ALTERATIONS IN AUTOFLUORESCENCE SIGNAL FROM RAT SKIN EX VIVO UNDER OPTICAL IMMERSION CLEARING

2010 ◽  
Vol 03 (03) ◽  
pp. 147-152 ◽  
Author(s):  
E. V. MIGACHEVA ◽  
A. B. PRAVDIN ◽  
V. V. TUCHIN
Keyword(s):  
Fluorescence Spectroscopy ◽  
Ex Vivo ◽  
Rat Skin ◽  
Optical Clearing ◽  
Spectral Curves ◽  
The Common ◽  
First Time ◽  
Tissue Optical Clearing ◽  
Optical Immersion

For the first time, the changes in autofluorescence spectra of ex vivo rat skin have been experimentally investigated using the combination of fluorescence spectroscopy and optical immersion clearing. The glucose, glycerol and propylene glycol solutions were used as clearing agents. The optical clearing was performed from the dermal side of skin imitating the in vivo injection of clearing agent under the dermal layers. In this contribution, the common properties of autofluorescence variation during optical immersion clearing were determined. The tendency of autofluorescence signal to decrease with reduction of scattering in tissue was noticed and discussed in detail. However, the differences in the shape of spectral curves under application of different clearing agents showed that optical clearing affects the autofluorescence properties of tissue differently depending on the type of clearing liquid. The results obtained are useful for the understanding of tissue optical clearing mechanisms and for improving techniques such as fluorescence spectroscopy.

Kinetics of Rat Skin Optical Clearing at Topical Application of 40%Glucose: Ex Vivo and In Vivo Studies

2019 ◽  
Vol 25 (1) ◽  
pp. 1-8 ◽  
Author(s):  
Daria K. Tuchina ◽  
Polina A. Timoshina ◽  
Valery V. Tuchin ◽  
Alexey N. Bashkatov ◽  
Elina A. Genina
Keyword(s):  
Topical Application ◽  
Ex Vivo ◽  
Rat Skin ◽  
Optical Clearing ◽  
Kinetics Of

scholarly journals Alkylated epidermal creatine kinase as a biomarker for sulfur mustard exposure: comparison to adducts of albumin and DNA in an in vivo rat study

2021 ◽  
Author(s):  
Dirk Steinritz ◽  
Robin Lüling ◽  
Markus Siegert ◽  
Julia Herbert ◽  
Harald Mückter ◽  
...  
Keyword(s):  
Creatine Kinase ◽  
Dna Adducts ◽  
Sulfur Mustard ◽  
Ex Vivo ◽  
Immunomagnetic Separation ◽  
Chemical Warfare Agent ◽  
Cysteine Residue ◽  
Islamic State ◽  
Rat Skin

AbstractSulfur mustard (SM) is a chemical warfare agent which use is banned under international law and that has been used recently in Northern Iraq and Syria by the so-called Islamic State. SM induces the alkylation of endogenous proteins like albumin and hemoglobin thus forming covalent adducts that are targeted by bioanalytical methods for the verification of systemic poisoning. We herein report a novel biomarker, namely creatine kinase (CK) B-type, suitable as a local biomarker for SM exposure on the skin. Human and rat skin were proven to contain CK B-type by Western blot analysis. Following exposure to SM ex vivo, the CK-adduct was extracted from homogenates by immunomagnetic separation and proteolyzed afterwards. The cysteine residue Cys282 was found to be alkylated by the SM-specific hydroxyethylthioethyl (HETE)-moiety detected as the biomarker tetrapeptide TC(-HETE)PS. A selective and sensitive micro liquid chromatography-electrospray ionization high-resolution tandem-mass spectrometry (µLC-ESI MS/HRMS) method was developed to monitor local CK-adducts in an in vivo study with rats percutaneously exposed to SM. CK-adduct formation was compared to already established DNA- and systemic albumin biomarkers. CK- and DNA-adducts were successfully detected in biopsies of exposed rat skin as well as albumin-adducts in plasma. Relative biomarker concentrations make the CK-adduct highly appropriate as a local dermal biomarker. In summary, CK or rather Cys282 in CK B-type was identified as a new, additional dermal target of local SM exposures. To our knowledge, it is also the first time that HETE-albumin adducts, and HETE-DNA adducts were monitored simultaneously in an in vivo animal study.

scholarly journals Extra-gastric expression of the proton pump H+/K+-ATPase in the gills and kidney of the teleost Oreochromis niloticus

2020 ◽  
Vol 223 (16) ◽  
pp. jeb214890
Author(s):  
Ebtesam Ali Barnawi ◽  
Justine E. Doherty ◽  
Patrícia Gomes Ferreira ◽  
Jonathan M. Wilson
Keyword(s):  
Oreochromis Niloticus ◽  
Cellular Localization ◽  
Ex Vivo ◽  
Uptake Inhibition ◽  
Custom Made ◽  
Potassium Regulation ◽  
Immunohistochemical Techniques ◽  
Collecting Tubules ◽  
First Time

ABSTRACTPotassium regulation is essential for the proper functioning of excitable tissues in vertebrates. The H+/K+-ATPase (HKA), which is composed of the HKα1 (gene: atp4a) and HKβ (gene: atp4b) subunits, has an established role in potassium and acid–base regulation in mammals and is well known for its role in gastric acidification. However, the role of HKA in extra-gastric organs such as the gill and kidney is less clear, especially in fishes. In the present study in Nile tilapia, Oreochromis niloticus, uptake of the K+ surrogate flux marker rubidium (Rb+) was demonstrated in vivo; however, this uptake was not inhibited with omeprazole, a potent inhibitor of the gastric HKA. This contrasts with gill and kidney ex vivo preparations, where tissue Rb+ uptake was significantly inhibited by omeprazole and SCH28080, another gastric HKA inhibitor. The cellular localization of this pump in both the gill and kidney was demonstrated using immunohistochemical techniques with custom-made antibodies specific for Atp4a and Atp4b. Antibodies against the two subunits showed the same apical ionocyte distribution pattern in the gill and collecting tubules/ducts in the kidney. Atp4a antibody specificity was confirmed by western blotting. RT-PCT was used to confirm the expression of both subunits in the gill and kidney. Taken together, these results indicate for the first time K+ (Rb+) uptake in O. niloticus and that HKA is implicated, as shown through the ex vivo uptake inhibition by omeprazole and SCH28080, verifying a role for HKA in K+ absorption in the gill's ionocytes and collecting tubule/duct segments of the kidney.

In Vivo and ex Vivo Approaches to Studying the Biomechanical Properties of Healing Wounds in Rat Skin

2013 ◽  
Vol 135 (10) ◽  
Author(s):  
Clare Y. L. Chao ◽  
Gabriel Y. F. Ng ◽  
Kwok-Kuen Cheung ◽  
Yong-Ping Zheng ◽  
Li-Ke Wang ◽  
...  
Keyword(s):  
Wound Healing ◽  
Ex Vivo ◽  
Normal Skin ◽  
Biomechanical Properties ◽  
Skin Wound ◽  
Sprague Dawley ◽  
Rat Skin ◽  
Mechanical Stiffness ◽  
Air Jet

An evaluation of wound mechanics is crucial in reflecting the wound healing status. The present study examined the biomechanical properties of healing rat skin wounds in vivo and ex vivo. Thirty male Sprague-Dawley rats, each with a 6 mm full-thickness circular punch biopsied wound at both posterior hind limbs were used. The mechanical stiffness at both the central and margins of the wound was measured repeatedly in five rats over the same wound sites to monitor the longitudinal changes over time of before wounding, and on days 0, 3, 7, 10, 14, and 21 after wounding in vivo by using an optical coherence tomography-based air-jet indentation system. Five rats were euthanized at each time point, and the biomechanical properties of the wound tissues were assessed ex vivo using a tensiometer. At the central wound bed region, the stiffness measured by the air-jet system increased significantly from day 0 (17.2%), peaked at day 7 (208.3%), and then decreased progressively until day 21 (40.2%) as compared with baseline prewounding status. The biomechanical parameters of the skin wound samples measured by the tensiometer showed a marked reduction upon wounding, then increased with time (all p < 0.05). On day 21, the ultimate tensile strength of the skin wound tissue approached 50% of the normal skin; while the stiffness of tissue recovered at a faster rate, reaching 97% of its prewounded state. Our results suggested that it took less time for healing wound tissues to recover their stiffness than their maximal strength in rat skin. The stiffness of wound tissues measured by air-jet could be an indicator for monitoring wound healing and contraction.

Abstract P283: Parkin Mediates Mitophagy in Cardioprotection

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Allen M Andres ◽  
Chengqun Huang ◽  
Eric P Ratliff ◽  
Genaro Hernandez ◽  
Pamela Lee ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Wild Type ◽  
Simulated Ischemia ◽  
Selective Targeting ◽  
Cardioprotective Effects ◽  
Mitochondrial Turnover ◽  
First Time

Autophagy-dependent mitochondrial turnover in response to cellular stress is necessary for maintaining cellular homeostasis. However, the mechanisms that govern the selective targeting of damaged mitochondria are poorly understood. Parkin, an E3 ubiquitin ligase, has been shown to be essential for the selective clearance of damaged mitochondria. Parkin is expressed in the heart, yet its function has not been investigated in the context of cardioprotection. We previously reported that autophagy is required for cardioprotection by ischemic preconditioning (IPC). In the present study, we used simulated ischemia in vitro and IPC in hearts (in vivo and ex vivo) to investigate the role of Parkin in mediating cardioprotection. In HL-1 cells, simulated ischemia induced Parkin translocation to mitochondria and mitochondrial elimination. Mitochondrial loss was blunted in Atg5-deficient cells, revealing the requirement for autophagy in mitochondrial elimination. Consistent with previous reports implicating p62/SQSTM1 in mitophagy, we found that downregulation of p62 attenuated mitophagy and exacerbated cell death in HL-1 cardiomyocytes subjected to simulated ischemia. While wild type mice showed p62 translocation to mitochondria after IPC, Parkin knockout mice exhibited attenuated translocation of p62 to mitochondria. Importantly, ablation of Parkin in mice abolished the cardioprotective effects of IPC. These results reveal for the first time the crucial role of Parkin and mitophagy in cardioprotection.

scholarly journals No cell is an island: circulating T cell:monocyte complexes are markers of immune perturbations

2018 ◽  
Author(s):  
Julie G. Burel ◽  
Mikhail Pomaznoy ◽  
Cecilia S. Lindestam Arlehamn ◽  
Daniela Weiskopf ◽  
Ricardo da Silva Antunes ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
T Cell ◽  
Immune Responses ◽  
Human Blood ◽  
Significant Proportion ◽  
Complex Frequency ◽  
The Common ◽  
First Time ◽  
High Resolution Microscopy

AbstractOur results highlight for the first time that a significant proportion of cell doublets in flow cytometry, previously believed to be the result of technical artefacts and thus ignored in data acquisition and analysis, are the result of true biological interaction between immune cells. In particular, we show that cell:cell doublets pairing a T cell and a monocyte can be directly isolated from human blood, and high resolution microscopy shows polarized distribution of LFA1/ICAM1 in many doublets, suggesting in vivo formation. Intriguingly, T cell:monocyte complex frequency and phenotype fluctuate with the onset of immune perturbations such as infection or immunization, reflecting expected polarization of immune responses. Overall these data suggest that cell doublets reflecting T cell-monocyte in vivo immune interactions can be detected in human blood and that the common approach in flow cytometry to avoid studying cell:cell complexes should be revisited.

scholarly journals Imaging depth extension of optical coherence tomography in rabbit eyes using optical clearing agents

2020 ◽  
Vol 245 (18) ◽  
pp. 1629-1636
Author(s):  
Ruiming Kong ◽  
Wenjuan Wu ◽  
Rui Qiu ◽  
Lei Gao ◽  
Fengxian Du ◽  
...  
Keyword(s):  
Optical Coherence Tomography ◽  
Ex Vivo ◽  
Light Attenuation ◽  
Anterior Segment ◽  
Posterior Segment ◽  
Glycerol Solution ◽  
Optical Coherence ◽  
Optical Clearing ◽  
Imaging Depth

Optical coherence tomography has become an indispensable diagnostic tool in ophthalmology for imaging the retina and the anterior segment of the eye. However, the imaging depth of optical coherence tomography is limited by light attenuation in tissues due to optical scattering and absorption. In this study of rabbit eye both ex vivo and in vivo, optical coherence tomography imaging depth of the anterior and posterior segments of the eye was extended by using optical clearing agents to reduce multiple scattering. The sclera, the iris, and the ciliary body were clearly visualized by direct application of glycerol at an incision on the conjunctiva, and the posterior boundary of sclera and even the deeper tissues were detected by submerging the posterior segment of eye in glycerol solution ex vivo or by retro-bulbar injection of glycerol in vivo. The ex vivo rabbit eyes recovered to their original state in 60 s after saline-wash treatment, and normal optical coherence tomography images of the posterior segment of the sample eyes proved the self-recovery of in vivo performance. Signal intensities of optical coherence tomography images obtained before and after glycerol treatment were compared to analysis of the effect of optical clearing. To the best of our knowledge, this is the first study for imaging depth extension of optical coherence tomography in both the anterior and posterior segments of eye by using optical clearing agents.

scholarly journals Candida albicans forms biofilms on the vaginal mucosa

Microbiology ◽  
2010 ◽  
Vol 156 (12) ◽  
pp. 3635-3644 ◽  
Author(s):  
M. M. Harriott ◽  
E. A. Lilly ◽  
T. E. Rodriguez ◽  
P. L. Fidel ◽  
M. C. Noverr
Keyword(s):  
Biofilm Formation ◽  
Ex Vivo ◽  
Microscopic Analysis ◽  
Vaginal Mucosa ◽  
First Time ◽  
Established Role ◽  
Type Strains

Current understanding of resistance and susceptibility to vulvovaginal candidiasis challenges existing paradigms of host defence against fungal infection. While abiotic biofilm formation has a clearly established role during systemic Candida infections, it is not known whether C. albicans forms biofilms on the vaginal mucosa and the possible role of biofilms in disease. In vivo and ex vivo murine vaginitis models were employed to examine biofilm formation by scanning electron and confocal microscopy. C. albicans strains included 3153A (lab strain), DAY185 (parental control strain), and mutants defective in morphogenesis and/or biofilm formation in vitro (efg1/efg1 and bcr1/bcr1). Both 3153A and DAY815 formed biofilms on the vaginal mucosa in vivo and ex vivo as indicated by high fungal burden and microscopic analysis demonstrating typical biofilm architecture and presence of extracellular matrix (ECM) co-localized with the presence of fungi. In contrast, efg1/efg1 and bcr1/bcr1 mutant strains exhibited weak or no biofilm formation/ECM production in both models compared to wild-type strains and complemented mutants despite comparable colonization levels. These data show for the first time that C. albicans forms biofilms in vivo on vaginal epithelium, and that in vivo biotic biofilm formation requires regulators of biofilm formation (BCR1) and morphogenesis (EFG1).

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