scholarly journals Spectral Characterization of Stem Cell-Derived Myelination within the Injured Adult PNS Using the Solvatochromic Dye Nile Red

Cells ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 189
Author(s):  
Joey Grochmal ◽  
Wulin Teo ◽  
Hardeep Gambhir ◽  
Ranjan Kumar ◽  
Jo Anne Stratton ◽  
...  
Keyword(s):  
Photophysical Properties ◽  
Spectral Characteristics ◽  
Nile Red ◽  
Emission Spectra ◽  
Membrane Composition ◽  
Myelin Membrane ◽  
Biochemical Processes ◽  
Solvatochromic Dye

Background: Myelin is an essential component of the peripheral and central nervous system, enabling fast axonal conduction and supporting axonal integrity; limited tools exist for analysis of myelin composition in-vivo. Objective: To demonstrate that the photophysical properties of myelin-incorporated solvatochromic dyes can be exploited to probe the biochemical composition of living peripheral nerve myelin at high spatial resolution. Methods: Using the myelin-incorporated fluorescent dye Nile Red we sequentially analyzed the spectral characteristics of remyelinating myelin membranes both in-vitro and in-vivo, including in living rats. Results: We demonstrated a consistent bi-phasic evolution of emission spectra during early remyelination, and visually report the reliable biochemical flux of myelin membrane composition in-vitro and in-vivo. Conclusions: Solvatochromic spectroscopy enables the analysis of myelin membrane maturity during remyelination, and can be performed in-vivo. As the formation of myelin during early-to-late remyelination likely incorporates fluctuating fractions of lipophilic components and changes in lateral membrane mobility, we propose that our spectrochemical data reflects the observation of these biochemical processes.

CD200 is a novel p53-target gene involved in apoptosis-associated immune tolerance

Blood ◽  
2004 ◽  
Vol 103 (7) ◽  
pp. 2691-2698 ◽  
Author(s):  
Michael D. Rosenblum ◽  
Edit Olasz ◽  
Jeffery E. Woodliff ◽  
Bryon D. Johnson ◽  
Marja C. Konkol ◽  
...  
Keyword(s):  
Target Gene ◽  
Molecular Mechanisms ◽  
Apoptotic Cell ◽  
Immune Reactivity ◽  
Biochemical Processes ◽  
The Face ◽  
P53 Target Gene ◽  
Self Antigens

Abstract During apoptotic cell death, biochemical processes modify self-proteins and create potential autoantigens. To maintain self-tolerance in the face of natural cell turnover, the immune system must prevent or control responses to apoptosis-associated autoantigens or risk autoimmunity. The molecular mechanisms governing this process remain largely unknown. Here, we show that expression of the immunoregulatory protein CD200 increases as murine dendritic cells (DCs) undergo apoptosis. We define CD200 as a p53-target gene and identify both p53- and caspase-dependent pathways that control CD200 expression during apoptosis. CD200 expression on apoptotic DCs diminishes proinflammatory cytokine production in response to self-antigens in vitro and is required for UVB-mediated tolerance to haptenated self-proteins in vivo. Up-regulation of CD200 may represent a novel mechanism, whereby immune reactivity to apoptosis-associated self-antigens is suppressed under steady state conditions. (Blood. 2004;103: 2691-2698)

scholarly journals Water-soluble cyclometalated platinum(ii) and iridium(iii) complexes: synthesis, tuning of the photophysical properties, and in vitro and in vivo phosphorescence lifetime imaging

RSC Advances ◽  
2018 ◽  
Vol 8 (31) ◽  
pp. 17224-17236 ◽  
Author(s):  
Anastasia I. Solomatina ◽  
Shih-Hao Su ◽  
Maria M. Lukina ◽  
Varvara V. Dudenkova ◽  
Vladislav I. Shcheslavskiy ◽  
...  
Keyword(s):  
Photophysical Properties ◽  
Water Soluble ◽  
Iridium Complexes ◽  
Hypoxia Imaging ◽  
Phosphorescence Lifetime ◽  
Lifetime Imaging ◽  
Phosphorescence Lifetime Imaging

Novel water-soluble iridium complexes with sulfonated diphosphine allow in vitro and in vivo lifetime hypoxia imaging.

161 LIPID CONTENT, RESISTANCE TO CRYOPRESERVATION, AND SEX RATIO OF IN VITRO BOVINE BLASTOCYSTS PRODUCED IN A SERUM-FREE SYSTEM

2006 ◽  
Vol 18 (2) ◽  
pp. 188
Author(s):  
F. George ◽  
C. Daniaux ◽  
G. Genicot ◽  
F. Focant ◽  
B. Verhaeghe ◽  
...  
Keyword(s):  
Sex Ratio ◽  
Lipid Content ◽  
Culture Medium ◽  
Nile Red ◽  
Embryo Cryopreservation ◽  
Hatching Rate ◽  
Free System ◽  
Serum Free

In vitro-produced (IVP) bovine blastocysts are known to be more sensitive to cryopreservation than their in vivo counterparts. Removing serum from the culture medium decreases sanitary risk and could improve embryo resistance to cryopreservation by preventing the accumulation of intracellular lipids. Our objectives were to evaluate the lipid content, resistance to cryopreservation, and sex ratio of IVP embryos cultured in a serum-free system. Oocytes from slaughterhouse ovaries were matured in a serum-free enriched medium (Donnay et al. 2004 Reprod. Fertil. Dev. 16, 274) and cultured in 5% O2 in modified SOF supplemented with 5% FCS (FCS) or with insulin-transferrin-selenium (ITS) and 0.1 mg/mL polyvinylpyrrolidone (PVP) (ITS-PVP) or 4 mg/mL BSA (ITS-BSA) (Daniaux et al. 2005 Reprod. Fertil. Dev. 17, 217). Day 5 morulae were stained with the fluorescent dye Nile Red in order to evaluate their lipid content (Genicot et al. 2005 Theriogenology 63, 1181). Day 7 blastocysts (diameter ≥160 µm) were selected, classified according to their size, and frozen in HEPES-SOF containing 1.5 M ethylene glycol, 0.1 M sucrose, and 1.8 mg/mL wheat peptones (George et al. 2002 Reproduction 29, 51). The lipid content was significantly lower in morulae cultured in ITS-BSA compared with the two other media (320 ± 10 arbitrary fluorescence units vs. 383 ± 12 in FCS and 406 ± 10 in ITS-PVP; n = 271; ANOVA2: P < 0.01). After cryopreservation, a higher total hatching rate was found 24 h post-thawing in blastocysts cultured in ITS-BSA and for both serum-free conditions at 48 h (Table 1). In particular, embryos ≤180 µm cultured in FCS were less resistant to cryopreservation than embryos of the same size produced without serum. Expanded blastocysts cultured in ITS-BSA were sexed by PCR (Grisart et al. 1995 Theriogenology 43, 1097) and a higher proportion of male embryos was found (62.7%; n = 51). In conclusion, a complete serum-free system was set up from oocyte maturation to embryo cryopreservation that gave high quality embryos resistant to cryo-preservation. Embryos produced in ITS-BSA presented a lower lipid content, but a shift of the expanded blastocyst sex ratio toward males was observed. Table 1. Hatching rates post-thawing as a function of the blastocyst size and the culture medium

201. Effects of relaxin deficiency on matrix metalloproteinase expression in the cervix and vagina of pregnant mice

2005 ◽  
Vol 17 (9) ◽  
pp. 76
Author(s):  
J. T. McGuane ◽  
H. M. Gehring ◽  
L. J. Parry
Keyword(s):  
Reproductive Tract ◽  
Late Gestation ◽  
Mrna Levels ◽  
Biochemical Processes ◽  
Fluorogenic Probes ◽  
Pregnant Mice ◽  
Ecm Degradation ◽  
Stromal Collagen

The major functions of relaxin are associated with female reproductive physiology, especially the regulation of biochemical processes involved in the remodelling of the reproductive tract at term. Studies in relaxin deficient mice (Rlx–/–) demonstrate that although females give birth to live young without apparent dystocia, they have abnormal cervices and vaginae. This phenotype is attributed to an increase in stromal collagen, but the mechanism(s) by which relaxin regulates extracellular matrix (ECM) production in reproductive tissues is poorly understood. In this study, we assessed the expression of matrix metalloproteinases (MMPs) in the cervix and vagina of pregnant wild-type (Rlx+/+) and Rlx–/– mice. Tissues were obtained from adult C57/Blk6J Rlx+/+ mice on days 7.5, 14.5, 17.5, 18.5 pc and Rlx–/– littermates on days 7.5, 14.5 and 18.5 pc. Real-time PCR using dual-labelled fluorogenic probes was performed in an Opticon 2 cycler (MJ Research) to quantify MMP-2, -3, -7, -9 and -13 gene expression. In the cervix and vagina of Rlx+/+ mice, only MMP-2 mRNA levels were significantly higher at term compared with earlier stages of gestation. There were significant decreases in MMP-7 and -13 expression at term, but no change in MMP-3 and -9. In contrast, MMP-3, -7, -9 and -13 mRNA levels were significantly higher in the cervix and vagina of late pregnant Rlx–/– mice. The expression of MMP-2 did not differ between Rlx+/+ and Rlx–/– mice at term. Despite the higher expression of the majority of MMPs we examined in Rlx–/– mice, there was no histological evidence of increased ECM degradation in the cervix and vagina in late gestation. Although previous in vitro studies suggest that relaxin positively regulates MMP activity, our data demonstrate that relaxin deficiency does not result in decreased MMP expression in the mouse cervix and vagina in vivo.

scholarly journals Lipophilic Dye Staining of Cryptococcus neoformans Extracellular Vesicles and Capsule

2009 ◽  
Vol 8 (9) ◽  
pp. 1373-1380 ◽  
Author(s):  
André Moraes Nicola ◽  
Susana Frases ◽  
Arturo Casadevall
Keyword(s):  
Cryptococcus Neoformans ◽  
Extracellular Vesicles ◽  
Capsular Polysaccharide ◽  
Complex Structure ◽  
Spectral Characteristics ◽  
Acid Dyes ◽  
Whole Cells ◽  
Direct Binding

ABSTRACT Cryptococcus neoformans is an encapsulated yeast that causes systemic mycosis in immunosuppressed individuals. Recent studies have determined that this fungus produces vesicles that are released to the extracellular environment both in vivo and in vitro. These vesicles contain assorted cargo that includes several molecules associated with virulence and implicated in host-pathogen interactions, such as capsular polysaccharides, laccase, urease, and other proteins. To date, visualization of extracellular vesicles has relied on transmission electron microscopy, a time-consuming technique. In this work we report the use of fluorescent membrane tracers to stain lipophilic structures in cryptococcal culture supernatants and capsules. Two dialkylcarbocyanine probes with different spectral characteristics were used to visualize purified vesicles by fluorescence microscopy and flow cytometry. Dual staining of vesicles with dialkylcarbocyanine and RNA-selective nucleic acid dyes suggested that a fraction of the vesicle population carried RNA. Use of these dyes to stain whole cells, however, was hampered by their possible direct binding to capsular polysaccharide. A fluorescent phospholipid was used as additional membrane tracer to stain whole cells, revealing punctate structures on the edge of the capsule which are consistent with vesicular trafficking. Lipophilic dyes provide new tools for the study of fungal extracellular vesicles and their content. The finding of hydrophobic regions in the capsule of C. neoformans adds to the growing evidence for a structurally complex structure composed of polysaccharide and nonpolysaccharide components.

scholarly journals The Novel Antitumor Compound HCA Promotes Glioma Cell Death by Inducing Endoplasmic Reticulum Stress and Autophagy

Cancers ◽  
2021 ◽  
Vol 13 (17) ◽  
pp. 4290
Author(s):  
Roberto Beteta-Göbel ◽  
Javier Fernández-Díaz ◽  
Laura Arbona-González ◽  
Raquel Rodríguez-Lorca ◽  
Manuel Torres ◽  
...  
Keyword(s):  
Cell Death ◽  
Primary Brain Tumor ◽  
Signal Pathways ◽  
Membrane Composition ◽  
Negative Effect ◽  
Human Gbm ◽  
In Vivo Growth ◽  
Cellular Metabolite

Glioblastoma (GBM) is the most common and aggressive type of primary brain tumor in adults, and the median survival of patients with GBM is 14.5 months. Melitherapy is an innovative therapeutic approach to treat different diseases, including cancer, and it is based on the regulation of cell membrane composition and structure, which modulates relevant signal pathways. Here, we have tested the effects of 2-hydroxycervonic acid (HCA) on GBM cells and xenograft tumors. HCA was taken up by cells and it compromised the survival of several human GBM cell lines in vitro, as well as the in vivo growth of xenograft tumors (mice) derived from these cells. HCA appeared to enhance ER stress/UPR signaling, which consequently induced autophagic cell death of the GBM tumor cells. This negative effect of HCA on GBM cells may be mediated by the JNK/c-Jun/CHOP/BiP axis, and it also seems to be provoked by the cellular metabolite of HCA, C21:5n-3 (heneicosapentaenoic acid). These results demonstrate the efficacy of the melitherapeutic treatment used and the potential of using C21:5n-3 as an efficacy biomarker for this treatment. Given the safety profile in animal models, the data presented here provide evidence that HCA warrants further clinical study as a potential therapy for GBM, currently an important unmet medical need.

Preparation of a Nile Red–Pd-based fluorescent CO probe and its imaging applications in vitro and in vivo

2018 ◽  
Vol 13 (5) ◽  
pp. 1020-1033 ◽  
Author(s):  
Keyin Liu ◽  
Xiuqi Kong ◽  
Yanyan Ma ◽  
Weiying Lin
Keyword(s):  
Nile Red ◽  
Co Probe

scholarly journals Changes in Spectral Fluorescence Properties of a Near-Infrared Photosensitizer in a Nanoform as a Coating of an Optical Fiber Neuroport

Photonics ◽  
2021 ◽  
Vol 8 (12) ◽  
pp. 556
Author(s):  
Yuliya Maklygina ◽  
Igor Romanishkin ◽  
Aleksej Skobeltsin ◽  
Dina Farrakhova ◽  
Sergej Kharnas ◽  
...  
Keyword(s):  
Immune System ◽  
Spectral Region ◽  
Near Infrared ◽  
Fiber Optic ◽  
Spectral Characteristics ◽  
Infrared Spectral Region ◽  
Advanced Therapies ◽  
Infrared Spectral

In this work, we tested a new approach to assess the presence of inflammatory process in the implant area using spectral methods and the technique of fiber fluorescence analysis of photosensitizers in nanoform. First of all, the spectral characteristics of the photosensitizer when interacting with the porous surface of the implant, based on hydroxyapatite under in vitro and in vivo conditions, were determined. Thus, it was shown that spectral characteristics of photosensitizers can be used for judgement on the process of inflammation in the implant area and thus on the local presence of the immunocompetent cells. The analysis was performed at a sufficient depth in the biotissue by using the near-infrared spectral region, as well as two different methods: fiber-based laser spectroscopy and fiber-optic neuroscopy, which served to monitor the process and regular fluorescence diagnosis of the studied area. Fluorescence spectroscopic analysis was performed on experimental animals in vivo, i.e., under conditions of active immune system intervention, as well as on cell cultures in vitro in order to judge the role of the immune system in the interaction with the implant in comparison. Thus, the aim of the study was to determine the relationship between the fluorescence signal of nanophotosensitizers in the near infrared spectral region and its parameters with the level of inflammation and the type of surface with which the photosensitizer interacts in the implant area. Thus, fiber-optic control opens up new approaches for further diagnosis and therapy in the implant area, making immune cells a prime target for advanced therapies.

scholarly journals Coumarin-Based Profluorescent and Fluorescent Substrates for Determining Xenobiotic-Metabolizing Enzyme Activities In Vitro

2020 ◽  
Vol 21 (13) ◽  
pp. 4708
Author(s):  
Hannu Raunio ◽  
Olli Pentikäinen ◽  
Risto O. Juvonen
Keyword(s):  
Enzyme Activities ◽  
Biological Activities ◽  
Photophysical Properties ◽  
Fluorescent Sensors ◽  
Enzyme Assays ◽  
Highly Sensitive ◽  
Metabolizing Enzyme ◽  
Transfer Properties

in vivo methods, such as spectrophotometric, fluorometric, mass spectrometric,and radioactivity-based techniques. In fluorescence-based assays, the reaction produces a fluorescentproduct from a nonfluorescent substrate or vice versa. Fluorescence-based enzyme assays areusually highly sensitive and specific, allowing measurements on small specimens of tissues withlow enzyme activities. Fluorescence assays are also amenable to miniaturization of the reactionmixtures and can thus be done in high throughput. 7-Hydroxycoumarin and its derivatives arewidely used as fluorophores due to their desirable photophysical properties. They possess a large -conjugated system with electron-rich and charge transfer properties. This conjugated structure leadsto applications of 7-hydroxycoumarins as fluorescent sensors for biological activities. We describe inthis review historical highlights and current use of coumarins and their derivatives in evaluatingactivities of the major types of xenobiotic-metabolizing enzyme systems. Traditionally, coumarinsubstrates have been used to measure oxidative activities of cytochrome P450 (CYP) enzymes. For thispurpose, profluorescent coumarins are very sensitive, but generally lack selectivity for individual CYPforms. With the aid of molecular modeling, we have recently described several new coumarin-basedsubstrates for measuring activities of CYP and conjugating enzymes with improved selectivity.

scholarly journals Evaluating Brightness and Spectral Properties of Click Beetle and Firefly Luciferases Using Luciferin Analogues: Identification of Preferred Pairings of Luciferase and Substrate for In Vivo Bioluminescence Imaging

2020 ◽  
Vol 22 (6) ◽  
pp. 1523-1531
Author(s):  
Giorgia Zambito ◽  
Natasa Gaspar ◽  
Yanto Ridwan ◽  
Mary P. Hall ◽  
Ce Shi ◽  
...  
Keyword(s):  
Near Infrared ◽  
Bioluminescence Imaging ◽  
Spectral Characteristics ◽  
Photon Emission ◽  
Ccd Camera ◽  
Spectral Unmixing ◽  
Tissue Imaging ◽  
Click Beetle

Abstract Purpose Currently, a variety of red and green beetle luciferase variants are available for bioluminescence imaging (BLI). In addition, new luciferin analogues providing longer wavelength luminescence have been developed that show promise for improved deep tissue imaging. However, a detailed assessment of these analogues (e.g., Akalumine-HCl, CycLuc1, and amino naphthyl luciferin (NH2-NpLH2)) combined with state of the art luciferases has not been performed. The aim of this study was to evaluate for the first time the in vivo brightness and spectral characteristics of firefly (Luc2), click beetle green (CBG99), click beetle red 2 (CBR2), and Akaluc luciferases when paired with different d-luciferin (d-LH2) analogues in vivo. Procedures Transduced human embryonic kidney (HEK 293T) cells expressing individual luciferases were analyzed both in vitro and in mice (via subcutaneous injection). Following introduction of the luciferins to cells or animals, the resulting bioluminescence signal and photon emission spectrum were acquired using a sensitive charge-coupled device (CCD) camera equipped with a series of band pass filters and spectral unmixing software. Results Our in vivo analysis resulted in four primary findings: (1) the best substrate for Luc2, CBG99, and CBR2 in terms of signal strength was d-luciferin; (2) the spectra for Luc2 and CBR2 were shifted to a longer wavelength when Akalumine-HCl was the substrate; (3) CBR2 gave the brightest signal with the near-infrared substrate, NH2-NpLH2; and (4) Akaluc was brighter when paired with either CycLuc1 or Akalumine-HCl when paired with d-LH2. Conclusion We believe that the experimental results described here should provide valuable guidance to end users for choosing the correct luciferin/luciferase pairs for a variety of BLI applications.

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