three orders of magnitude faster than Staudinger ligation suggesting further applications for both bioluminescence and specific molecular targeting in vivo. PMID:23463944

  16. Utilization of leukapheresis and CD4 positive selection in Treg isolation and the ex-vivo expansion for a clinical application in transplantation and autoimmune disorders

    PubMed Central

    Gołąb, Karolina; Grose, Randall; Trzonkowski, Piotr; Wickrema, Amittha; Tibudan, Martin; Marek-Trzonkowska, Natalia; Matosz, Sabrina; Solomina, Julia; Ostrega, Diane; Millis, J. Michael; Witkowski, Piotr

    2016-01-01

    Adoptive transfer of T regulatory cells (Tregs) is of great interest as a novel immunosuppressive therapy in autoimmune disorders and transplantation. Obtaining a sufficient number of stable and functional Tregs generated according to current Good Manufacturing Practice (cGMP) requirements has been a major challenge in introducing Tregs as a clinical therapy. Here, we present a protocol involving leukapheresis and CD4+ cell pre-enrichment prior to Treg sorting, which allows a sufficient number of Tregs for a clinical application to be obtained. With this method there is a decreased requirement for ex-vivo expansion. The protocol was validated in cGMP conditions. Our final Treg product passed all release criteria set for clinical applications. Moreover, during expansion Tregs presented their stable phenotype: percentage of CD4+CD25hiCD127− and CD4+FoxP3+ Tregs was > 95% and > 80%, respectively, and Tregs maintained proper immune suppressive function in vitro. Our results suggest that utilization of leukapheresis and CD4 positive selection during Treg isolation improves the likelihood of obtaining a sufficient number of high quality Treg cells during subsequent ex-vivo expansion and they can be applied clinically. PMID:27821811

  17. Feasibility Study of Glass Dosimeter for In Vivo Measurement: Dosimetric Characterization and Clinical Application in Proton Beams

    SciTech Connect

    Rah, Jeong-Eun; Oh, Do Hoon; Kim, Jong Won; Kim, Dae-Hyun; Suh, Tae-Suk; Ji, Young Hoon; Shin, Dongho; Lee, Se Byeong; Kim, Dae Yong; Park, Sung Yong

    2012-10-01

    Purpose: To evaluate the suitability of the GD-301 glass dosimeter for in vivo dose verification in proton therapy. Methods and Materials: The glass dosimeter was analyzed for its dosimetrics characteristic in proton beam. Dosimeters were calibrated in a water phantom using a stairlike holder specially designed for this study. To determine the accuracy of the glass dosimeter in proton dose measurements, we compared the glass dosimeter and thermoluminescent dosimeter (TLD) dose measurements using a cylindrical phantom. We investigated the feasibility of the glass dosimeter for the measurement of dose distributions near the superficial region for proton therapy plans with a varying separation between the target volume and the surface of 6 patients. Results and Discussion: Uniformity was within 1.5%. The dose-response has good linearity. Dose-rate, fading, and energy dependence were found to be within 3%. The beam profile measured using the glass dosimeter was in good agreement with the profile obtained from the ionization chamber. Depth-dose distributions in nonmodulated and modulated proton beams obtained with the glass dosimeter were estimated to be within 3%, which was lower than those with the ionization chamber. In the phantom study, the difference of isocenter dose between the delivery dose calculated by the treatment planning system and that measured by the glass dosimeter was within 5%. With in vivo dosimetry, the calculated surface doses overestimated measurements by 4%-16% using glass dosimeter and TLD. Conclusion: It is recommended that bolus be added for these clinical cases. We also believe that the glass dosimeter has considerable potential for use with in vivo patient proton dosimetry.

  18. Development and application of a membrane cyclone reactor for in vivo NMR spectroscopy with high microbial cell densities.

    PubMed

    Hartbrich, A; Schmitz, G; Weuster-Botz, D; de Graaf, A A; Wandrey, C

    1996-09-20

    A new bioreactor system has been developed for in vivo NMR spectroscopy of microorganisms under defined physiological conditions. This cyclone reactor with an integrated NMR flow cell is continuously operated in the magnet of a 400-MHz wide-bore NMR spectrometer system. The residence times of medium and cells are decoupled by a circulation-integrated cross-flow microfiltration module to achieve higher cell densities as compared to continuous fermentations without cell retention (increase in cell density up to a factor of 10 in steady state). Volumetric mass transfer coefficients k(L)a of more than 1.0 s(-1) are possible in the membrane cyclone reactor, ensuring adequate oxygen supply [oxygen transfer rate >15,000 mg O(2) .(L h)(-1)] of high cell densities. With the aid of the membrane cyclone reactor we were able to show, using continuous in vivo (31)P NMR spectroscopy of anaerobic glucose fermentation by Zymomonas mobilis, that the NMR signal intensity was directly proportional to the cell concentration in the reactor. The concentration profiles of intracellular inorganic phosphate, NAD(H), NDP, NTP, UDP-sugar, a cyclic pyrophosphate, two sugar phosphate pools, and extracellular inorganic phosphate were recorded after a shift from one steady state to another. The intracellular cyclic pyrophosphate had not been detected before in in vitro measurements of Zymomonas mobilis extracts due to the high instability of this compound. Using continuous in vivo (13)C NMR spectroscopy of aerobic glucose utilization by Corynebacterium glutamicum at a density of 25 g(cell dry weight) . L(-1), the membrane cyclone reactor served to measure the different dynamics of labeling in the carbon atoms of L-lactate, L-glutamate, succinate, and L-lysine with a time resolution of 10 min after impressing a [1-(13)C]-glucose pulse.

  19. Application of wide-field optical coherence tomography to monitoring of viability of rat brain in vivo

    NASA Astrophysics Data System (ADS)

    Sato, Manabu; Nishidate, Izumi

    2014-05-01

    We investigated the feasibility of OCT in monitoring the viability of the brain. It was confirmed that after an overdose of pentobarbital sodium salt for an euthanasia, the OCT signal intensity increased before cardiac arrest and finally became 2.7 times, and by periodically changing the tissue temperature from 20 to 32 °C in vivo, average correlation coefficients between the ratio of signal intensity (RSI) and temperature were determined to be -0:42 to -0:50. RSI reversibly changed with subsequent variations of temperatures and finally increased rapidly just before cardiac arrest. These results indicate that RSI could correspond to decreases in viability.

  20. The application of in vivo laser confocal microscopy to the diagnosis and evaluation of meibomian gland dysfunction

    PubMed Central

    Matsumoto, Yukihiro; Sato, Enrique Adan; Ibrahim, Osama M.A.; Tsubota, Kazuo

    2008-01-01

    Purpose To evaluate the morphological changes of the meibomian glands (MG) in patients with meibomian gland dysfunction (MGD) compared to normal subjects by in vivo confocal microscopy and to investigate the relation of these changes to the clinical ocular surface findings and tear functions. Methods Twenty MGD patients and 15 normal subjects were recruited into this prospective study. Patients and controls underwent slit lamp examinations, tear film break-up time (BUT) measurements, fluorescein and Rose-Bengal stainings, Schirmer test I without anesthesia, tear evaporation rate assessment (TEROS), tear film lipid layer interferometry (DR-1), transillumination of the lids (meibography), MG expressibility test, and in vivo laser confocal microscopy of the lids (HRTII-RCM). Results The BUT, DR-1 tear film lipid layer interferometry grades, fluorescein and Rose-Bengal staining scores, MG drop out grade in meibography, and MG expressibility grades were significantly worse in MGD patients compared to normal controls (p<0.05). The severity of both MG dropout and MG expressibility related significantly with the BUT, DR-1 grades, and TEROS (p<0.05). The mean density of acinar units of MGs as measured by HRTII-RCM was significantly lower in MGD patients (47.6±26.6/mm2) than in control subjects (101.3±33.8/mm2; p<0.05). The mean acinar unit diameter as determined by HRTII-RCM was significantly larger in MGD patients (98.2±53.3 μm) than in controls (41.6±11.9 μm; p<0.05). Both the density and diameter of MG acinar units related significantly with the severity of MG dropout and MG expression grades (p<0.05). Conclusions In vivo confocal microscopy can effectively demonstrate the morphological changes of the MG in patients with MGD. Glandular acinar density and acinar unit diameter seemed to be promising new parameters of in vivo confocal microscopy, which is significantly related to the clinical ocular surface and tear function findings of MGD. PMID:18618006

  1. Application of Antrodia camphorata Promotes Rat's Wound Healing In Vivo and Facilitates Fibroblast Cell Proliferation In Vitro.

    PubMed

    Amin, Zahra A; Ali, Hapipah M; Alshawsh, Mohammed A; Darvish, Pouya H; Abdulla, Mahmood A

    2015-01-01

    Antrodia camphorata is a parasitic fungus from Taiwan, it has been documented to possess a variety of pharmacological and biological activities. The present study was undertaken to evaluate the potential of Antrodia camphorata ethanol extract to accelerate the rate of wound healing closure and histology of wound area in experimental rats. The safety of Antrodia camphorata was determined in vivo by the acute toxicity test and in vitro by fibroblast cell proliferation assay. The scratch assay was used to evaluate the in vitro wound healing in fibroblast cells and the excision model of wound healing was tested in vivo using four groups of adult Sprague Dawley rats. Our results showed that wound treated with Antrodia camphorata extract and intrasite gel significantly accelerates the rate of wound healing closure than those treated with the vehicle. Wounds dressed with Antrodia camphorata extract showed remarkably less scar width at wound closure and granulation tissue contained less inflammatory cell and more fibroblast compared to wounds treated with the vehicle. Masson's trichrom stain showed granulation tissue containing more collagen and less inflammatory cell in Antrodia camphorata treated wounds. In conclusion, Antrodia camphorata extract significantly enhanced the rate of the wound enclosure in rats and promotes the in vitro healing through fibroblast cell proliferation.

  2. Application of Antrodia camphorata Promotes Rat's Wound Healing In Vivo and Facilitates Fibroblast Cell Proliferation In Vitro

    PubMed Central

    Amin, Zahra A.; Ali, Hapipah M.; Alshawsh, Mohammed A.; Darvish, Pouya H.; Abdulla, Mahmood A.

    2015-01-01

    Antrodia camphorata is a parasitic fungus from Taiwan, it has been documented to possess a variety of pharmacological and biological activities. The present study was undertaken to evaluate the potential of Antrodia camphorata ethanol extract to accelerate the rate of wound healing closure and histology of wound area in experimental rats. The safety of Antrodia camphorata was determined in vivo by the acute toxicity test and in vitro by fibroblast cell proliferation assay. The scratch assay was used to evaluate the in vitro wound healing in fibroblast cells and the excision model of wound healing was tested in vivo using four groups of adult Sprague Dawley rats. Our results showed that wound treated with Antrodia camphorata extract and intrasite gel significantly accelerates the rate of wound healing closure than those treated with the vehicle. Wounds dressed with Antrodia camphorata extract showed remarkably less scar width at wound closure and granulation tissue contained less inflammatory cell and more fibroblast compared to wounds treated with the vehicle. Masson's trichrom stain showed granulation tissue containing more collagen and less inflammatory cell in Antrodia camphorata treated wounds. In conclusion, Antrodia camphorata extract significantly enhanced the rate of the wound enclosure in rats and promotes the in vitro healing through fibroblast cell proliferation. PMID:26557855

  3. Development of multifunctional optical coherence tomography and application to mouse myocardial infarction model in vivo (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Jang, Sun-Joo; Park, Taejin; Shin, Inho; Park, Hyun Sang; Shin, Paul; Oh, Wang-Yuhl

    2016-02-01

    Optical coherence tomography (OCT) is a useful imaging method for in vivo tissue imaging with deep penetration and high spatial resolution. However, imaging of the beating mouse heart is still challenging due to limited temporal resolution or penetration depth. Here, we demonstrate a multifunctional OCT system for a beating mouse heart, providing various types of visual information about heart pathophysiology with high spatiotemporal resolution and deep tissue imaging. Angiographic imaging and polarization-sensitive (PS) imaging were implemented with the electrocardiogram (ECG)-triggered beam scanning scheme on the high-speed OCT platform (A-line rate: 240 kHz). Depth-resolved local birefringence and the local orientation of the mouse myocardial fiber were visualized from the PS-OCT. ECG-triggered angiographic OCT (AOCT) with the custom-built motion stabilization imaging window provided myocardial vasculature of a beating mouse heart. Mice underwent coronary artery ligation to derive myocardial infarction (MI) and were imaged with the multifunctional OCT system at multiple time points. AOCT and PS-OCT visualize change of functionality of coronary vessels and myocardium respectively at different phases (acute and chronic) of MI in an ischemic mouse heart. Taken together, the integrated imaging of PS-OCT and AOCT would play an important role in study of MI providing multi-dimensional information of the ischemic mouse heart in vivo.

  4. Application of the critical angle method to refractive index measurement of human skin in vivo under partial contact.

    PubMed

    Yoshida, Kenichiro; Ohkubo, Kohji; Ojima, Nobutoshi; Iwata, Kayoko

    2013-03-01

    We adapted the critical angle method for measuring rough surfaces under partial contact to acquire an in vivo skin refractive index (RI). Assuming that the total reflection is the simple sum of reflection from areas that are in contact and reflection from those that are not in contact, the RI can be estimated even for partial contact with a rough surface. We found that cheek skin is sufficiently soft that a sufficiently large area can be in contact and that the critical angle was detectable. The RIs of the cheeks of adult females were measured. The RI range was about 1.51 to 1.53, at a wavelength of 550 nm, without considering systematic errors. The RIs of cheeks are significantly correlated with their conductance, which corresponds to their water content. We determined the relationship between the RI and conductance within the variation of skin under normal conditions; this relationship was theoretically obtained in previous studies. In the present study, a direct in vivo measurement method was developed that enabled us to measure the RI in daily life, although this method contains errors for several reasons, including disregarding absorption.

  5. Adhesive strength of bone-implant interfaces and in-vivo degradation of PHB composites for load-bearing applications.

    PubMed

    Meischel, M; Eichler, J; Martinelli, E; Karr, U; Weigel, J; Schmöller, G; Tschegg, E K; Fischerauer, S; Weinberg, A M; Stanzl-Tschegg, S E

    2016-01-01

    Aim of this study was to evaluate the response of bone to novel biodegradable polymeric composite implants in the femora of growing rats. Longitudinal observation of bone reaction at the implant site (BV/TV) as well as resorption of the implanted pins were monitored using in vivo micro-focus computed tomography (µCT). After 12, 24 and 36 weeks femora containing the implants were explanted, scanned with high resolution ex vivo µCT, and the surface roughness of the implants was measured to conclude on the ingrowth capability for bone tissue. Scanning electron microscope (SEM) and energy dispersive X-ray spectroscopy (EDX) were used to observe changes on the surface of Polyhydroxybutyrate (PHB) during degradation and cell ingrowth. Four different composites with zirconium dioxide (ZrO2) and Herafill(®) were compared. After 36 weeks in vivo, none of the implants did show significant degradation. The PHB composite with ZrO2 and a high percentage (30%) of Herafill® as well as the Mg-alloy WZ21 showed the highest values of bone accumulation (increased BV/TV) around the implant. The lowest value was measured in PHB with 3% ZrO2 containing no Herafill®. Roughness measurements as well as EDX and SEM imaging could not reveal any changes on the PHB composites׳ surfaces. Biomechanical parameters, such as the adhesion strength between bone and implant were determined by measuring the shear strength as well as push-out energy of the bone-implant interface. The results showed that improvement of these mechanical properties of the studied PHBs P3Z, P3Z10H and P3Z30H is necessary in order to obtain appropriate load-bearing material. The moduli of elasticity, tensile strength and strain properties of the PHB composites are close to that of bone and thus promising. Compared to clinically used PLGA, PGA and PLA materials, their additional benefit is an unchanged local pH value during degradation, which makes them well tolerated by cells and immune system. They might be used

  6. Potential application of in vivo imaging of impaired lymphatic duct to evaluate the severity of pressure ulcer in mouse model

    NASA Astrophysics Data System (ADS)

    Kasuya, Akira; Sakabe, Jun-Ichi; Tokura, Yoshiki

    2014-02-01

    Ischemia-reperfusion (IR) injury is a cause of pressure ulcer. However, a mechanism underlying the IR injury-induced lymphatic vessel damage remains unclear. We investigated the alterations of structure and function of lymphatic ducts in a mouse cutaneous IR model. And we suggested a new method for evaluating the severity of pressure ulcer. Immunohistochemistry showed that lymphatic ducts were totally vanished by IR injury, while blood vessels were relatively preserved. The production of harmful reactive oxygen species (ROS) was increased in injured tissue. In vitro study showed a high vulnerability of lymphatic endothelial cells to ROS. Then we evaluated the impaired lymphatic drainage using an in vivo imaging system for intradermally injected indocyanine green (ICG). The dysfunction of ICG drainage positively correlated with the severity of subsequent cutaneous changes. Quantification of the lymphatic duct dysfunction by this imaging system could be a useful strategy to estimate the severity of pressure ulcer.

  7. In Vivo Noninvasive Analysis of Human Forearm Muscle Function and Fatigue: Applications to EVA Operations and Training Maneuvers

    NASA Technical Reports Server (NTRS)

    Fotedar, L. K.; Marshburn, T.; Quast, M. J.; Feeback, D. L.

    1999-01-01

    Forearm muscle fatigue is one of the major limiting factors affecting endurance during performance of deep-space extravehicular activity (EVA) by crew members. Magnetic resonance (MR) provides in vivo noninvasive analysis of tissue level metabolism and fluid exchange dynamics in exercised forearm muscles through the monitoring of proton magnetic resonance imaging (MRI) and phosphorus magnetic resonance spectroscopy (P-31-MRS) parameter variations. Using a space glove box and EVA simulation protocols, we conducted a preliminary MRS/MRI study in a small group of human test subjects during submaximal exercise and recovery and following exhaustive exercise. In assessing simulated EVA-related muscle fatigue and function, this pilot study revealed substantial changes in the MR image longitudinal relaxation times (T2) as an indicator of specific muscle activation and proton flux as well as changes in spectral phosphocreatine-to-phosphate (PCr/Pi) levels as a function of tissue bioenergetic potential.

  8. Carotid artery wall motion analysis from B-mode ultrasound using adaptive block matching: in silico evaluation and in vivo application

    NASA Astrophysics Data System (ADS)

    Gastounioti, A.; Golemati, S.; Stoitsis, J. S.; Nikita, K. S.

    2013-12-01

    Valid risk stratification for carotid atherosclerotic plaques represents a crucial public health issue toward preventing fatal cerebrovascular events. Although motion analysis (MA) provides useful information about arterial wall dynamics, the identification of motion-based risk markers remains a significant challenge. Considering that the ability of a motion estimator (ME) to handle changes in the appearance of motion targets has a major effect on accuracy in MA, we investigated the potential of adaptive block matching (ABM) MEs, which consider changes in image intensities over time. To assure the validity in MA, we optimized and evaluated the ABM MEs in the context of a specially designed in silico framework. ABMFIRF2, which takes advantage of the periodicity characterizing the arterial wall motion, was the most effective ABM algorithm, yielding a 47% accuracy increase with respect to the conventional block matching. The in vivo application of ABMFIRF2 revealed five potential risk markers: low movement amplitude of the normal part of the wall adjacent to the plaques in the radial (RMAPWL) and longitudinal (LMAPWL) directions, high radial motion amplitude of the plaque top surface (RMAPTS), and high relative movement, expressed in terms of radial strain (RSIPL) and longitudinal shear strain (LSSIPL), between plaque top and bottom surfaces. The in vivo results were reproduced by OFLK(WLS) and ABMKF-K2, MEs previously proposed by the authors and with remarkable in silico performances, thereby reinforcing the clinical values of the markers and the potential of those MEs. Future in vivo studies will elucidate with confidence the full potential of the markers.

  9. Non-contact respiration monitoring for in-vivo murine micro computed tomography: characterization and imaging applications

    NASA Astrophysics Data System (ADS)

    Burk, Laurel M.; Lee, Yueh Z.; Wait, J. Matthew; Lu, Jianping; Zhou, Otto Z.

    2012-09-01

    A cone beam micro-CT has previously been utilized along with a pressure-tracking respiration sensor to acquire prospectively gated images of both wild-type mice and various adult murine disease models. While the pressure applied to the abdomen of the subject by this sensor is small and is generally without physiological effect, certain disease models of interest, as well as very young animals, are prone to atelectasis with added pressure, or they generate too weak a respiration signal with this method to achieve optimal prospective gating. In this work we present a new fibre-optic displacement sensor which monitors respiratory motion of a subject without requiring physical contact. The sensor outputs an analogue signal which can be used for prospective respiration gating in micro-CT imaging. The device was characterized and compared against a pneumatic air chamber pressure sensor for the imaging of adult wild-type mice. The resulting images were found to be of similar quality with respect to physiological motion blur; the quality of the respiration signal trace obtained using the non-contact sensor was comparable to that of the pressure sensor and was superior for gating purposes due to its better signal-to-noise ratio. The non-contact sensor was then used to acquire in-vivo micro-CT images of a murine model for congenital diaphragmatic hernia and of 11-day-old mouse pups. In both cases, quality CT images were successfully acquired using this new respiration sensor. Despite the presence of beam hardening artefacts arising from the presence of a fibre-optic cable in the imaging field, we believe this new technique for respiration monitoring and gating presents an opportunity for in-vivo imaging of disease models which were previously considered too delicate for established animal handling methods.

  10. Application of both a physical theory and statistical procedure in the analyses of an in vivo study of aerosol deposition

    SciTech Connect

    Cheng, K.H.; Swift, D.L.; Yang, Y.H.

    1995-12-01

    Regional deposition of inhaled aerosols in the respiratory tract is a significant factor in assessing the biological effects from exposure to a variety of environmental particles. Understanding the deposition efficiency of inhaled aerosol particles in the nasal and oral airways can help evaluate doses to the extrathoracic region as well as to the lung. Dose extrapolation from laboratory animals to humans has been questioned due to significant physiological and anatomical variations. Although human studies are considered ideal for obtaining in vivo toxicity information important in risk assessment, the number of subjects in the study is often small compared to epidemiological and animal studies. This study measured in vivo the nasal airway dimensions and the extrathoracic deposition of ultrafine aerosols in 10 normal adult males. Variability among individuals was significant. The nasal geometry of each individual was characterized at a resolution of 3 mm using magnetic resonance imaging (MRI) and acoustic rhinometry (AR). The turbulent diffusion theory was used to describe the nonlinear nature of extrathoracic aerosol deposition. To determine what dimensional features of the nasal airway were responsible for the marked differences in particle deposition, the MIXed-effects NonLINear Regression (MIXNLIN) procedure was used to account for the random effort of repeated measurements on the same subject. Using both turbulent diffusion theory and MIXNLIN, the ultrafine particle deposition is correlated with nasal dimensions measured by the surface area, minimum cross-sectional area, and complexity of the airway shape. The combination of MRI and AR is useful for characterizing both detailed nasal dimensions and temporal changes in nasal patency. We conclude that a suitable statistical procedure incorporated with existing physical theories must be used in data analyses for experimental studies of aerosol deposition that involve a relatively small number of human subjects.

  11. In vivo application of ( sup 111 In-DTPA-D-Phe sup 1 )-octreotide for detection of somatostatin receptor-positive tumors in rats

    SciTech Connect

    Bakker, W.H.; Krenning, E.P.; Reubi, J.C.; Breeman, W.A.P.; Setyono-Han, B.; de Jong, M.; Kooij, P.P.M.; Bruns, C.; van Hagen, P.M.; Marbach, P.; Visser, T.J.; Pless, J.; Lamberts, S.W.J. Sandoz Research Inst., Berne Dr. Daniel den Hoed Cancer Centre, Rotterdam Sandoz Pharma AG, Basel )

    1991-01-01

    In this study the authors investigated its in vivo application in the visualization of somatostatin receptor-positive tumors in rats. The distribution of the radiopharmaceutical was investigated after intravenous injection in normal rats and in rats bearing the somatostatin receptor-positive rat pancreatic carcinoma CA 20948. Ex vivo autoradiographic studies showed that specific accumulation of radioactivity occurred in somatostatin receptor-containing tissue (anterior pituitary gland). However, in contrast to the adrenals and pituitary, the tracer accumulation in the kidneys was not mediated by somatostatin receptors. Increasing radioactivity over the somatostatin receptor-positive tumors was measured rapidly after injection and the tumors were clearly visualized by gamma camera scintigraphy. In rats pretreated with 1 mg octreotide accumulation of ({sup 111}In-DPTA-D-Phe{sup 1})-octreotide in the tumors was prevented. Because of its relatively long effective half-life, ({sup 111}In-DTPA-D-Phe{sup 1})-octreotide is a radionuclide-coupled somatostatin analogue which can be used to visualize somatostatin receptor-bearing tumors efficiently after 24 hr, when interfering background radioactivity is minimized by renal clearance.

  12. Functional applications of novel Semliki Forest virus vectors are limited by vector toxicity in cultures of primary neurons in vitro and in the substantia nigra in vivo.

    PubMed

    Lingor, Paul; Schöll, Ulrike; Bähr, Mathias; Kügler, Sebastian

    2005-03-01

    The Semliki Forest virus (SFV) system has been shown to be highly efficient in transduction of cell lines and primary cells. We employed a novel "noncytotoxic" SFV(PD) vector for transduction of primary ventral midbrain floor cultures in vitro and rat substantia nigra in vivo. Rapid protein expression was noted with preferential transduction of neuronal cells including the dopaminergic subpopulation. To examine the suitability of the SFV vector system for functional gene expression, SFV(PD) vectors encoding for antiapoptotic proteins Bcl-X(L) and XIAP were designed. Despite effective transgene expression, SFV(PD) vectors were unable to rescue dopaminergic neurons from MPP+-induced apoptosis. In vivo, virus injection into substantia nigra resulted in fast onset of transgene expression, but elicited an activation of microglia and an inflammation response. We conclude that the use of novel SFV(PD) vectors is currently limited by persistent neurotoxicity of the vector system. Although SFV(PD) vectors may be useful for protein localization studies in dopaminergic neurons, functional applications will require the development of even less cytopathic vector systems.

  13. An in vivo validation of the application of acoustic radiation force to enhance the diagnostic utility of molecular imaging using 3-d ultrasound.

    PubMed

    Gessner, Ryan C; Streeter, Jason E; Kothadia, Roshni; Feingold, Steven; Dayton, Paul A

    2012-04-01

    For more than a decade, the application of acoustic radiation force (ARF) has been proposed as a mechanism to increase ultrasonic molecular imaging (MI) sensitivity in vivo. Presented herein is the first noninvasive in vivo validation of ARF-enhanced MI with an unmodified clinical system. First, an in vitro optical-acoustical setup was used to optimize system parameters and ensure sufficient microbubble translation when exposed to ARF. 3-D ARF-enhanced MI was then performed on 7 rat fibrosarcoma tumors using microbubbles targeted to α(v)β₃ and nontargeted microbubbles. Low-amplitude (<25 kPa) 3-D ARF pulse sequences were tested and compared with passive targeting studies in the same animal. Our results demonstrate that a 78% increase in image intensity from targeted microbubbles can be achieved when using ARF relative to the passive targeting studies. Furthermore, ARF did not significantly increase image contrast when applied to nontargeted agents, suggesting that ARF did not increase nonspecific adhesion.

  14. Determination of steady-state protein breakdown rate in vivo by the disappearance of protein-bound tracer-labeled amino acids: a method applicable in humans.

    PubMed

    Holm, Lars; O'Rourke, Bruce; Ebenstein, David; Toth, Michael J; Bechshoeft, Rasmus; Holstein-Rathlou, Niels-Henrik; Kjaer, Michael; Matthews, Dwight E

    2013-04-15

    A method to determine the rate of protein breakdown in individual proteins was developed and tested in rats and confirmed in humans, using administration of deuterium oxide and incorporation of the deuterium into alanine that was subsequently incorporated into body proteins. Measurement of the fractional breakdown rate of proteins was determined from the rate of disappearance of deuterated alanine from the proteins. The rate of disappearance of deuterated alanine from the proteins was calculated using an exponential decay, giving the fractional breakdown rate (FBR) of the proteins. The applicability of this protein-specific FBR approach is suitable for human in vivo experimentation. The labeling period of deuterium oxide administration is dependent on the turnover rate of the protein of interest.

  15. Special conference of the American Association for Cancer Research on molecular imaging in cancer: linking biology, function, and clinical applications in vivo.

    PubMed

    Luker, Gary D

    2002-04-01

    The AACR Special Conference on Molecular Imaging in Cancer: Linking Biology, Function, and Clinical Applications In Vivo, was held January 23-27, 2002, at the Contemporary Hotel, Walt Disney World, Orlando, FL. Co-Chairs David Piwnica-Worms, Patricia Price and Thomas Meade brought together researchers with diverse expertise in molecular biology, gene therapy, chemistry, engineering, pharmacology, and imaging to accelerate progress in developing and applying technologies for imaging specific cellular and molecular signals in living animals and humans. The format of the conference was the presentation of research that focused on basic and translational biology of cancer and current state-of-the-art techniques for molecular imaging in animal models and humans. This report summarizes the special conference on molecular imaging, highlighting the interfaces of molecular biology with animal models, instrumentation, chemistry, and pharmacology that are essential to convert the dreams and promise of molecular imaging into improved understanding, diagnosis, and management of cancer.

  16. Folic acid-functionalized up-conversion nanoparticles: toxicity studies in vivo and in vitro and targeted imaging applications

    NASA Astrophysics Data System (ADS)

    Sun, Lining; Wei, Zuwu; Chen, Haige; Liu, Jinliang; Guo, Jianjian; Cao, Ming; Wen, Tieqiao; Shi, Liyi

    2014-07-01

    Folate receptors (FRs) are overexpressed on a variety of human cancer cells and tissues, including cancers of the breast, ovaries, endometrium, and brain. This over-expression of FRs can be used to target folate-linked imaging specifically to FR-expressing tumors. Fluorescence is emerging as a powerful new modality for molecular imaging in both the diagnosis and treatment of disease. Combining innovative molecular biology and chemistry, we prepared three kinds of folate-targeted up-conversion nanoparticles as imaging agents (UCNC-FA: UCNC-Er-FA, UCNC-Tm-FA, and UCNC-Er,Tm-FA). In vivo and in vitro toxicity studies showed that these nanoparticles have both good biocompatibility and low toxicity. Moreover, the up-conversion luminescence imaging indicated that they have good targeting to HeLa cells and can therefore serve as potential fluorescent contrast agents.Folate receptors (FRs) are overexpressed on a variety of human cancer cells and tissues, including cancers of the breast, ovaries, endometrium, and brain. This over-expression of FRs can be used to target folate-linked imaging specifically to FR-expressing tumors. Fluorescence is emerging as a powerful new modality for molecular imaging in both the diagnosis and treatment of disease. Combining innovative molecular biology and chemistry, we prepared three kinds of folate-targeted up-conversion nanoparticles as imaging agents (UCNC-FA: UCNC-Er-FA, UCNC-Tm-FA, and UCNC-Er,Tm-FA). In vivo and in vitro toxicity studies showed that these nanoparticles have both good biocompatibility and low toxicity. Moreover, the up-conversion luminescence imaging indicated that they have good targeting to HeLa cells and can therefore serve as potential fluorescent contrast agents. Electronic supplementary information (ESI) available: Up-conversion luminescence spectra of UCNC-Er and UCNC-Er-FA, UCNC-Tm and UCNC-Tm-FA. Confocal luminescence imaging data collected as a series along the Z optical axis. See DOI: 10.1039/c4nr02312a

  17. Preparation and In Vitro/Ex Vivo Evaluation of Moxifloxacin-Loaded PLGA Nanosuspensions for Ophthalmic Application

    PubMed Central

    Mudgil, Meetali; Pawar, Pravin K.

    2013-01-01

    The aim of the present investigation was to prepare a colloidal ophthalmic formulation to improve the residence time of moxifloxacin. Moxifloxacin-loaded poly(dl-lactide-co-glycolide) (PLGA) nanosuspensions were prepared by using the solvent evaporation technique. The nanosuspensions were characterised physically by using different techniques like particle size, zeta potential, FTIR, DSC, and XRD analysis. In vitro and ex vivo studies of nanosuspensions were carried out using a modified USP dissolution apparatus and all-glass Franz diffusion cells, respectively. The antibacterial activities of the nanosuspension and marketed formulations were performed against S. aureus and P. aeroginosa. The moxifloxacin-loaded PLGA nanosuspensions showed uniform particle size, ranging between 164–490 nm with negative zeta potential for all batches. The percentage entrapment efficiency of the drug-loaded nano-suspension was found to be between 84.09 to 92.05%. In vitro drug release studies suggest that all of the formulations showed extended drug release profiles and follow Korsemeyer-Peppas release kinetics. In vitro corneal permeability was found to be comparable with that of the marketed formulation across isolated goat cornea, indicating the suitability of the nanosuspension formulation in the ophthalmic delivery of moxifloxacin. The optimised nano-suspension was found to be more active against S. aureus and P. aeruginosa compared to the marketed eye drops. PMID:23833723

  18. In vivo biocompatibility assessment of (PTFE–PVDF–PP) terpolymer-based membrane with potential application for glaucoma treatment

    PubMed Central

    Leszczynski, Rafał; Stodolak, Ewa; Wieczorek, Jarosław; Orlowska-Heitzman, Jolanta; Gumula, Teresa

    2010-01-01

    The aim of the work was to evaluate the in vivo biological behaviour of polymeric membrane materials for glaucoma implants. The base material was biostable synthetic terpolymer (PTFE–PVDF–PP) with proved biocompability (PN-EN ISO 10993). The samples manufactured in the form a membrane were subjected to chemical and physical treatment to create an open pore system within the polymer matrix. As a porogenic phase biodegradable natrium alginate in a fibrous form was employed. The non-perforating deep sclerectomy technique was performed in a rabbit model. The clinical observations were made after 14 and 30 days. During the study clinical symptoms of a moderate degree were observed, and histopathological changes were typical for foreign body implantation. At the end stage of the study no significant difference in histopathological assessment was found between control and experimental group. Similarities observed in both groups and relatively mild histopathological changes in the tissue surrounding the implant indicate that the observed symptoms come from a deep scleral trauma caused by surgery, and not by the presence of the implant itself. PMID:20652824

  19. In Vivo Osseointegration Performance of Titanium Dioxide Coating Modified Polyetheretherketone Using Arc Ion Plating for Spinal Implant Application

    PubMed Central

    Tsou, Hsi-Kai; Chi, Meng-Hui; Hung, Yi-Wen; Chung, Chi-Jen; He, Ju-Liang

    2015-01-01

    Polyetheretherketone (PEEK), which has biomechanical performance similar to that of human cancellous bone, is used widely as a spinal implant material. However, its bioinertness and hydrophobic surface properties result in poor osseointegration. This study applies a novel modification method, arc ion plating (AIP), that produces a highly osteoblast compatible titanium dioxide (TiO2) coatings on a PEEK substrate. This PEEK with TiO2 coating (TiO2/PEEK) was implanted into the femurs of New Zealand white male rabbits to evaluate its in vivo performance by the push-out test and histological observation. Analytical results show that AIP can prepare TiO2 coatings on bullet-shaped PEEK substrates as implant materials. After prolonged implantation in rabbits, no signs of inflammation existed. Newly regenerated bone formed more prominently with the TiO2/PEEK implant by histological observation. The shear strength of the bone/implant interface increases as implantation period increases. Most importantly, bone bonding performance of the TiO2/PEEK implant was superior to that of bare PEEK. The rutile-TiO2 coatings achieved better osseointegration than the anatase-TiO2 coatings. Therefore, AIP-TiO2 can serve as a novel surface modification method on PEEK for spinal interbody fusion cages. PMID:26504800

  20. Modelling and testing of a piezoelectric ultrasonic micro-motor suitable for in vivo micro-robotic applications

    NASA Astrophysics Data System (ADS)

    Watson, B.; Friend, J.; Yeo, L.

    2010-11-01

    A piezoelectric ultrasonic resonant micro-motor is developed with a stator diameter of 241 μm and an overall diameter of 400 μm. The motor is shown to produce a start-up torque of 1.2 nN m and a peak output power of 0.25 μW as designed, with a preload of 46.6 μN. An increase in preload to 2264 μN improved the performance to a start-up torque of 29 nN m and a peak output power of 9.1 μW. The motor is five times smaller than the current smallest piezoelectric ultrasonic resonant motor produced by Kanda et al. The motor is designed to operate at approximately 771 kHz, matching the fundamental axial, second harmonic torsional and electro-mechanical resonant frequencies. This is achieved through the use of a novel design process that uses scaling theories to greatly reduce the computational time to design the device. The resultant size and performance of the motor make it the first motor design capable of meeting the requirements of a drive system in a tetherless swimming in vivo micro-robot.

  1. Ex vivo application of carbon monoxide in UW solution prevents transplant-induced renal ischemia/reperfusion injury in pigs.

    PubMed

    Yoshida, J; Ozaki, K S; Nalesnik, M A; Ueki, S; Castillo-Rama, M; Faleo, G; Ezzelarab, M; Nakao, A; Ekser, B; Echeverri, G J; Ross, M A; Stolz, D B; Murase, N

    2010-04-01

    I/R injury is a major deleterious factor of successful kidney transplantation (KTx). Carbon monoxide (CO) is an endogenous gaseous regulatory molecule, and exogenously delivered CO in low concentrations provides potent cytoprotection. This study evaluated efficacies of CO exposure to excised kidney grafts to inhibit I/R injury in the pig KTx model. Porcine kidneys were stored for 48 h in control UW or UW supplemented with CO (CO-UW) and autotransplanted in a 14-day follow-up study. In the control UW group, animal survival was 80% (4/5) with peak serum creatinine levels of 12.0 +/- 5.1 mg/dL. CO-UW showed potent protection, and peak creatinine levels were reduced to 6.9 +/- 1.4 mg/dL with 100% (5/5) survival without any noticeable adverse event or abnormal COHb value. Control grafts at 14 days showed significant tubular damages, focal fibrotic changes and numerous infiltrates. The CO-UW group showed significantly less severe histopathological changes with less TGF-beta and p-Smad3 expression. Grafts in CO-UW also showed significantly lower early mRNA levels for proinflammatory cytokines and less lipid peroxidation. CO in UW provides significant protection against renal I/R injury in the porcine KTx model. Ex vivo exposure of kidney grafts to CO during cold storage may therefore be a safe strategy to reduce I/R injury.

  2. Automated Segmentation and Object Classification of CT Images: Application to In Vivo Molecular Imaging of Avian Embryos

    PubMed Central

    Schmidt, Jana; Zimmermann, Johannes; Saluz, Hans Peter

    2013-01-01

    Background. Although chick embryogenesis has been studied extensively, there has been growing interest in the investigation of skeletogenesis. In addition to improved poultry health and minimized economic loss, a greater understanding of skeletal abnormalities can also have implications for human medicine. True in vivo studies require noninvasive imaging techniques such as high-resolution microCT. However, the manual analysis of acquired images is both time consuming and subjective. Methods. We have developed a system for automated image segmentation that entails object-based image analysis followed by the classification of the extracted image objects. For image segmentation, a rule set was developed using Definiens image analysis software. The classification engine was implemented using the WEKA machine learning tool. Results. Our system reduces analysis time and observer bias while maintaining high accuracy. Applying the system to the quantification of long bone growth has allowed us to present the first true in ovo data for bone length growth recorded in the same chick embryos. Conclusions. The procedures developed represent an innovative approach for the automated segmentation, classification, quantification, and visualization of microCT images. MicroCT offers the possibility of performing longitudinal studies and thereby provides unique insights into the morpho- and embryogenesis of live chick embryos. PMID:23997760

  3. In vivo fluence rate measurements during Foscan®-mediated photodynamic therapy of persistent and recurrent nasopharyngeal carcinomas using a dedicated light applicator

    NASA Astrophysics Data System (ADS)

    van Veen, R. L. P.; Nyst, H.; Indrasari, S. R.; Yudharto, M. A.; Robinson, D. J.; Tan, I. B.; Meewis, C.; Peters, R.; Spaniol, Stefan B.; Stewart, Fiona A.; Levendag, P. C.; Sterenborg, Henricus J. C. M.

    2006-07-01

    The objective of this study was to evaluate the performance of a dedicated light applicator for light delivery and fluence rate monitoring during Foscan®-mediated photodynamic therapy of nasopharyngeal carcinoma in a clinical phase I/II study. We have developed a flexible silicone applicator that can be inserted through the mouth and fixed in the nasopharyngeal cavity. Three isotropic fibers, for measuring of the fluence (rate) during therapy, were located within the nasopharyngeal tumor target area and one was manually positioned to monitor structures at risk in the shielded area. A flexible black silicon patch tailored to the patient's anatomy is attached to the applicator to shield the soft palate and oral cavity from the 652-nm laser light. Fourteen patients were included in the study, resulting in 26 fluence rate measurements in the risk volume (two failures). We observed a systematic reduction in fluence rate during therapy in 20 out of 26 illuminations, which may be related to photodynamic therapy-induced increased blood content, decreased oxygenation, or reduced scattering. Our findings demonstrate that the applicator was easily inserted into the nasopharynx. The average light distribution in the target area was reasonably uniform over the length of the applicator, thus giving an acceptably homogeneous illumination throughout the cavity. Shielding of the risk area was adequate. Large interpatient variations in fluence rate stress the need for in vivo dosimetry. This enables corrections to be made for differences in optical properties and geometry resulting in comparable amounts of light available for Foscan® absorption.

  4. Clinical Application of In-Room Positron Emission Tomography for In Vivo Treatment Monitoring in Proton Radiation Therapy

    SciTech Connect

    Min, Chul Hee; Zhu, Xuping; Winey, Brian A.; Grogg, Kira; Testa, Mauro; El Fakhri, Georges; Bortfeld, Thomas R.; Paganetti, Harald; Shih, Helen A.

    2013-05-01

    Purpose: The purpose of this study is to evaluate the potential of using in-room positron emission tomography (PET) for treatment verification in proton therapy and for deriving suitable PET scan times. Methods and Materials: Nine patients undergoing passive scattering proton therapy underwent scanning immediately after treatment with an in-room PET scanner. The scanner was positioned next to the treatment head after treatment. The Monte Carlo (MC) method was used to reproduce PET activities for each patient. To assess the proton beam range uncertainty, we designed a novel concept in which the measured PET activity surface distal to the target at the end of range was compared with MC predictions. The repositioning of patients for the PET scan took, on average, approximately 2 minutes. The PET images were reconstructed considering varying scan times to test the scan time dependency of the method. Results: The measured PET images show overall good spatial correlations with MC predictions. Some discrepancies could be attributed to uncertainties in the local elemental composition and biological washout. For 8 patients treated with a single field, the average range differences between PET measurements and computed tomography (CT) image-based MC results were <5 mm (<3 mm for 6 of 8 patients) and root-mean-square deviations were 4 to 11 mm with PET-CT image co-registration errors of approximately 2 mm. Our results also show that a short-length PET scan of 5 minutes can yield results similar to those of a 20-minute PET scan. Conclusions: Our first clinical trials in 9 patients using an in-room PET system demonstrated its potential for in vivo treatment monitoring in proton therapy. For a quantitative range prediction with arbitrary shape of target volume, we suggest using the distal PET activity surface.

  5. In vivo complementation of complex I by the yeast Ndi1 enzyme. Possible application for treatment of Parkinson disease.

    PubMed

    Seo, Byoung Boo; Nakamaru-Ogiso, Eiko; Flotte, Terence R; Matsuno-Yagi, Akemi; Yagi, Takao

    2006-05-19

    Recent studies suggest that dysfunction of the NADH-quinone oxidoreductase (complex I) is associated with a number of human diseases, including neurodegenerative disorders such as Parkinson disease. We have shown previously that the single subunit rotenone-insensitive NADH-quinone oxidoreductase (Ndi1) of Saccharomyces cerevisiae mitochondria can restore NADH oxidation in complex I-deficient mammalian cells. The Ndi1 enzyme is insensitive to complex I inhibitors such as rotenone and 1-methyl-4-phenylpyridinium ion, known as a metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). To test the possible use of the NDI1 gene as a therapeutic agent in vivo, we chose a mouse model of Parkinson disease. The NDI1-recombinant adeno-associated virus particles (rAAV-NDI1) were injected unilaterally into the substantia nigra of mice. The animals were then subjected to treatment with MPTP. The degree of neurodegeneration in the nigrostriatal system was assessed immunohistochemically through the analysis of tyrosine hydroxylase and glial fibrillary acidic protein. It was evident that the substantia nigra neurons on the side used for injection of rAAV-NDI1 retained a high level of tyrosine hydroxylase-positive cells, and the ipsilateral striatum exhibited significantly less denervation than the contralateral striatum. Furthermore, striatal concentrations of dopamine and its metabolites in the hemisphere that received rAAV-NDI1 were substantially higher than those of the untreated hemisphere, reaching more than 50% of the normal levels. These results indicate that the expressed Ndi1 protein elicits resistance to MPTP-induced neuronal injury. The present study is the first successful demonstration of complementation of complex I by the Ndi1 enzyme in animals.

  6. Applications of In Vivo Functional Testing of the Rat Tibialis Anterior for Evaluating Tissue Engineered Skeletal Muscle Repair

    PubMed Central

    Mintz, Ellen L.; Passipieri, Juliana A.; Lovell, Daniel Y.; Christ, George J.

    2016-01-01

    Despite the regenerative capacity of skeletal muscle, permanent functional and/or cosmetic deficits (e.g., volumetric muscle loss (VML) resulting from traumatic injury, disease and various congenital, genetic and acquired conditions are quite common. Tissue engineering and regenerative medicine technologies have enormous potential to provide a therapeutic solution. However, utilization of biologically relevant animal models in combination with longitudinal assessments of pertinent functional measures are critical to the development of improved regenerative therapeutics for treatment of VML-like injuries. In that regard, a commercial muscle lever system can be used to measure length, tension, force and velocity parameters in skeletal muscle. We used this system, in conjunction with a high power, bi-phase stimulator, to measure in vivo force production in response to activation of the anterior crural compartment of the rat hindlimb. We have previously used this equipment to assess the functional impact of VML injury on the tibialis anterior (TA) muscle, as well as the extent of functional recovery following treatment of the injured TA muscle with our tissue engineered muscle repair (TEMR) technology. For such studies, the left foot of an anaesthetized rat is securely anchored to a footplate linked to a servomotor, and the common peroneal nerve is stimulated by two percutaneous needle electrodes to elicit muscle contraction and dorsiflexion of the foot. The peroneal nerve stimulation-induced muscle contraction is measured over a range of stimulation frequencies (1-200 Hz), to ensure an eventual plateau in force production that allows for an accurate determination of peak tetanic force. In addition to evaluation of the extent of VML injury as well as the degree of functional recovery following treatment, this methodology can be easily applied to study diverse aspects of muscle physiology and pathophysiology. Such an approach should assist with the more rational

  7. An innovative application of a small-scale motion analysis technique to quantify human skin deformation in vivo.

    PubMed

    Mahmud, Jamaluddin; Holt, Cathy A; Evans, Sam L

    2010-03-22

    This study highlights a new experimental method developed to measure full-field deformation of human skin in vivo. The technique uses a small-scale Qualisys (Sweden) 3D motion capture system and an array of reflective markers placed on the forearm of five healthy volunteers. A load of up to 1.5N was applied to induce skin deformation by pulling a fine wire attached to the centre of the marker configuration. Loading and marker displacements were recorded simultaneously. 3D marker trajectory data was generated for three different load directions. Tests were repeated to investigate accuracy and repeatability. Calibration results indicate the accuracy of the motion capture system with an average residual of 0.05 mm. The procedure was found to be repeatable and accurate for five repeated tests of measured displacements with a maximum variance of 5%. Experimental data are presented to demonstrate robustness and the ability to produce significant outputs. For all five subjects, at 1N load, the mean and standard deviations of skin axial and lateral displacements were found to be 11.7+/-1.6mm and 12.3+/-3.3mm, respectively. The axial displacements ratio (u(90)/u(0)) ranges from 0.63 to 1.45 with mean+/-standard deviation of 0.982+/-0.34 and 0.982+/-0.32 for left and right arms, respectively. The experiments generated useful and accurate data that can be used to study the viscoelastic, hyperelastic or anisotropic behaviour of human skin. The measured displacements will be analysed further to determine the mechanical properties of skin using inverse Finite Element Analysis and Ogden model.

  8. In vitro and in vivo evaluation of the chitosan/Tur composite film for wound healing applications.

    PubMed

    Zou, Qin; Cai, Bin; Li, Junfeng; Li, Jidong; Li, Yubao

    2017-05-01

    We have developed tourmaline/chitosan (Tur/CS) composite films for wound healing applications. The characteristics of composite films were studied by optical microscope, infrared spectra and X-ray diffraction. Tur particles were uniformly distributed in the CS film and the crystal structure of CS was not remarkably changed except the decrease of crystallinity. The influence of Tur on wound healing applications was characterized by modulating Tur concentrations in the Tur/CS composite film prepared by loading Tur powder into CS matrix with different proportion (0, 1/40 and 1/10). Then L929 cells were co-cultured on the composite films to access the cytotoxicity in vitro. Tur concentrations strongly influenced cell process extension. Tur/CS composite film with 1/40 mass ratio could promote the cell adhesion and proliferation. Fewer and shorter processes were observed at high Tur density. When the composite films were transplanted on porcine full-thickness burn wounds, histological results demonstrated that the Tur/CS group with 1/40 mass ratio had a significantly higher number of newly-formed and mature blood vessels, and fastest regeneration of dermis. Based on the observed facts these films can be tailored for their potential utilization in wound healing and skin tissue engineering applications.

  9. The application of micro-CT in monitoring bone alterations in tail-suspended rats in vivo

    NASA Astrophysics Data System (ADS)

    Luan, Hui-Qin; Sun, Lian-Wen; Huang, Yun-Fei; Wang, Ying; McClean, Colin J.; Fan, Yu-Bo

    2014-06-01

    Osteopenia is a pathological process that affects human skeletal health not only on earth but also in long-time spaceflight. Micro-computed tomography (micro-CT) is a nondestructive method for assessing both bone quantity and bone quality. To investigate the characteristics of micro-CT on evaluating the microgravity-induced osteopenia (e.g. early detection time and the sensitive parameters), the bone loss process of tail-suspended rats was monitored by micro-CT in this study. 8-Week-old female Sprague Dawley rats were divided into two groups: tail suspension (TS) and control (CON). Volumetric bone mineral density (vBMD) and microstructure of the femur and tibia were evaluated in vivo by micro-CT at 0, 7, 14, 22 days. Biomechanical properties of the femur and tibia were determined by three-point bending test. The ash weight of bone was also investigated. The results showed that (1) bone loss in the proximal tibia appeared earlier than in the distal femur. (2) On day 7, the percent bone volume (BV/TV) of the tibia 15.44% decreased significantly, and the trabecular separation (Tb.Sp) 30.29% increased significantly in TS group, both of which were detected earlier than other parameters. (3) Biomechanical properties (e.g. femur, -22.4% maximum load and -23.75% Young’s modulus vs. CON) and ash weight of the femur and tibia decreased significantly in the TS group in comparison to CON group. (4) vBMD of the femur and tibia were clearly related to bone ash and dry weight (r = 0.75-0.87, p < 0.05). (5) BV/TV of both femur and tibia were clearly related to maximum load and Young’s modulus (r = 0.66-0.87, p < 0.05). Similarly, trabecular vBMD and BV/TV of the femur and tibia were clearly related to Young’s modulus (r = 0.73-0.89, p < 0.05). These indicated that BV/TV and Tb.Sp were more sensitive than other parameters for evaluating bone loss induced by tail suspension, moreover, trabecular vBMD and other parameters might be used to evaluate bone strength. Therefore

  10. Application of laser-induced autofluorescence spectra detection system in human colorectal cancer in-vivo screening

    NASA Astrophysics Data System (ADS)

    Chia, Teck Chee; Fu, Sheng; Chia, Yee Hong; Kwek, Leong Chuan; Tang, Choong Leong

    2005-09-01

    This study aimed at applying Laser induced-autofluorescence (LIAF) diagnostics method as an in-vivo screening of colorectal polyplcancer. The spectrum algorithm based on the ratio of autofluorescence intensity was used to identify the diseased tissues from the normal tissues as it was generally performed better than an algorithm based only simply on the intensity of the spectrum. Histopathological biopsy results were compared with the detected AF spectra characteristics for different kinds of polyps. 73 patients had been examined via the LIAF spectroscopy detection system during their colonoscopy screening in Endoscopy Center, Singapore General Hospital. The autofluorescence from the surface of the colorectal tissues under 405 nm laser light excitation was detected using our detecting system. In the experimental investigation two groups of patients were involved. One group was "abnormal" group. There were 25 patients belonging to this group since polyps or carcinoma was found in their colorectal tract during colonoscopy. The histopathology reports confirm the group classification. Total 36 polyps' AF spectra and 9 carcinoma' AF spectra were detected from 25 patients of the abnormal group during their regular endoscopy examination. The intensity ratios RI-680/I-500 and RI-630/I-500 of polyps/cancerous AF spectra and intensity ratios of corresponding normal colorectal AF spectra were calculated. Two critical intensity ratios for separating the AF intensity ratios RI-680/I-500 and RI-630/I-500 of normal and abnormal colorectal tissues were defined as 0.5 and 0.6 respectively. Using the critical intensity ratio values, 48 "normal" group patients' rectums were checked via the LIAF detection system. There were 20 patients (41.7%) whose AF spectra of colorectal tract mucosa belonging to abnormal spectra. However, these 20 patients had not been found under white light via traditional endoscopy. For small diseased area like small plat polyp disease and carcinoma, it was

  11. Application of Wnt Pathway Inhibitor Delivering Scaffold for Inhibiting Fibrosis in Urethra Strictures: In Vitro and in Vivo Study

    PubMed Central

    Zhang, Kaile; Guo, Xuran; Zhao, Weixin; Niu, Guoguang; Mo, Xiumei; Fu, Qiang

    2015-01-01

    Objective: To evaluate the mechanical property and biocompatibility of the Wnt pathway inhibitor (ICG-001) delivering collagen/poly(l-lactide-co-caprolactone) (P(LLA-CL)) scaffold for urethroplasty, and also the feasibility of inhibiting the extracellular matrix (ECM) expression in vitro and in vivo. Methods: ICG-001 (1 mg (2 mM)) was loaded into a (P(LLA-CL)) scaffold with the co-axial electrospinning technique. The characteristics of the mechanical property and drug release fashion of scaffolds were tested with a mechanical testing machine (Instron) and high-performance liquid chromatography (HPLC). Rabbit bladder epithelial cells and the dermal fibroblasts were isolated by enzymatic digestion method. (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay) and scanning electron microscopy (SEM) were used to evaluate the viability and proliferation of the cells on the scaffolds. Fibrolasts treated with TGF-β1 and ICG-001 released medium from scaffolds were used to evaluate the anti-fibrosis effect through immunofluorescence, real time PCR and western blot. Urethrography and histology were used to evaluate the efficacy of urethral implantation. Results: The scaffold delivering ICG-001 was fabricated, the fiber diameter and mechanical strength of scaffolds with inhibitor were comparable with the non-drug scaffold. The SEM and MTT assay showed no toxic effect of ICG-001 to the proliferation of epithelial cells on the collagen/P(LLA-CL) scaffold with ICG-001. After treatment with culture medium released from the drug-delivering scaffold, the expression of Collagen type 1, 3 and fibronectin of fibroblasts could be inhibited significantly at the mRNA and protein levels. In the results of urethrography, urethral strictures and fistulas were found in the rabbits treated with non-ICG-001 delivering scaffolds, but all the rabbits treated with ICG-001-delivering scaffolds showed wide caliber in urethras. Histology results showed less collagen but more

  12. The application of P-gp inhibiting phospholipids as novel oral bioavailability enhancers - An in vitro and in vivo comparison.

    PubMed

    Weinheimer, Manuel; Fricker, Gert; Burhenne, Jürgen; Mylius, Patricia; Schubert, Rolf

    2016-08-30

    The efflux transporter P-glycoprotein (P-gp) significantly modulates drug transport across the intestinal mucosa, strongly reducing the systemic absorption of various active pharmaceutical ingredients. P-gp inhibitors could serve as helpful tools to enhance the oral bioavailability of those substances. As a membrane-associated protein P-gp is surrounded and influenced by phospholipids. Some synthetic phospholipids have been found to strongly reduce P-gp's activity. In this study two representative phospholipids, 1,2-dioctanoyl-sn-glycero-3-phosphocholine (8:0 PC) and 1,2-didecanoyl-sn-glycero-3-phosphocholine (10:0 PC), were compared with Tween® 80 and Cremophor® EL, both commonly used surfactants with P-gp inhibitory properties. Their influence on the cellular transport of the P-gp substrate rhodamine 123 (RH123) was examined using Caco-2 cell layers. In addition, fluorescence anisotropy measurements were performed in order to investigate their effect on membrane fluidity. Finally, we compared the phospholipids with Tween® 80 and the competitive P-gp inhibitor verapamil in an in vivo study, testing their effects on the oral bioavailability of the P-gp substrate drug ritonavir. Both phospholipids not only led to the strongest absorption of RH123, but a permeability enhancing effect was detected in addition to the P-gp inhibition. Their effects on membrane fluidity were not consistent with their P-gp inhibiting effects, and therefore suggested a more complex mode of action. Both phospholipids significantly increased the area under the ritonavir plasma level curve (AUC) within 150min by more than tenfold, but were inferior to Tween® 80, which showed superior solubilizing effects. Finally, these phospholipids represent a novel substance class showing a high permeabilization potential for P-gp substrates. Because of their physiological structure and intestinal degradability, good tolerability without systemic absorption is expected. Formulating P-gp substrates with

  13. In vitro and preliminary in vivo toxicity screening of high-surface-area TiO2-chondroitin-4-sulfate nanocomposites for bone regeneration application.

    PubMed

    Kandiah, Kavitha; Venkatachalam, Rajendran; Wang, Chunyan; Valiyaveettil, Suresh; Ganesan, Kumaresan

    2015-04-01

    The goal of this study was to prepare nontoxic, biomimetic TiO2/chondroitin-4-sulfate nanocomposites with osteointegration ability for biomedical applications. Nanocomposites with higher surface area were subjected to bioactivity study and obtained bone-like layer with stoichiometric Ca/P ratio of 1.64 and 1.66. The susceptibility of nanocomposites against Staphylococcus aureus (∼16 mm) and Escherichia coli (∼12 mm) is favorable in preventing the risk of bone diseases and postoperative infections. Adequate swelling and degradations properties were favorably achieved to reduce the risk of nanoparticle accumulation in cell organelles. Moreover, the toxicity in AGS cell line and biocompatibility in osteoblast-like MG-63 cell line showed no significant mitochondrial damage. In addition, the in vitro expression of osteoblast inducing genes (OCN, OPN, ALP and COL 1) and their up-regulation, and 20% of increased hatching rate in preliminary in vivo (zebrafish) analysis were favorable for the nanocomposite at the ratio of 2:0.50 than pure TiO2. Hence, it can be concluded that among the prepared nanocomposites TCs.5 is a promising biomimetic biomaterial that can be used for advanced orthopedic research and other applications.

  14. In-Vivo Characterization of Glassy Carbon Micro-Electrode Arrays for Neural Applications and Histological Analysis of the Brain Tissue

    NASA Astrophysics Data System (ADS)

    Vomero, Maria

    The aim of this work is to fabricate and characterize glassy carbon Microelectrode Arrays (MEAs) for sensing and stimulating neural activity, and conduct histological analysis of the brain tissue after the implant to determine long-term performance. Neural applications often require robust electrical and electrochemical response over a long period of time, and for those applications we propose to replace the commonly used noble metals like platinum, gold and iridium with glassy carbon. We submit that such material has the potential to improve the performances of traditional neural prostheses, thanks to better charge transfer capabilities and higher electrochemical stability. Great interest and attention is given in this work, in particular, to the investigation of tissue response after several weeks of implants in rodents' brain motor cortex and the associated materials degradation. As part of this work, a new set of devices for Electrocorticography (ECoG) has been designed and fabricated to improve durability and quality of the previous generation of devices, designed and manufactured by the same research group in 2014. In-vivo long-term impedance measurements and brain activity recordings were performed to test the functionality of the neural devices. In-vitro electrical characterization of the carbon electrodes, as well as the study of the adhesion mechanisms between glassy carbon and different substrates is also part of the research described in this book.

  15. Transcutaneous application of carbon dioxide (CO2) induces mitochondrial apoptosis in human malignant fibrous histiocytoma in vivo.

    PubMed

    Onishi, Yasuo; Kawamoto, Teruya; Ueha, Takeshi; Kishimoto, Kenta; Hara, Hitomi; Fukase, Naomasa; Toda, Mitsunori; Harada, Risa; Minoda, Masaya; Sakai, Yoshitada; Miwa, Masahiko; Kurosaka, Masahiro; Akisue, Toshihiro

    2012-01-01

    Mitochondria play an essential role in cellular energy metabolism and apoptosis. Previous studies have demonstrated that decreased mitochondrial biogenesis is associated with cancer progression. In mitochondrial biogenesis, peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α) regulates the activities of multiple nuclear receptors and transcription factors involved in mitochondrial proliferation. Previously, we showed that overexpression of PGC-1α leads to mitochondrial proliferation and induces apoptosis in human malignant fibrous histiocytoma (MFH) cells in vitro. We also demonstrated that transcutaneous application of carbon dioxide (CO(2)) to rat skeletal muscle induces PGC-1α expression and causes an increase in mitochondrial proliferation. In this study, we utilized a murine model of human MFH to determine the effect of transcutaneous CO(2) exposure on PGC-1α expression, mitochondrial proliferation and cellular apoptosis. PGC-1α expression was evaluated by quantitative real-time PCR, while mitochondrial proliferation was assessed by immunofluorescence staining and the relative copy number of mitochondrial DNA (mtDNA) was assessed by real-time PCR. Immunofluorescence staining and DNA fragmentation assays were used to examine mitochondrial apoptosis. We also evaluated the expression of mitochondrial apoptosis related proteins, such as caspases, cytochorome c and Bax, by immunoblot analysis. We show that transcutaneous application of CO(2) induces PGC-1α expression, and increases mitochondrial proliferation and apoptosis of tumor cells, significantly reducing tumor volume. Proteins involved in the mitochondrial apoptotic cascade, including caspase 3 and caspase 9, were elevated in CO(2) treated tumors compared to control. We also observed an enrichment of cytochrome c in the cytoplasmic fraction and Bax protein in the mitochondrial fraction of CO(2) treated tumors, highlighting the involvement of mitochondria in apoptosis. These

  16. Evaluation of Drosophila metabolic labeling strategies for in vivo quantitative proteomic analyses with applications to early pupa formation and amino acid starvation.

    PubMed

    Chang, Ying-Che; Tang, Hong-Wen; Liang, Suh-Yuen; Pu, Tsung-Hsien; Meng, Tzu-Ching; Khoo, Kay-Hooi; Chen, Guang-Chao

    2013-05-03

    Although stable isotope labeling by amino acids in cell culture (SILAC)-based quantitative proteomics was first developed as a cell culture-based technique, stable isotope-labeled amino acids have since been successfully introduced in vivo into select multicellular model organisms by manipulating the feeding diets. An earlier study by others has demonstrated that heavy lysine labeled Drosophila melanogaster can be derived by feeding with an exclusive heavy lysine labeled yeast diet. In this work, we have further evaluated the use of heavy lysine and/or arginine for metabolic labeling of fruit flies, with an aim to determine its respective quantification accuracy and versatility. In vivo conversion of heavy lysine and/or heavy arginine to several nonessential amino acids was observed in labeled flies, leading to distorted isotope pattern and underestimated heavy to light ratio. These quantification defects can nonetheless be rectified at protein level using the normalization function. The only caveat is that such a normalization strategy may not be suitable for every biological application, particularly when modified peptides need to be individually quantified at peptide level. In such cases, we showed that peptide ratios calculated from the summed intensities of all isotope peaks are less affected by the heavy amino acid conversion and therefore less sequence-dependent and more reliable. Applying either the single Lys8 or double Lys6/Arg10 metabolic labeling strategy to flies, we quantitatively mapped the proteomic changes during the onset of metamorphosis and upon amino acid deprivation. The expression of a number of steroid hormone 20-hydroxyecdysone regulated proteins was found to be changed significantly during larval-pupa transition, while several subunits of the V-ATPase complex and components regulating actomyosin were up-regulated under starvation-induced autophagy conditions.

  17. Ultrapure laser-synthesized Si-based nanomaterials for biomedical applications: in vivo assessment of safety and biodistribution

    NASA Astrophysics Data System (ADS)

    Baati, Tarek; Al-Kattan, Ahmed; Esteve, Marie-Anne; Njim, Leila; Ryabchikov, Yury; Chaspoul, Florence; Hammami, Mohamed; Sentis, Marc; Kabashin, Andrei V.; Braguer, Diane

    2016-05-01

    Si/SiOx nanoparticles (NPs) produced by laser ablation in deionized water or aqueous biocompatible solutions present a novel extremely promising object for biomedical applications, but the interaction of these NPs with biological systems has not yet been systematically examined. Here, we present the first comprehensive study of biodistribution, biodegradability and toxicity of laser-synthesized Si-SiOx nanoparticles using a small animal model. Despite a relatively high dose of Si-NPs (20 mg/kg) administered intravenously in mice, all controlled parameters (serum, enzymatic, histological etc.) were found to be within safe limits 3 h, 24 h, 48 h and 7 days after the administration. We also determined that the nanoparticles are rapidly sequestered by the liver and spleen, then further biodegraded and directly eliminated in urine without any toxicity effects. Finally, we found that intracellular accumulation of Si-NPs does not induce any oxidative stress damage. Our results evidence a huge potential in using these safe and biodegradable NPs in biomedical applications, in particular as vectors, contrast agents and sensitizers in cancer therapy and diagnostics (theranostics).

  18. Ultrapure laser-synthesized Si-based nanomaterials for biomedical applications: in vivo assessment of safety and biodistribution

    PubMed Central

    Baati, Tarek; Al-Kattan, Ahmed; Esteve, Marie-Anne; Njim, Leila; Ryabchikov, Yury; Chaspoul, Florence; Hammami, Mohamed; Sentis, Marc; Kabashin, Andrei V.; Braguer, Diane

    2016-01-01

    Si/SiOx nanoparticles (NPs) produced by laser ablation in deionized water or aqueous biocompatible solutions present a novel extremely promising object for biomedical applications, but the interaction of these NPs with biological systems has not yet been systematically examined. Here, we present the first comprehensive study of biodistribution, biodegradability and toxicity of laser-synthesized Si-SiOx nanoparticles using a small animal model. Despite a relatively high dose of Si-NPs (20 mg/kg) administered intravenously in mice, all controlled parameters (serum, enzymatic, histological etc.) were found to be within safe limits 3 h, 24 h, 48 h and 7 days after the administration. We also determined that the nanoparticles are rapidly sequestered by the liver and spleen, then further biodegraded and directly eliminated in urine without any toxicity effects. Finally, we found that intracellular accumulation of Si-NPs does not induce any oxidative stress damage. Our results evidence a huge potential in using these safe and biodegradable NPs in biomedical applications, in particular as vectors, contrast agents and sensitizers in cancer therapy and diagnostics (theranostics). PMID:27151839

  19. In vivo evaluation of whey protein-based biofilms as scaffolds for cutaneous cell cultures and biomedical applications.

    PubMed

    Rouabhia, Mahmoud; Gilbert, Vanessa; Wang, Hongxum; Subirade, Muriel

    2007-03-01

    This study evaluated the toxicity, biodegradability and immunogenicity of newly developed whey protein-based biofilms for possible use as biomaterials for medical applications. Biofilms were prepared using (A) a whey protein isolate plasticized with either diethylene glycol (DEG) or glycerol (GLY), and (B) beta-lactoglobulin (betaLGA) plasticized with DEG. The biofilms were implanted subcutaneously into Balb/c mice. Analyses were performed at various time points. At 15, 30 and 60 days post-implantation, no necrotic zones or exudates were present at the recipient sites. The biofilms began to degrade as early as 15 days post-implantation, as evidenced by erosion and crumbling. The macroscopic observations were supported by tissue analyses revealing no tissue necrosis or degradation and confirming that the biodegradation of the biofilms began as early as 15 days post-implantation and was almost complete after 60 days. The biodegradation was accompanied by significant leukocyte infiltration at 15 days which significantly decreased at 60 days. The absence of splenomagaly in the implanted mice confirms that these biofilms were not immunogenic. Whey protein-based biofilms are biocompatible and biodegradable and may be of interest for medical applications such as scaffolds for cutaneous cell cultures and skin recovery in burn patients.

  20. Application of FLIM-FIDSAM for the in vivo analysis of hormone competence of different cell types.

    PubMed

    Elgass, Kirstin; Caesar, Katharina; Wanke, Dierk; Harter, Klaus; Meixner, Alfred J; Schleifenbaum, Frank

    2010-11-01

    Background fluorescence derived from subcellular compartments is a major drawback in high-resolution live imaging, especially of plant cells. A novel technique for contrast enhancement of fluorescence images of living cells expressing fluorescent fusion proteins termed fluorescence intensity decay shape analysis microscopy (FIDSAM) has been recently published and is applied here to plant cells expressing wild-type levels of a low-abundant membrane protein (BRI1-EGFP), demonstrating the applicability of FIDSAM to samples exhibiting about 80% autofluorescence. Furthermore, the combination of FIDSAM and fluorescence lifetime imaging microscopy enables the simultaneous determination and quantification of different ligand-specific responses in living cells with high spatial and temporal resolution even in samples with high autofluorescence background. Correlation of different responses can be used to determine the hormone ligand competence of different cell types as demonstrated here in BRI1-EGFP-expressing root and hypocotyl cells.

  1. Application of direct current electric fields to cells and tissues in vitro and modulation of wound electric field in vivo.

    PubMed

    Song, Bing; Gu, Yu; Pu, Jin; Reid, Brian; Zhao, Zhiqiang; Zhao, Min

    2007-01-01

    It has long been known that cells can be induced to migrate by the application of small d.c. electric fields (EFs), a phenomenon referred to as galvanotaxis. We recently reported some significant effects of electric signals of physiological strength in guiding cell migration and wound healing. We present here protocols to apply an EF to cells or tissues cultured in an electrotactic chamber. The chamber can be built to allow controlled medium flow to prevent the potential development of chemical gradients generated by the EFs. It can accommodate cells on planar culture or tissues in 3D gels. Mounted on an inverted microscope, this setup allows close and well-controlled observation of cellular responses to electric signals. As similar EFs are widely present during development and wound healing, this experimental system can be used to simulate and study cellular and molecular responses to electric signals in these events.

  2. Review: in vivo optical spectral tissue sensing-how to go from research to routine clinical application?

    PubMed

    de Boer, Lisanne L; Spliethoff, Jarich W; Sterenborg, Henricus J C M; Ruers, Theo J M

    2017-04-01

    Innovations in optical spectroscopy have helped the technology reach a point where performance previously seen only in laboratory settings can be translated and tested in real-world applications. In the field of oncology, spectral tissue sensing (STS) by means of optical spectroscopy is considered to have major potential for improving diagnostics and optimizing treatment outcome. The concept has been investigated for more than two decades and yet spectral tissue sensing is not commonly employed in routine medical practice. It is therefore important to understand what is needed to translate technological advances and insights generated through basic scientific research in this field into clinical practice. The aim of the discussion presented here is not to provide a comprehensive review of all work published over the last decades but rather to highlight some of the challenges found in literature and encountered by our group in the quest to translate optical technologies into useful clinical tools. Furthermore, an outlook is proposed on how translational researchers could proceed to eventually have STS incorporated in the process of clinical decision-making.

  3. Expanding the Versatility of Mesoporous Silica Nanoparticles towards Drug Delivery for In-vitro, In-vivo and Clinical Applications

    NASA Astrophysics Data System (ADS)

    Ferris, Daniel Patrick

    The work covered in this thesis focuses on research developments in the mesoporous silica nanoparticle platform as a drug delivery vehicle for containment and controlled release of therapeutic agents to inhibit disease. Mesoporous silica is a very versatile material with a very robust structure that is easily modified both internally and externally to change its physical properties. Once modified, the silica nanoparticies can be loaded with therapeutic agents that can be isolated from interacting with their surroundings until an on command delivery signal is received. In this dissertation, first, application of a noninvasive externally controlled means of activation such as light activation and magnetically based heating have been investigated and achieved. Next, by altering the structure of rotaxanes based on azobenzene, steps towards a self-sealing light activated full rotaxane system have been developed. Then, through the manipulation of the particle structure as well as the internal pore environment of silica particle, the interaction between guest drug molecules and the particles has been better understood towards optimizing drug loading and release efficiency. Finally, surface modification of silica nanoparticles with biomolcules has been achieved and observed to increase the efficacy of the silica nanoparticle system in the cellular environment. A combination of all these areas of research results in the advancement of the mesoporous silica nanoparticle drug delivery system towards utilization within living organisms.

  4. Optical imaging in vivo with a focus on paediatric disease: technical progress, current preclinical and clinical applications and future perspectives

    PubMed Central

    Napp, Joanna; Mathejczyk, Julia E.

    2011-01-01

    To obtain information on the occurrence and location of molecular events as well as to track target-specific probes such as antibodies or peptides, drugs or even cells non-invasively over time, optical imaging (OI) technologies are increasingly applied. Although OI strongly contributes to the advances made in preclinical research, it is so far, with the exception of optical coherence tomography (OCT), only very sparingly applied in clinical settings. Nevertheless, as OI technologies evolve and improve continuously and represent relatively inexpensive and harmful methods, their implementation as clinical tools for the assessment of children disease is increasing. This review focuses on the current preclinical and clinical applications as well as on the future potential of OI in the clinical routine. Herein, we summarize the development of different fluorescence and bioluminescence imaging techniques for microscopic and macroscopic visualization of microstructures and biological processes. In addition, we discuss advantages and limitations of optical probes with distinct mechanisms of target-detection as well as of different bioluminescent reporter systems. Particular attention has been given to the use of near-infrared (NIR) fluorescent probes enabling observation of molecular events in deeper tissue. PMID:21221568

  5. In vivo application of short-lag spatial coherence and harmonic spatial coherence imaging in fetal ultrasound.

    PubMed

    Kakkad, Vaibhav; Dahl, Jeremy; Ellestad, Sarah; Trahey, Gregg

    2015-04-01

    Fetal scanning is one of the most common applications of ultrasound imaging and serves as a source of vital information about maternal and fetal health. Visualization of clinically relevant structures, however, can be severely compromised in difficult-to-image patients due to poor resolution and the presence of high levels of acoustical noise or clutter. We have developed novel coherence-based beamforming methods called Short-Lag Spatial Coherence (SLSC) imaging and Harmonic Spatial Coherence imaging (HSCI), and applied them to suppress the effects of clutter in fetal imaging. This method is used to create images of the spatial coherence of the backscattered ultrasound as opposed to images of echo magnitude. We present the results of a patient study to assess the benefits of coherence-based beamforming in the context of first trimester fetal exams. Matched fundamental B-mode, SLSC, harmonic B-mode, and HSCI images were generated using raw radio frequency data collected on 11 volunteers in the first trimester of pregnancy. The images were compared for qualitative differences in image texture and target conspicuity as well as using quantitative imaging metrics such as signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR), and contrast. SLSC and HSCI showed statistically significant improvements across all imaging metrics compared with B-mode and harmonic B-mode, respectively. These improvements were greatest for poor quality B-mode images where contrast of anechoic targets was improved from 15 dB in fundamental B-mode to 27 dB in SLSC and 17 dB in harmonic B-mode to 30 dB in HSCI. CNR improved from 1.4 to 2.5 in the fundamental images and 1.4 to 3.1 in the harmonic case. These results exhibit the potential of coherence-based beamforming to improve image quality and target detectability, especially in high noise environments.

  6. Ex vivo assessment of polyol coated-iron oxide nanoparticles for MRI diagnosis applications: toxicological and MRI contrast enhancement effects

    NASA Astrophysics Data System (ADS)

    Bomati-Miguel, Oscar; Miguel-Sancho, Nuria; Abasolo, Ibane; Candiota, Ana Paula; Roca, Alejandro G.; Acosta, Milena; Schwartz, Simó; Arus, Carles; Marquina, Clara; Martinez, Gema; Santamaria, Jesus

    2014-03-01

    Polyol synthesis is a promising method to obtain directly pharmaceutical grade colloidal dispersion of superparamagnetic iron oxide nanoparticles (SPIONs). Here, we study the biocompatibility and performance as T2-MRI contrast agents (CAs) of high quality magnetic colloidal dispersions (average hydrodynamic aggregate diameter of 16-27 nm) consisting of polyol-synthesized SPIONs (5 nm in mean particle size) coated with triethylene glycol (TEG) chains (TEG-SPIONs), which were subsequently functionalized to carboxyl-terminated meso-2-3-dimercaptosuccinic acid (DMSA) coated-iron oxide nanoparticles (DMSA-SPIONs). Standard MTT assays on HeLa, U87MG, and HepG2 cells revealed that colloidal dispersions of TEG-coated iron oxide nanoparticles did not induce any loss of cell viability after 3 days incubation with dose concentrations below 50 μg Fe/ml. However, after these nanoparticles were functionalized with DMSA molecules, an increase on their cytotoxicity was observed, so that particles bearing free terminal carboxyl groups on their surface were not cytotoxic only at low concentrations (<10 μg Fe/ml). Moreover, cell uptake assays on HeLa and U87MG and hemolysis tests have demonstrated that TEG-SPIONs and DMSA-SPIONs were well internalized by the cells and did not induce any adverse effect on the red blood cells at the tested concentrations. Finally, in vitro relaxivity measurements and post mortem MRI studies in mice indicated that both types of coated-iron oxide nanoparticles produced higher negative T2-MRI contrast enhancement than that measured for a similar commercial T2-MRI CAs consisting in dextran-coated ultra-small iron oxide nanoparticles (Ferumoxtran-10). In conclusion, the above attributes make both types of as synthesized coated-iron oxide nanoparticles, but especially DMSA-SPIONs, promising candidates as T2-MRI CAs for nanoparticle-enhanced MRI diagnosis applications.

  7. Time-resolved singlet oxygen luminescence detection under photodynamic therapy relevant conditions: comparison of ex vivo application of two photosensitizer formulations

    NASA Astrophysics Data System (ADS)

    Schlothauer, Jan C.; Hackbarth, Steffen; Jäger, Lutz; Drobniewski, Kai; Patel, Hemantbhai; Gorun, Sergiu M.; Röder, Beate

    2012-11-01

    Singlet oxygen plays a crucial role in photo-dermatology and photodynamic therapy (PDT) of cancer. Its direct observation by measuring the phosphorescence at 1270 nm, however, is still challenging due to the very low emission probability. It is especially challenging for the time-resolved detection of singlet oxygen kinetics in vivo which is of special interest for biomedical applications. Photosensitized generation of singlet oxygen, in pig ear skin as model for human skin, is investigated here. Two photosensitizers (PS) were topically applied to the pig ear skin and examined in a comparative study, which include the amphiphilic pheophorbide-a and the highly hydrophobic perfluoroalkylated zinc phthalocyanine (F64PcZn). Fluorescence microscopy indicates the exclusive accumulation of pheophorbide-a in the stratum corneum, while F64PcZn can also accumulate in deeper layers of the epidermis of the pig ear skin. The kinetics obtained with phosphorescence measurements show the singlet oxygen interaction with the PS microenvironment. Different generation sites of singlet oxygen correlate with the luminescence kinetics. The results show that singlet oxygen luminescence detection can be used as a diagnostic tool, not only for research, but also during treatment. The detection methodology is suitable for the monitoring of chemical quenchers' oxidation as well as O2 saturation at singlet oxygen concentration levels relevant to PDT treatment protocols.

  8. In vivo assessments of bioabsorbable AZ91 magnesium implants coated with nanostructured fluoridated hydroxyapatite by MAO/EPD technique for biomedical applications.

    PubMed

    Razavi, Mehdi; Fathi, Mohammadhossein; Savabi, Omid; Vashaee, Daryoosh; Tayebi, Lobat

    2015-03-01

    Although magnesium (Mg) is a unique biodegradable metal which possesses mechanical property similar to that of the natural bone and can be an attractive material to be used as orthopedic implants, its quick corrosion rate restricts its actual clinical applications. To control its rapid degradation, we have modified the surface of magnesium implant using fluoridated hydroxyapatite (FHA: Ca10(PO4)6OH2-xFx) through the combined micro-arc oxidation (MAO) and electrophoretic deposition (EPD) techniques, which was presented in our previous paper. In this article, the biocompatibility examinations were conducted on the coated AZ91 magnesium alloy by implanting it into the greater trochanter area of rabbits. The results of the in vivo animal test revealed a significant enhancement in the biocompatibility of FHA/MAO coated implant compared to the uncoated one. By applying the FHA/MAO coating on the AZ91 implant, the amount of weight loss and magnesium ion release in blood plasma decreased. According to the histological results, the formation of the new bone increased and the inflammation decreased around the implant. In addition, the implantation of the uncoated AZ91 alloy accompanied by the release of hydrogen gas around the implant; this release was suppressed by applying the coated implant. Our study exemplifies that the surface coating of magnesium implant using a bioactive ceramic such as fluoridated hydroxyapatite may improve the biocompatibility of the implant to make it suitable as a commercialized biomedical product.

  9. Resistance after Chronic Application of the HDAC-Inhibitor Valproic Acid Is Associated with Elevated Akt Activation in Renal Cell Carcinoma In Vivo

    PubMed Central

    Juengel, Eva; Makarević, Jasmina; Tsaur, Igor; Bartsch, Georg; Nelson, Karen; Haferkamp, Axel; Blaheta, Roman A.

    2013-01-01

    Targeted drugs have significantly improved the therapeutic options for advanced renal cell carcinoma (RCC). However, resistance often develops, negating the benefit of these agents. In the present study, the molecular mechanisms of acquired resistance towards the histone deacetylase (HDAC) inhibitor valproic acid (VPA) in a RCC in vivo model were investigated. NMRI:nu/nu mice were transplanted with Caki-1 RCC cells and then treated with VPA (200 mg/kg/day). Controls remained untreated. Based on tumor growth dynamics, the mice were divided into “responders” and “non-responders” to VPA. Histone H3 and H4 acetylation and expression of cell signaling and cell cycle regulating proteins in the RCC mouse tumors were evaluated by Western blotting. Tumor growth of VPA responders was significantly diminished, whereas that of VPA non-responders even exceeded control values. Cdk1, 2 and 4 proteins were strongly enhanced in the non-responders. Importantly, Akt expression and activity were massively up-regulated in the tumors of the VPA non-responders. Chronic application (12 weeks) of VPA to Caki-1 cells in vitro evoked a distinct elevation of Akt activity and cancer cells no longer responded with cell growth reduction, compared to the short 2 week treatment. We assume that chronic use of an HDAC-inhibitor is associated with (re)-activation of Akt, which may be involved in resistance development. Consequently, combined blockade of both HDAC and Akt may delay or prevent drug resistance in RCC. PMID:23372654

  10. Time-resolved singlet oxygen luminescence detection under photodynamic therapy relevant conditions: comparison of ex vivo application of two photosensitizer formulations.

    PubMed

    Schlothauer, Jan C; Hackbarth, Steffen; Jäger, Lutz; Drobniewski, Kai; Patel, Hemantbhai; Gorun, Sergiu M; Röder, Beate

    2012-11-01

    Singlet oxygen plays a crucial role in photo-dermatology and photodynamic therapy (PDT) of cancer. Its direct observation by measuring the phosphorescence at 1270 nm, however, is still challenging due to the very low emission probability. It is especially challenging for the time-resolved detection of singlet oxygen kinetics in vivo which is of special interest for biomedical applications. Photosensitized generation of singlet oxygen, in pig ear skin as model for human skin, is investigated here. Two photosensitizers (PS) were topically applied to the pig ear skin and examined in a comparative study, which include the amphiphilic pheophorbide-a and the highly hydrophobic perfluoroalkylated zinc phthalocyanine (F64PcZn). Fluorescence microscopy indicates the exclusive accumulation of pheophorbide-a in the stratum corneum, while F64PcZn can also accumulate in deeper layers of the epidermis of the pig ear skin. The kinetics obtained with phosphorescence measurements show the singlet oxygen interaction with the PS microenvironment. Different generation sites of singlet oxygen correlate with the luminescence kinetics. The results show that singlet oxygen luminescence detection can be used as a diagnostic tool, not only for research, but also during treatment. The detection methodology is suitable for the monitoring of chemical quenchers’ oxidation as well as saturation at singlet oxygen concentration levels relevant to PDT treatment protocols.

  11. Hydrogen sulfide activates the carotid body chemoreceptors in cat, rabbit and rat ex vivo preparations.

    PubMed

    Jiao, Yingfu; Li, Qian; Sun, Biying; Zhang, Guohua; Rong, Weifang

    2015-03-01

    We and others previously reported experimental evidence suggesting an important role for hydrogen sulfide (H2S) in oxygen sensing in murine carotid body chemoreceptors. More recent data implicated abnormal H2S-mediated chemoreceptor signaling in pathological conditions such as chronic heart failure and hypertension. However, the idea of H2S as a mediator of oxygen-sensing in chemoreceptors has been challenged. In particular, it was shown that exogenous H2S inhibited the release of neurotransmitters (ACh and ATP) from the cat carotid body, raising the possibility that there exists significant species difference in H2S-mediated signaling in chemoreceptors. This study was designed specifically to determine the effect of H2S on chemoreceptors in different species. We conducted multiunit extracellular recordings of the sinus nerve in the ex vivo carotid body preparation taken from the rat, the cat and the rabbit. As observed in the mouse carotid body, H2S donors (NaHS or Na2S) evoked qualitatively similar excitatory responses of the afferent sinus nerves of the species studied here. The excitatory effects of the H2S donors were concentration-dependent and reversible. The sinus nerve responses to H2S donors were prevented by blockade of the transmission between type I cells and the afferent terminals, as was the response to hypoxia. These results demonstrate that exogenous H2S exerts qualitatively similar excitatory effects on chemoreceptor afferents of different species. The role of endogenous H2S-mediated signaling in carotid body function in different species awaits further investigation.

  12. Relaxation-compensated CEST-MRI at 7 T for mapping of creatine content and pH--preliminary application in human muscle tissue in vivo.

    PubMed

    Rerich, Eugenia; Zaiss, Moritz; Korzowski, Andreas; Ladd, Mark E; Bachert, Peter

    2015-11-01

    The small biomolecule creatine is involved in energy metabolism. Mapping of the total creatine (mostly PCr and Cr) in vivo has been done with chemical shift imaging. Chemical exchange saturation transfer (CEST) allows an alternative detection of creatine via water MRI. Living tissue exhibits CEST effects from different small metabolites, including creatine, with four exchanging protons of its guanidinium group resonating about 2 ppm from the water peak and hence contributing to the amine proton CEST peak. The intermediate exchange rate (≈ 1000 Hz) of the guanidinium protons requires high RF saturation amplitude B1. However, strong B1 fields also label semi-solid magnetization transfer (MT) effects originating from immobile protons with broad linewidths (~kHz) in the tissue. Recently, it was shown that endogenous CEST contrasts are strongly affected by the MT background as well as by T1 relaxation of the water protons. We show that this influence can be corrected in the acquired CEST data by an inverse metric that yields the apparent exchange-dependent relaxation (AREX). AREX has some useful linearity features that enable preparation of both concentration, and--by using the AREX-ratio of two RF irradiation amplitudes B1--purely exchange-rate-weighted CEST contrasts. These two methods could be verified in phantom experiments with different concentration and pH values, but also varying water relaxation properties. Finally, results from a preliminary application to in vivo CEST imaging data of the human calf muscle before and after exercise are presented. The creatine concentration increases during exercise as expected and as confirmed by (31)P NMR spectroscopic imaging. However, the estimated concentrations obtained by our method were higher than the literature values: cCr,rest=24.5±3.74mM to cCr,ex=38.32±13.05mM. The CEST-based pH method shows a pH decrease during exercise, whereas a slight increase was observed by (31)P NMR spectroscopy.

  13. Effects of fluoride-ion-implanted titanium surface on the cytocompatibility in vitro and osseointegatation in vivo for dental implant applications.

    PubMed

    Wang, Xue-jin; Liu, Hui-ying; Ren, Xiang; Sun, Hui-yan; Zhu, Li-ying; Ying, Xiao-xia; Hu, Shu-hai; Qiu, Ze-wen; Wang, Lang-ping; Wang, Xiao-feng; Ma, Guo-wu

    2015-12-01

    As an attractive technique for the improvement of biomaterials, Plasma immersion ion implantation (PIII) has been applied to modifying the titanium material for dental implant application. The present study investigated the cytocompatibility and early osseointegration of fluoride-ion-implanted titanium (F-Ti) surface and implants, both characterizing in their composition of titanium oxide and titanium fluoride. The cytocompatibility of F-Ti was evaluated in vitro by using scanning electron microscope, Cell Counting Kit-8 assay, alkaline phosphatase activity assay, and quantitative real-time polymerase chain reaction. The results showed that the F-Ti weakened the effects that Porphyromonas gingivalis exerted on the MG-63 cells in terms of morphology, proliferation, differentiation, and genetic expression when MG-63 cells and Porphyromonas gingivalis were co-cultured on the surface of F-Ti. Meanwhile, the osteogenic activity of F-Ti implants was assessed in vivo via evaluating the histological morphology and estimating histomorphometric parameters. The analysis of toluidine blue staining indicated that the new bone was more mature in subjects with F-Ti group, which exhibited the Haversian system, and the mean bone-implant contact value of F-Ti group was slightly higher than that of cp-Ti group (p>0.05). Fluorescence bands were wider and brighter in the F-Ti group, and the intensity of fluorochromes deposited at the sites of mineralized bone formation was significantly higher for F-Ti surfaces than for cp-Ti surfaces, within the 2nd, 3rd and 4th weeks (p<0.05). An indication is that the fluoride modified titanium can promote cytocompatibility and early osseointegration, thus providing a promising alternative for clinical use.

  14. Comparison of Diffusion MRI Acquisition Protocols for the In Vivo Characterization of the Mouse Spinal Cord: Variability Analysis and Application to an Amyotrophic Lateral Sclerosis Model

    PubMed Central

    Marcuzzo, Stefania; Bonanno, Silvia; Padelli, Francesco; Moreno-Manzano, Victoria; García-Verdugo, José Manuel; Bernasconi, Pia; Mantegazza, Renato; Bruzzone, Maria Grazia; Zucca, Ileana

    2016-01-01

    Diffusion-weighted Magnetic Resonance Imaging (dMRI) has relevant applications in the microstructural characterization of the spinal cord, especially in neurodegenerative diseases. Animal models have a pivotal role in the study of such diseases; however, in vivo spinal dMRI of small animals entails additional challenges that require a systematical investigation of acquisition parameters. The purpose of this study is to compare three acquisition protocols and identify the scanning parameters allowing a robust estimation of the main diffusion quantities and a good sensitivity to neurodegeneration in the mouse spinal cord. For all the protocols, the signal-to-noise and contrast-to noise ratios and the mean value and variability of Diffusion Tensor metrics were evaluated in healthy controls. For the estimation of fractional anisotropy less variability was provided by protocols with more diffusion directions, for the estimation of mean, axial and radial diffusivity by protocols with fewer diffusion directions and higher diffusion weighting. Intermediate features (12 directions, b = 1200 s/mm2) provided the overall minimum inter- and intra-subject variability in most cases. In order to test the diagnostic sensitivity of the protocols, 7 G93A-SOD1 mice (model of amyotrophic lateral sclerosis) at 10 and 17 weeks of age were scanned and the derived diffusion parameters compared with those estimated in age-matched healthy animals. The protocols with an intermediate or high number of diffusion directions provided the best differentiation between the two groups at week 17, whereas only few local significant differences were highlighted at week 10. According to our results, a dMRI protocol with an intermediate number of diffusion gradient directions and a relatively high diffusion weighting is optimal for spinal cord imaging. Further work is needed to confirm these results and for a finer tuning of acquisition parameters. Nevertheless, our findings could be important for the

  15. ITRAQ MASS SPECTROMETRIC PROTEOMIC APPLICATIONS FOR IN VIVO TOXICOLOGY STUDIES OF AMPHIBIAN SPECIES: DATA HANDLING AND INTERPRETATION USING PEPTIDE-TAGGING SOFTWARE

    EPA Science Inventory

    This addresses the USEPA's need for a cost effective, non-mammalian screening assay for thyroid axis disrupting chemicals; a multi-endpoint strategy combining molecular and in vivo protocols in an amphibian model is being applied at MED Duluth.

  16. Ex vivo lung perfusion

    PubMed Central

    Machuca, Tiago N.

    2014-01-01

    Lung transplantation (LTx) is an established treatment option for eligible patients with end-stage lung disease. Nevertheless, the imbalance between suitable donor lungs available and the increasing number of patients considered for LTx reflects in considerable waitlist mortality. Among potential alternatives to address this issue, ex vivo lung perfusion (EVLP) has emerged as a modern preservation technique that allows for more accurate lung assessment and also improvement of lung function. Its application in high-risk donor lungs has been successful and resulted in safe expansion of the donor pool. This article will: (I) review the technical details of EVLP; (II) the rationale behind the method; (III) report the worldwide clinical experience with the EVLP, including the Toronto technique and others; (IV) finally, discuss the growing literature on EVLP application for donation after cardiac death (DCD) lungs. PMID:25132972

  17. Application of biorelevant dissolution tests to the prediction of in vivo performance of diclofenac sodium from an oral modified-release pellet dosage form.

    PubMed

    Jantratid, Ekarat; De Maio, Vincenzo; Ronda, Emanuela; Mattavelli, Valentina; Vertzoni, Maria; Dressman, Jennifer B

    2009-06-28

    In vitro biorelevant dissolution tests enabling the prediction of in vivo performance of an oral modified-release (MR) dosage form were developed in this study. In vitro dissolution of MR diclofenac sodium pellets containing 100mg active ingredient was evaluated under simulated pre- and postprandial conditions using USP Apparatus 3 (reciprocating cylinder, Bio-Dis) and 4 (flow-through cell) and results compared with compendial methods using USP Apparatus 1 (basket) and 2 (paddle). In vivo, the effects of food on the absorption of diclofenac sodium from the pellet dosage form were investigated by administering the product to 16 healthy volunteers pre- and postprandially in a crossover-design study. The in vitro results were compared with the in vivo data by means of Level A in vitro-in vivo correlation (IVIVC) and Weibull distribution analysis. The compendial dissolution tests were not able to predict food effects. The biorelevant dissolution tests predicted correctly that the release (and hence absorption) of diclofenac sodium would be slower in the fed state than in the fasted state. No significant differences in extent of absorption due to changes in extent of release were predicted or observed. The results demonstrate good correlations between in vitro drug release and in vivo drug absorption in both pre- and postprandial states using the biorelevant dissolution test methods.

  18. Development of a biphasic dissolution test for Deferasirox dispersible tablets and its application in establishing an in vitro-in vivo correlation.

    PubMed

    Al Durdunji, Amal; AlKhatib, Hatim S; Al-Ghazawi, Mutasim

    2016-05-01

    In a biphasic dissolution medium, the integration of the in vitro dissolution of a drug in an aqueous phase and its subsequent partitioning into an organic phase is hypothesized to simulate the in vivo drug absorption. Such a methodology is expected to improve the probability of achieving a successful in vitro-in vivo correlation. Dissolution of Dispersible tablets of Deferasirox, a biopharmaceutics classification system type II compound, was studied in a biphasic dissolution medium using a flow-through dissolution apparatus coupled to a paddle apparatus. The experimental parameters associated with dissolution were optimized to discriminate between Deferasirox dispersible tablets of different formulations. The dissolution profiles obtained from this system were subsequently used to construct a level A in vitro-in vivo correlation.

  19. Development and in vivo testing of a high frequency endoscopic Raman spectroscopy system for potential applications in the detection of early colonic neoplasia.

    PubMed

    Short, Michael A; Wang, Wenbo; Tai, Isabella T; Zeng, Haishan

    2016-01-01

    The objective of this study was to build and test an adjunct system to a colonoscope for in vivo measurement of Raman spectra from colon tissue for potentially improving the detection of early cancers. The novelty of this system was that low cost fibre optic probes were used, without the addition of expensive optical filters. Good quality in vivo Raman spectra were successfully obtained with a 1 s integration time in the high frequency (HF) range from normal tissue and polyps of patients during a colonoscopy. The polyps were subsequently removed, and their pathology determined. The acquired in vivo Raman spectra showed clear changes between tissue with normal and tubular adenoma pathology. Further clinical study with this low cost HF Raman probe is warranted to fully test its clinical utility.

  20. Clinical applications of a real-time scanning-slit confocal microscope designed for real-time observations of the in-vivo human cornea

    NASA Astrophysics Data System (ADS)

    Masters, Barry R.

    1995-05-01

    We describe a new, real-time, flying slit confocal microscope, that has unique features and imaging characteristics for in vivo human ocular imaging. In vivo real-time confocal microscopy is currently used to investigate the tear film, renewal of the ocular surface, the role of epithelial innervation in epithelial cell proliferation, wound healing, kinetics of drug penetration, the effects of laser refractive surgery on the keratocyte activation and distribution in the stroma, and the nature of endothelial defects. The following clinical examples will be presented and discussed: confocal microscopy of normal human basal and wing cells in the epithelium, confocal microscopy of lamellar and penetrating corneal grafts, confocal microscopy of corneal ulcer, confocal microscopy of scar formation after herpes keratitis, and confocal microscopy of corneal innervation. The use of scanning slit confocal microscopes has unique advantages over other instrumental systems based on pinhole-containing Nipkow disks (tandem-scanning confocal microscopes) for clinical in vivo confocal microscopy.

  1. Feasibility and application of a retronasal aroma-trapping device to study in vivo aroma release during the consumption of model wine-derived beverages

    PubMed Central

    Muñoz-González, Carolina; Rodríguez-Bencomo, Juan José; Moreno-Arribas, Maria Victoria; Pozo-Bayón, Maria Ángeles

    2014-01-01

    New types of wine-derived beverages are now in the market. However, little is known about the impact of ingredient formulation on aroma release during consumption, which is directly linked to consumer preferences and liking. In this study, the optimization and validation of a retronasal aroma-trapping device (RATD) for the in vivo monitoring of aroma release was carried out. This device was applied to assess the impact of two main ingredients (sugar and ethanol) in these types of beverages on in vivo aroma release. Two aroma-trapping materials (Lichrolut and Tenax) were firstly assayed. Tenax provided higher recovery and lower intra- and inter-trap variability. In in vivo conditions, RATD provided an adequate linear range (R2 > 0.91) between 0 and 50 mg L−1 of aroma compounds. Differences in the total aroma release were observed in equally trained panelists. It was proven that the addition of sugar (up to 150 mg kg−1) did not have effect on aroma release, while ethanol (up to 40 mg L−1) enhanced the aroma release during drinking. The RATD is a useful tool to collect real in vivo data to extract reliable conclusions about the effect of beverage components on aroma release during consumption. The concentration of ethanol should be taken into consideration for the formulation of wine-derived beverages. PMID:25473493

  2. Feasibility and application of a retronasal aroma-trapping device to study in vivo aroma release during the consumption of model wine-derived beverages.

    PubMed

    Muñoz-González, Carolina; Rodríguez-Bencomo, Juan José; Moreno-Arribas, Maria Victoria; Pozo-Bayón, Maria Ángeles

    2014-07-01

    New types of wine-derived beverages are now in the market. However, little is known about the impact of ingredient formulation on aroma release during consumption, which is directly linked to consumer preferences and liking. In this study, the optimization and validation of a retronasal aroma-trapping device (RATD) for the in vivo monitoring of aroma release was carried out. This device was applied to assess the impact of two main ingredients (sugar and ethanol) in these types of beverages on in vivo aroma release. Two aroma-trapping materials (Lichrolut and Tenax) were firstly assayed. Tenax provided higher recovery and lower intra- and inter-trap variability. In in vivo conditions, RATD provided an adequate linear range (R (2) > 0.91) between 0 and 50 mg L(-1) of aroma compounds. Differences in the total aroma release were observed in equally trained panelists. It was proven that the addition of sugar (up to 150 mg kg(-1)) did not have effect on aroma release, while ethanol (up to 40 mg L(-1)) enhanced the aroma release during drinking. The RATD is a useful tool to collect real in vivo data to extract reliable conclusions about the effect of beverage components on aroma release during consumption. The concentration of ethanol should be taken into consideration for the formulation of wine-derived beverages.

  3. Application of an in vitro DDASS to evaluate oral absorption of two chemicals simultaneously: establishment of a level A in vitro-in vivo correlation.

    PubMed

    Hou, Jipeng; He, Xin; Xu, Xuefang; Shi, Xiaoyan; Xu, Yanyan; Liu, Changxiao

    2012-11-01

    The aim of this study was to evaluate the oral absorption of two chemicals simultaneously using a drug dissolution/absorption simulating system (DDASS), and to establish a correlation between DDASS and in vivo absorption to clarify the prediction of this in vitro model. Ferulic acid (FA) and tetrahydropalmatine (THP), the components of Angelicae Sinensis Radix and Corydalis Yanhusuo Rhizoma, respectively, were chosen as model compounds. Three groups including FA, THP, and FA and THP together (FA + THP) were studied in DDASS. The corresponding in vivo pharmacokinetics study was performed in rats. Then the correlation was analysed between DDASS permeation in vitro and rat absorption data in vivo. A strong level A correlation (r > 0.84) was obtained after a correlation coefficient test (p < 0.05 or 0.01). Moreover, when FA and THP were used together in DDASS, the cumulative permeation of FA increased by 38.5%, while THP permeation decreased by 25.8%. In rats, the area under the concentration-time curve from time to infinity for FA increased 2.6-fold, while THP decreased 19.6%. The changes in rat intestinal permeation modeled by the DDASS were consistent with the absorption changes in rats. We conclude that DDASS is a valid in vitro model to evaluate oral absorption of two drug components simultaneously and reflect the in vivo characteristics of drug absorption accurately.

  4. Microsomal and cytosolic scaling factors in dog and human kidney cortex and application for in vitro-in vivo extrapolation of renal metabolic clearance.

    PubMed

    Scotcher, Daniel; Billington, Sarah; Brown, Jay; Jones, Christopher; Brown, Colin D A; Rostami-Hodjegan, Amin; Galetin, Aleksandra

    2017-03-07

    In vitro-in vivo extrapolation of drug metabolism data obtained in enriched preparations of sub-cellular fractions rely upon robust estimates of physiologically relevant scaling factors for prediction of clearance in vivo. The purpose of the current study was to measure the microsomal and cytosolic protein per gram of kidney (MPPGK and CPPGK) in dog and human kidney cortex using appropriate protein recovery markers, and evaluate functional activity of human cortex microsomes. Cytochrome P450 (CYP) content and glucose-6-phosphatase activity were used as microsomal protein markers, whereas glutathione-S-transferase activity was a cytosolic marker. Functional activity of human microsomal samples was assessed by measuring mycophenolic acid glucuronidation. MPPGK was 33.9 and 44.0 mg/g dog kidney cortex, and 41.1 and 63.6 mg/g dog liver (n=17), using CYP content and glucose-6-phosphatase activity, respectively. There were no trends between kidney, liver and intestinal scalars from the same animals. Species differences were evident, as human MPPGK and CPPGK were 26.2 and 53.3 mg/g kidney cortex (n=38), respectively. MPPGK was 2-fold higher than the commonly used in vitro-in vivo extrapolation scalar; difference was mainly attributed to tissue source (mixed kidney regions vs cortex). Robust human MPPGK and CPPGK scalars were measured for the first time. The work emphasized the importance of regional differences (cortex vs. whole kidney specific MPPGK, tissue weight and blood flow) and a need to account for these to improve assessment of renal metabolic clearance and its extrapolation to in vivo.

  5. Oxygen sensing characteristics of individual ZnO nanowire transistors

    SciTech Connect

    Li, Q.H.; Liang, Y.X.; Wan, Q.; Wang, T.H.

    2004-12-27

    Individual ZnO nanowire transistors are fabricated, and their sensing properties are investigated. The transistors show a carrier density of 2300 {mu}m{sup -1} and mobility up to 6.4 cm{sup 2}/V s, which are obtained from the I{sub SD}-V{sub G} curves. The threshold voltage shifts in the positive direction and the source-drain current decreases as ambient oxygen concentration increases. However, the opposite occurs when the transistors are under illumination. Surface adsorbates on the ZnO nanowires affect both the mobility and the carrier density. Our data are helpful in understanding the sensing mechanism of the gas sensors.

  6. A Pyrene@Micelle Sensor for Fluorescent Oxygen Sensing

    PubMed Central

    Yuan, Yan-xia; Peng, Hong-shang; Ping, Jian-tao; Wang, Xiao-hui; You, Fang-tian

    2015-01-01

    For most fluorescent oxygen sensors developed today, their fabrication process is either time-consuming or needs specialized knowledge. In this work, a robust fluorescent oxygen sensor is facilely constructed by dissolving pyrene molecules into CTAB aqueous solution. The as-prepared pyrene@micelle sensors have submicron-sized diameter, and the concentration of utilized pyrene can be reduced as low as 0.8 mM but still can exhibit dominant excimer emission. The excimer fluorescence is sensitive to dissolved oxygen in both intensity and lifetime, and the respective Stern-Volmer plot follows a nonlinear behavior justified by a two-site model. Because of the merits of large Stokes shift (~140 nm), easy fabrication, and robustness, the pyrene@micelle sensors are very attractive for practical determination of oxygen. PMID:26539471

  7. Oxygen-sensing by arterial chemoreceptors: Mechanisms and medical translation.

    PubMed

    López-Barneo, José; Ortega-Sáenz, Patricia; González-Rodríguez, Patricia; Fernández-Agüera, M Carmen; Macías, David; Pardal, Ricardo; Gao, Lin

    2016-01-01

    Acute O2 sensing is necessary for the activation of cardiorespiratory reflexes (hyperventilation and sympathetic activation), which permit the survival of individuals under hypoxic environments (e.g. high altitude) or medical conditions presenting with reduced capacity for gas exchange between the lung alveoli and the blood. Changes in blood O2 tension are detected by the arterial chemoreceptors, in particular the carotid body (CB), which act in concert with the adrenal medulla (AM) to facilitate rapid adaptations to hypoxia. The field of arterial chemoreception has undergone a considerable expansion in recent years, with many of the fundamental observations made at the molecular and cellular levels serving to improve our understanding of the pathogenesis of numerous medical disorders, and even to propose advances in the treatment strategies. In this review, after a short historical preface, we describe the current model of chemosensory transduction based on the modulation of membrane K(+) channels by O2 in specialized chemoreceptor cells. Recent progress in elucidating the molecular mechanisms underlying the modulation of ion channels by O2 tension, which involves mitochondrial complex I, is also discussed. The discovery in the last few years of a specific population of neural crest-derived stem cells in the CB explains the reversible growth of this organ, an intriguing and unusual property of this type of neuronal tissue that contributes to acclimatization under chronic hypoxia. The essential homeostatic role of the CB-AM axis is clearly evident in newly generated mouse models that reach adulthood, albeit with CB and AM atrophy. These animals exhibit a marked intolerance to even mild hypoxia. CB inhibition or over-activation can have important medical consequences. Respiratory depression by general anesthetics or by opioid use is a common clinical condition that frequently causes death in susceptible individuals. An exaggerated sympathetic outflow due to over-activation of the CB-AM axis may contribute to the pathogenesis of several highly prevalent medical conditions, such as chronic heart failure, obstructive sleep apnea, obesity, metabolic syndrome, and diabetes. A detailed understanding of the molecular mechanisms underlying acute O2 sensing may help in the design of more efficient therapeutic approaches to combat these disorders.

  8. Carotid body oxygen sensing and adaptation to hypoxia.

    PubMed

    López-Barneo, José; Macías, David; Platero-Luengo, Aida; Ortega-Sáenz, Patricia; Pardal, Ricardo

    2016-01-01

    The carotid body (CB) is the principal arterial chemoreceptor that mediates the hyperventilatory response to hypoxia. Our understanding of CB function and its role in disease mechanisms has progressed considerably in the last decades, particularly in recent years. The sensory elements of the CB are the neuron-like glomus cells, which contain numerous transmitters and form synapses with afferent sensory fibers. The activation of glomus cells under hypoxia mainly depends on the modulation of O2-sensitive K(+) channels which leads to cell depolarization and the opening of Ca(2+) channels. This model of sensory transduction operates in all mammalian species studied thus far, including man. However, the molecular mechanisms underlying the modulation of ion channel function by changes in the O2 level are as yet unknown. The CB plays a fundamental role in acclimatization to sustained hypoxia. Mice with CB atrophy or patients who have undergone CB resection due to surgical treatments show a marked intolerance to even mild hypoxia. CB growth under hypoxia is supported by the existence of a resident population of neural crest-derived stem cells of glia-like phenotype. These stem cells are not highly affected by exposure to low O2 tension; however, there are abundant synapse-like contacts between the glomus cells and stem cells (chemoproliferative synapses), which may be needed to trigger progenitor cell proliferation and differentiation under hypoxia. CB hypo- or hyper-activation may also contribute to the pathogenesis of several prevalent human diseases.

  9. Evaluation of the in vivo and ex vivo optical properties in a mouse ear model

    NASA Astrophysics Data System (ADS)

    Salomatina, E.; Yaroslavsky, A. N.

    2008-06-01

    Determination of in vivo optical properties is a challenging problem. Absorption and scattering measured ex vivo are often used for in vivo applications. To investigate the validity of this approach, we have obtained and compared the optical properties of mouse ears in vivo and ex vivo in the spectral range from 370 to 1650 nm. Integrating sphere spectrophotometry in combination with the inverse Monte Carlo technique was employed to determine absorption coefficients, μa, scattering coefficients, μs, and anisotropy factors, g. Two groups of mice were used for the study. The first group was measured in vivo and ex vivo within 5-10 min post mortem. The second group was measured in vivo and ex vivo every 24 h for up to 72 h after sacrifice. Between the measurements the tissues were kept at 4 °C wrapped in a gauze moistened with saline solution. Then the specimens were frozen at -25 °C for 40 min, thawed and measured again. The results indicate that the absorption coefficients determined in vivo and ex vivo within 5-10 min post mortem differed considerably only in the spectral range dominated by hemoglobin. These changes can be attributed to rapid deoxygenation of tissue and blood post mortem. Absorption coefficients determined ex vivo up to 72 h post mortem decreased gradually with time in the spectral regions dominated by hemoglobin and water, which can be explained by the continuing loss of blood. Absorption properties of the frozen-thawed ex vivo tissues showed increase in oxygenation, which is likely caused by the release of hemoglobin from hemolyzed erythrocytes. Scattering of the ex vivo tissues decreased gradually with time in the entire spectral range due to the continuing loss of blood and partial cell damage. Anisotropy factors did not change considerably.

  10. Motion-artifact-robust, polarization-resolved second-harmonic-generation microscopy based on rapid polarization switching with electro-optic Pockells cell and its application to in vivo visualization of collagen fiber orientation in human facial skin.

    PubMed

    Tanaka, Yuji; Hase, Eiji; Fukushima, Shuichiro; Ogura, Yuki; Yamashita, Toyonobu; Hirao, Tetsuji; Araki, Tsutomu; Yasui, Takeshi

    2014-04-01

    Polarization-resolved second-harmonic-generation (PR-SHG) microscopy is a powerful tool for investigating collagen fiber orientation quantitatively with low invasiveness. However, the waiting time for the mechanical polarization rotation makes it too sensitive to motion artifacts and hence has hampered its use in various applications in vivo. In the work described in this article, we constructed a motion-artifact-robust, PR-SHG microscope based on rapid polarization switching at every pixel with an electro-optic Pockells cell (PC) in synchronization with step-wise raster scanning of the focus spot and alternate data acquisition of a vertical-polarization-resolved SHG signal and a horizontal-polarization-resolved one. The constructed PC-based PR-SHG microscope enabled us to visualize orientation mapping of dermal collagen fiber in human facial skin in vivo without the influence of motion artifacts. Furthermore, it implied the location and/or age dependence of the collagen fiber orientation in human facial skin. The robustness to motion artifacts in the collagen orientation measurement will expand the application scope of SHG microscopy in dermatology and collagen-related fields.

  11. Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications.

    PubMed

    Spronken, Monique I; Short, Kirsty R; Herfst, Sander; Bestebroer, Theo M; Vaes, Vincent P; van der Hoeven, Barbara; Koster, Abraham J; Kremers, Gert-Jan; Scott, Dana P; Gultyaev, Alexander P; Sorell, Erin M; de Graaf, Miranda; Bárcena, Montserrat; Rimmelzwaan, Guus F; Fouchier, Ron A

    2015-01-01

    Bioluminescent and fluorescent influenza A viruses offer new opportunities to study influenza virus replication, tropism and pathogenesis. To date, several influenza A reporter viruses have been described. These strategies typically focused on a single reporter gene (either bioluminescent or fluorescent) in a single virus backbone. However, whilst bioluminescence is suited to in vivo imaging, fluorescent viruses are more appropriate for microscopy. Therefore, the idea l reporter virus varies depending on the experiment in question, and it is important that any reporter virus strategy can be adapted accordingly. Herein, a strategy was developed to create five different reporter viruses in a single virus backbone. Specifically, enhanced green fluorescent protein (eGFP), far-red fluorescent protein (fRFP), near-infrared fluorescent protein (iRFP), Gaussia luciferase (gLUC) and firefly luciferase (fLUC) were inserted into the PA gene segment of A/PR/8/34 (H1N1). This study provides a comprehensive characterisation of the effects of different reporter genes on influenza virus replication and reporter activity. In vivo reporter gene expression, in lung tissues, was only detected for eGFP, fRFP and gLUC expressing viruses. In vitro, the eGFP-expressing virus displayed the best reporter stability and could be used for correlative light electron microscopy (CLEM). This strategy was then used to create eGFP-expressing viruses consisting entirely of pandemic H1N1, highly pathogenic avian influenza (HPAI) H5N1 and H7N9. The HPAI H5N1 eGFP-expressing virus infected mice and reporter gene expression was detected, in lung tissues, in vivo. Thus, this study provides new tools and insights for the creation of bioluminescent and fluorescent influenza A reporter viruses.

  12. Endothelial progenitor and mesenchymal stem cell-derived cells persist in tissue-engineered patch in vivo: application of green and red fluorescent protein-expressing retroviral vector.

    PubMed

    Sales, Virna L; Mettler, Bret A; Lopez-Ilasaca, Marco; Johnson, John A; Mayer, John E

    2007-03-01

    An unresolved question regarding tissue-engineered (TE) cardiac valves and vessels is the fate of the transplanted cells in vivo. We have developed a strategy to track the anatomic location of seeded cells within TE constructs and neighboring tissues using a retroviral vector system encoding green and red fluorescent proteins (GFPs and RFPs, respectively) in ovine circulating endothelial progenitor cells (EPCs) and bone marrow-derived mesenchymal stem cells (BMSCs). We demonstrate that stable transduction ex vivo with high-titer Moloney murine leukemia virus-based retroviral vector yields transduction efficiency of greater than 97% GFP(+) EPC- and RFP(+) mesenchymal stem cell (MSC)-derived cells. Cellular phenotype and transgene expression were also maintained through 25 subsequent passages. Using a retroviral vector system to distinguish our pre-seeded cells from tissue-resident progenitor cells and circulating endothelial and marrow-derived precursors, we simultaneously co-seeded 2 x 10(6) GFP(+) EPCs and 2 x 10(5) RFP(+) MSCs onto the TE patches. In a series of ovine pulmonary artery patch augmentation studies, transplanted GFP(+) EPC- and RFP(+) MSC-derived cells persisted within the TE patch 7 to 14 days after implantation, as identified using immunofluorescence. Analysis showed 81% luminal coverage of the TE patches before implantation with transduced cells, increasing to 96% at day 7 and decreasing to 67% at day 14 post-implantation. This suggests a temporal association between retroviral expression of progenitor cells and mediating effects of these cells on the physiological remodeling and maturation of the TE constructs. To our knowledge, this is the first cardiovascular tissue-engineering in vivo study using a double-labeling method to demonstrate a direct evidence of the source, persistence, and incorporation into a TE vascular patch of co-cultured and simultaneously pre-seeded adult progenitor cells.

  13. Pharmacokinetic interaction between warfarin and a uricosuric agent, bucolome: application of In vitro approaches to predicting In vivo reduction of (S)-warfarin clearance.

    PubMed

    Takahashi, H; Kashima, T; Kimura, S; Murata, N; Takaba, T; Iwade, K; Abe, T; Tainaka, H; Yasumori, T; Echizen, a H

    1999-10-01

    A uricosuric agent, bucolome, has been shown to intensify the anticoagulant effect of warfarin. The aims of the present study were to clarify its mechanism(s) and to apply in vitro approaches for predicting this potentially life-threatening in vivo interaction. An in vivo study revealed that Japanese patients given warfarin with bucolome (300 mg/day, n = 21) showed a 1.5-fold greater international normalized ratio than those given warfarin alone (n = 34) despite that the former received a 58% smaller warfarin dose than the latter. Enantioselective assays revealed that bucolome increased plasma unbound fractions of (S)- and (R)-warfarin by 2-fold (p <.01), reduced unbound oral clearances of (S)- and (R)-warfarin by 84 (p <.01) and 26% (p <.05), respectively, and inhibited the unbound formation clearance for (S)-warfarin 7-hydroxylation by 89% (p <.01). In contrast, bucolome elicited no appreciable changes in the plasma unbound (S)-warfarin concentration versus the international normalized ratio relationship. In vitro studies with recombinant human cytochrome P-450 2C9 and liver microsomes showed that bucolome was a potent mixed-type inhibitor for (S)-warfarin 7-hydroxylation, with K(i) of 8.2 and 20.2 microM, respectively. An in vitro model incorporating maximum unbound bucolome concentration in the liver estimated as a sum of hepatic artery and portal vein concentrations and in vitro K(i) made an acceptable prediction for bucolome-induced reductions in in vivo total (bound + unbound) oral clearance, unbound oral clearance, and unbound formation clearance for (S)-warfarin. In conclusion, the augmented anticoagulant effect of warfarin by bucolome due to the metabolic inhibition for pharmacologically more potent (S)-warfarin may be predictable from in vitro data.

  14. Single-layer MnO2 nanosheets suppressed fluorescence of 7-hydroxycoumarin: mechanistic study and application for sensitive sensing of ascorbic acid in vivo.

    PubMed

    Zhai, Wanying; Wang, Chunxia; Yu, Ping; Wang, Yuexiang; Mao, Lanqun

    2014-12-16

    In this study, we systematically investigate the mechanism of single-layer MnO2 nanosheets suppressing fluorescence of 7-hydroxycoumarin and, based on this, demonstrate a new fluorescent method for in vivo sensing of ascorbic acid (AA) in rat brain. The mechanism for the fluorescence suppression is attributed to a combination of inner filter effect (IFE) and static quenching effect (SQE), which is different from those reported for the traditional two-dimensional nanosheets, and Förster resonant energy transfer (FRET) mechanism reported for MnO2 nanosheets. The combination of IFE and SQE leads to an exponential decay in fluorescence intensity of 7-hydroxycoumarin with increasing concentration of MnO2 nanosheets in solution. Such a property allows optimization of the concentration of MnO2 nanosheets in such a way that the addition of reductive analyte (e.g., AA) will to the greatest extent restore the MnO2 nanosheets-suppressed fluorescence of 7-hydroxycoumarin through the redox reaction between AA and MnO2 nanosheets. On the basis of this feature, we demonstrate a fluorescent method for in vivo sensing of AA in the cerebral systems with an improved sensitivity. Compared with the turn-on fluorescent method through first decreasing the fluorescence to the lowest level by adding concentrated MnO2 nanosheets, the method demonstrated here possesses a higher sensitivity, lower limit of detection, and wider linear range. Upon the use of ascorbate oxidase to achieve the selectivity for AA, the turn-on fluorescence method demonstrated here can be used for in vivo sensing of AA in a simple but reliable way.

  15. In Vivo Application of Bacteriophage as a Potential Therapeutic Agent To Control OXA-66-Like Carbapenemase-Producing Acinetobacter baumannii Strains Belonging to Sequence Type 357

    PubMed Central

    Jeon, Jongsoo; Ryu, Choong-Min; Lee, Jun-Young; Park, Jong-Hwan; Lee, Kyungwon

    2016-01-01

    ABSTRACT The increasing prevalence of carbapenem-resistant Acinetobacter baumannii (CRAB) strains in intensive care units has caused major problems in public health worldwide. Our aim was to determine whether this phage could be used as an alternative therapeutic agent against multidrug-resistant bacterial strains, specifically CRAB clinical isolates, using a mouse model. Ten bacteriophages that caused lysis in CRAB strains, including blaOXA-66-like genes, were isolated. YMC13/01/C62 ABA BP (phage Bϕ-C62), which showed the strongest lysis activity, was chosen for further study by transmission electron microscopy (TEM), host range test, one-step growth and phage adsorption rate, thermal and pH stability, bacteriolytic activity test, genome sequencing and bioinformatics analysis, and therapeutic effect of phage using a mouse intranasal infection model. The phage Bϕ-C62 displayed high stability at various temperatures and pH values and strong cell lysis activity in vitro. The phage Bϕ-C62 genome has a double-stranded linear DNA with a length of 44,844 bp, and known virulence genes were not identified in silico. In vivo study showed that all mice treated with phage Bϕ-C62 survived after intranasal bacterial challenge. Bacterial clearance in the lung was observed within 3 days after bacterial challenge, and histologic damage also improved significantly; moreover, no side effects were observed. IMPORTANCE In our study, the novel A. baumannii phage Bϕ-C62 was characterized and evaluated in vitro, in silico, and in vivo. These results, including strong lytic activities and the improvement of survival rates, showed the therapeutic potential of the phage Bϕ-C62 as an antimicrobial agent. This study reports the potential of a novel phage as a therapeutic candidate or nontoxic disinfectant against CRAB clinical isolates in vitro and in vivo. PMID:27208124

  16. In vivo application of 3D-line skeleton graph analysis (LSGA) technique with high-resolution magnetic resonance imaging of trabecular bone structure.

    PubMed

    Pothuaud, Laurent; Newitt, David C; Lu, Ying; MacDonald, Brian; Majumdar, Sharmila

    2004-05-01

    Over the last several years magnetic resonance (MR) imaging has emerged as a means of measuring in vivo 3D trabecular bone structure. In particular, MR based diagnosis could be used to complement standard bone mineral density (BMD) methods for assessing osteoporosis and evaluating longitudinal changes. The aim of this study was to demonstrate the feasibility of using the 3D-LSGA technique for the evaluation of trabecular bone structure of high-resolution MR images, particularly for assessing longitudinal changes, in vivo. First, the reproducibility of topological 3D-LSGA based measurements was evaluated in a set of seven volunteers, and coefficients of variations ranged from 3.5% to 6%. Second, high-resolution MR images of the radius in 30 postmenopausal women from a placebo controlled drug study (Idoxifene), divided into placebo ( n=9) and treated ( n=21) groups, were obtained at baseline (BL) and after 1 year of treatment (follow-up, FU). In addition, dual X-ray absorptiometry (DXA) measures of BMD were obtained in the distal radius. Standard morphological measurements based on the mean intercept length (MIL) technique as well as 3D-LSGA based measurements were applied to the 3D MR images. Significant changes from BL to FU were detected, in the treated group, using the topological 3D-LSGA based measurements, morphological measures of volume of connected trabeculae and App Tb.N from MIL analysis. The duration of the study was short, and the number of patients remaining in the study was small, hence these results cannot be interpreted with regard to a true therapeutic response. Furthermore, the site (wrist) and the drug (idoxifene) are not optimal for follow-up study. However, this paper demonstrated the feasibility of using 3D-LSGA based evaluation coupled with in vivo high-resolution MR imaging as a complementary approach for the monitoring of trabecular bone changes in individual subjects.

  17. The application of anti-ESAT-6 monoclonal antibody fluorescent probe in ex vivo near-infrared fluorescence imaging in mice with pulmonary tuberculosis.

    PubMed

    Feng, Feng; Zhang, Haoling; Zhu, Zhaoqin; Li, Cong; Shi, Yuxin; Zhang, Zhiyong

    2014-09-01

    Here, we aimed to assess the feasibility of anti-ESAT-6 monoclonal antibody (mAb) coupling with IR783 and rhodamine fluorescent probe in the detection of ESAT-6 expression in tuberculosis tissue of mice using near-infrared fluorescence imaging. IR783 and rhodamine were conjugated to the anti-ESAT-6 mAb or IgG. Mice in the experimental group were injected with fluorescence-labeled mAb probe, and mice in the control group were injected with fluorescence-labeled non-specific IgG antibody. Twenty-four hours later, the lung tissue of mice was examined using ex vivo near-infrared fluorescence imaging. In addition, the contrast-to-noise ratio (CNR) was calculated by measuring the signal intensities of the pulmonary lesions, normal lung tissue and background noise. The frozen lung tissue section was examined under fluorescence microscopy and compared with hemoxylin and eosin (HE) staining. The ex vivo near-infrared fluorescence imaging showed that the fluorescence signal in the lung tuberculosis lesions in the experimental group was significantly enhanced, whereas there was only a weak fluorescence signal or even no fluorescence signal in the control group. CNR values were 64.40 ± 7.02 (n = 6) and 8.75 ± 3.87 (n = 6), respectively (t = 17.01, p < 0.001). The fluorescence accumulation distribution detected under fluorescence microscopy was consistent with HE staining of the tuberculosis region. In conclusion, anti-ESAT-6 mAb fluorescent probe could target and be applied in specific ex vivo imaging of mice tuberculosis, and may be of further use in tuberculosis in living mice.

  18. Optimisations and Challenges Involved in the Creation of Various Bioluminescent and Fluorescent Influenza A Virus Strains for In Vitro and In Vivo Applications

    PubMed Central

    Herfst, Sander; Bestebroer, Theo M.; Vaes, Vincent P.; van der Hoeven, Barbara; Koster, Abraham J.; Kremers, Gert-Jan; Scott, Dana P.; Gultyaev, Alexander P.; Sorell, Erin M.; de Graaf, Miranda; Bárcena, Montserrat; Rimmelzwaan, Guus F.; Fouchier, Ron A.

    2015-01-01

    Bioluminescent and fluorescent influenza A viruses offer new opportunities to study influenza virus replication, tropism and pathogenesis. To date, several influenza A reporter viruses have been described. These strategies typically focused on a single reporter gene (either bioluminescent or fluorescent) in a single virus backbone. However, whilst bioluminescence is suited to in vivo imaging, fluorescent viruses are more appropriate for microscopy. Therefore, the idea l reporter virus varies depending on the experiment in question, and it is important that any reporter virus strategy can be adapted accordingly. Herein, a strategy was developed to create five different reporter viruses in a single virus backbone. Specifically, enhanced green fluorescent protein (eGFP), far-red fluorescent protein (fRFP), near-infrared fluorescent protein (iRFP), Gaussia luciferase (gLUC) and firefly luciferase (fLUC) were inserted into the PA gene segment of A/PR/8/34 (H1N1). This study provides a comprehensive characterisation of the effects of different reporter genes on influenza virus replication and reporter activity. In vivo reporter gene expression, in lung tissues, was only detected for eGFP, fRFP and gLUC expressing viruses. In vitro, the eGFP-expressing virus displayed the best reporter stability and could be used for correlative light electron microscopy (CLEM). This strategy was then used to create eGFP-expressing viruses consisting entirely of pandemic H1N1, highly pathogenic avian influenza (HPAI) H5N1 and H7N9. The HPAI H5N1 eGFP-expressing virus infected mice and reporter gene expression was detected, in lung tissues, in vivo. Thus, this study provides new tools and insights for the creation of bioluminescent and fluorescent influenza A reporter viruses. PMID:26241861

  19. Diffuse optical tomography with an amplified ultrafast laser and a single-shot streak camera: application to real-time in vivo songbird neuro-imaging

    NASA Astrophysics Data System (ADS)

    Guillet de Chatellus, Hugues; Vignal, Clémentine; Ramstein, Stéphane; Verjat, Nicolas; Mathevon, Nicolas; Mottin, Stéphane

    2005-10-01

    A new ultrafast Diffuse Optical Tomography (DOT) has been developed for real time in vivo brain metabolism monitoring in songbird. The technique is based on space resolved time of flight measurements of the photons across the brain tissues. A three dimensional reconstruction of the brain activity is foreseeable by means of a double space and time sampling of the reflectance signal. The setup and the treatment procedure are described in depth and promising preliminary results showing the response of brain tissues to hypercapnia stimulations (increase of CO2) are presented.

  20. Behavior of tip-steerable needles in ex vivo and in vivo tissue.

    PubMed

    Majewicz, Ann; Marra, Steven P; van Vledder, Mark G; Lin, MingDe; Choti, Michael A; Song, Danny Y; Okamura, Allison M

    2012-10-01

    Robotic needle steering is a promising technique to improve the effectiveness of needle-based clinical procedures, such as biopsies and ablation, by computer-controlled, curved insertions of needles within solid organs. In this paper, we explore the capabilities, challenges, and clinical relevance of asymmetric-tip needle steering through experiments in ex vivo and in vivo tissue. We evaluate the repeatability of needle insertion in inhomogeneous biological tissue and compare ex vivo and in vivo needle curvature and insertion forces. Steerable needles curved more in kidney than in liver and prostate, likely due to differences in tissue properties. Pre-bent needles produced higher insertion forces in liver and more curvature in vivo than ex vivo. When compared to straight stainless steel needles, steerable needles did not cause a measurable increase in tissue damage and did not exert more force during insertion. The minimum radius of curvature achieved by prebent needles was 5.23 cm in ex vivo tissue, and 10.4 cm in in vivo tissue. The curvatures achieved by bevel tip needles were negligible for in vivo tissue. The minimum radius of curvature for bevel tip needles in ex vivo tissue was 16.4 cm; however, about half of the bevel tip needles had negligible curvatures. We also demonstrate a potential clinical application of needle steering by targeting and ablating overlapping regions of cadaveric canine liver.

  1. Behavior of Tip-Steerable Needles in ex vivo and in vivo Tissue

    PubMed Central

    Majewicz, Ann; Marra, Steven P.; van Vledder, Mark G.; Lin, MingDe; Choti, Michael A.; Song, Danny Y.; Okamura, Allison M.

    2012-01-01

    Robotic needle steering is a promising technique to improve the effectiveness of needle-based clinical procedures, such as biopsies and ablation, by computer-controlled, curved insertions of needles within solid organs. In this paper, we explore the capabilities, challenges, and clinical relevance of asymmetric-tip needle steering though experiments in ex vivo and in vivo tissue. We evaluate the repeatability of needle insertion in inhomogeneous biological tissue and compare ex vivo and in vivo needle curvature and insertion forces. Steerable needles curved more in kidney than in liver and prostate, likely due to differences in tissue properties. Pre-bent needles produced higher insertion forces in liver and more curvature in vivo than ex vivo. When compared to straight stainless steel needles, steerable needles did not cause a measurable increase in tissue damage and did not exert more force during insertion. The minimum radius of curvature achieved by pre-bent needles was 5.23 cm in ex vivo tissue, and 10.4 cm in in vivo tissue. The curvatures achieved by bevel tip needles were negligible for in vivo tissue. The minimum radius of curvature for bevel tip needles in ex vivo tissue was 16.4 cm; however, about half of the bevel tip needles had negligible curvatures. We also demonstrate a potential clinical application of needle steering by targeting and ablating overlapping regions of cadaveric canine liver. PMID:22711767

  2. Implanted, inductively-coupled, radiofrequency coils fabricated on flexible polymeric material: Application to in vivo rat brain MRI at 7 T

    NASA Astrophysics Data System (ADS)

    Ginefri, J.-C.; Rubin, A.; Tatoulian, M.; Woytasik, M.; Boumezbeur, F.; Djemaï, B.; Poirier-Quinot, M.; Lethimonnier, F.; Darrasse, L.; Dufour-Gergam, E.

    2012-11-01

    Combined with high-field MRI scanners, small implanted coils allow for high resolution imaging with locally improved SNR, as compared to external coils. Small flexible implantable coils dedicated to in vivo MRI of the rat brain at 7 T were developed. Based on the Multi-turn Transmission Line Resonator design, they were fabricated with a Teflon substrate using copper micromolding process and a specific metal-polymer adhesion treatment. The implanted coils were made biocompatible by PolyDimethylSiloxane (PDMS) encapsulation. The use of low loss tangent material achieves low dielectric losses within the substrate and the use of the PDMS layer reduces the parasitic coupling with the surrounding media. An implanted coil was implemented in a 7 T MRI system using inductive coupling and a dedicated external pick-up coil for signal transmission. In vivo images of the rat brain acquired with in plane resolution of (150 μm)2 thanks to the implanted coil revealed high SNR near the coil, allowing for the visualization of fine cerebral structures.

  3. Simultaneous quantification of VX and its toxic metabolite in blood and plasma samples and its application for in vivo and in vitro toxicological studies.

    PubMed

    Reiter, Georg; Mikler, John; Hill, Ira; Weatherby, Kendal; Thiermann, Horst; Worek, Franz

    2011-09-15

    The present study was initiated to develop a sensitive and highly selective method for the simultaneous quantification of the nerve agent VX (O-ethyl S-[2(diisopropylamino)ethyl] methylphosphonothioate) and its toxic metabolite (EA-2192) in blood and plasma samples in vivo and in vitro. For the quantitative detection of VX and EA-2192 the resolution was realized on a HYPERCARB HPLC phase. A specific procedure was developed to isolate both toxic analytes from blood and plasma samples. The limit of detection was 0.1 pg/ml and the absolute recovery of the overall sample preparation procedure was 74% for VX and 69% for EA-2192. After intravenous and percutaneous administration of a supralethal doses of VX in anaesthetised swine both VX and EA-2192 could be quantified over 540 min following exposure. This study is the first to verify the in vivo formation of the toxic metabolite EA-2192 after poisoning with the nerve agent VX. Further toxicokinetic and therapeutic studies are required in order to determine the impact of EA-2192 on the treatment of acute VX poisoning.

  4. Development of a disposable maglev centrifugal blood pump intended for one-month support in bridge-to-bridge applications: in vitro and initial in vivo evaluation.

    PubMed

    Someya, Takeshi; Kobayashi, Mariko; Waguri, Satoshi; Ushiyama, Tomohiro; Nagaoka, Eiki; Hijikata, Wataru; Shinshi, Tadahiko; Arai, Hirokuni; Takatani, Setsuo

    2009-09-01

    MedTech Dispo, a disposable maglev centrifugal blood pump with two degrees of freedom magnetic suspension and radial magnetic coupling rotation, has been developed for 1-month extracorporeal circulatory support. As the first stage of a two-stage in vivo evaluation, 2-week evaluation of a prototype MedTech Dispo was conducted. In in vitro study, the pump could produce 5 L/min against 800 mm Hg and the normalized index of hemolysis was 0.0054 +/- 0.0008 g/100 L. In in vivo study, the pump, with its blood-contacting surface coated with biocompatible 2-methacryloyloxyethyl phosphorylcholine polymer, was implanted in seven calves in left heart bypass. Pump performance was stable with a mean flow of 4.49 +/- 0.38 L/min at a mean speed of 2072.1 +/- 64.5 rpm. The maglev control revealed its stability in rotor position during normal activity by the calves. During 2 weeks of operation in two calves which survived the intended study period, no thrombus formation was seen inside the pump and levels of plasma free hemoglobin were maintained below 4 mg/dL. Although further experiments are required, the pump demonstrated the potential for sufficient and reliable performance and biocompatibility in meeting the requirements for cardiopulmonary bypass and 1-week circulatory support.

  5. Applicability of the Rayleigh equation for enantioselective metabolism of chiral xenobiotics by microsomes, hepatocytes and in-vivo retention in rabbit tissues.

    PubMed

    Jammer, Shifra; Gelman, Faina; Lev, Ovadia

    2016-03-29

    In this study we propose a new approach for analyzing the enantioselective biodegradation of some antidepressant drugs mediated by human and rat liver microsomes by using the Rayleigh equation to describe the enantiomeric enrichment-conversion dependencies. Analysis of reported degradation data of additional six pesticides, an alpha blocker and a flame retardant by microsomes or hepatocytes in vitro reaffirmed the universality of the approach. In all the in vitro studied cases that involved enantioselective degradation, a Rayleigh dependence of the enantiomeric enrichment was observed. Published data regarding in vivo retention of myclobutanil in liver, kidney, muscle and brain tissues of rabbits following injection of the racemate were remodeled showing prevalence of the Rayleigh law for the chiral enrichment of the fungicide in the various tissues. This approach will revolutionize data organization in metabolic pathway research of target xenobiotics by either liver microsomes, hepatocytes or their organ-specific in vivo retention. The fact that the enantiomeric enrichment as a function of the conversion can be described by a single quantifier, will pave the road for the use of structure activity predictors of the enantiomeric enrichment and for mechanistic discrimination based on parametric dependence of the quantifier.

  6. Applicability of the Rayleigh equation for enantioselective metabolism of chiral xenobiotics by microsomes, hepatocytes and in-vivo retention in rabbit tissues

    PubMed Central

    Jammer, Shifra; Gelman, Faina; Lev, Ovadia

    2016-01-01

    In this study we propose a new approach for analyzing the enantioselective biodegradation of some antidepressant drugs mediated by human and rat liver microsomes by using the Rayleigh equation to describe the enantiomeric enrichment−conversion dependencies. Analysis of reported degradation data of additional six pesticides, an alpha blocker and a flame retardant by microsomes or hepatocytes in vitro reaffirmed the universality of the approach. In all the in vitro studied cases that involved enantioselective degradation, a Rayleigh dependence of the enantiomeric enrichment was observed. Published data regarding in vivo retention of myclobutanil in liver, kidney, muscle and brain tissues of rabbits following injection of the racemate were remodeled showing prevalence of the Rayleigh law for the chiral enrichment of the fungicide in the various tissues. This approach will revolutionize data organization in metabolic pathway research of target xenobiotics by either liver microsomes, hepatocytes or their organ-specific in vivo retention. The fact that the enantiomeric enrichment as a function of the conversion can be described by a single quantifier, will pave the road for the use of structure activity predictors of the enantiomeric enrichment and for mechanistic discrimination based on parametric dependence of the quantifier. PMID:27021918

  7. Biocompatible nanogenerators through high piezoelectric coefficient 0.5Ba(Zr0.2Ti0.8)O3-0.5(Ba0.7Ca0.3)TiO3 nanowires for in-vivo applications.

    PubMed

    Yuan, Miaomiao; Cheng, Li; Xu, Qi; Wu, Weiwei; Bai, Suo; Gu, Long; Wang, Zhe; Lu, Jun; Li, Huanping; Qin, Yong; Jing, Tao; Wang, Zhong Lin

    2014-11-26

    Lead-free BZT-BCT (0.5Ba(Zr0.2Ti0.8)O3-0.5(Ba0.7Ca0.3)TiO3) nanowires with a high piezoelectric coefficient are synthesized and nanogenerators (NGs) composed of them are successfully developed. The studied in vitro and in vivo biocompatibility of the NGs shows great potential for their application as in vivo power sources.

  8. Clinical application of in vivo treatment delivery verification based on PET/CT imaging of positron activity induced at high energy photon therapy

    NASA Astrophysics Data System (ADS)

    Janek Strååt, Sara; Andreassen, Björn; Jonsson, Cathrine; Noz, Marilyn E.; Maguire, Gerald Q., Jr.; Näfstadius, Peder; Näslund, Ingemar; Schoenahl, Frederic; Brahme, Anders

    2013-08-01

    The purpose of this study was to investigate in vivo verification of radiation treatment with high energy photon beams using PET/CT to image the induced positron activity. The measurements of the positron activation induced in a preoperative rectal cancer patient and a prostate cancer patient following 50 MV photon treatments are presented. A total dose of 5 and 8 Gy, respectively, were delivered to the tumors. Imaging was performed with a 64-slice PET/CT scanner for 30 min, starting 7 min after the end of the treatment. The CT volume from the PET/CT and the treatment planning CT were coregistered by matching anatomical reference points in the patient. The treatment delivery was imaged in vivo based on the distribution of the induced positron emitters produced by photonuclear reactions in tissue mapped on to the associated dose distribution of the treatment plan. The results showed that spatial distribution of induced activity in both patients agreed well with the delivered beam portals of the treatment plans in the entrance subcutaneous fat regions but less so in blood and oxygen rich soft tissues. For the preoperative rectal cancer patient however, a 2 ± (0.5) cm misalignment was observed in the cranial-caudal direction of the patient between the induced activity distribution and treatment plan, indicating a beam patient setup error. No misalignment of this kind was seen in the prostate cancer patient. However, due to a fast patient setup error in the PET/CT scanner a slight mis-position of the patient in the PET/CT was observed in all three planes, resulting in a deformed activity distribution compared to the treatment plan. The present study indicates that the induced positron emitters by high energy photon beams can be measured quite accurately using PET imaging of subcutaneous fat to allow portal verification of the delivered treatment beams. Measurement of the induced activity in the patient 7 min after receiving 5 Gy involved count rates which were about

  9. In Vivo and In Vitro Detection of Luminescent and Fluorescent Lactobacillus reuteri and Application of Red Fluorescent mCherry for Assessing Plasmid Persistence

    PubMed Central

    Karimi, Shokoufeh; Ahl, David; Vågesjö, Evelina; Holm, Lena; Phillipson, Mia; Jonsson, Hans; Roos, Stefan

    2016-01-01

    Lactobacillus reuteri is a symbiont that inhabits the gastrointestinal (GI) tract of mammals, and several strains are used as probiotics. After introduction of probiotic strains in a complex ecosystem like the GI tract, keeping track of them is a challenge. The main objectives of this study were to introduce reporter proteins that would enable in vivo and in vitro detection of L. reuteri and increase knowledge about its interactions with the host. We describe for the first time cloning of codon-optimized reporter genes encoding click beetle red luciferase (CBRluc) and red fluorescent protein mCherry in L. reuteri strains ATCC PTA 6475 and R2LC. The plasmid persistence of mCherry-expressing lactobacilli was evaluated by both flow cytometry (FCM) and conventional plate count (PC), and the plasmid loss rates measured by FCM were lower overall than those determined by PC. Neutralization of pH and longer induction duration significantly improved the mCherry signal. The persistency, dose-dependent signal intensity and localization of the recombinant bacteria in the GI tract of mice were studied with an in vivo imaging system (IVIS), which allowed us to detect fluorescence from 6475-CBRluc-mCherry given at a dose of 1×1010 CFU and luminescence signals at doses ranging from 1×105 to 1×1010 CFU. Both 6475-CBRluc-mCherry and R2LC-CBRluc were localized in the colon 1 and 2 h after ingestion, but the majority of the latter were still found in the stomach, possibly reflecting niche specificity for R2LC. Finally, an in vitro experiment showed that mCherry-producing R2LC adhered efficiently to the intra cellular junctions of cultured IPEC-J2 cells. In conclusion, the two reporter genes CBRluc and mCherry were shown to be suitable markers for biophotonic imaging (BPI) of L. reuteri and may provide useful tools for future studies of in vivo and in vitro interactions between the bacteria and the host. PMID:27002525

  10. In vivo transport of a dynorphin-like analgesic peptide, E-2078, through the blood-brain barrier: an application of brain microdialysis.

    PubMed

    Terasaki, T; Deguchi, Y; Sato, H; Hirai, K; Tsuji, A

    1991-07-01

    In vivo transport through the blood-brain barrier (BBB) has been demonstrated for a dynorphin-like analgesic peptide, CH3-[125I]Tyr-Gly-Gly-Phe-Leu-Arg-CH3Arg-D-Leu-NHC2H5 ( [125I]E-2078). A remarkable time-dependent increase in the distribution volume of [125I]E-2078 in the brain parenchyma separated from blood vessels and capillaries was observed during a brain perfusion. The distribution volume of [125I]E-2078 in the brain parenchyma after 20 min of perfusion was 2.18 +/- 0.09 microliters/g brain (mean +/- SE) and was significantly greater than the distribution volume of [3H]inulin (0.994 +/- 0.138 microliters/g brain), providing in vivo evidence for the penetration of [125I]E-2078 into the brain parenchyma. Brain microdialysis was carried out to collect directly the brain interstitial fluid (ISF) during the brain perfusion of [125I]E-2078. No metabolite of [125I]E-2078 in the brain ISF was found by high-performance liquid chromatographic analysis of the brain dialysate. The concentrations of [125I]E-2078 and [14C]sucrose in the brain ISF were estimated based on an in vitro evaluation of dialysis clearance. The concentration ratio of [125I]E-2078 between the brain ISF and the brain perfusate was determined to be 2.92 x 10(-1) +/- 0.50 x 10(-1) and was approximately 100 times higher than that of [14C]sucrose (2.71 x 10(-3) +/- 1.43 x 10(-3), demonstrating transport of [125I]E-2078 through the BBB in vivo. On the other hand, no remarkable difference in the cerebrospinal fluid (CSF)-to-perfusate concentration ratios of [125I]E-2078 and [14C]sucrose was observed, indicating little contribution of the blood-CSF barrier (BCSF barrier) transport to the penetration of [125I]E-2078 into the brain.

  11. Terahertz pulsed imaging in vivo

    NASA Astrophysics Data System (ADS)

    Pickwell-MacPherson, E.

    2011-03-01

    Terahertz (1012 Hz) pulsed imaging is a totally non-destructive and non-ionising imaging modality and thus potential applications in medicine are being investigated. In this paper we present results using our hand-held terahertz probe that has been designed for in vivo use. In particular, we use the terahertz probe to perform reflection geometry in vivo measurements of human skin. The hand-held terahertz probe gives more flexibility than a typical flat-bed imaging system, but it also results in noisier data and requires existing processing methods to be improved. We describe the requirements and limitations of system geometry, data acquisition rate, image resolution and penetration depth and explain how various factors are dependent on each other. We show how some of the physical limitations can be overcome using novel data processing methods.

  12. Patient-derived Models of Human Breast Cancer: Protocols for In vitro and In vivo Applications in Tumor Biology and Translational Medicine

    PubMed Central

    DeRose, Yoko S.; Gligorich, Keith M.; Wang, Guoying; Georgelas, Ann; Bowman, Paulette; Courdy, Samir J.; Welm, Alana L.; Welm, Bryan E.

    2013-01-01

    Research models that replicate the diverse genetic and molecular landscape of breast cancer are critical for developing the next generation therapeutic entities that can target specific cancer subtypes. Patient-derived tumorgrafts, generated by transplanting primary human tumor samples into immune-compromised mice, are a valuable method to model the clinical diversity of breast cancer in mice, and are a potential resource in personalized medicine. Primary tumorgrafts also enable in vivo testing of therapeutics and make possible the use of patient cancer tissue for in vitro screens. Described in this unit are a variety of protocols including tissue collection, biospecimen tracking, tissue processing, transplantation, and 3-dimensional culturing of xenografted tissue, that enable use of bona fide uncultured human tissue in designing and validating cancer therapies. PMID:23456611

  13. High-field MRI of single histological slices using an inductively coupled, self-resonant microcoil: application to ex vivo samples of patients with Alzheimer's disease.

    PubMed

    Nabuurs, Rob J A; Hegeman, Ingrid; Natté, Remco; van Duinen, Sjoerd G; van Buchem, Mark A; van der Weerd, Louise; Webb, Andrew G

    2011-05-01

    A simple inductively coupled microcoil has been designed to image tissue samples placed on a microscope slide, samples which can subsequently be stained histologically. As the exact same tissue is used for MRI and histology, the two data sets can be compared without the need for complicated image registration techniques. The design can be integrated into any MRI system using existing commercial hardware. Compared with a commercial 25-mm-diameter birdcage, the signal-to-noise ratio was increased by a factor of 3.8, corresponding to an approximate 15-fold reduction in the data acquisition time. An example is shown of ex vivo samples from patients with Alzheimer's disease, in which the coregistration of highly sensitive iron staining and amyloid-β deposits is confirmed.

  14. Development of ultrafast laser-based x-ray in-vivo phase-contrast micro-CT beamline for biomedical applications at Advanced Laser Light Source (ALLS).

    PubMed

    Kincaid, Russell; Krol, Andrzej; Fourmaux, Sylvain; Kieffer, Jean-Claude; Serbanescu, Cristina; Servol, Marina; Vogelsang, Levon; Wilkins, Steve; Stevenson, Andrew; Nesterets, Yakov; Lipson, Edward; Ye, Hongwei; Pogany, Andrew

    2008-01-01

    We are developing and exploring the imaging performance of, an in vivo, in-line holography, x-ray phase-contrast, micro-CT system with an ultrafast laser-based x-ray (ULX) source. By testing and refining our system, and by performing computer simulations, we plan to improve system performance in terms of contrast resolution and multi-energy imaging to a level beyond what can be obtained using a conventional microfocal x-ray tube. Initial CT projection sets at single energy (Mo K(alpha) and K(beta) lines) were acquired in the Fresnel regime and reconstructed for phantoms and a euthanized mouse. We also performed computer simulations of phase-contrast micro-CT scans for low-contrast, soft-tissue, tumor imaging. We determined that, in order to perform a phase-contrast, complete micro-CT scan using ULX, the following conditions must be met: (i) the x-ray source needs to be stable during the scan; (ii) the laser focal spot size needs to be less than 10 mum for source-to-object distance greater than 30 cm; (iii) the laser light intensity on the target needs to be in the range of 5 x 10(17) to 5 x 10(19) W/cm(2); (iv) the ablation protection system needs to allow uninterrupted scans; (v) the laser light focusing on the target needs to remain accurate during the entire scan; (vi) a fresh surface of the target must be exposed to consecutive laser shots during the entire scan; (vii) the effective detector element size must be less than 12 mum. Based on the results obtained in this research project, we anticipate that the new 10 Hz, 200 TW laser with 50 W average power that is being commissioned at ALLS will allow us practical implementation of in vivo x-ray phase-contrast micro-CT.

  15. Effects of In Vitro Low Oxygen Tension Preconditioning of Adipose Stromal Cells on Their In Vivo Chondrogenic Potential: Application in Cartilage Tissue Repair

    PubMed Central

    Gauthier, Olivier; Lesoeur, Julie; Sourice, Sophie; Masson, Martial; Fellah, Borhane Hakim; Geffroy, Olivier; Lallemand, Elodie; Weiss, Pierre

    2013-01-01

    Purpose Multipotent stromal cell (MSC)-based regenerative strategy has shown promise for the repair of cartilage, an avascular tissue in which cells experience hypoxia. Hypoxia is known to promote the early chondrogenic differentiation of MSC. The aim of our study was therefore to determine whether low oxygen tension could be used to enhance the regenerative potential of MSC for cartilage repair. Methods MSC from rabbit or human adipose stromal cells (ASC) were preconditioned in vitro in control or chondrogenic (ITS and TGF-β) medium and in 21 or 5% O2. Chondrogenic commitment was monitored by measuring COL2A1 and ACAN expression (real-time PCR). Preconditioned rabbit and human ASC were then incorporated into an Si-HPMC hydrogel and injected (i) into rabbit articular cartilage defects for 18 weeks or (ii) subcutaneously into nude mice for five weeks. The newly formed tissue was qualitatively and quantitatively evaluated by cartilage-specific immunohistological staining and scoring. The phenotype of ASC cultured in a monolayer or within Si-HPMC in control or chondrogenic medium and in 21 or 5% O2 was finally evaluated using real-time PCR. Results/Conclusions 5% O2 increased the in vitro expression of chondrogenic markers in ASC cultured in induction medium. Cells implanted within Si-HPMC hydrogel and preconditioned in chondrogenic medium formed a cartilaginous tissue, regardless of the level of oxygen. In addition, the 3D in vitro culture of ASC within Si-HPMC hydrogel was found to reinforce the pro-chondrogenic effects of the induction medium and 5% O2. These data together indicate that although 5% O2 enhances the in vitro chondrogenic differentiation of ASC, it does not enhance their in vivo chondrogenesis. These results also highlight the in vivo chondrogenic potential of ASC and their potential value in cartilage repair. PMID:23638053

  16. The design of a double-tuned two-port surface resonator and its application to in vivo Hydrogen- and Sodium-MRI

    NASA Astrophysics Data System (ADS)

    Wetterling, Friedrich; Högler, Miroslav; Molkenthin, Ute; Junge, Sven; Gallagher, Lindsay; Mhairi Macrae, I.; Fagan, Andrew J.

    2012-04-01

    The design and construction of a two-port surface transceiver resonator for both 1H- and 23Na-MRI in the rodent brain at 7 T is described. Double-tuned resonators are required for accurately co-registering multi-nuclei data sets, especially when the time courses of 1H and 23Na signals are of interest as, for instance, when investigating the pathological progression of ischaemic stroke tissue in vivo. In the current study, a single-element two-port surface resonator was developed wherein both frequency components were measured with the same detector element but with each frequency signal routed along different output channels. This was achieved by using the null spot technique, allowing for optimal variable tuning and matching of each channel in situ within the MRI scanner. The 23Na signal to noise ratio, measured in the ventricles of the rat brain, was increased by a factor of four compared to recent state-of-the-art rat brain studies reported in the literature. The resonator's performance was demonstrated in an in vivo rodent stroke model, where regional variations in 1H apparent diffusion coefficient maps and the 23Na signal were recorded in an interleaved fashion as a function of time in the acute phase of the stroke without having to exchange, re-adjust, or re-connect resonators between scans. Using the practical construction steps described in this paper, this coil design can be easily adapted for MRI of other X-nuclei, such as 17O, 13C, 39K, and 43Ca at various field strengths.

  17. Redox imaging of skeletal muscle using in vivo DNP-MRI and its application to an animal model of local inflammation.

    PubMed

    Eto, Hinako; Hyodo, Fuminori; Kosem, Nutavutt; Kobayashi, Ryoma; Yasukawa, Keiji; Nakao, Motonao; Kiniwa, Mamoru; Utsumi, Hideo

    2015-12-01

    Disorders of skeletal muscle are often associated with inflammation and alterations in redox status. A non-invasive technique that could localize and evaluate the severity of skeletal muscle inflammation based on its redox environment would be useful for disease identification and monitoring, and for the development of treatments; however, no such technique currently exists. We describe a method for redox imaging of skeletal muscle using dynamic nuclear polarization magnetic resonance imaging (DNP-MRI), and apply this method to an animal model of local inflammation. Female C57/BL6 mice received injections of 0.5% bupivacaine into their gastrocnemius muscles. Plasma biomarkers, myeloperoxidase activity, and histological sections were assessed at 4 and 24h after bupivacaine injection to measure the inflammatory response. In vivo DNP-MRI was performed with the nitroxyl radicals carbamoyl-PROXYL (cell permeable) and carboxy-PROXYL (cell impermeable) as molecular imaging probes at 4 and 24h after bupivacaine administration. The images obtained after carbamoyl-PROXYL administration were confirmed with the results of L-band EPR spectroscopy. The plasma biomarkers, myeloperoxidase activity, and histological findings indicated that bupivacaine injection caused acute muscle damage and inflammation. DNP-MRI images of mice treated with carbamoyl-PROXYL or carboxy-PROXYL at 4 and 24h after bupivacaine injection showed similar increases in image intensity and decay rate was significantly increased at 24h. In addition, reduction rates in individual mice at 4h and 24h showed faster trends with bupivacaine injection than in their contralateral sides by image-based analysis. These findings indicate that in vivo DNP-MRI with nitroxyl radicals can non-invasively detect changes in the focal redox status of muscle resulting from locally-induced inflammation.

  18. Application of dynamic metabolomics to examine in vivo skeletal muscle glucose metabolism in the chronically high-fat fed mouse

    SciTech Connect

    Kowalski, Greg M.; De Souza, David P.; Burch, Micah L.; Hamley, Steven; Kloehn, Joachim; Selathurai, Ahrathy; Tull, Dedreia; O'Callaghan, Sean; McConville, Malcolm J.; Bruce, Clinton R.

    2015-06-19

    Rationale: Defects in muscle glucose metabolism are linked to type 2 diabetes. Mechanistic studies examining these defects rely on the use of high fat-fed rodent models and typically involve the determination of muscle glucose uptake under insulin-stimulated conditions. While insightful, they do not necessarily reflect the physiology of the postprandial state. In addition, most studies do not examine aspects of glucose metabolism beyond the uptake process. Here we present an approach to study rodent muscle glucose and intermediary metabolism under the dynamic and physiologically relevant setting of the oral glucose tolerance test (OGTT). Methods and results: In vivo muscle glucose and intermediary metabolism was investigated following oral administration of [U-{sup 13}C] glucose. Quadriceps muscles were collected 15 and 60 min after glucose administration and metabolite flux profiling was determined by measuring {sup 13}C mass isotopomers in glycolytic and tricarboxylic acid (TCA) cycle intermediates via gas chromatography–mass spectrometry. While no dietary effects were noted in the glycolytic pathway, muscle from mice fed a high fat diet (HFD) exhibited a reduction in labelling in TCA intermediates. Interestingly, this appeared to be independent of alterations in flux through pyruvate dehydrogenase. In addition, our findings suggest that TCA cycle anaplerosis is negligible in muscle during an OGTT. Conclusions: Under the dynamic physiologically relevant conditions of the OGTT, skeletal muscle from HFD fed mice exhibits alterations in glucose metabolism at the level of the TCA cycle. - Highlights: • Dynamic metabolomics was used to investigate muscle glucose metabolism in vivo. • Mitochondrial TCA cycle metabolism is altered in muscle of HFD mice. • This defect was not pyruvate dehydrogenase mediated, as has been previously thought. • Mitochondrial TCA cycle anaplerosis in muscle is virtually absent during the OGTT.

  19. In vitro and in vivo application of RNA interference for targeting genes involved in peritrophic matrix synthesis in a lepidopteran system.

    PubMed

    Toprak, Umut; Baldwin, Doug; Erlandson, Martin; Gillott, Cedric; Harris, Stephanie; Hegedus, Dwayne D

    2013-02-01

    The midgut of most insects is lined with a semipermeable acellular tube, the peritrophic matrix (PM), composed of chitin and proteins. Although various genes encoding PM proteins have been characterized, our understanding of their roles in PM structure and function is very limited. One promising approach for obtaining functional information is RNA interference, which has been used to reduce the levels of specific mRNAs using double-stranded RNAs administered to larvae by either injection or feeding. Although this method is well documented in dipterans and coleopterans, reports of its success in lepidopterans are varied. In the current study, the silencing midgut genes encoding PM proteins (insect intestinal mucin 1, insect intestinal mucin 4, PM protein 1) and the chitin biosynthetic or modifying enzymes (chitin synthase-B and chitin deacetylase 1) in a noctuid lepidopteran, Mamestra configurata, was examined in vitro and in vivo. In vitro studies in primary midgut epithelial cell preparations revealed an acute and rapid silencing (by 24 h) for the gene encoding chitin deacetylase 1 and a slower rate of silencing (by 72 h) for the gene encoding PM protein 1. Genes encoding insect intestinal mucins were slightly silenced by 72 h, whereas no silencing was detected for the gene encoding chitin synthase-B. In vivo experiments focused on chitin deacetylase 1, as the gene was silenced to the greatest extent in vitro. Continuous feeding of neonates and fourth instar larvae with double-stranded RNA resulted in silencing of chitin deacetylase 1 by 24 and 36 h, respectively. Feeding a single dose to neonates also resulted in silencing by 24 h. The current study demonstrates that genes encoding PM proteins can be silenced and outlines conditions for RNA interference by per os feeding in lepidopterans.

  20. The Antisense RNA Approach: a New Application for In Vivo Investigation of the Stress Response of Oenococcus oeni, a Wine-Associated Lactic Acid Bacterium

    PubMed Central

    Darsonval, Maud; Msadek, Tarek; Alexandre, Hervé

    2015-01-01

    Oenococcus oeni is a wine-associated lactic acid bacterium mostly responsible for malolactic fermentation in wine. In wine, O. oeni grows in an environment hostile to bacterial growth (low pH, low temperature, and ethanol) that induces stress response mechanisms. To survive, O. oeni is known to set up transitional stress response mechanisms through the synthesis of heat stress proteins (HSPs) encoded by the hsp genes, notably a unique small HSP named Lo18. Despite the availability of the genome sequence, characterization of O. oeni genes is limited, and little is known about the in vivo role of Lo18. Due to the lack of genetic tools for O. oeni, an efficient expression vector in O. oeni is still lacking, and deletion or inactivation of the hsp18 gene is not presently practicable. As an alternative approach, with the goal of understanding the biological function of the O. oeni hsp18 gene in vivo, we have developed an expression vector to produce antisense RNA targeting of hsp18 mRNA. Recombinant strains were exposed to multiple stresses inducing hsp18 gene expression: heat shock and acid shock. We showed that antisense attenuation of hsp18 affects O. oeni survival under stress conditions. These results confirm the involvement of Lo18 in heat and acid tolerance of O. oeni. Results of anisotropy experiments also confirm a membrane-protective role for Lo18, as previous observations had already suggested. This study describes a new, efficient tool to demonstrate the use of antisense technology for modulating gene expression in O. oeni. PMID:26452552

  1. A novel near-infrared fluorescent probe for H2O2 in alkaline environment and the application for H2O2 imaging in vitro and in vivo.

    PubMed

    Liu, Keyin; Shang, Huiming; Kong, Xiuqi; Ren, Mingguang; Wang, Jian-Yong; Liu, Yong; Lin, Weiying

    2016-09-01

    H2O2 as one of the most important ROS (Reactive Oxygen Species) has more attack activity to biomolecules such as DNA, RNA, protein and enzyme in alkaline environment and leads to a series of disease. However, no attention has been paid to the fluorescent detection of H2O2 in alkaline environment in the past. Herein, we reported the first ratiometric near-infrared fluorescent probe based on a boric acid derivative of Changsha near-infrared dye (CSBOH) for H2O2 detection in alkaline condition and the application for H2O2 imaging in vivo. ICT (intra-molecular charge transfer) mechanism was used in CSBOH to modulate the fluorescence change. The photophysical change of CSBOH was investigated by comparison with a phenol derivative of Changsha near-infrared dye (CSOH), a structural analogue bearing phenol group. In the presence of H2O2, CSBOH exhibited remarkably different fluorescence change at 650 nm and 720 nm when excited by 560 nm and 670 nm light respectively in alkaline buffer and showed high selectivity toward H2O2. Cellular experiments demonstrate that CSBOH can image endogenously generated H2O2 in macrophages and A431 cells. In vivo experiment demonstrates that both CSOH and CSBOH can be used for bio-imaging, and CSBOH can image H2O2 in living animal successfully.

  2. Immobilized Cytochrome P450 2C9 (CYP2C9): Applications for Metabolite Generation, Monitoring Protein-Protein Interactions, and Improving In-vivo Predictions Using Enhanced In-vitro Models

    NASA Astrophysics Data System (ADS)

    Wollenberg, Lance A.

    Cytochrome P450 (P450) enzymes are a family of oxoferroreductase enzymes containing a heme moiety and are well known to be involved in the metabolism of a wide variety of endogenous and xenobiotic materials. It is estimated that roughly 75% of all pharmaceutical compounds are metabolized by these enzymes. Traditional reconstituted in-vitro incubation studies using recombinant P450 enzymes are often used to predict in-vivo kinetic parameters of a drug early in development. However, in many cases, these reconstituted incubations are prone to aggregation which has been shown to affect the catalytic activity of an enzyme. Moreover, the presence of other isoforms of P450 enzymes present in a metabolic incubation, as is the case with microsomal systems, may affect the catalytic activity of an enzyme through isoform-specific protein-protein interactions. Both of these effects may result in inaccurate prediction of in-vivo drug metabolism using in-vitro experiments. Here we described the development of immobilized P450 constructs designed to elucidate the effects of aggregation and protein-protein interactions between P450 isoforms on catalytic activities. The long term objective of this project is to develop a system to control the oligomeric state of Cytochrome P450 enzymes to accurately elucidate discrepancies between in vitro reconstituted systems and actual in vivo drug metabolism for the precise prediction of metabolic activity. This approach will serve as a system to better draw correlations between in-vivo and in-vitro drug metabolism data. The central hypothesis is that Cytochrome P450 enzymes catalytic activity can be altered by protein-protein interactions occurring between Cytochrome P450 enzymes involved in drug metabolism, and is dependent on varying states of protein aggregation. This dissertation explains the details of the construction and characterization of a nanostructure device designed to control the state of aggregation of a P450 enzyme. Moreover

  3. Multimodal Mn-doped I-III-VI quantum dots for near infrared fluorescence and magnetic resonance imaging: from synthesis to in vivo application

    NASA Astrophysics Data System (ADS)

    Sitbon, Gary; Bouccara, Sophie; Tasso, Mariana; Francois, Aurélie; Bezdetnaya, Lina; Marchal, Frédéric; Beaumont, Marine; Pons, Thomas

    2014-07-01

    The development of sensitive multimodal contrast agents is a key issue to provide better global, multi-scale images for diagnostic or therapeutic purposes. Here we present the synthesis of Zn-Cu-In-(S, Se)/Zn1-xMnxS core-shell quantum dots (QDs) that can be used as markers for both near-infrared fluorescence imaging and magnetic resonance imaging (MRI). We first present the synthesis of Zn-Cu-In-(S, Se) cores coated with a thick ZnS shell doped with various proportions of Mn. Their emission wavelengths can be tuned over the NIR optical window suitable for deep tissue imaging. The incorporation of manganese ions (up to a few thousand ions per QD) confers them a paramagnetic character, as demonstrated by structural analysis and electron paramagnetic resonance spectroscopy. These QDs maintain their optical properties after transfer to water using ligand exchange. They exhibit T1-relaxivities up to 1400 mM-1 [QD] s-1 at 7 T and 300 K. We finally show that these QDs are suitable multimodal in vivo probes and demonstrate MRI and NIR fluorescence detection of regional lymph nodes in mice.The development of sensitive multimodal contrast agents is a key issue to provide better global, multi-scale images for diagnostic or therapeutic purposes. Here we present the synthesis of Zn-Cu-In-(S, Se)/Zn1-xMnxS core-shell quantum dots (QDs) that can be used as markers for both near-infrared fluorescence imaging and magnetic resonance imaging (MRI). We first present the synthesis of Zn-Cu-In-(S, Se) cores coated with a thick ZnS shell doped with various proportions of Mn. Their emission wavelengths can be tuned over the NIR optical window suitable for deep tissue imaging. The incorporation of manganese ions (up to a few thousand ions per QD) confers them a paramagnetic character, as demonstrated by structural analysis and electron paramagnetic resonance spectroscopy. These QDs maintain their optical properties after transfer to water using ligand exchange. They exhibit T1-relaxivities

  4. The Combined Application of the Caco-2 Cell Bioassay Coupled with In Vivo (Gallus gallus) Feeding Trial Represents an Effective Approach to Predicting Fe Bioavailability in Humans

    PubMed Central

    Tako, Elad; Bar, Haim; Glahn, Raymond P.

    2016-01-01

    Research methods that predict Fe bioavailability for humans can be extremely useful in evaluating food fortification strategies, developing Fe-biofortified enhanced staple food crops and assessing the Fe bioavailability of meal plans that include such crops. In this review, research from four recent poultry (Gallus gallus) feeding trials coupled with in vitro analyses of Fe-biofortified crops will be compared to the parallel human efficacy studies which used the same varieties and harvests of the Fe-biofortified crops. Similar to the human studies, these trials were aimed to assess the potential effects of regular consumption of these enhanced staple crops on maintenance or improvement of iron status. The results demonstrate a strong agreement between the in vitro/in vivo screening approach and the parallel human studies. These observations therefore indicate that the in vitro/Caco-2 cell and Gallus gallus models can be integral tools to develop varieties of staple food crops and predict their effect on iron status in humans. The cost-effectiveness of this approach also means that it can be used to monitor the nutritional stability of the Fe-biofortified crop once a variety has released and integrated into the food system. These screening tools therefore represent a significant advancement to the field for crop development and can be applied to ensure the sustainability of the biofortification approach. PMID:27869705

  5. Development of a liquid chromatographic method for the quantification of paromomycin. Application to in vitro release and ex vivo permeation studies

    NASA Astrophysics Data System (ADS)

    Pujol-Brugués, A.; Calpena-Campmany, A. C.; Riera-Lizandra, C.; Halbaut-Bellowa, L.; Clares-Naveros, B.

    2014-12-01

    We have developed a reversed phase high performance liquid chromatography pulsed amperometric detection (RPHPLC-PAD) method for the determination of paromomycin. It is sensitive, repeatable, and selective without the pretreatment step. Trifluoroacetic acid-water was utilized as the eluent and detected by PAD under NaOH alkaline conditions. The paromomycin detection limit (S/N = 3.3) was 2 μg mL-1 and the quantification limit (S/N = 10) was 6 μg mL-1. Coefficients of linear regression were higher than 0.99 for concentrations between 6.25 and 200 μg mL-1. The intra and inter-day precision (RSD) was less than 6.5%. The average recoveries were 97.53-102.01%. The proposed HPLC-PAD method presented advantageous performance characteristics and it can be considered suitable for the evaluation of paromomycin loaded nanogel formulation in ex vivo permeation and in vitro release studies.

  6. The Application of Contrast Media for In Vivo Feature Enhancement in X-Ray Computed Tomography of Soil-Grown Plant Roots.

    PubMed

    Keyes, Samuel D; Gostling, Neil J; Cheung, Jessica H; Roose, Tiina; Sinclair, Ian; Marchant, Alan

    2017-03-21

    The use of in vivo X-ray microcomputed tomography (μCT) to study plant root systems has become routine, but is often hampered by poor contrast between roots, soil, soil water, and soil organic matter. In clinical radiology, imaging of poorly contrasting regions is frequently aided by the use of radio-opaque contrast media. In this study, we present evidence for the utility of iodinated contrast media (ICM) in the study of plant root systems using μCT. Different dilutions of an ionic and nonionic ICM (Gastrografin 370 and Niopam 300) were perfused into the aerial vasculature of juvenile pea plants via a leaf flap (Pisum sativum). The root systems were imaged via μCT, and a variety of image-processing approaches used to quantify and compare the magnitude of the contrast enhancement between different regions. Though the treatment did not appear to significantly aid extraction of full root system architectures from the surrounding soil, it did allow the xylem and phloem units of seminal roots and the vascular morphology within rhizobial nodules to be clearly visualized. The nonionic, low-osmolality contrast agent Niopam appeared to be well tolerated by the plant, whereas Gastrografin showed evidence of toxicity. In summary, the use of iodine-based contrast media allows usually poorly contrasting root structures to be visualized nondestructively using X-ray μCT. In particular, the vascular structures of roots and rhizobial nodules can be clearly visualized in situ.

  7. Intact Endogenous Metabolite Analysis of Mice Liver by Probe Electrospray Ionization/Triple Quadrupole Tandem Mass Spectrometry and Its Preliminary Application to in Vivo Real-Time Analysis.

    PubMed

    Zaitsu, Kei; Hayashi, Yumi; Murata, Tasuku; Ohara, Tomomi; Nakagiri, Kenta; Kusano, Maiko; Nakajima, Hiroki; Nakajima, Tamie; Ishikawa, Tetsuya; Tsuchihashi, Hitoshi; Ishii, Akira

    2016-04-05

    Probe electrospray ionization (PESI) is a recently developed ionization technique that enables the direct detection of endogenous compounds like metabolites without sample preparation. In this study, we have demonstrated the first combination use of PESI with triple quadrupole tandem mass spectrometry (MS/MS), which was then applied to intact endogenous metabolite analysis of mice liver, achieving detection of 26 metabolites including amino acids, organic acids, and sugars. To investigate its practicality, metabolic profiles of control and CCl4-induced acute hepatic injury mouse model were measured by the developed method. Results showed clear separation of the two groups in score plots of principal component analysis and identified taurine as the primary contributor to group separation. The results were further validated by the established gas chromatography/MS/MS method, demonstrating the present method's usefulness. In addition, we preliminarily applied the method to real-time analysis of an intact liver of a living mouse. We successfully achieved monitoring of the real-time changes of two tricarboxylic acid cycle intermediates, α-ketoglutaric acid and fumaric acid, in the liver immediately after pyruvic acid injection via a cannulated tube to the portal vein. The present method achieved an intact analysis of metabolites in liver without sample preparation, and it also demonstrates future possibility to establish in vivo real-time metabolome analysis of living animals by PESI/MS/MS.

  8. The application of hyaluronic acid-derivatized carbon nanotubes in hematoporphyrin monomethyl ether-based photodynamic therapy for in vivo and in vitro cancer treatment

    PubMed Central

    Shi, Jinjin; Ma, Rourou; Wang, Lei; Zhang, Jing; Liu, Ruiyuan; Li, Lulu; Liu, Yan; Hou, Lin; Yu, Xiaoyuan; Gao, Jun; Zhang, Zhenzhong

    2013-01-01

    Carbon nanotubes (CNTs) have shown great potential in both photothermal therapy and drug delivery. In this study, a CNT derivative, hyaluronic acid-derivatized CNTs (HA-CNTs) with high aqueous solubility, neutral pH, and tumor-targeting activity, were synthesized and characterized, and then a new photodynamic therapy agent, hematoporphyrin monomethyl ether (HMME), was adsorbed onto the functionalized CNTs to develop HMME-HA-CNTs. Tumor growth inhibition was investigated both in vivo and in vitro by a combination of photothermal therapy and photodynamic therapy using HMME-HA-CNTs. The ability of HMME-HA-CNT nanoparticles to combine local specific photodynamic therapy with external near-infrared photothermal therapy significantly improved the therapeutic efficacy of cancer treatment. Compared with photodynamic therapy or photothermal therapy alone, the combined treatment demonstrated a synergistic effect, resulting in higher therapeutic efficacy without obvious toxic effects to normal organs. Overall, it was demonstrated that HMME-HA-CNTs could be successfully applied to photodynamic therapy and photothermal therapy simultaneously in future tumor therapy. PMID:23843694

  9. A new multiple quantum filter design procedure for use on strongly coupled spin systems found in vivo: its application to glutamate.

    PubMed

    Thompson, R B; Allen, P S

    1998-05-01

    A numerical procedure is outlined that is appropriate for the design of multiple quantum filter sequences targeted for the strongly coupled, multiple spin systems that occur in metabolites present in brain. The procedure uses numerical methods of solution of the density matrix equations, first, to establish the most appropriate resonance to target with the filter; second, to provide contour plots of a performance index of the filter in terms of critical sequence parameters; and third, to produce the response signals of the target and the background metabolites to the optimized filter. The procedure is exemplified for the AMNPQ spin system of the amino acid glutamate at a field strength of 3 T. The 2.3 ppm peak of the PQ multiplet of glutamate was identified as the target resonance, and the performance of the filter so derived was evaluated experimentally on phantom solutions and in human brain. These experiments clearly demonstrate that a linewidth of vivo, the 2.3 ppm peak of glutamate dominates the filter response and thereby removes a significant cause of uncertainty in measuring changes in glutamate by eliminating most of the background observed in unedited spectra obtained using PRESS or STEAM.

  10. In vivo application of chitosan to improve bioavailability of cyanocobalamin, a form of vitamin B12, following intraintestinal administration in rats.

    PubMed

    Goto, Yuko; Masuda, Ayumi; Aiba, Tetsuya

    2015-04-10

    The effect of chitosan on the intestinal absorption of cyanocobalamin (VB12), a stable form of vitamin B12, was investigated in vivo in rats, with the aim of improving the oral bioavailability of VB12 for anemia treatment in patients with gastrectomy. The bioavailability was evaluated based on the plasma concentration profile of VB12 following intraintestinal administration of the VB12 solution containing chitosan at various concentrations. The bioavailability of VB12 was 0.6±0.2% when the chitosan-free VB12 solution was administered, while it increased to 10.5±3.3% when chitosan was dissolved in the VB12 solution at a concentration of 1%. The bioavailability of VB12 increases with the chitosan concentration, in which chitosan seems to augment the amount of VB12 absorbed without affecting the absorption rate constant of VB12. It was also shown that the bioavailability of VB12 does not increase further when the degree of chitosan deacetylation is increased from 83 to 100% by substitutively employing the fully deacetylated chitosan. These findings suggest that the oral administration of VB12 with readily available chitosan may be a practical approach for anemia treatment in patients with gastrectomy.

  11. Very small embryonic-like stem-cell optimization of isolation protocols: an update of molecular signatures and a review of current in vivo applications

    PubMed Central

    Shin, Dong-Myung; Suszynska, Malwina; Mierzejewska, Kasia; Ratajczak, Janina; Ratajczak, Mariusz Z

    2013-01-01

    As the theory of stem cell plasticity was first proposed, we have explored an alternative hypothesis for this phenomenon: namely that adult bone marrow (BM) and umbilical cord blood (UCB) contain more developmentally primitive cells than hematopoietic stem cells (HSCs). In support of this notion, using multiparameter sorting we were able to isolate small Sca1+Lin−CD45− cells and CD133+Lin−CD45− cells from murine BM and human UCB, respectively, which were further enriched for the detection of various early developmental markers such as the SSEA antigen on the surface and the Oct4 and Nanog transcription factors in the nucleus. Similar populations of cells have been found in various organs by our team and others, including the heart, brain and gonads. Owing to their primitive cellular features, such as the high nuclear/cytoplasm ratio and the presence of euchromatin, they are called very small embryonic-like stem cells (VSELs). In the appropriate in vivo models, VSELs differentiate into long-term repopulating HSCs, mesenchymal stem cells (MSCs), lung epithelial cells, cardiomyocytes and gametes. In this review, we discuss the most recent data from our laboratory and other groups regarding the optimal isolation procedures and describe the updated molecular characteristics of VSELs. PMID:24232255

  12. Cultivation of keratinocytes and fibroblasts in a three-dimensional bovine collagen-elastin matrix (Matriderm®) and application for full thickness wound coverage in vivo.

    PubMed

    Killat, Jasper; Reimers, Kerstin; Choi, Claudia Y; Jahn, Sabrina; Vogt, Peter M; Radtke, Christine

    2013-07-11

    New skin substitutes for burn medicine or reconstructive surgery pose an important issue in plastic surgery. Matriderm® is a clinically approved three-dimensional bovine collagen-elastin matrix which is already used as a dermal substitute of full thickness burn wounds. The drawback of an avital matrix is the limited integration in full thickness skin defects, depending on the defect size. To further optimize this process, Matriderm® has also been studied as a matrix for tissue engineering of skin albeit long-term cultivation of the matrix with cells has been difficult. Cells have generally been seeded onto the matrix with high cell loss and minimal time-consuming migration. Here we developed a cell seeded skin equivalent after microtransfer of cells directly into the matrix. First, cells were cultured, and microinjected into Matriderm®. Then, cell viability in the matrix was determined by histology in vitro. As a next step, the skin substitute was applied in vivo into a full thickness rodent wound model. The wound coverage and healing was observed over a period of two weeks followed by histological examination assessing cell viability, proliferation and integration into the host. Viable and proliferating cells could be found throughout the entire matrix. The presented skin substitute resembles healthy skin in morphology and integrity. Based on this study, future investigations are planned to examine behaviour of epidermal stem cells injected into a collagen-elastin matrix under the aspects of establishment of stem cell niches and differentiation.

  13. Noninvasive photoacoustic microscopy of methemoglobin in vivo

    PubMed Central

    Tang, Min; Zhou, Yong; Zhang, Ruiying; Wang, Lihong V.

    2015-01-01

    Abstract. Due to the various causes of methemoglobinemia and its potential to be confused with other diseases, in vivo measurements of methemoglobin have significant applications in the clinic. Using photoacoustic microscopy (PAM), we quantified the average and the distributed percentage of methemoglobin both in vitro and in vivo. Based on the absorption spectra of methemoglobin, oxyhemoglobin, and deoxyhemoglobin, three wavelengths were chosen to differentiate methemoglobin from the others. The methemoglobin concentrations calculated from the photoacoustic signals agreed well with the preset concentrations. Then we imaged the methemoglobin percentage in microtubes that mimicked blood vessels. Average percentages calculated for five samples with different methemoglobin concentrations also agreed well with the preset values. Finally, we demonstrated the ability of PAM to detect methemoglobin in vivo in a mouse ear. Our results show that PAM can quantitatively image methemoglobin distribution in vivo. PMID:25760655

  14. EDITORIAL: Nanotechnology in vivo Nanotechnology in vivo

    NASA Astrophysics Data System (ADS)

    Demming, Anna

    2010-04-01

    -imaging labels [4]. A surface hydroxyl group renders silicon quantum dots soluble in water and the photoluminescence can be made stable with oxygen-passivation. In addition, researchers in Japan have demonstrated how the initially modest yield in the preparation of silicon quantum dots can be improved to tens of milligrams per batch, thus further promoting their application in bio-imaging [5]. In the search for non-toxic quantum dots, researchers at the Amrita Centre for Nanoscience in India have prepared heavy metal-free quantum dot bio-probes based on single phase ZnS [6]. The quantum dots are selectively doped with metals, transition metals and halides to provide tuneable luminescence properties, and they are surface conjugated with folic acid for cancer targeting. The quantum dots were demonstrated to be water-soluble, non-toxic in normal and cancer cell lines, and have bright, tuneable luminescence. So far most of the quantum dots developed for bio-imaging have had excitation and emission wavelengths in the visible spectrum, which is highly absorbed by tissue. This limits imaging with these quantum dots to superficial tissues. This week, researchers in China and the US reported work developing functionalized dots for in vivo tumour vasculature in the infrared part of the spectrum [7]. In addition the quantum dots were functionalised with glycine-aspartic acid (RGD) peptides, which target the vasculature of almost all types of growing tumours, unlike antibody- or aptamer-mediated targeting strategies that are specific to a particular cancer type. In this issue, researchers in China and the US demonstrate a novel type of contrast agent for ultrasonic tumour imaging [8]. Contrast-enhanced ultrasonic tumour imaging extends the diagnostic and imaging capabilities of traditional techniques. The use of nanoparticles as ultrasound contrast agents exploits the presence of open pores in the range of 380 to 780 nm in tumour blood vessels, which enhance the permeability and retention

  15. Silicate, borosilicate, and borate bioactive glass scaffolds with controllable degradation rate for bone tissue engineering applications. II. In vitro and in vivo biological evaluation.

    PubMed

    Fu, Qiang; Rahaman, Mohamed N; Bal, B Sonny; Bonewald, Lynda F; Kuroki, Keiichi; Brown, Roger F

    2010-10-01

    In Part I, the in vitro degradation of bioactivAR52115e glass scaffolds with a microstructure similar to that of human trabecular bone, but with three different compositions, was investigated as a function of immersion time in a simulated body fluid. The glasses consisted of a silicate (13-93) composition, a borosilicate composition (designated 13-93B1), and a borate composition (13-93B3), in which one-third or all of the SiO2 content of 13-93 was replaced by B2O3, respectively. This work is an extension of Part I, to investigate the effect of the glass composition on the in vitro response of osteogenic MLO-A5 cells to these scaffolds, and on the ability of the scaffolds to support tissue infiltration in a rat subcutaneous implantation model. The results of assays for cell viability and alkaline phosphatase activity showed that the slower degrading silicate 13-93 and borosilicate 13-93B1 scaffolds were far better than the borate 13-93B3 scaffolds in supporting cell proliferation and function. However, all three groups of scaffolds showed the ability to support tissue infiltration in vivo after implantation for 6 weeks. The results indicate that the required bioactivity and degradation rate may be achieved by substituting an appropriate amount of SiO2 in 13-93 glass with B2O3, and that these trabecular glass scaffolds could serve as substrates for the repair and regeneration of contained bone defects.

  16. Characterization of Fast-Scan Cyclic Voltammetric Electrodes Using Paraffin as an Effective Sealant with In Vitro and In Vivo Applications

    PubMed Central

    Ramsson, Eric S.; Cholger, Daniel; Dionise, Albert; Poirier, Nicholas; Andrus, Avery; Curtiss, Randi

    2015-01-01

    Fast-scan cyclic voltammetry (FSCV) is a powerful technique for measuring sub-second changes in neurotransmitter levels. A great time-limiting factor in the use of FSCV is the production of high-quality recording electrodes; common recording electrodes consist of cylindrical carbon fiber encased in borosilicate glass. When the borosilicate is heated and pulled, the molten glass ideally forms a tight seal around the carbon fiber cylinder. It is often difficult, however, to guarantee a perfect seal between the glass and carbon. Indeed, much of the time spent creating electrodes is in an effort to find a good seal. Even though epoxy resins can be useful in this regard, they are irreversible (seals are permanent), wasteful (epoxy cannot be reused once hardener is added), hazardous (hardeners are often caustic), and require curing. Herein we characterize paraffin as an electrode sealant for FSCV microelectrodes. Paraffin boasts the advantages of near-immediate curing times, simplicity in use, long shelf-life and stable waterproof seals capable of withstanding extended cycling. Borosilicate electrode tips were left intact or broken and dipped in paraffin embedding wax. Excess wax was removed from the carbon surface with xyelenes or by repeated cycling at an extended waveform (-0.4 to 1.4V, 400 V/s, 60 Hz). Then, the waveform was switched to a standard waveform (-0.4 to 1.3V, 400 V/s, 10 Hz) and cycled until stable. Wax-sealing does not inhibit electrode sensitivity, as electrodes detected linear changes in dopamine before and after wax (then xylenes) exposure. Paraffin seals are intact after 11 days of implantation in the mouse, and still capable of measuring transient changes in in vivo dopamine. From this it is clear that paraffin wax is an effective sealant for FSCV electrodes that provides a convenient substitute to epoxy sealants. PMID:26505195

  17. Application of fast-scan cyclic voltammetry for the in vivo characterization of optically evoked dopamine in the olfactory tubercle of the rat brain.

    PubMed

    Wakabayashi, Ken T; Bruno, Michael J; Bass, Caroline E; Park, Jinwoo

    2016-06-21

    The olfactory tubercle (OT), as a component of the ventral striatum, serves as an important multisensory integration center for reward-related processes in the brain. Recent studies show that dense dopaminergic innervation from the ventral tegmental area (VTA) into the OT may play an outsized role in disorders such as psychostimulant addiction and disorders of motivation, increasing recent scientific interest in this brain region. However, due to its anatomical inaccessibility, relative small size, and proximity to other dopamine-rich structures, neurochemical assessments using conventional methods cannot be readily employed. Here, we investigated dopamine (DA) regulation in the OT of urethane-anesthetized rats using in vivo fast-scan voltammetry (FSCV) coupled with carbon-fiber microelectrodes, following optogenetic stimulation of the VTA. The results were compared with DA regulation in the nucleus accumbens (NAc), a structure located adjacent to the OT and which also receives dense DA innervation from the VTA. FSCV coupled with optically evoked release allowed us to investigate the spatial distribution of DA in the OT and characterize OT DA dynamics (release and clearance) with subsecond temporal and micrometer spatial resolution for the first time. In this study, we demonstrated that DA transporters play an important role in regulating DA in the OT. However, the control of extracellular DA by uptake in the OT was less than in the NAc. The difference in DA transmission in the terminal fields of the OT and NAc may be involved in region-specific responses to drugs of abuse and contrasting roles in mediating reward-related behavior.

  18. Optimization of In Vivo Confocal Autofluorescence Imaging of the Ocular Fundus in Mice and Its Application to Models of Human Retinal Degeneration

    PubMed Central

    Issa, Peter Charbel; Singh, Mandeep S.; Lipinski, Daniel M.; Chong, Ngaihang V.; Delori, François C.; Barnard, Alun R.; MacLaren, Robert E.

    2012-01-01

    Purpose. To investigate the feasibility and to identify sources of experimental variability of quantitative and qualitative fundus autofluorescence (AF) assessment in mice. Methods. Blue (488 nm) and near-infrared (790 nm) fundus AF imaging was performed in various mouse strains and disease models (129S2, C57Bl/6, Abca4−/−, C3H-Pde6brd1/rd1, Rho−/−, and BALB/c mice) using a commercially available scanning laser ophthalmoscope. Gray-level analysis was used to explore factors influencing fundus AF measurements. Results. A contact lens avoided cataract development and resulted in consistent fundus AF recordings. Fundus illumination and magnification were sensitive to changes of the camera position. Standardized adjustment of the recorded confocal plane and consideration of the pupil area allowed reproducible recording of fundus AF from the retinal pigment epithelium with an intersession coefficient of repeatability of ±22%. Photopigment bleaching occurred during the first 1.5 seconds of exposure to 488 nm blue light (∼10 mW/cm2), resulting in an increase of fundus AF. In addition, there was a slight decrease in fundus AF during prolonged blue light exposure. Fundus AF at 488 nm was low in animals with an absence of a normal visual cycle, and high in BALB/c and Abca4−/− mice. Degenerative alterations in Pde6brd1/rd1 and Rho−/− were reminiscent of findings in human retinal disease. Conclusions. Investigation of retinal phenotypes in mice is possible in vivo using standardized fundus AF imaging. Correlation with postmortem analysis is likely to lead to further understanding of human disease phenotypes and of retinal degenerations in general. Fundus AF imaging may be useful as an outcome measure in preclinical trials, such as for monitoring effects aimed at lowering lipofuscin accumulation in the retinal pigment epithelium. PMID:22169101

  19. Local application of L- threo-hydroxyaspartate and malonate in rats in vivo induces rigidity and damages neurons of the substantia nigra, pars compacta.

    PubMed

    Loopuijt, L D

    2002-10-01

    In order to study neuronal death in Parkinson's disease, neurons of the substantia nigra, pars compacta in rats were exposed to elevated levels of glutamate and decreased levels of energy in vivo and consequences for behavior and neuronal morphology were studied. Thus, repeated local injections (9x) of the glutamate uptake inhibitor L- threo-hydroxyaspartate (L-THA; 833 microM in 0.3 microl) in the presence or absence of the succinate dehydrogenase inhibitor malonate (25 mM in 0.3 microl) were applied during three weeks. 24 h after injection, rigidity and catalepsy were measured, as well as, at the end of the three week period, locomotion, rearing and exploratory behavior. Thereafter, the cytoarchitecture of the substantia nigra, pars compacta of the brains of these rats was described. The L-THA plus malonate injected rats did not differ in their behavior from carrier injected rats, except for rigidity: their scores were higher than that of carrier and L-THA injected rats (P < 0.05), while L-THA injected rats did not differ from carrier injected controls. Observations on cresyl violet sections revealed, that, although many neurons with a shrunken nucleolus and faintly stained cytoplasm were present in both L-THA and L-THA plus malonate treated rats, the ventral edge of the substantia nigra, pars compacta containing modified cells was longer in L-THA plus malonate than in L-THA injected rats (P < 0.05). This indicates, that a minimum amount of damage to neurons in the ventral part of the substantia nigra, pars compacta might be required for the expression of rigidity.

  20. Relevance of sunscreen application method, visible light and sunlight intensity to free-radical protection: A study of ex vivo human skin.

    PubMed

    Haywood, Rachel

    2006-01-01

    With the continued rise in skin cancers worldwide there is a need for effective skin protection against sunlight damage. It was shown previously that sunscreens, which claimed UVA protection (SPF 20+), provided limited protection against UV-induced ascorbate radicals in human skin. Here the results of an electron spin resonance (ESR) investigation to irradiate ex vivo human skin with solar-simulated light are reported. The ascorbate radical signal in the majority of skin samples was directly proportional to the irradiance over relevant sunlight intensities (0.9-2.9 mW cm(-2)). Radical production (substratum-corneum) by UV (wavelengths < 400 nm) and visible components (> 400 nm) was approximately 67% and 33% respectively. Ascorbate radicals were in steady state concentration at low irradiance (approximately 1 mW cm(-2) equivalent to UK sunlight), but at higher irradiance (approximately 3 mW cm(-2)) decreased with time, suggesting ascorbate depletion. Radical protection by a four star-rated sunscreen (with UVA protection) was optimal when applied as a thin film (40-60% at 2 mg cm(-2)) but less so when rubbed into the skin (37% at 4 mg cm(-2) and no significant protection at 2 mg cm(-2)), possibly due to cream filling crevices, which reduced film thickness. This study validates ESR determinations of the ascorbate radical for quantitative protection measurements. Visible light contribution to radical production, and loss of protection when sunscreen is rubbed into skin, has implications for sunscreen design and use for the prevention of free-radical damage.

  1. Characterization of Fast-Scan Cyclic Voltammetric Electrodes Using Paraffin as an Effective Sealant with In Vitro and In Vivo Applications.

    PubMed

    Ramsson, Eric S; Cholger, Daniel; Dionise, Albert; Poirier, Nicholas; Andrus, Avery; Curtiss, Randi

    2015-01-01

    Fast-scan cyclic voltammetry (FSCV) is a powerful technique for measuring sub-second changes in neurotransmitter levels. A great time-limiting factor in the use of FSCV is the production of high-quality recording electrodes; common recording electrodes consist of cylindrical carbon fiber encased in borosilicate glass. When the borosilicate is heated and pulled, the molten glass ideally forms a tight seal around the carbon fiber cylinder. It is often difficult, however, to guarantee a perfect seal between the glass and carbon. Indeed, much of the time spent creating electrodes is in an effort to find a good seal. Even though epoxy resins can be useful in this regard, they are irreversible (seals are permanent), wasteful (epoxy cannot be reused once hardener is added), hazardous (hardeners are often caustic), and require curing. Herein we characterize paraffin as an electrode sealant for FSCV microelectrodes. Paraffin boasts the advantages of near-immediate curing times, simplicity in use, long shelf-life and stable waterproof seals capable of withstanding extended cycling. Borosilicate electrode tips were left intact or broken and dipped in paraffin embedding wax. Excess wax was removed from the carbon surface with xyelenes or by repeated cycling at an extended waveform (-0.4 to 1.4V, 400 V/s, 60 Hz). Then, the waveform was switched to a standard waveform (-0.4 to 1.3V, 400 V/s, 10 Hz) and cycled until stable. Wax-sealing does not inhibit electrode sensitivity, as electrodes detected linear changes in dopamine before and after wax (then xylenes) exposure. Paraffin seals are intact after 11 days of implantation in the mouse, and still capable of measuring transient changes in in vivo dopamine. From this it is clear that paraffin wax is an effective sealant for FSCV electrodes that provides a convenient substitute to epoxy sealants.

  2. Degradation pattern of porous CaCO3 and hydroxyapatite microspheres in vitro and in vivo for potential application in bone tissue engineering.

    PubMed

    Zhong, Qiwei; Li, Wenhua; Su, Xiuping; Li, Geng; Zhou, Ying; Kundu, Subhas C; Yao, Juming; Cai, Yurong

    2016-07-01

    Despite superior clinical handling, excellent biocompatibility, biodegradation property of calcium phosphate needs to be improved to coincide with the rate of new bone formation. In this study, spherical CaCO3 are fabricated in the presence of the silk sericin and then transformed into porous hydroxyapatite (HAP) microspheres via hydrothermal method. The degradation behavior of obtained CaCO3, HAP and their mixture is first investigated in vitro. The result demonstrates that the weight loss of HAP microspheres are almost 24.3% after immersing in pH 7.40 Tris-HCl buffer solution for 12 weeks, which is far slower than that of spherical CaCO3 (97.5%). The degradation speed of the mixtures depends on the proportion of CaCO3 and HAP. The mixture with higher content of CaCO3 possesses a quicker degradation speed. The obtained CaCO3 and HAP microspheres are injected into subcutaneous tissue of ICR mice with the assistance of sodium alginate. The result in vivo also shows an obvious difference of degradation speed between the obtained CaCO3 and HAP microspheres, implying it is feasible to modulate the degradation property of the mixture through changing the proportion of CaCO3 and HAP The good cytocompatibility of the two kinds of microspheres is proved and a mild inflammation response is observed only at early stage of implantation. The job offers a simple method to modify the degradation properties of biomaterial for potential use in bone tissue engineering.

  3. Fibrin gel-immobilized primary osteoblasts in calcium phosphate bone cement: in vivo evaluation with regard to application as injectable biological bone substitute.

    PubMed

    Kneser, U; Voogd, A; Ohnolz, J; Buettner, O; Stangenberg, L; Zhang, Y H; Stark, G B; Schaefer, D J

    2005-01-01

    Osteogenic injectable bone substitutes may be useful for many applications. We developed a novel injectable bone substitute based on osteoblast-fibrin glue suspension and calcium phosphate bone cement (BC). Human osteoblasts were isolated from trabecular bone samples and cultured under standard conditions. Osteoblasts were suspended in fibrinogen solution (FS). BC was cured with thrombin solution. 8 x 4 mm injectable bone discs were prepared using silicon molds and a custom-made applicator device. Discs containing BC, BC/FS, or BC/FS/osteoblasts were implanted subcutaneously into athymic nude mice. After 3, 9 and 24 weeks, specimens were explanted and subjected to morphologic and biomechanical evaluation. In vitro fibrin gel-embedded osteoblasts displayed a differentiated phenotype as evidenced by alkaline phosphatase, collagen type 1 and von Kossa stains. A proportion of osteoblasts appeared morphologically intact over a 3-day in vitro period following application into the BC. BC/FS and BC/FS/osteoblast discs were sparsely infiltrated with vascularized connective tissue. There was no bone formation in implants from all groups. However, positive von Kossa staining only in BC/FS/osteoblast groups suggests engraftment of at least some of the transplanted cells. Biomechanical evaluation demonstrated initial stability of the composites. Young's modulus and maximal load did not differ significantly in the BC/FS and BC/FS/osteoblast groups. The practicability of osteoblast-containing injectable bone could be demonstrated. The dense microstructure and the suboptimal initial vascularization of the composites may explain the lack of bone formation. Modifications with regard to enhanced osteoblast survival are mandatory for a possible application as injectable osteogenic bone replacement system.

  4. Local topological analysis at the distal radius by HR-pQCT: Application to in vivo bone microarchitecture and fracture assessment in the OFELY study.

    PubMed

    Pialat, J B; Vilayphiou, N; Boutroy, S; Gouttenoire, P J; Sornay-Rendu, E; Chapurlat, R; Peyrin, F

    2012-09-01

    High-resolution peripheral quantitative computed tomography (HR-pQCT) is an in-vivo technique used to analyze the distal radius and tibia. It provides a voxel size of 82μm. In addition to providing the usual microarchitecture parameters, local topological analysis (LTA) depicting rod- and plate-like trabeculae may improve prediction of bone fragility. Thirty-three women with prevalent wrist fractures from the OFELY cohort were compared with age-matched controls. Bone microarchitecture, including the structural model index (SMI), was assessed by HR-pQCT, and micro-finite element analysis (μFE) was computed on trabecular bone images of the distal radius (XtremeCT, Scanco Medical AG). A new LTA method was applied to label each bone voxel as a rod, plate or node. Then the bone volume fraction (BV/TV*), the rod, plate and node ratios over bone volume (RV/BV*, PV/BV*, NV/BV*) or total volume (RV/TV*, PV/TV*, NV/TV*) and the rod to plate ratio (RV/PV*) were calculated. Associations between LTA parameters and wrist fractures were computed in a conditional logistic regression model. Multivariate models were tested to predict the μFE-derived trabecular bone stiffness. RV/TV* (OR=4.41 [1.05-18.62]) and BV/TV* (OR=6.45 [1.06-39.3]), were significantly associated with prevalent wrist fracture, after adjustment for ultra distal radius aBMD. Multivariate linear models including PV/TV* or BV/TV*+RV/PV* predicted trabecular stiffness with the same magnitude as those including SMI. Conversion from plates into rods was significantly associated with bone fragility, with a negative correlation between RV/PV* and trabecular bone stiffness (r=-0.63, p<0.0001). We conclude that our local topological analysis is feasible for a voxel size of 82μm. After further validation, it may improve bone fragility description.

  5. In vitro and in vivo evaluation of biologically synthesized silver nanoparticles for topical applications: effect of surface coating and loading into hydrogels

    PubMed Central

    Mekkawy, Aml I; El-Mokhtar, Mohamed A; Nafady, Nivien A; Yousef, Naeima; Hamad, Mostafa A; El-Shanawany, Sohair M; Ibrahim, Ehsan H; Elsabahy, Mahmoud

    2017-01-01

    In the present study, silver nanoparticles (AgNPs) were synthesized via biological reduction of silver nitrate using extract of the fungus Fusarium verticillioides (green chemistry principle). The synthesized nanoparticles were spherical and homogenous in size. AgNPs were coated with polyethylene glycol (PEG) 6000, sodium dodecyl sulfate (SDS), and β-cyclodextrin (β-CD). The averaged diameters of AgNPs were 19.2±3.6, 13±4, 14±4.4, and 15.7±4.8 nm, for PEG-, SDS-, and β-CD-coated and uncoated AgNPs, respectively. PEG-coated AgNPs showed greater stability as indicated by a decreased sedimentation rate of particles in their water dispersions. The antibacterial activities of different AgNPs dispersions were investigated against Gram-positive bacteria (methicillin-sensitive and methicillin-resistant Staphylococcus aureus) and Gram-negative bacteria (Escherichia coli) by determination of the minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs). MIC and MBC values were in the range of 0.93–7.5 and 3.75–15 µg/mL, respectively, which were superior to the reported values in literature. AgNPs-loaded hydrogels were prepared from the coated-AgNPs dispersions using several gelling agents (sodium carboxymethyl cellulose [Na CMC], sodium alginate, hydroxypropylmethyl cellulose, Pluronic F-127, and chitosan). The prepared formulations were evaluated for their viscosity, spreadability, in vitro drug release, and antibacterial activity, and the combined effect of the type of surface coating and the polymers utilized to form the gel was studied. The in vivo wound-healing activity and antibacterial efficacy of Na CMC hydrogel loaded with PEG-coated AgNPs in comparison to the commercially available silver sulfadiazine cream (Dermazin®) were evaluated. Superior antibacterial activity and wound-healing capability, with normal skin appearance and hair growth, were demonstrated for the hydrogel formulations, as compared to the silver

  6. In vitro and in vivo evaluation of biologically synthesized silver nanoparticles for topical applications: effect of surface coating and loading into hydrogels.

    PubMed

    Mekkawy, Aml I; El-Mokhtar, Mohamed A; Nafady, Nivien A; Yousef, Naeima; Hamad, Mostafa A; El-Shanawany, Sohair M; Ibrahim, Ehsan H; Elsabahy, Mahmoud

    2017-01-01

    In the present study, silver nanoparticles (AgNPs) were synthesized via biological reduction of silver nitrate using extract of the fungus Fusarium verticillioides (green chemistry principle). The synthesized nanoparticles were spherical and homogenous in size. AgNPs were coated with polyethylene glycol (PEG) 6000, sodium dodecyl sulfate (SDS), and β-cyclodextrin (β-CD). The averaged diameters of AgNPs were 19.2±3.6, 13±4, 14±4.4, and 15.7±4.8 nm, for PEG-, SDS-, and β-CD-coated and uncoated AgNPs, respectively. PEG-coated AgNPs showed greater stability as indicated by a decreased sedimentation rate of particles in their water dispersions. The antibacterial activities of different AgNPs dispersions were investigated against Gram-positive bacteria (methicillin-sensitive and methicillin-resistant Staphylococcus aureus) and Gram-negative bacteria (Escherichia coli) by determination of the minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs). MIC and MBC values were in the range of 0.93-7.5 and 3.75-15 µg/mL, respectively, which were superior to the reported values in literature. AgNPs-loaded hydrogels were prepared from the coated-AgNPs dispersions using several gelling agents (sodium carboxymethyl cellulose [Na CMC], sodium alginate, hydroxypropylmethyl cellulose, Pluronic F-127, and chitosan). The prepared formulations were evaluated for their viscosity, spreadability, in vitro drug release, and antibacterial activity, and the combined effect of the type of surface coating and the polymers utilized to form the gel was studied. The in vivo wound-healing activity and antibacterial efficacy of Na CMC hydrogel loaded with PEG-coated AgNPs in comparison to the commercially available silver sulfadiazine cream (Dermazin(®)) were evaluated. Superior antibacterial activity and wound-healing capability, with normal skin appearance and hair growth, were demonstrated for the hydrogel formulations, as compared to the silver

  7. One-pot hydrothermal synthesis of lanthanide ions doped one-dimensional upconversion submicrocrystals and their potential application in vivo CT imaging

    NASA Astrophysics Data System (ADS)

    Gao, Guo; Zhang, Chunlei; Zhou, Zhijun; Zhang, Xin; Ma, Jiebing; Li, Chao; Jin, Weilin; Cui, Daxiang

    2012-12-01

    Multi-functional rare-earth Yb3+ and Ln3+ (Ln = Er, Tm and Ho) ions doped one-dimensional (1-D) upconversion submicrocrystals (NaYF4 and NaGdF4) possessing upconversion luminescence, biocompatibility and magnetic properties have been synthesized by a one-pot hydrothermal method. Rare-earth Yb3+ and Ln3+ ions doped NaYF4 microrods (~1 μm in diameter, 3-5 μm in length) exhibit porous properties, and the average pore sizes are ~28.2 nm. They show paramagnetism in the magnetic range of -60 to -2 kOe and 2 to 60 kOe at 300 K, and exhibit near superparamagnetic behaviour at the magnetic range of -2 to 2 kOe. Saturation magnetization was ~12.1 emu g-1 at 2 K. The Yb3+ and Ln3+ ions doped NaGdF4 submicrocrystals (~100 nm in diameter, 200-300 nm in length) show paramagnetism at 300 K, and exhibit superparamagnetic behaviour with a saturation magnetization of 129.2 emu g-1 at 2 K. The magnetic properties of Yb3+ and Ln3+ ions doped 1-D upconversion submicrocrystals indicate they can be used for drug targeting under a magnetic field. Their unique upconversion emission (green for Yb3+/Er3+ and blue for Yb3+/Tm3+) under 980 nm laser excitation indicate that they could be used for specific luminescent immunolabeling and imaging. MTT assays reveal that 1-D upconversion submicrocrystals have satisfactory bio-affinity, where the viability keeps in good state even at a concentration of 500 μg mL-1, which is much higher than the concentration usually used in cell labelling. Luminescent microscopy images show that the morphologies of the cytoskeleton and cell nucleus are well maintained after incubating different concentrations of 1-D upconversion submicrocrystals. After injecting upconversion submicrocrystals into the mice (tumor sites or back normal tissue), a clearly distinguished CT signal was observed, indicating the synthesized 1-D submicrocrystals are effective for CT imaging in vivo.Multi-functional rare-earth Yb3+ and Ln3+ (Ln = Er, Tm and Ho) ions doped one-dimensional (1

  8. THz Medical Imaging: in vivo Hydration Sensing

    PubMed Central

    Taylor, Zachary D.; Singh, Rahul S.; Bennett, David B.; Tewari, Priyamvada; Kealey, Colin P.; Bajwa, Neha; Culjat, Martin O.; Stojadinovic, Alexander; Lee, Hua; Hubschman, Jean-Pierre; Brown, Elliott R.; Grundfest, Warren S.

    2015-01-01

    The application of THz to medical imaging is experiencing a surge in both interest and federal funding. A brief overview of the field is provided along with promising and emerging applications and ongoing research. THz imaging phenomenology is discussed and tradeoffs are identified. A THz medical imaging system, operating at ~525 GHz center frequency with ~125 GHz of response normalized bandwidth is introduced and details regarding principles of operation are provided. Two promising medical applications of THz imaging are presented: skin burns and cornea. For burns, images of second degree, partial thickness burns were obtained in rat models in vivo over an 8 hour period. These images clearly show the formation and progression of edema in and around the burn wound area. For cornea, experimental data measuring the hydration of ex vivo porcine cornea under drying is presented demonstrating utility in ophthalmologic applications. PMID:26085958

  9. Exploring in vivo violacein biosynthesis by application of multivariate curve resolution on fused UV-VIS absorption, fluorescence, and liquid chromatography-mass spectrometry data.

    PubMed

    Dantas, Clecio; Tauler, Romà; Ferreira, Márcia Miguel Castro

    2013-02-01

    In this work, the application of multivariate curve resolution-alternating least squares (MCR-ALS) is proposed for extracting information from multitechnique fused multivariate data (UV-VIS absorption, fluorescence, and liquid chromatography-mass spectrometry) gathered during the biosynthesis of violacein pigment. Experimental data sets were pretreated and arranged in a row-wise augmented data matrix before their chemometric investigation. Five different chemical components were resolved. Kinetic and spectral information about these components were obtained and their relationship with violacein biosynthesis was established. Three new chemical compounds with molar masses of 453, 465, and 479 u, until now not reported in the literature, were identified and proposed as intermediates in the biosynthesis of other indolocarbazoles. The precursor (tryptophan), one intermediate (deoxyviolacein), and the final product (violacein) of violacein biosynthesis were identified and characterized using the proposed approach. The chemometric procedure based on the MCR-ALS method has proved to be a powerful tool to investigate violacein biosynthesis and its application can be easily extended to the study of other bioprocesses.

  10. Radical protection in the visible and infrared by a hyperforin-rich cream--in vivo versus ex vivo methods.

    PubMed

    Arndt, Sophia; Haag, Stefan F; Kleemann, Anke; Lademann, Juergen; Meinke, Martina C

    2013-05-01

    The formation of radicals plays an important role in the development of atopic eczema or barrier-disrupted skin. We evaluated the radical scavenging effect of a cream containing a Hypericum perforatum extract rich in hyperforin in a double-blind placebo-controlled study on 11 healthy volunteers. Electron paramagnetic resonance spectroscopy was applied to determine radical formation during VIS/NIR irradiation of the inner forearm. The results were compared to ex vivo investigations on excised porcine ear skin after a single application of the creams. The non-treated skin was measured as control. The absolute values and the kinetics are not comparable for ex vivo and in vivo radical formation. Whereas in vivo, the radical production decreases with time, it remains stable ex vivo over the investigated timescale. Nevertheless, ex vivo methods could be developed to estimate the protection efficiency of creams. In vivo as well as ex vivo, the radical formation could be reduced by almost 80% when applying the hyperforin-rich cream onto the skin, whereas placebo resulted in about 60%. In vivo, a daylong protection effect could be validated after a 4-week application time of the cream indicating that a regular application is necessary to obtain the full effect.

  11. Optimization of modified scanning protocol based correlation mapping optical coherence tomography at 200 kHz VCSEL source for in vivo microcirculation imaging applications

    NASA Astrophysics Data System (ADS)

    Lal, Cerine; McGrath, James; Subhash, Hrebesh; Leahy, Martin

    2016-03-01

    Optical Coherence Tomography (OCT) is a non-invasive 3 dimensional optical imaging modality that enables high resolution cross sectional imaging in biological tissues and materials. Unlike other 3 D medical imaging modalities, OCT provides high axial and lateral resolution combined with high sensitivity, imaging depth and wide field of view which makes it suitable for wide variety of medical imaging applications1. Apart from analysing the morphological characteristics of the biological organs with micron scale axial and lateral resolution, OCT also provides functional information from the biological sample. Among the various functional extensions of OCT, angiographic OCT that enables visualization of lumens of blood vessels from the acquired OCT B scan images has been of high research interest in the recent past.

  12. Evaluation of wrought Zn-Al alloys (1, 3, and 5 wt % Al) through mechanical and in vivo testing for stent applications.

    PubMed

    Bowen, Patrick K; Seitz, Jan-Marten; Guillory, Roger J; Braykovich, Jacob P; Zhao, Shan; Goldman, Jeremy; Drelich, Jaroslaw W

    2017-01-27

    Special high grade zinc and wrought zinc-aluminum (Zn-Al) alloys containing up to 5.5 wt % Al were processed, characterized, and implanted in rats in search of a new family of alloys with possible applications as bioabsorbable endovascular stents. These materials retained roll-induced texture with an anisotropic distribution of the second-phase Al precipitates following hot-rolling, and changes in lattice parameters were observed with respect to Al content. Mechanical properties for the alloys fell roughly in line with strength (190-240 MPa yield strength; 220-300 MPa ultimate tensile strength) and elongation (15-30%) benchmarks, and favorable elastic ranges (0.19-0.27%) were observed. Intergranular corrosion was observed during residence of Zn-Al alloys in the murine aorta, suggesting a different corrosion mechanism than that of pure zinc. This mode of failure needs to be avoided for stent applications because the intergranular corrosion caused cracking and fragmentation of the implants, although the composition of corrosion products was roughly identical between non- and Al-containing materials. In spite of differences in corrosion mechanisms, the cross-sectional reduction of metals in murine aorta was nearly identical at 30-40% and 40-50% after 4.5 and 6 months, respectively, for pure Zn and Zn-Al alloys. Histopathological analysis and evaluation of arterial tissue compatibility around Zn-Al alloys failed to identify areas of necrosis, though both chronic and acute inflammatory indications were present. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2017.

  13. Application of the principles of systems biology and Wiener's cybernetics for analysis of regulation of energy fluxes in muscle cells in vivo.

    PubMed

    Guzun, Rita; Saks, Valdur

    2010-03-08

    The mechanisms of regulation of respiration and energy fluxes in the cells are analyzed based on the concepts of systems biology, non-equilibrium steady state kinetics and applications of Wiener's cybernetic principles of feedback regulation. Under physiological conditions cardiac function is governed by the Frank-Starling law and the main metabolic characteristic of cardiac muscle cells is metabolic homeostasis, when both workload and respiration rate can be changed manifold at constant intracellular level of phosphocreatine and ATP in the cells. This is not observed in skeletal muscles. Controversies in theoretical explanations of these observations are analyzed. Experimental studies of permeabilized fibers from human skeletal muscle vastus lateralis and adult rat cardiomyocytes showed that the respiration rate is always an apparent hyperbolic but not a sigmoid function of ADP concentration. It is our conclusion that realistic explanations of regulation of energy fluxes in muscle cells require systemic approaches including application of the feedback theory of Wiener's cybernetics in combination with detailed experimental research. Such an analysis reveals the importance of limited permeability of mitochondrial outer membrane for ADP due to interactions of mitochondria with cytoskeleton resulting in quasi-linear dependence of respiration rate on amplitude of cyclic changes in cytoplasmic ADP concentrations. The system of compartmentalized creatine kinase (CK) isoenzymes functionally coupled to ANT and ATPases, and mitochondrial-cytoskeletal interactions separate energy fluxes (mass and energy transfer) from signalling (information transfer) within dissipative metabolic structures - intracellular energetic units (ICEU). Due to the non-equilibrium state of CK reactions, intracellular ATP utilization and mitochondrial ATP regeneration are interconnected by the PCr flux from mitochondria. The feedback regulation of respiration occurring via cyclic fluctuations of

  14. Application of the Principles of Systems Biology and Wiener’s Cybernetics for Analysis of Regulation of Energy Fluxes in Muscle Cells in Vivo

    PubMed Central

    Guzun, Rita; Saks, Valdur

    2010-01-01

    The mechanisms of regulation of respiration and energy fluxes in the cells are analyzed based on the concepts of systems biology, non-equilibrium steady state kinetics and applications of Wiener’s cybernetic principles of feedback regulation. Under physiological conditions cardiac function is governed by the Frank-Starling law and the main metabolic characteristic of cardiac muscle cells is metabolic homeostasis, when both workload and respiration rate can be changed manifold at constant intracellular level of phosphocreatine and ATP in the cells. This is not observed in skeletal muscles. Controversies in theoretical explanations of these observations are analyzed. Experimental studies of permeabilized fibers from human skeletal muscle vastus lateralis and adult rat cardiomyocytes showed that the respiration rate is always an apparent hyperbolic but not a sigmoid function of ADP concentration. It is our conclusion that realistic explanations of regulation of energy fluxes in muscle cells require systemic approaches including application of the feedback theory of Wiener’s cybernetics in combination with detailed experimental research. Such an analysis reveals the importance of limited permeability of mitochondrial outer membrane for ADP due to interactions of mitochondria with cytoskeleton resulting in quasi-linear dependence of respiration rate on amplitude of cyclic changes in cytoplasmic ADP concentrations. The system of compartmentalized creatine kinase (CK) isoenzymes functionally coupled to ANT and ATPases, and mitochondrial-cytoskeletal interactions separate energy fluxes (mass and energy transfer) from signalling (information transfer) within dissipative metabolic structures – intracellular energetic units (ICEU). Due to the non-equilibrium state of CK reactions, intracellular ATP utilization and mitochondrial ATP regeneration are interconnected by the PCr flux from mitochondria. The feedback regulation of respiration occurring via cyclic fluctuations

  15. A ruthenium(II) complex as turn-on Cu(II) luminescent sensor based on oxidative cyclization mechanism and its application in vivo

    PubMed Central

    Zhang, Yunfei; Liu, Zonglun; Yang, Kui; Zhang, Yi; Xu, Yongqian; Li, Hongjuan; Wang, Chaoxia; Lu, Aiping; Sun, Shiguo

    2015-01-01

    Copper ions play a vital role in a variety of fundamental physiological processes not only in human beings and plants, but also for extensive insects and microorganisms. In this paper, a novel water-soluble ruthenium(II) complex as a turn-on copper(II) ions luminescent sensor based on o-(phenylazo)aniline was designed and synthesized. The azo group would undergo a specific oxidative cyclization reaction with copper(II) ions and turn into high luminescent benzotriazole, triggering significant luminescent increasements which were linear to the concentrations of copper(II) ions. The sensor distinguished by its high sensitivity (over 80-fold luminescent switch-on response), good selectivity (the changes of the emission intensity in the presence of other metal ions or amino acids were negligible) and low detection limit (4.42 nM) in water. Moreover, the copper(II) luminescent sensor exhibited good photostability under light irradiation. Furthermore, the applicability of the proposed sensor in biological samples assay was also studied and imaged copper(II) ions in living pea aphids successfully. PMID:25640000

  16. Application of structure-based drug design and parallel chemistry to identify selective, brain penetrant, in vivo active phosphodiesterase 9A inhibitors.

    PubMed

    Claffey, Michelle M; Helal, Christopher J; Verhoest, Patrick R; Kang, Zhijun; Fors, Kristina S; Jung, Stanley; Zhong, Jiaying; Bundesmann, Mark W; Hou, Xinjun; Lui, Shenping; Kleiman, Robin J; Vanase-Frawley, Michelle; Schmidt, Anne W; Menniti, Frank; Schmidt, Christopher J; Hoffman, William E; Hajos, Mihaly; McDowell, Laura; O'Connor, Rebecca E; Macdougall-Murphy, Mary; Fonseca, Kari R; Becker, Stacey L; Nelson, Frederick R; Liras, Spiros

    2012-11-08

    Phosphodiesterase 9A inhibitors have shown activity in preclinical models of cognition with potential application as novel therapies for treating Alzheimer's disease. Our clinical candidate, PF-04447943 (2), demonstrated acceptable CNS permeability in rats with modest asymmetry between central and peripheral compartments (free brain/free plasma = 0.32; CSF/free plasma = 0.19) yet had physicochemical properties outside the range associated with traditional CNS drugs. To address the potential risk of restricted CNS penetration with 2 in human clinical trials, we sought to identify a preclinical candidate with no asymmetry in rat brain penetration and that could advance into development. Merging the medicinal chemistry strategies of structure-based design with parallel chemistry, a novel series of PDE9A inhibitors was identified that showed improved selectivity over PDE1C. Optimization afforded preclinical candidate 19 that demonstrated free brain/free plasma ≥ 1 in rat and reduced microsomal clearance along with the ability to increase cyclic guanosine monophosphosphate levels in rat CSF.

  17. Long-Term Cultures of Human Cornea Limbal Explants Form 3D Structures Ex Vivo - Implications for Tissue Engineering and Clinical Applications.

    PubMed

    Szabó, Dóra Júlia; Noer, Agate; Nagymihály, Richárd; Josifovska, Natasha; Andjelic, Sofija; Veréb, Zoltán; Facskó, Andrea; Moe, Morten C; Petrovski, Goran

    2015-01-01

    Long-term cultures of cornea limbal epithelial stem cells (LESCs) were developed and characterized for future tissue engineering and clinical applications. The limbal tissue explants were cultivated and expanded for more than 3 months in medium containing serum as the only growth supplement and without use of scaffolds. Viable 3D cell outgrowth from the explants was observed within 4 weeks of cultivation. The outgrowing cells were examined by immunofluorescent staining for putative markers of stemness (ABCG2, CK15, CK19 and Vimentin), proliferation (p63α, Ki-67), limbal basal epithelial cells (CK8/18) and differentiated cornea epithelial cells (CK3 and CK12). Morphological and immunostaining analyses revealed that long-term culturing can form stratified 3D tissue layers with a clear extracellular matrix deposition and organization (collagen I, IV and V). The LESCs showed robust expression of p63α, ABCG2, and their surface marker fingerprint (CD117/c-kit, CXCR4, CD146/MCAM, CD166/ALCAM) changed over time compared to short-term LESC cultures. Overall, we provide a model for generating stem cell-rich, long-standing 3D cultures from LESCs which can be used for further research purposes and clinical transplantation.

  18. Long-Term Cultures of Human Cornea Limbal Explants Form 3D Structures Ex Vivo – Implications for Tissue Engineering and Clinical Applications

    PubMed Central

    Nagymihály, Richárd; Josifovska, Natasha; Andjelic, Sofija; Veréb, Zoltán; Facskó, Andrea; Moe, Morten C.; Petrovski, Goran

    2015-01-01

    Long-term cultures of cornea limbal epithelial stem cells (LESCs) were developed and characterized for future tissue engineering and clinical applications. The limbal tissue explants were cultivated and expanded for more than 3 months in medium containing serum as the only growth supplement and without use of scaffolds. Viable 3D cell outgrowth from the explants was observed within 4 weeks of cultivation. The outgrowing cells were examined by immunofluorescent staining for putative markers of stemness (ABCG2, CK15, CK19 and Vimentin), proliferation (p63α, Ki-67), limbal basal epithelial cells (CK8/18) and differentiated cornea epithelial cells (CK3 and CK12). Morphological and immunostaining analyses revealed that long-term culturing can form stratified 3D tissue layers with a clear extracellular matrix deposition and organization (collagen I, IV and V). The LESCs showed robust expression of p63α, ABCG2, and their surface marker fingerprint (CD117/c-kit, CXCR4, CD146/MCAM, CD166/ALCAM) changed over time compared to short-term LESC cultures. Overall, we provide a model for generating stem cell-rich, long-standing 3D cultures from LESCs which can be used for further research purposes and clinical transplantation. PMID:26580800

  19. A ruthenium(II) complex as turn-on Cu(II) luminescent sensor based on oxidative cyclization mechanism and its application in vivo

    NASA Astrophysics Data System (ADS)

    Zhang, Yunfei; Liu, Zonglun; Yang, Kui; Zhang, Yi; Xu, Yongqian; Li, Hongjuan; Wang, Chaoxia; Lu, Aiping; Sun, Shiguo

    2015-02-01

    Copper ions play a vital role in a variety of fundamental physiological processes not only in human beings and plants, but also for extensive insects and microorganisms. In this paper, a novel water-soluble ruthenium(II) complex as a turn-on copper(II) ions luminescent sensor based on o-(phenylazo)aniline was designed and synthesized. The azo group would undergo a specific oxidative cyclization reaction with copper(II) ions and turn into high luminescent benzotriazole, triggering significant luminescent increasements which were linear to the concentrations of copper(II) ions. The sensor distinguished by its high sensitivity (over 80-fold luminescent switch-on response), good selectivity (the changes of the emission intensity in the presence of other metal ions or amino acids were negligible) and low detection limit (4.42 nM) in water. Moreover, the copper(II) luminescent sensor exhibited good photostability under light irradiation. Furthermore, the applicability of the proposed sensor in biological samples assay was also studied and imaged copper(II) ions in living pea aphids successfully.

  20. Enhanced in Vitro and in Vivo Performance of Mg-Zn-Y-Nd Alloy Achieved with APTES Pretreatment for Drug-Eluting Vascular Stent Application.

    PubMed

    Liu, Jing; Zheng, Bo; Wang, Pei; Wang, Xingang; Zhang, Bin; Shi, Qiuping; Xi, Tingfei; Chen, Ming; Guan, Shaokang

    2016-07-20

    Bioabsorbable magnesium alloys are becoming prominent as temporary functional implants, as they avoid the risks generated by permanent metallic implants such as persistent inflammation and late restenosis. Nevertheless, the overfast corrosion of Mg alloys under physiological conditions hinders their wider application as medical implant materials. Here we investigate a simple one-step process to introduce a cross-linked 3-amino-propyltrimethoxysilane (APTES) silane physical barrier layer on the surface of Mg-Zn-Y-Nd alloys prior to electrostatic spraying with rapamycin-eluting poly(lactic-co-glycolic acid) (PLGA) layer. Surface microstructure was characterized by scanning electron microscope and Fourier transform infrared spectroscopy. Nanoscratch test verified the superior adhesion strength of PLGA coating in the group pretreated with APTES. Electrochemical tests combined with long-term immersion results suggested that the preferable in vitro anticorrosion behavior could be achieved by dense APTES barrier. Cell morphology and proliferation data demonstrated that APTES pretreated group resulted in remarkably preferable compatibility for both human umbilical vein endothelial cells and vascular smooth muscle cells. On the basis of excellent in vitro mechenical property, the animal study on the APTES pretreated Mg-Zn-Y-Nd stent implanted into porcine coronary arteries confirmed benign tissue compatibility as well as re-endothelialization without thrombogenesis or in-stent restenosis at six-month followup.

  1. Cubosomes for in vivo fluorescence lifetime imaging

    NASA Astrophysics Data System (ADS)

    Biffi, Stefania; Andolfi, Laura; Caltagirone, Claudia; Garrovo, Chiara; Falchi, Angela M.; Lippolis, Vito; Lorenzon, Andrea; Macor, Paolo; Meli, Valeria; Monduzzi, Maura; Obiols-Rabasa, Marc; Petrizza, Luca; Prodi, Luca; Rosa, Antonella; Schmidt, Judith; Talmon, Yeshayahu; Murgia, Sergio

    2017-02-01

    Herein we provided the first proof of principle for in vivo fluorescence optical imaging application using monoolein-based cubosomes in a healthy mouse animal model. This formulation, administered at a non-cytotoxic concentration, was capable of providing both exogenous contrast for NIR fluorescence imaging with very high efficiency and chemospecific information upon lifetime analysis. Time-resolved measurements of fluorescence after the intravenous injection of cubosomes revealed that the dye rapidly accumulated mainly in the liver, while lifetimes profiles obtained in vivo allowed for discriminating between free dye or dye embedded within the cubosome nanostructure after injection.

  2. Cubosomes for in vivo fluorescence lifetime imaging.

    PubMed

    Biffi, Stefania; Andolfi, Laura; Caltagirone, Claudia; Garrovo, Chiara; Falchi, Angela M; Lippolis, Vito; Lorenzon, Andrea; Macor, Paolo; Meli, Valeria; Monduzzi, Maura; Obiols-Rabasa, Marc; Petrizza, Luca; Prodi, Luca; Rosa, Antonella; Schmidt, Judith; Talmon, Yeshayahu; Murgia, Sergio

    2017-02-03

    Herein we provided the first proof of principle for in vivo fluorescence optical imaging application using monoolein-based cubosomes in a healthy mouse animal model. This formulation, administered at a non-cytotoxic concentration, was capable of providing both exogenous contrast for NIR fluorescence imaging with very high efficiency and chemospecific information upon lifetime analysis. Time-resolved measurements of fluorescence after the intravenous injection of cubosomes revealed that the dye rapidly accumulated mainly in the liver, while lifetimes profiles obtained in vivo allowed for discriminating between free dye or dye embedded within the cubosome nanostructure after injection.

  3. Ex vivo lung perfusion.

    PubMed

    Reeb, Jeremie; Cypel, Marcelo

    2016-03-01

    Lung transplantation is an established life-saving therapy for patients with end-stage lung disease. Unfortunately, greater success in lung transplantation is hindered by a shortage of lung donors and the relatively poor early-, mid-, and long-term outcomes associated with severe primary graft dysfunction. Ex vivo lung perfusion has emerged as a modern preservation technique that allows for a more accurate lung assessment and improvement in lung quality. This review outlines the: (i) rationale behind the method; (ii) techniques and protocols; (iii) Toronto ex vivo lung perfusion method; (iv) devices available; and (v) clinical experience worldwide. We also highlight the potential of ex vivo lung perfusion in leading a new era of lung preservation.

  4. IN VIVO EVALUATION OF SKIN IRRITATION POTENTIAL, MELASMA AND SEBUM CONTENT FOLLOWING LONG TERM APPLICATION OF SKIN CARE CREAM IN HEALTHY ADULTS, USING NON-INVASIVE BIOMETROLOGICAL TECHNIQUES.

    PubMed

    Arshad, Atif I; Khan, Shoaib H M; Akhtar, Naveed; Mahmood, Asif; Sarfraz, Rai Muhammad

    2016-01-01

    The present investigation was conducted to evaluate non-invasively, various functional skin parameters i.e., irritation potential, melasma and sebum contents following long term application of topical cream (w/o) loaded with 2% methanolic extract of Ananas comosus L. versus placebo control (base) in healthy adults. Healthy human volunteers (n = 11, aged 20-30 years) were recruited for investigation and written informed consent was taken from each volunteer. In this single blinded study every volunteer applied formulation on one side of face and placebo on the other side of face twice daily for a period of 12 weeks (three months). Different skin parameters i.e., skin irritancy, melasma, and sebum contents were measured on both sides of face at baseline and after two weeks interval, using photometric device Mexameter and Sebumeter in a draught free room with modulated conditions of temperature (22-25°C) and humidity (55-60%). It was evident from the results that no primary skin irritancy was observed with patch test. Besides, statistical interpretation indicates that treatment with formulation is superior to placebo because it significantly (p ≤ 0.05) reduced the skin irritancy, melasma and sebum secretions throughout the study and reaching maximum -20.76 ± 0.89, -54.2 ± 0.37 and -40.71 ± 0.75%, respectively, at the end of study period. Antioxidant activity of extract was 92% compared to standard antioxidant. Conclusively, active cream loaded with fruit extract was well tolerated by all the volunteers and suitable to treat contact dermatitis, greasy skin, acne and seborrheic dermatitis and augmenting beauty and attraction by depigmentation of human skin. So, in the future, there is need to clinically evaluate these formulations in patients with compromised skin functions i.e., contact dermatitis, melasma, and acne vulgaris in order to explore the actual potential of this fruit.

  5. Ex Vivo Application of Secreted Metabolites Produced by Soil-Inhabiting Bacillus spp. Efficiently Controls Foliar Diseases Caused by Alternaria spp.

    PubMed Central

    El-Sayed, Ashraf S. A.; Patel, Jaimin S.; Green, Kari B.; Ali, Mohammad; Brennan, Mary; Norman, David

    2015-01-01

    Bacterial biological control agents (BCAs) are largely used as live products to control plant pathogens. However, due to variable environmental and ecological factors, live BCAs usually fail to produce desirable results against foliar pathogens. In this study, we investigated the potential of cell-free culture filtrates of 12 different bacterial BCAs isolated from flower beds for controlling foliar diseases caused by Alternaria spp. In vitro studies showed that culture filtrates from two isolates belonging to Bacillus subtilis and Bacillus amyloliquefaciens displayed strong efficacy and potencies against Alternaria spp. The antimicrobial activity of the culture filtrate of these two biological control agents was effective over a wider range of pH (3.0 to 9.0) and was not affected by autoclaving or proteolysis. Comparative liquid chromatography-mass spectrometry (LC-MS) analyses showed that a complex mixture of cyclic lipopeptides, primarily of the fengycin A and fengycin B families, was significantly higher in these two BCAs than inactive Bacillus spp. Interaction studies with mixtures of culture filtrates of these two species revealed additive activity, suggesting that they produce similar products, which was confirmed by LC-tandem MS analyses. In in planta pre- and postinoculation trials, foliar application of culture filtrates of B. subtilis reduced lesion sizes and lesion frequencies caused by Alternaria alternata by 68 to 81%. Taken together, our studies suggest that instead of live bacteria, culture filtrates of B. subtilis and B. amyloliquefaciens can be applied either individually or in combination for controlling foliar diseases caused by Alternaria species. PMID:26519395

  6. Development of In Vitro-In Vivo Correlation for Amorphous Solid Dispersion Immediate-Release Suvorexant Tablets and Application to Clinically Relevant Dissolution Specifications and In-Process Controls.

    PubMed

    Kesisoglou, Filippos; Hermans, Andre; Neu, Colleen; Yee, Ka Lai; Palcza, John; Miller, Jessica

    2015-09-01

    Although in vitro-in vivo correlations (IVIVCs) are commonly pursued for modified-release products, there are limited reports of successful IVIVCs for immediate-release (IR) formulations. This manuscript details the development of a Multiple Level C IVIVC for the amorphous solid dispersion formulation of suvorexant, a BCS class II compound, and its application to establishing dissolution specifications and in-process controls. Four different 40 mg batches were manufactured at different tablet hardnesses to produce distinct dissolution profiles. These batches were evaluated in a relative bioavailability clinical study in healthy volunteers. Although no differences were observed for the total exposure (AUC) of the different batches, a clear relationship between dissolution and Cmax was observed. A validated Multiple Level C IVIVC against Cmax was developed for the 10, 15, 20, 30, and 45 min dissolution time points and the tablet disintegration time. The relationship established between tablet tensile strength and dissolution was subsequently used to inform suitable tablet hardness ranges within acceptable Cmax limits. This is the first published report for a validated Multiple Level C IVIVC for an IR solid dispersion formulation demonstrating how this approach can facilitate Quality by Design in formulation development and help toward clinically relevant specifications and in-process controls.

  7. Improving in vivo calibration phantoms

    SciTech Connect

    Lynch, T.P.; Olsen, P.C.

    1991-10-01

    Anthropomorphic phantoms have been the basis for quantification of radioactive material in the body using in vivo measurements. The types of phantoms used and the degree of anthropomorphic detail vary depending on the counting application, the radioactive material to be measured, phantom availability and cost. Consequently, measurement results for the same types of radioactive material from different facilities are not always comparable. At a February 1990 meeting at the National Institute of Standards and Technology (NIST) the need to develop the gold standards'' or primary reference standards for in vivo phantoms was discussed in detail. The consensus of the attendees at the meeting was that the state of the art in phantoms was adequate as a starting point and that there was no need to start phantom development from scratch. In particular, the torso phantom developed at the Lawrence Livermore National Laboratory (LLNL) and its commercial progeny, the bottle manikin absorption (BOMAB) phantom and the American National Standards Institute (ANSI) Standard N44.3 thyroid phantom, were identified as the starting points for the development of the primary reference standards. Working groups at the meeting subsequently recommended design improvements for the existing phantom designs. The implementation of these recommendations is the subject of this paper.

  8. Noninvasive photoacoustic microscopy of methemoglobin in vivo

    NASA Astrophysics Data System (ADS)

    Tang, Min; Zhou, Yong; Zhang, Ruiying; Wang, Lihong V.

    2015-03-01

    Various causes can lead to methemoglobinemia, and it has the potential to be confused with other diseases. In vivo measurements of methemoglobin have significant applications in the clinics. We quantified the average and the distributed percentage of methemoglobin both in vitro and in vivo using photoacoustic microscopy (PAM). Based on the absorption spectra of methemoglobin, oxyhemoglobin, and deoxyhemoglobin, three wavelengths were chosen to differentiate methemoglobin from the others. We imaged the methemoglobin percentage in microtubes that mimicked blood vessels as a phantom experiment. The methemoglobin concentrations calculated from the photoacoustic signals were in accordance with the preset concentrations. We also demonstrated the ability of PAM to quantitatively image methemoglobin distribution in vivo in a mouse ear.

  9. Bioluminescence imaging of myeloperoxidase activity in vivo

    PubMed Central

    Gross, Shimon; Gammon, Seth T; Moss, Britney L; Rauch, Daniel; Harding, John; Heinecke, Jay W; Ratner, Lee; Piwnica-Worms, David

    2010-01-01

    The myeloperoxidase (MPO) system of activated phagocytes is central to normal host defense mechanisms, and dysregulated MPO contributes to the pathogenesis of inflammatory disease states ranging from atherosclerosis to cancer. Here we show that upon systemic administration, the small molecule luminol enables noninvasive bioluminescence imaging (BLI) of MPO activity in vivo. Luminol-BLI allowed quantitative longitudinal monitoring of MPO activity in animal models of acute dermatitis, mixed allergic contact hypersensitivity, focal arthritis and spontaneous large granular lymphocytic tumors. Bioluminescence colocalized with histological sites of inflammation and was totally abolished in gene-deleted Mpo−/− mice, despite massive tissue infiltration of neutrophils and activated eosinophils, indicating that eosinophil peroxidase did not contribute to luminol-BLI in vivo. Thus, luminol-BLI provides a noninvasive, specific and highly sensitive optical readout of phagocyte-mediated MPO activity in vivo and may enable new diagnostic applications in a wide range of acute and chronic inflammatory conditions. PMID:19305414

  10. Bioluminescence imaging of myeloperoxidase activity in vivo.

    PubMed

    Gross, Shimon; Gammon, Seth T; Moss, Britney L; Rauch, Daniel; Harding, John; Heinecke, Jay W; Ratner, Lee; Piwnica-Worms, David

    2009-04-01

    The myeloperoxidase (MPO) system of activated phagocytes is central to normal host defense mechanisms, and dysregulated MPO contributes to the pathogenesis of inflammatory disease states ranging from atherosclerosis to cancer. Here we show that upon systemic administration, the small molecule luminol enables noninvasive bioluminescence imaging (BLI) of MPO activity in vivo. Luminol-BLI allowed quantitative longitudinal monitoring of MPO activity in animal models of acute dermatitis, mixed allergic contact hypersensitivity, focal arthritis and spontaneous large granular lymphocytic tumors. Bioluminescence colocalized with histological sites of inflammation and was totally abolished in gene-deleted Mpo(-/-) mice, despite massive tissue infiltration of neutrophils and activated eosinophils, indicating that eosinophil peroxidase did not contribute to luminol-BLI in vivo. Thus, luminol-BLI provides a noninvasive, specific and highly sensitive optical readout of phagocyte-mediated MPO activity in vivo and may enable new diagnostic applications in a wide range of acute and chronic inflammatory conditions.

  11. In-vivo optical investigation of psoriasis

    NASA Astrophysics Data System (ADS)

    Kapsokalyvas, Dimitrios; Cicchi, Riccardo; Bruscino, Nicola; Alfieri, Domenico; Massi, Daniela; Lotti, Torello; Pavone, Francesco S.

    2011-03-01

    Psoriasis is an autoimmune disease of the skin characterized by hyperkeratosis, hyperproliferation of the epidermis, inflammatory cell accumulation and increased dilatation of dermal papillary blood vessels. Cases of psoriasis were investigated in vivo with optical means in order to evaluate the potential of in vivo optical biopsy. A Polarization Multispectral Dermoscope was employed for the macroscopic observation. Features such as the 'dotted' blood vessels pattern was observed with high contrast. The average size of dot vessels in Psoriasis was measured to be 974 μm2 which is much higher compared to healthy skin. High resolution image sections of the epidermis and the dermis were produced with a custom made Multiphoton Microscope. Imaging extended from the surface of the lesion down to the papillary dermis, at a depth of 200 μm. In the epidermis, a characteristic morphology of the stratum corneum found only in Psoriasis was revealed. Additionally, the cytoplasmic area of the cells in the stratum spinosum layer was found to be smaller than normal. In the dermis the morphological features were more pronounced, where the elongated dermal papillae dominated the papillary layer. Their length exceeds 100μm, which is a far greater value compared to that of healthy skin. These in vivo observations are consistent with the ex vivo histopathological observations, supporting both the applicability and potentiality of multispectral dermoscopy and multiphoton microscopy in the field of in vivo optical investigation and biopsy of skin.

  12. Bi-layer composite dressing of gelatin nanofibrous mat and poly vinyl alcohol hydrogel for drug delivery and wound healing application: in-vitro and in-vivo studies.

    PubMed

    Jaiswal, Maneesh; Gupta, Asheesh; Agrawal, Ashwini K; Jassal, Manjeet; Dinda, Amit Kr; Koul, Veena

    2013-09-01

    Present investigation involves the development of a bi-layer dressing of gelatin nanofibrous mat loaded with epigallocatechin gallate (EGCG)/poly vinyl alcohol (PVA) hydrogel and its in-vivo evaluation on full-thickness excision wounds in experimental Wistar rats. Nanomorphological observation, porosity, effect of crosslinking on tensile strength, physical stability and drug release profile in phosphate buffer and biocompatibility aspects of electrospun nanomat were investigated by various physico-chemical tools. EGCGa release profile was found to increase from 2-4 days with decreasing crosslinking time from 15 to 5 min. PVA hydrogels were prepared by freeze-thaw method and has been utilized as a protective and hydrating outer layer of the bi-layer dressing. Topical application of bi-layer composite dressing loaded with EGCG improve the healing rate in experimental rats as acute wounds model which was evidenced by significant increase in DNA (approximately 42%), total protein (approximately 32%), hydroxyproline (approximately 26%) and hexosamine approximately 24%) contents. A faster wound contraction was observed in wounds treated with composite dressing from approximately 14% to 47%. Histopathological examination revealed significant improvement in angiogenesis, re-epithelialization and less inflammatory response in comparison to control. Van-Gieson's collagen stains revealed matured, compact and parallel deposition of collagen fibrils on day 12. These results were supported by up-regulated expressions of matrix metalloproteinase (MMPs-2 and 9) by gelatin zymography. Control release of EGCG, 3D porous architecture of nanofibrous scaffolds as well as moist microenvironment provides ideal conditions for uninterrupted wound healing.

  13. Comparison of In Vivo and Ex Vivo MRI for the Detection of Structural Abnormalities in a Mouse Model of Tauopathy

    PubMed Central

    Holmes, Holly E.; Powell, Nick M.; Ma, Da; Ismail, Ozama; Harrison, Ian F.; Wells, Jack A.; Colgan, Niall; O'Callaghan, James M.; Johnson, Ross A.; Murray, Tracey K.; Ahmed, Zeshan; Heggenes, Morten; Fisher, Alice; Cardoso, M. Jorge; Modat, Marc; O'Neill, Michael J.; Collins, Emily C.; Fisher, Elizabeth M. C.; Ourselin, Sébastien; Lythgoe, Mark F.

    2017-01-01

    With increasingly large numbers of mouse models of human disease dedicated to MRI studies, compromises between in vivo and ex vivo MRI must be fully understood in order to inform the choice of imaging methodology. We investigate the application of high resolution in vivo and ex vivo MRI, in combination with tensor-based morphometry (TBM), to uncover morphological differences in the rTg4510 mouse model of tauopathy. The rTg4510 mouse also offers a novel paradigm by which the overexpression of mutant tau can be regulated by the administration of doxycycline, providing us with a platform on which to investigate more subtle alterations in morphology with morphometry. Both in vivo and ex vivo MRI allowed the detection of widespread bilateral patterns of atrophy in the rTg4510 mouse brain relative to wild-type controls. Regions of volume loss aligned with neuronal loss and pathological tau accumulation demonstrated by immunohistochemistry. When we sought to investigate more subtle structural alterations in the rTg4510 mice relative to a subset of doxycycline-treated rTg4510 mice, ex vivo imaging enabled the detection of more regions of morphological brain changes. The disadvantages of ex vivo MRI may however mitigate this increase in sensitivity: we observed a 10% global shrinkage in brain volume of the post-mortem tissues due to formalin fixation, which was most notable in the cerebellum and olfactory bulbs. However, many central brain regions were not adversely affected by the fixation protocol, perhaps due to our “in-skull” preparation. The disparity between our TBM findings from in vivo and ex vivo MRI underlines the importance of appropriate study design, given the trade-off between these two imaging approaches. We support the utility of in vivo MRI for morphological phenotyping of mouse models of disease; however, for subtler phenotypes, ex vivo offers enhanced sensitivity to discrete morphological changes.

  14. In vivo Noninvasive Small Animal Molecular Imaging

    PubMed Central

    Youn, Hyewon; Hong, Kee-Jong

    2012-01-01

    The remarkable efforts that are made on molecular imaging technologies demonstrate its potential importance and range of applications. The generation of disease-specific animal models, and the developments of target-specific probes and genetically encoded reporters are another important component. Continued improvements in the instrumentation, the identification of novel targets and genes, and the availability of improved imaging probes should be made. Multimodal imaging probes should provide easier transitions between laboratory studies, including small animal studies and clinical applications. Here, we reviewed basic strategies of noninvasive in vivo imaging methods in small animals to introducing the concept of molecular imaging. PMID:24159487

  15. Semiconductor quantum dot toxicity in a mouse in vivo model

    NASA Astrophysics Data System (ADS)

    Bozrova, Svetlana V.; Baryshnikova, Maria A.; Nabiev, Igor; Sukhanova, Alyona

    2017-01-01

    Quantum dots (QDs) are increasingly widely used in clinical medicine. Their most promising potential applications are cancer diagnosis, including in vivo tumour imaging and targeted drug delivery. In this connection, the main questions are whether or not QDs are toxic for humans and, if they are, what concentration is relatively harmless. We have carried out in vivo experiments with CdSe/ZnS fluorescent semiconductor core/shell QDs, which are currently the most widely used in research.

  16. In vivo dosimetry for IMRT

    SciTech Connect

    Vial, Philip

    2011-05-05

    In vivo dosimetry has a well established role in the quality assurance of 2D radiotherapy and 3D conformal radiotherapy. The role of in vivo dosimetry for IMRT is not as well established. IMRT introduces a range of technical issues that complicate in vivo dosimetry. The first decade or so of IMRT implementation has largely relied upon pre-treatment phantom based dose verification. During that time, several new devices and techniques for in vivo dosimetry have emerged with the promise of providing the ultimate form of IMRT dose verification. Solid state dosimeters continue to dominate the field of in vivo dosimetry in the IMRT era. In this report we review the literature on in vivo dosimetry for IMRT, with an emphasis on clinical evidence for different detector types. We describe the pros and cons of different detectors and techniques in the IMRT setting and the roles that they are likely to play in the future.

  17. Electrodeposition of platinum-iridium coatings and nanowires for neurostimulating applications: Fabrication, characterization and in-vivo retinal stimulation/recording. EIS studies of hexavalent and trivalent chromium based military coating systems

    NASA Astrophysics Data System (ADS)

    Petrossians, Artin

    The studies presented in this thesis are composed of two different projects demonstrated in two different parts. The first part of this thesis represents an electrochemical approach to possible improvements of the interface between an implantable device and biological tissue. The second part of this thesis represents electrochemical impedance spectroscopy (EIS) studies on the corrosion resistance behavior of different types of polymer coated Al2024 alloys. In the first part of this thesis, a broad range of investigations on the development of an efficient and reproducible electrochemical deposition method for fabrication of thin-film platinum-iridium alloys were performed. The developed method for production of dense films was then modified to produce very high surface area coatings with ultra-low electrochemical impedance characteristics. The high-surface area platinum-iridium coating was applied on microelectrode arrays for chronic in-vitro stimulation. Using the same method of producing dense films, platinum-iridium nanowires were fabricated using Anodized Aluminum Oxide (AAO) templates for hermetic packaging applications to be used in implantable microelectronics. The implantable microelectronics will be used to perform data reception and transmission management, power recovery, digital processing and analog output of stimulus current. Finally, in-vivo electrical stimulation tests were performed on an animal retina using high surface-area platinum-iridium coated single microelectrodes to verify the charge transfer characteristics of the coatings. In the second part of this thesis, three different sets of samples with different combinations of pretreatments, primers with the same type of topcoat were tested in 0.5 N NaCl for period of 30 days. The surface changes measured by EIS as a function of time were then analyzed. The analysis of the fit parameters of the impedance spectra showed that the different primers had the most effect on the corrosion protection

  18. Ex vivo expansion of hematopoietic stem cells.

    PubMed

    Xie, JingJing; Zhang, ChengCheng

    2015-09-01

    Ex vivo expansion of hematopoietic stem cells (HSCs) would benefit clinical applications in several aspects, to improve patient survival, utilize cord blood stem cells for adult applications, and selectively propagate stem cell populations after genetic manipulation. In this review we summarize and discuss recent advances in the culture systems of mouse and human HSCs, which include stroma/HSC co-culture, continuous perfusion and fed-batch cultures, and those supplemented with extrinsic ligands, membrane transportable transcription factors, complement components, protein modification enzymes, metabolites, or small molecule chemicals. Some of the expansion systems have been tested in clinical trials. The optimal condition for ex vivo expansion of the primitive and functional human HSCs is still under development. An improved understanding of the mechanisms for HSC cell fate determination and the HSC culture characteristics will guide development of new strategies to overcome difficulties. In the future, development of a combination treatment regimen with agents that enhance self-renewal, block differentiation, and improve homing will be critical. Methods to enhance yields and lower cost during collection and processing should be employed. The employment of an efficient system for ex vivo expansion of HSCs will facilitate the further development of novel strategies for cell and gene therapies including genome editing.

  19. In vivo and ex vivo evaluation of cosmetic properties of seedcakes.

    PubMed

    Ratz-Łyko, Anna; Arct, Jacek; Pytkowska, Katarzyna; Majewski, Sławomir

    2015-04-01

    The seedcakes are a potential source of natural bioactive substances: antioxidants, protein, and carbohydrates. Thus, they may scavenge free radicals and have an effect on the stratum corneum hydration and epidermal barrier function. The aim of the study was to evaluate the in vivo and ex vivo properties of emulsions with the seedcake extracts using the pH meter, corneometer, tewameter, methyl nicotinate model of micro-inflammation in human skin, and tape stripping of the stratum corneum. The in vivo and ex vivo studies showed that the emulsions with Oenothera biennis, Borago officinalis, and Nigella sativa seedcake extracts have anti-inflammatory and antioxidant activity. The 6-week topical application of the emulsions with the B. officinalis and N. sativa seedcakes significantly reduced skin irritation and influenced the improvement of the skin hydration and epidermal barrier function compared with placebo. The seedcakes due to their antioxidant and anti-inflammatory activities have potential application in anti-aging, moisturizing, mitigating, and protective cosmetics.

  20. Disposable Fluidic Actuators for Miniature In-Vivo Surgical Robotics.

    PubMed

    Pourghodrat, Abolfazl; Nelson, Carl A

    2017-03-01

    Fusion of robotics and minimally invasive surgery (MIS) has created new opportunities to develop diagnostic and therapeutic tools. Surgical robotics is advancing from externally actuated systems to miniature in-vivo robotics. However, with miniaturization of electric-motor-driven surgical robots, there comes a trade-off between the size of the robot and its capability. Slow actuation, low load capacity, sterilization difficulties, leaking electricity and transferring produced heat to tissues, and high cost are among the key limitations of the use of electric motors in in-vivo applications. Fluid power in the form of hydraulics or pneumatics has a long history in driving many industrial devices and could be exploited to circumvent these limitations. High power density and good compatibility with the in-vivo environment are the key advantages of fluid power over electric motors when it comes to in-vivo applications. However, fabrication of hydraulic/pneumatic actuators within the desired size and pressure range required for in-vivo surgical robotic applications poses new challenges. Sealing these types of miniature actuators at operating pressures requires obtaining very fine surface finishes which is difficult and costly. The research described here presents design, fabrication, and testing of a hydraulic/pneumatic double-acting cylinder, a limited-motion vane motor, and a balloon-actuated laparoscopic grasper. These actuators are small, seal-less, easy to fabricate, disposable, and inexpensive, thus ideal for single-use in-vivo applications. To demonstrate the ability of these actuators to drive robotic joints, they were modified and integrated in a robotic arm. The design and testing of this surgical robotic arm are presented to validate the concept of fluid-power actuators for in-vivo applications.

  1. In vivo Cytotoxicity Studies of Amaryllidaceae Alkaloids.

    PubMed

    Nair, Jerald J; Bastida, Jaume; van Staden, Johannes

    2016-01-01

    The plant family Amaryllidaceae is recognizable for its esthetic floral characteristics, its widespread usage in traditional medicine as well as its unique alkaloid principles. Few alkaloid-producing families rival the Amaryllidaceae in terms of the diversity of its structures as well as their wide applicability on the biological landscape. In particular, cytotoxic effects have come to be a dominant theme in the biological properties of Amaryllidacea alkaloids. To this extent, a significant number of structures have been subjected to in vitro studies in numerous cell lines from which several targets have been identified as promising chemotherapeutics. By contrast, in vivo models of study involving these alkaloids have been carried out to a lesser extent and should prove crucial in the continued development of a clinical target such as pancratistatin. This survey examines the cytotoxic effects of Amaryllidaceae alkaloids in vivo and contrasts these against the corresponding in vitro effects.

  2. Bioluminescent bacterial imaging in vivo.

    PubMed

    Baban, Chwanrow K; Cronin, Michelle; Akin, Ali R; O'Brien, Anne; Gao, Xuefeng; Tabirca, Sabin; Francis, Kevin P; Tangney, Mark

    2012-11-04

    This video describes the use of whole body bioluminesce imaging (BLI) for the study of bacterial trafficking in live mice, with an emphasis on the use of bacteria in gene and cell therapy for cancer. Bacteria present an attractive class of vector for cancer therapy, possessing a natural ability to grow preferentially within tumors following systemic administration. Bacteria engineered to express the lux gene cassette permit BLI detection of the bacteria and concurrently tumor sites. The location and levels of bacteria within tumors over time can be readily examined, visualized in two or three dimensions. The method is applicable to a wide range of bacterial species and tumor xenograft types. This article describes the protocol for analysis of bioluminescent bacteria within subcutaneous tumor bearing mice. Visualization of commensal bacteria in the Gastrointestinal tract (GIT) by BLI is also described. This powerful, and cheap, real-time imaging strategy represents an ideal method for the study of bacteria in vivo in the context of cancer research, in particular gene therapy, and infectious disease. This video outlines the procedure for studying lux-tagged E. coli in live mice, demonstrating the spatial and temporal readout achievable utilizing BLI with the IVIS system.

  3. Applications

    NASA Astrophysics Data System (ADS)

    Stern, Arthur M.

    1986-07-01

    Economic incentives have spurred numerous applications of genetically engineered organisms in manufacture of pharmaceuticals and industrial chemicals. These successes, involving a variety of methods of genetic manipulation, have dispelled early fears that genetic engineering could not be handled safely, even in the laboratory. Consequently, the potential for applications in the wider environment without physical containment is being considered for agriculture, mining, pollution control, and pest control. These proposed applications range from modest extensions of current plant breeding techniques for new disease-resistant species to radical combinations of organisms (for example, nitrogen-fixing corn plants). These applications raise concerns about potential ecological impacts (see chapter 5), largely because of adverse experiences with both deliberate and inadvertent introductions of nonindigenous species.

  4. Tacrine: In vivo veritas.

    PubMed

    Jarrott, Bevyn

    2017-02-01

    Tacrine was initially synthesised in 1945 as part of a project seeking antibacterial drugs to treat infected wounds in soldiers. However, it was inactive in vitro against common strains of bacteria. Serendipitously, it was injected in vivo into dogs anaesthetised with chloroform and morphine and noted to immediately counter the respiratory rate depression caused by morphine but not block analgesia. Subsequent studies showed that tacrine was an acetylcholinesterase inhibitor. When combined with morphine in ampoules it was possible to inject larger doses of morphine without causing respiratory depression and it was marketed for 10 years in Australia. Tacrine was also used alone for treating acute anticholinergic syndrome in the 1980s. Shortly after this, it was hypothesised by William Summers that it could be of benefit in treating the early stages of Alzheimer's dementia and an IND was granted by the US Food and Drug Administration and a use patent awarded to Summers. It was the first of four anticholinesterases to be approved for treating this condition although its variable pharmacokinetics was a disadvantage.

  5. In vivo microscopy✩

    PubMed Central

    Peti-Peterdi, János

    2016-01-01

    This article summarizes the past, present, and future promise of multiphoton excitation fluorescence microscopy for intravital kidney imaging. During the past 15 years, several high-power visual research approaches have been developed using multiphoton imaging to study the normal functions of the healthy, intact, living kidney, and the various molecular and cellular mechanisms of the development of kidney diseases. In this review, the main focus will be on intravital multiphoton imaging of the glomerulus, the structure and function of the glomerular filtration barrier, especially the podocyte. Examples will be given for the combination of two powerful research tools, in vivo multiphoton imaging and mouse genetics using commercially available whole animal models for the detailed characterization of glomerular cell types, their function and fate, and for the better understanding of the molecular mechanisms of glomerular pathologies. One of the new modalities of multiphoton imaging, serial imaging of the same glomerulus in the same animal over several days will be emphasized for its potential for further advancing the field of nephrology research. PMID:26968479

  6. In vivo microscopy.

    PubMed

    Peti-Peterdi, János

    2016-04-01

    This article summarizes the past, present, and future promise of multiphoton excitation fluorescence microscopy for intravital kidney imaging. During the past 15years, several high-power visual research approaches have been developed using multiphoton imaging to study the normal functions of the healthy, intact, living kidney, and the various molecular and cellular mechanisms of the development of kidney diseases. In this review, the main focus will be on intravital multiphoton imaging of the glomerulus, the structure and function of the glomerular filtration barrier, especially the podocyte. Examples will be given for the combination of two powerful research tools, in vivo multiphoton imaging and mouse genetics using commercially available whole animal models for the detailed characterization of glomerular cell types, their function and fate, and for the better understanding of the molecular mechanisms of glomerular pathologies. One of the new modalities of multiphoton imaging, serial imaging of the same glomerulus in the same animal over several days will be emphasized for its potential for further advancing the field of nephrology research.

  7. A method to study in vivo stability of DNA nanostructures.

    PubMed

    Surana, Sunaina; Bhatia, Dhiraj; Krishnan, Yamuna

    2013-11-01

    DNA nanostructures are rationally designed, synthetic, nanoscale assemblies obtained from one or more DNA sequences by their self-assembly. Due to the molecularly programmable as well as modular nature of DNA, such designer DNA architectures have great potential for in cellulo and in vivo applications. However, demonstrations of functionality in living systems necessitates a method to assess the in vivo stability of the relevant nanostructures. Here, we outline a method to quantitatively assay the stability and lifetime of various DNA nanostructures in vivo. This exploits the property of intact DNA nanostructures being uptaken by the coelomocytes of the multicellular model organism Caenorhabditis elegans. These studies reveal that the present fluorescence based assay in coelomocytes of C. elegans is an useful in vivo test bed for measuring DNA nanostructure stability.

  8. In vivo imaging of hydrogen peroxide with chemiluminescent nanoparticles.

    PubMed

    Lee, Dongwon; Khaja, Sirajud; Velasquez-Castano, Juan C; Dasari, Madhuri; Sun, Carrie; Petros, John; Taylor, W Robert; Murthy, Niren

    2007-10-01

    The overproduction of hydrogen peroxide is implicated in the development of numerous diseases and there is currently great interest in developing contrast agents that can image hydrogen peroxide in vivo. In this report, we demonstrate that nanoparticles formulated from peroxalate esters and fluorescent dyes can image hydrogen peroxide in vivo with high specificity and sensitivity. The peroxalate nanoparticles image hydrogen peroxide by undergoing a three-component chemiluminescent reaction between hydrogen peroxide, peroxalate esters and fluorescent dyes. The peroxalate nanoparticles have several attractive properties for in vivo imaging, such as tunable wavelength emission (460-630 nm), nanomolar sensitivity for hydrogen peroxide and excellent specificity for hydrogen peroxide over other reactive oxygen species. The peroxalate nanoparticles were capable of imaging hydrogen peroxide in the peritoneal cavity of mice during a lipopolysaccharide-induced inflammatory response. We anticipate numerous applications of peroxalate nanoparticles for in vivo imaging of hydrogen peroxide, given their high specificity and sensitivity and deep-tissue-imaging capability.

  9. The Expanding Toolbox of In Vivo Bioluminescent Imaging

    PubMed Central

    Xu, Tingting; Close, Dan; Handagama, Winode; Marr, Enolia; Sayler, Gary; Ripp, Steven

    2016-01-01

    In vivo bioluminescent imaging (BLI) permits the visualization of engineered bioluminescence from living cells and tissues to provide a unique perspective toward the understanding of biological processes as they occur within the framework of an authentic in vivo environment. The toolbox of in vivo BLI includes an inventory of luciferase compounds capable of generating bioluminescent light signals along with sophisticated and powerful instrumentation designed to detect and quantify these light signals non-invasively as they emit from the living subject. The information acquired reveals the dynamics of a wide range of biological functions that play key roles in the physiological and pathological control of disease and its therapeutic management. This mini review provides an overview of the tools and applications central to the evolution of in vivo BLI as a core technology in the preclinical imaging disciplines. PMID:27446798

  10. In vivo NMR imaging of deuterium

    NASA Astrophysics Data System (ADS)

    Müller, S.; Seelig, J.

    D 2O is used as a contrast agent for studying anatomical images and flow in vivo by deuterium NMR. A deuterium image of the head of a living rat after administration of D 2O (5% v/v) in the drinking water is shown. It was obtained in 14 min with a surface coil and has a spatial resolution of about one millimeter. The application of D 2O as a tracer is discussed and the inflow of heavy water into the brain of a rat is recorded in a time series of deuterium images. Spatially resolved inflow time constants have been determined.

  11. In vivo confocal microscopy of toxic keratopathy

    PubMed Central

    Chen, Y; Le, Q; Hong, J; Gong, L; Xu, J

    2017-01-01

    Purpose To explore the morphological characteristics of toxic keratopathy (TK), which clinically presented as superficial punctate keratopathy (SPK), with the application of in vivo laser-scanning confocal microscopy (LSCM), and evaluate its potential in the early diagnosis of TK. Patients and methods This was a cross-sectional study involving 16 patients with TK and 16 patients with dry eye (DE), demonstrating SPK under slit-lamp observation, and 10 normal eyes were enrolled in the study. All participants underwent history interviews, fluorescein staining, tear film break-up time (BUT) tests, Schirmer tests, and in vivo LSCM. Results The area grading of corneal fluorescein punctate staining was higher in the TK group than the DE group. Measured by in vivo LSCM, superficial epithelial cell density was lower in the TK group than that of DE group. The sub-basal nerve presented lower density and tortuosity in the TK group than the DE group. Most notably, deposits with a snow-like appearance were observed in the epithelium in 12/16 (75.0%) of the TK cases, but none in the DE group (P<0.05). Conclusion The SPK in TK patients was characterized by more widespread punctate staining, a lower density of superficial epithelial cells and sub-basal nerves, and a typical snow-like pattern of deposits in the epithelium by LSCM. These features might facilitate early diagnosis of TK from other disorders manifested as SPK. PMID:27740620

  12. Type I cell ROS kinetics under hypoxia in the intact mouse carotid body ex vivo: a FRET-based study.

    PubMed

    Bernardini, A; Brockmeier, U; Metzen, E; Berchner-Pfannschmidt, U; Harde, E; Acker-Palmer, A; Papkovsky, D; Acker, H; Fandrey, J

    2015-01-01

    Reactive oxygen species (ROS) mainly originating from NADPH oxidases have been shown to be involved in the carotid body (CB) oxygen-sensing cascade. For measuring ROS kinetics, type I cells of the mouse CB in an ex vivo preparation were transfected with the ROS sensor construct FRET-HSP33. After 2 days of tissue culture, type I cells expressed FRET-HSP33 as shown by immunohistochemistry. In one population of CBs, 5 min of hypoxia induced a significant and reversible decrease of type I cell ROS levels (n = 9 CBs; P < 0.015), which could be inhibited by 4-(2-aminoethyl)benzensulfonylfluorid (AEBSF), a highly specific inhibitor of the NADPH oxidase subunits p47(phox) and p67(phox). In another population of CBs, however, 5 min of hypoxia induced a significant and reversible increase of ROS levels in type I cells (n = 8 CBs; P < 0.05), which was slightly enhanced by administration of 3 mM AEBSF. These different ROS kinetics seemed to coincide with different mice breeding conditions. Type I cells of both populations showed a typical hypoxia-induced membrane potential (MP) depolarization, which could be inhibited by 3 mM AEBSF. ROS and MP closely followed the hypoxic decrease in CB tissue oxygen as measured with an O2-sensitive dye. We conclude that attenuated p47(phox) subunit activity of the NADPH oxidase under hypoxia is the physiological trigger for type I cell MP depolarization probably due to ROS decrease, whereas the observed ROS increase has no influence on type I cell MP kinetics under hypoxia.

  13. In vivo photoacoustic imaging of mouse embryos

    NASA Astrophysics Data System (ADS)

    Laufer, Jan; Norris, Francesca; Cleary, Jon; Zhang, Edward; Treeby, Bradley; Cox, Ben; Johnson, Peter; Scambler, Pete; Lythgoe, Mark; Beard, Paul

    2012-06-01

    The ability to noninvasively image embryonic vascular anatomy in mouse models is an important requirement for characterizing the development of the normal cardiovascular system and malformations in the heart and vascular supply. Photoacoustic imaging, which can provide high resolution non invasive images of the vasculature based upon optical absorption by endogenous hemoglobin, is well suited to this application. In this study, photoacoustic images of mouse embryos were obtained ex vivo and in vivo. The images show intricate details of the embryonic vascular system to depths of up to 10 mm, which allowed whole embryos to be imaged in situ. To achieve this, an all-optical photoacoustic scanner and a novel time reversal image reconstruction algorithm, which provide deep tissue imaging capability while maintaining high spatial resolution and contrast were employed. This technology may find application as an imaging tool for preclinical embryo studies in developmental biology as well as more generally in preclinical and clinical medicine for studying pathologies characterized by changes in the vasculature.

  14. In Vivo Ischemia Detection by Luminescent Nanothermometers.

    PubMed

    Ximendes, Erving Clayton; Rocha, Uéslen; Del Rosal, Blanca; Vaquero, Alberto; Sanz-Rodríguez, Francisco; Monge, Luis; Ren, Fuqiang; Vetrone, Fiorenzo; Ma, Dongling; García-Solé, José; Jacinto, Carlos; Jaque, Daniel; Fernández, Nuria

    2017-02-01

    There is an urgent need to develop new diagnosis tools for real in vivo detection of first stages of ischemia for the early treatment of cardiovascular diseases and accidents. However, traditional approaches show low sensitivity and a limited penetration into tissues, so they are only applicable for the detection of surface lesions. Here, it is shown how the superior thermal sensing capabilities of near infrared-emitting quantum dots (NIR-QDs) can be efficiently used for in vivo detection of subcutaneous ischemic tissues. In particular, NIR-QDs make possible ischemia detection by high penetration transient thermometry studies in a murine ischemic hindlimb model. NIR-QDs nanothermometers are able to identify ischemic tissues by means of their faster thermal dynamics. In addition, they have shown to be capable of monitoring both the revascularization and damage recovery processes of ischemic tissues. This work demonstrates the applicability of fluorescence nanothermometry for ischemia detection and treatment, as well as a tool for early diagnosis of cardiovascular disease.

  15. In vivo versus ex vivo CRISPR therapies for retinal dystrophy

    PubMed Central

    Bakondi, Benjamin

    2017-01-01

    SUMMARY Two therapeutic paths have been proposed to treat inherited retinal dystrophy using clustered regularly interspaced short palindromic repeats (CRISPR). One strategy is to genetically correct patient cells ex vivo for autologous transplant, whereas the second is to modify cells in vivo by delivering CRISPR effectors to the retina. The feasibility of both editing strategies has been demonstrated within three years of CRISPR’s adaptation to mammalian systems. However, the functional integration of transplanted cells into host retinae has been a long-standing challenge that currently represents the 2025 moonshot of the National Eye Institute’s Audacious Goals Initiative. The clinical translatability of each path is discussed with regard to current investigations and whether cell replacement can be circumvented by in vivo editing. PMID:28163772

  16. Tailoring vessel morphology in vivo

    NASA Astrophysics Data System (ADS)

    Gould, Daniel Joseph

    Tissue engineering is a rapidly growing field which seeks to provide alternatives to organ transplantation in order to address the increasing need for transplantable tissues. One huge hurdle in this effort is the provision of thick tissues; this hurdle exists because currently there is no way to provide prevascularized or rapidly vascularizable scaffolds. To design thick, vascularized tissues, scaffolds are needed that can induce vessels which are similar to the microvasculature found in normal tissues. Angiogenic biomaterials are being developed to provide useful scaffolds to address this problem. In this thesis angiogenic and cell signaling and adhesion factors were incorporated into a biomimetic poly(ethylene glycol) (PEG) hydrogel system. The composition of these hydrogels was precisely tuned to induce the formation of differing vessel morphology. To sensitively measure induced microvascular morphology and to compare it to native microvessels in several tissues, this thesis developed an image-based tool for quantification of scale invariant and classical measures of vessel morphology. The tool displayed great utility in the comparison of native vessels and remodeling vessels in normal tissues. To utilize this tool to tune the vessel response in vivo, Flk1::myr-mCherry fluorescently labeled mice were implanted with Platelet Derived Growth Factor-BB (PDGF-BB) and basic Fibroblast Growth Factor (FGF-2) containing PEG-based hydrogels in a modified mouse corneal angiogenesis assay. Resulting vessels were imaged with confocal microscopy, analyzed with the image based tool created in this thesis to compare morphological differences between treatment groups, and used to create a linear relationship between space filling parameters and dose of growth factor release. Morphological parameters of native mouse tissue vessels were then compared to the linear fit to calculate the dose of growth factors needed to induce vessels similar in morphology to native vessels

  17. Spectral characteristics of two-photon autofluorescence and second harmonic generation from human skin in vivo

    NASA Astrophysics Data System (ADS)

    Breunig, Hans G.; König, Karsten

    2011-03-01

    We performed multiphoton imaging of human skin and recorded in combination the complete spectral content of the signals in vivo. The spectra represent the integration of multiphoton signals over the investigated regions of the epidermis and dermis. They are used to study depth-resolved in vivo emission characteristics of main endogenous skin fluorophores like keratin, NAD(P)H, collagen and elastin. The identification of the specific fluorophores is supported by analysis of additional in vivo fluorescence lifetime imaging. Furthermore, as a potential application of spectrally selective imaging the possibility to investigate the penetration of nanoparticles from sunscreen lotion into skin in vivo is discussed.

  18. Modeling disease in vivo with CRISPR/Cas9

    PubMed Central

    Dow, Lukas E.

    2015-01-01

    The recent advent of CRISPR/Cas9-mediated genome editing has created a wave of excitement across the scientific research community, carrying the promise of simple and effective genomic manipulation of nearly any cell type. CRISPR has quickly become the preferred tool for genetic manipulation, and shows incredible promise as a platform for studying gene function in vivo. Here, I discuss the current application of CRISPR technology to create new in vivo disease models, with a particular focus on how these tools, derived from an adaptive bacterial immune system, are helping us better model the complexity of human cancer. PMID:26432018

  19. Modeling Disease In Vivo With CRISPR/Cas9.

    PubMed

    Dow, Lukas E

    2015-10-01

    The recent advent of CRISPR/Cas9-mediated genome editing has created a wave of excitement across the scientific research community, carrying the promise of simple and effective genomic manipulation of nearly any cell type. CRISPR has quickly become the preferred tool for genetic manipulation, and shows incredible promise as a platform for studying gene function in vivo. I discuss the current application of CRISPR technology to create new in vivo disease models, with a particular focus on how these tools, derived from an adaptive bacterial immune system, are helping us to better model the complexity of human cancer.

  20. In vitro and ex vivo strategies for intracellular delivery

    NASA Astrophysics Data System (ADS)

    Stewart, Martin P.; Sharei, Armon; Ding, Xiaoyun; Sahay, Gaurav; Langer, Robert; Jensen, Klavs F.

    2016-10-01

    Intracellular delivery of materials has become a critical component of genome-editing approaches, ex vivo cell-based therapies, and a diversity of fundamental research applications. Limitations of current technologies motivate development of next-generation systems that can deliver a broad variety of cargo to diverse cell types. Here we review in vitro and ex vivo intracellular delivery approaches with a focus on mechanisms, challenges and opportunities. In particular, we emphasize membrane-disruption-based delivery methods and the transformative role of nanotechnology, microfluidics and laboratory-on-chip technology in advancing the field.

  1. Monitoring protein stability in vivo.

    PubMed

    Ignatova, Zoya

    2005-08-24

    Reduced protein stability in vivo is a prerequisite to aggregation. While this is merely a nuisance factor in recombinant protein production, it holds a serious impact for man. This review focuses on specific approaches to selectively determine the solubility and/or stability of a target protein within the complex cellular environment using different detection techniques. Noninvasive techniques mapping folding/misfolding events on a fast time scale can be used to unravel the complexity and dynamics of the protein aggregation process and factors altering protein solubility in vivo. The development of approaches to screen for folding and solubility in vivo should facilitate the identification of potential components that improve protein solubility and/or modulate misfolding and aggregation and may provide a therapeutic benefit.

  2. Experimental Cryosurgery Investigations In Vivo

    PubMed Central

    Gage, A.A.; Baust, J.M.; Baust, J.G.

    2009-01-01

    Cryosurgery is the use of freezing temperatures to elicit an ablative response in a targeted tissue. This review provides a global overview of experimentation in vivo which has been the basis of advancement of this widely applied therapeutic option. The cellular and tissue-related events that underlie the mechanisms of destruction, including direct cell injury (cryolysis), vascular stasis, apoptosis and necrosis, are described and are related to the optimal methods of technique of freezing to achieve efficacious therapy. In vivo experiments with major organs, including wound healing, the putative immunological response following thawing, and the use of cryoadjunctive strategies to enhance cancer cell sensitivity to freezing, are described. PMID:19833119

  3. In vivo Raman spectroscopy of cervix cancers

    NASA Astrophysics Data System (ADS)

    Rubina, S.; Sathe, Priyanka; Dora, Tapas Kumar; Chopra, Supriya; Maheshwari, Amita; Krishna, C. Murali

    2014-03-01

    Cervix-cancer is the third most common female cancer worldwide. It is the leading cancer among Indian females with more than million new diagnosed cases and 50% mortality, annually. The high mortality rates can be attributed to late diagnosis. Efficacy of Raman spectroscopy in classification of normal and pathological conditions in cervix cancers on diverse populations has already been demonstrated. Our earlier ex vivo studies have shown the feasibility of classifying normal and cancer cervix tissues as well as responders/non-responders to Concurrent chemoradiotherapy (CCRT). The present study was carried out to explore feasibility of in vivo Raman spectroscopic methods in classifying normal and cancerous conditions in Indian population. A total of 182 normal and 132 tumor in vivo Raman spectra, from 63 subjects, were recorded using a fiberoptic probe coupled HE-785 spectrometer, under clinical supervision. Spectra were acquired for 5 s and averaged over 3 times at 80 mW laser power. Spectra of normal conditions suggest strong collagenous features and abundance of non-collagenous proteins and DNA in case of tumors. Preprocessed spectra were subjected to Principal Component-Linear Discrimination Analysis (PCLDA) followed by leave-one-out-cross-validation. Classification efficiency of ~96.7% and 100% for normal and cancerous conditions respectively, were observed. Findings of the study corroborates earlier studies and suggest applicability of Raman spectroscopic methods in combination with appropriate multivariate tool for objective, noninvasive and rapid diagnosis of cervical cancers in Indian population. In view of encouraging results, extensive validation studies will be undertaken to confirm the findings.

  4. Development and Application of a dapB-Based In Vivo Expression Technology System To Study Colonization of Rice by the Endophytic Nitrogen-Fixing Bacterium Pseudomonas stutzeri A15

    PubMed Central

    Rediers, Hans; Bonnecarrère, Victoria; Rainey, Paul B.; Hamonts, Kelly; Vanderleyden, Jos; De Mot, René

    2003-01-01

    Pseudomonas stutzeri A15 is a nitrogen-fixing bacterium isolated from paddy rice. Strain A15 is able to colonize and infect rice roots. This strain may provide rice plants with fixed nitrogen and hence promote plant growth. In this article, we describe the use of dapB-based in vivo expression technology to identify P. stutzeri A15 genes that are specifically induced during colonization and infection (cii). We focused on the identification of P. stutzeri A15 genes that are switched on during rice root colonization and are switched off during free-living growth on synthetic medium. Several transcriptional fusions induced in the rice rhizosphere were isolated. Some of the corresponding genes are involved in the stress response, chemotaxis, metabolism, and global regulation, while others encode putative proteins with unknown functions or without significant homology to known proteins. PMID:14602651

  5. In Vivo Bioluminescence Imaging of Intratumoral Bacteria.

    PubMed

    Cronin, Michelle; Akin, Ali R; Francis, Kevin P; Tangney, Mark

    2016-01-01

    This chapter describes the use of whole-body bioluminescent imaging (BLI) for the study of bacterial trafficking in live mice, with an emphasis on the use of bacteria in therapy of cancer. Bacteria present an attractive class of vector for cancer therapy, possessing a natural ability to grow preferentially within tumors following systemic administration. Bacteria engineered to express the lux gene cassette permit BLI detection of the bacteria and tumor sites concurrently. The location and levels of bacteria within tumors over time can be readily examined, visualized in two or three dimensions. The method is applicable to a wide range of bacterial species and tumor xenograft types. This article describes the protocol for analysis of bioluminescent bacteria within subcutaneous tumor-bearing mice. This powerful, and inexpensive, real-time imaging strategy represents an ideal method for the study of bacteria in vivo in the context of cancer research. This protocol outlines the procedure for studying lux-tagged Escherichia coli and Bifidobacterium breve in mice, demonstrating the spatial and temporal readout from 2D and 3D BLI achievable with whole-body in vivo luminescence imaging.

  6. Comparison of in vivo and ex vivo imaging of the microvasculature with 2-photon fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Steinman, Joe; Koletar, Margaret; Stefanovic, Bojana; Sled, John G.

    2016-03-01

    This study evaluates 2-Photon fluorescence microscopy of in vivo and ex vivo cleared samples for visualizing cortical vasculature. Four mice brains were imaged with in vivo 2PFM. Mice were then perfused with a FITC gel and cleared in fructose. The same regions imaged in vivo were imaged ex vivo. Vessels were segmented automatically in both images using an in-house developed algorithm that accounts for the anisotropic and spatially varying PSF ex vivo. Through non-linear warping, the ex vivo image and tracing were aligned to the in vivo image. The corresponding vessels were identified through a local search algorithm. This enabled comparison of identical vessels in vivo/ex vivo. A similar process was conducted on the in vivo tracing to determine the percentage of vessels perfused. Of all the vessels identified over the four brains in vivo, 98% were present ex vivo. There was a trend towards reduced vessel diameter ex vivo by 12.7%, and the shrinkage varied between specimens (0% to 26%). Large diameter surface vessels, through a process termed 'shadowing', attenuated in vivo signal from deeper cortical vessels by 40% at 300 μm below the cortical surface, which does not occur ex vivo. In summary, though there is a mean diameter shrinkage ex vivo, ex vivo imaging has a reduced shadowing artifact. Additionally, since imaging depths are only limited by the working distance of the microscope objective, ex vivo imaging is more suitable for imaging large portions of the brain.

  7. Tracking immune cells in vivo using magnetic resonance imaging.

    PubMed

    Ahrens, Eric T; Bulte, Jeff W M

    2013-10-01

    The increasing complexity of in vivo imaging technologies, coupled with the development of cell therapies, has fuelled a revolution in immune cell tracking in vivo. Powerful magnetic resonance imaging (MRI) methods are now being developed that use iron oxide- and ¹⁹F-based probes. These MRI technologies can be used for image-guided immune cell delivery and for the visualization of immune cell homing and engraftment, inflammation, cell physiology and gene expression. MRI-based cell tracking is now also being applied to evaluate therapeutics that modulate endogenous immune cell recruitment and to monitor emerging cellular immunotherapies. These recent uses show that MRI has the potential to be developed in many applications to follow the fate of immune cells in vivo.

  8. Novel chemical attempts at ex vivo hematopoietic stem cell expansion.

    PubMed

    Zhang, Yu; Gao, Yingdai

    2016-05-01

    Hematopoietic stem cells (HSCs) are the most extensively studied stem cell type in adults, and the only stem cell type with proof of clinical utility. However, the greatest challenge for the broader use of HSCs remains the true expansion of the stem cells ex vivo. The development of researches on small-molecule compounds that support the safe and efficient ex vivo expansion of HSCs would help to promote the clinical application of HSCs. In recent years, several novel small-molecule compounds have been reported to improve ex vivo HSC expansion by promoting self-renewal, delaying differentiation, increasing homing, and inhibiting apoptosis. Here, we review recent chemical developments in stem cell research and the mechanisms underlying these compounds' effects.

  9. Progress Toward In Vivo Use of siRNAs-II

    PubMed Central

    Rettig, Garrett R; Behlke, Mark A

    2012-01-01

    RNA interference (RNAi) has been extensively employed for in vivo research since its use was first demonstrated in mammalian cells 10 years ago. Design rules have improved, and it is now routinely possible to obtain reagents that suppress expression of any gene desired. At the same time, increased understanding of the molecular basis of unwanted side effects has led to the development of chemical modification strategies that mitigate these concerns. Delivery remains the single greatest hurdle to widespread adoption of in vivo RNAi methods. However, exciting advances have been made and new delivery systems under development may help to overcome these barriers. This review discusses advances in RNAi biochemistry and biology that impact in vivo use and provides an overview of select publications that demonstrate interesting applications of these principles. Emphasis is placed on work with synthetic, small interfering RNAs (siRNAs) published since the first installment of this review which appeared in 2006. PMID:22186795

  10. In-vivo morphologic and spectroscopic investigation of Psoriasis

    NASA Astrophysics Data System (ADS)

    Kapsokalyvas, Dimitrios; Cicchi, Riccardo; Bruscino, Nicola; Alfieri, Domenico; Massi, Daniela; Lotti, Torello; Pavone, Francesco S.

    2011-07-01

    Psoriasis is an autoimmune disease of the skin characterized by hyperkeratosis, hyperproliferation of the epidermis, inflammatory cell accumulation and increased dilatation of dermal papillary blood vessels. Cases of psoriasis were investigated in vivo with optical means in order to evaluate the potential of in vivo optical biopsy. A Polarization Multispectral Dermoscope was employed for the macroscopic observation. Features such as the 'dotted' blood vessels pattern was observed with high contrast. High resolution image sections of the epidermis and the dermis were produced with a custom made Multiphoton Microscope. Imaging extended from the surface of the lesion down to the papillary dermis, at a depth of 200 μm. In the epidermis, a characteristic morphology of the stratum corneum found only in Psoriasis was revealed. Additionally, the cytoplasmic area of the cells in the stratum spinosum layer was found to be smaller than normal. In the dermis the morphological features were more pronounced, where the elongated dermal papillae dominated the papillary layer. Their length exceeds 100μm, which is a far greater value compared to that of healthy skin. These in vivo observations are consistent with the ex vivo histopathological observations, supporting both the applicability and potentiality of multispectral dermoscopy and multiphoton microscopy in the field of in vivo optical investigation and biopsy of skin.

  11. Quantifying drug-protein binding in vivo.

    SciTech Connect

    Buchholz, B; Bench, G; Keating III, G; Palmblad, M; Vogel, J; Grant, P G; Hillegonds, D

    2004-02-17

    Accelerator mass spectrometry (AMS) provides precise quantitation of isotope labeled compounds that are bound to biological macromolecules such as DNA or proteins. The sensitivity is high enough to allow for sub-pharmacological (''micro-'') dosing to determine macromolecular targets without inducing toxicities or altering the system under study, whether it is healthy or diseased. We demonstrated an application of AMS in quantifying the physiologic effects of one dosed chemical compound upon the binding level of another compound in vivo at sub-toxic doses [4].We are using tissues left from this study to develop protocols for quantifying specific binding to isolated and identified proteins. We also developed a new technique to quantify nanogram to milligram amounts of isolated protein at precisions that are comparable to those for quantifying the bound compound by AMS.

  12. 3D Ultrafast Ultrasound Imaging In Vivo

    PubMed Central

    Provost, Jean; Papadacci, Clement; Arango, Juan Esteban; Imbault, Marion; Gennisson, Jean-Luc; Tanter, Mickael; Pernot, Mathieu

    2014-01-01

    Very high frame rate ultrasound imaging has recently allowed for the extension of the applications of echography to new fields of study such as the functional imaging of the brain, cardiac electrophysiology, and the quantitative real-time imaging of the intrinsic mechanical properties of tumors, to name a few, non-invasively and in real time. In this study, we present the first implementation of Ultrafast Ultrasound Imaging in three dimensions based on the use of either diverging or plane waves emanating from a sparse virtual array located behind the probe. It achieves high contrast and resolution while maintaining imaging rates of thousands of volumes per second. A customized portable ultrasound system was developed to sample 1024 independent channels and to drive a 32×32 matrix-array probe. Its capability to track in 3D transient phenomena occurring in the millisecond range within a single ultrafast acquisition was demonstrated for 3-D Shear-Wave Imaging, 3-D Ultrafast Doppler Imaging and finally 3D Ultrafast combined Tissue and Flow Doppler. The propagation of shear waves was tracked in a phantom and used to characterize its stiffness. 3-D Ultrafast Doppler was used to obtain 3-D maps of Pulsed Doppler, Color Doppler, and Power Doppler quantities in a single acquisition and revealed, for the first time, the complex 3-D flow patterns occurring in the ventricles of the human heart during an entire cardiac cycle, and the 3-D in vivo interaction of blood flow and wall motion during the pulse wave in the carotid at the bifurcation. This study demonstrates the potential of 3-D Ultrafast Ultrasound Imaging for the 3-D real-time mapping of stiffness, tissue motion, and flow in humans in vivo and promises new clinical applications of ultrasound with reduced intra- and inter-observer variability. PMID:25207828

  13. 3D ultrafast ultrasound imaging in vivo.

    PubMed

    Provost, Jean; Papadacci, Clement; Arango, Juan Esteban; Imbault, Marion; Fink, Mathias; Gennisson, Jean-Luc; Tanter, Mickael; Pernot, Mathieu

    2014-10-07

    Very high frame rate ultrasound imaging has recently allowed for the extension of the applications of echography to new fields of study such as the functional imaging of the brain, cardiac electrophysiology, and the quantitative imaging of the intrinsic mechanical properties of tumors, to name a few, non-invasively and in real time. In this study, we present the first implementation of Ultrafast Ultrasound Imaging in 3D based on the use of either diverging or plane waves emanating from a sparse virtual array located behind the probe. It achieves high contrast and resolution while maintaining imaging rates of thousands of volumes per second. A customized portable ultrasound system was developed to sample 1024 independent channels and to drive a 32  ×  32 matrix-array probe. Its ability to track in 3D transient phenomena occurring in the millisecond range within a single ultrafast acquisition was demonstrated for 3D Shear-Wave Imaging, 3D Ultrafast Doppler Imaging, and, finally, 3D Ultrafast combined Tissue and Flow Doppler Imaging. The propagation of shear waves was tracked in a phantom and used to characterize its stiffness. 3D Ultrafast Doppler was used to obtain 3D maps of Pulsed Doppler, Color Doppler, and Power Doppler quantities in a single acquisition and revealed, at thousands of volumes per second, the complex 3D flow patterns occurring in the ventricles of the human heart during an entire cardiac cycle, as well as the 3D in vivo interaction of blood flow and wall motion during the pulse wave in the carotid at the bifurcation. This study demonstrates the potential of 3D Ultrafast Ultrasound Imaging for the 3D mapping of stiffness, tissue motion, and flow in humans in vivo and promises new clinical applications of ultrasound with reduced intra--and inter-observer variability.

  14. 3D ultrafast ultrasound imaging in vivo

    NASA Astrophysics Data System (ADS)

    Provost, Jean; Papadacci, Clement; Esteban Arango, Juan; Imbault, Marion; Fink, Mathias; Gennisson, Jean-Luc; Tanter, Mickael; Pernot, Mathieu

    2014-10-01

    Very high frame rate ultrasound imaging has recently allowed for the extension of the applications of echography to new fields of study such as the functional imaging of the brain, cardiac electrophysiology, and the quantitative imaging of the intrinsic mechanical properties of tumors, to name a few, non-invasively and in real time. In this study, we present the first implementation of Ultrafast Ultrasound Imaging in 3D based on the use of either diverging or plane waves emanating from a sparse virtual array located behind the probe. It achieves high contrast and resolution while maintaining imaging rates of thousands of volumes per second. A customized portable ultrasound system was developed to sample 1024 independent channels and to drive a 32  ×  32 matrix-array probe. Its ability to track in 3D transient phenomena occurring in the millisecond range within a single ultrafast acquisition was demonstrated for 3D Shear-Wave Imaging, 3D Ultrafast Doppler Imaging, and, finally, 3D Ultrafast combined Tissue and Flow Doppler Imaging. The propagation of shear waves was tracked in a phantom and used to characterize its stiffness. 3D Ultrafast Doppler was used to obtain 3D maps of Pulsed Doppler, Color Doppler, and Power Doppler quantities in a single acquisition and revealed, at thousands of volumes per second, the complex 3D flow patterns occurring in the ventricles of the human heart during an entire cardiac cycle, as well as the 3D in vivo interaction of blood flow and wall motion during the pulse wave in the carotid at the bifurcation. This study demonstrates the potential of 3D Ultrafast Ultrasound Imaging for the 3D mapping of stiffness, tissue motion, and flow in humans in vivo and promises new clinical applications of ultrasound with reduced intra—and inter-observer variability.

  15. Carbon Nanomaterials Interfacing with Neurons: An In vivo Perspective

    PubMed Central

    Baldrighi, Michele; Trusel, Massimo; Tonini, Raffaella; Giordani, Silvia

    2016-01-01

    Developing new tools that outperform current state of the art technologies for imaging, drug delivery or electrical sensing in neuronal tissues is one of the great challenges in neurosciences. Investigations into the potential use of carbon nanomaterials for such applications started about two decades ago. Since then, numerous in vitro studies have examined interactions between these nanomaterials and neurons, either by evaluating their compatibility, as vectors for drug delivery, or for their potential use in electric activity sensing and manipulation. The results obtained indicate that carbon nanomaterials may be suitable for medical therapies. However, a relatively small number of in vivo studies have been carried out to date. In order to facilitate the transformation of carbon nanomaterial into practical neurobiomedical applications, it is essential to identify and highlight in the existing literature the strengths and weakness that different carbon nanomaterials have displayed when probed in vivo. Unfortunately the current literature is sometimes sparse and confusing. To offer a clearer picture of the in vivo studies on carbon nanomaterials in the central nervous system, we provide a systematic and critical review. Hereby we identify properties and behavior of carbon nanomaterials in vivo inside the neural tissues, and we examine key achievements and potentially problematic toxicological issues. PMID:27375413

  16. In vivo static field perturbations in magnetic resonance

    NASA Astrophysics Data System (ADS)

    Koch, Kevin Matthew

    2007-12-01

    Fundamental magnetic resonance (MR) theory assumes the spatial homogeneity of a dominating static magnetic field B = B 0ẑ. When this assumption is violated, a myriad of artifacts and compromising factors are introduced to MR spectra and images. Though in vivo nuclear magnetic resonance (NMR) is one of the most widely used scientific and diagnostic tools in medicine and biology, it remains haunted by the continual and persistant ghost of B0 inhomogeneity. An inclusive list of in vivo NMR applications severely impacted by B0 inhomogeneity could go on ad infinitum. Examples of such applications include neurosurgical utility in functional magnetic resonance imaging (fMRI), cerebral metabolic flux mapping, cerebral diffusion tractography, and abdominal diagnostic imaging. Given this wide impact on in vivo NMR, significant effort has been exerted in developing methods of compensating B0 inhomogeneity. Complicating this task is the sample-specific nature of in vivo B 0 inhomogeneity and its exacerbation with ever increasing B 0 field strengths. State of the art B 0 inhomogeneity compensation is currently at a critical juncture where homogenization demands are overwhelming the outer capabilities of existing technology and methods. This thesis addresses the B 0 inhomogeneity problem in the mammalian brain and presents novel solutions to the homogenization technology stalemate.

  17. In vitro/in vivo correlations in transdermal product development.

    PubMed

    Ghosh, Priyanka; Milewski, Mikolaj; Paudel, Kalpana

    2015-01-01

    As per the US FDA's guidance for industry entitled 'Extended Release Oral Dosage Forms: Development, Evaluation, and Application of In Vitro/In Vivo Correlations', in vitro-in vivo correlations (IVIVC) can be used to establish a dissolution test as a surrogate for human bioequivalence studies and certain scale-up and postapproval changes. However, at the present time, establishment of a transdermal IVIVC is not used to support biowaiver claims in late phases of clinical development or postapproval changes (major formulation changes, i.e., >10% changes in inactive ingredients) to the best of the authors' knowledge. The value of developing an IVIVC for percutaneous drugs lies mainly in facilitating permeation testing of transdermal drug candidates and formulation performance optimization at much lower cost as compared with carrying out multiple in vivo studies. The present article will introduce the concept of transdermal IVIVC, outlining certain limitations to its applicability, in vitro and in vivo methods, regulatory product development requirements and the most common approaches to establish an IVIVC for a transdermal drug. Additionally, this article will also summarize some challenges and recent advancements in this field, along with selected academic examples of transdermal IVIVCs.

  18. Comparative real-time study of cellular uptake of a formulated conjugated linolenic acid rich nano and conventional macro emulsions and their bioactivity in ex vivo models for parenteral applications.

    PubMed

    Paul, Debjyoti; Mukherjee, Sayani; Chakraborty, Rajarshi; Mallick, Sanjaya K; Dhar, Pubali

    2015-02-01

    The objective of the present study was to fabricate and monitor real-time, impact of a stable conjugated linolenic acid, α-eleostearic acid (ESA) rich nanoemulsion (NE) formulation (d < 200 nm) vis-à-vis ESA conventional emulsion (CE) system in ex vivo systems against both endogenous and exogenous reactive oxygen species (ROS). Accordingly, stable nanoemulsion formulation of ESA was engineered with the aid of bitter melon seed oil and non-toxic excipients. Morphology and particle size of the emulsion formulations were studied to validate stability. The real-time rapid uptake of the ESA NE and its increased prophylactic efficacy against induced endogenous and exogenous ROS in terms of cell viability and membrane integrity was evaluated flow-cytometrically and with fluorescence microscopic analysis of different primary cells. It was found that the fabricated non-toxic ESA NE had stable parameters (hydrodynamic mean diameter, particle size distribution and zeta potential) for over 12 weeks. Further, ESA NE at a concentration of ∼ 70 μM exhibited maximum efficacy in protecting cells from oxidative damage against both endogenous and exogenous ROS in lymphocytes and hepatocytes as compared to its corresponding presence in the CE formulation. This study provides a real-time empirical evidence on the influence of nano formulation in enhancing bioavailability and antioxidative properties of ESA.

  19. The in vivo activation of persistent nanophosphors for optical imaging of vascularization, tumours and grafted cells

    NASA Astrophysics Data System (ADS)

    Maldiney, Thomas; Bessière, Aurélie; Seguin, Johanne; Teston, Eliott; Sharma, Suchinder K.; Viana, Bruno; Bos, Adrie J. J.; Dorenbos, Pieter; Bessodes, Michel; Gourier, Didier; Scherman, Daniel; Richard, Cyrille

    2014-04-01

    Optical imaging for biological applications requires more sensitive tools. Near-infrared persistent luminescence nanoparticles enable highly sensitive in vivo optical detection and complete avoidance of tissue autofluorescence. However, the actual generation of persistent luminescence nanoparticles necessitates ex vivo activation before systemic administration, which prevents long-term imaging in living animals. Here, we introduce a new generation of optical nanoprobes, based on chromium-doped zinc gallate, whose persistent luminescence can be activated in vivo through living tissues using highly penetrating low-energy red photons. Surface functionalization of this photonic probe can be adjusted to favour multiple biomedical applications such as tumour targeting. Notably, we show that cells can endocytose these nanoparticles in vitro and that, after intravenous injection, we can track labelled cells in vivo and follow their biodistribution by a simple whole animal optical detection, opening new perspectives for cell therapy research and for a variety of diagnosis applications.

  20. In vivo imaging of sulfotransferases

    DOEpatents

    Barrio, Jorge R; Kepe, Vladimir; Small, Gary W; Satyamurthy, Nagichettiar

    2013-02-12

    Radiolabeled tracers for sulfotransferases (SULTs), their synthesis, and their use are provided. Included are substituted phenols, naphthols, coumarins, and flavones radiolabeled with .sup.18F, .sup.123I, .sup.124I, .sup.125I, or .sup.11C. Also provided are in vivo techniques for using these and other tracers as analytical and diagnostic tools to study sulfotransferase distribution and activity, in health and disease, and to evaluate therapeutic interventions.