Ex vivo expansion of bovine corneal endothelial cells in xeno-free medium supplemented with platelet releasate.

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Ming-Li Chou

Full Text Available Clinical-grade ex vivo expansion of corneal endothelial cells can increase the availability of corneal tissues for transplantation and treatment of corneal blindness. However, these cells have very limited proliferative capacity. Successful propagation has required so far to use very complex growth media supplemented with fetal bovine serum and other xenocomponents. We hypothesized that human platelet releasates rich in multiple growth factors, and in particular neurotrophins, could potentially be a useful supplement for ex vivo expansion of corneal endothelium cells due to their neural crest origin. Platelet releasates were prepared by calcium salt activation of apheresis platelet concentrates, subjected or not to complement inactivation by heat treatment at 56°C for 30 minutes. Platelet releasates were characterized for their content in proteins and were found to contain high amount of growth factors including platelet-derived growth factor-AB (30.56 to 39.08 ng/ml and brain-derived neurotrophic factor (30.57 to 37.11 ng/ml neurotrophins. We compared the growth and viability of corneal endothelium cells in DMEM-F12 medium supplemented with different combinations of components, including 2.5%∼10% of the platelet releasates. Corneal endothelium cells expanded in platelet releasates exhibited good adhesion and a typical hexagonal morphology. Their growth and viability were enhanced when using the complement-inactivated platelet releasate at a concentration of 10%. Immunostaining and Western blots showed that CECs maintained the expressions of four important membrane markers: Na-K ATPase α1, zona occludens-1, phospho-connexin 43 and N-cadherin. In conclusion, our study provides the first proof-of-concept that human platelet releasates can be used for ex vivo expansion of corneal endothelium cells. These findings open a new paradigm for ex vivo propagation protocols of corneal endothelium cells in compliance with good tissue culture practices

PASSIVE CAVITATION DETECTION DURING PULSED HIFU EXPOSURES OF EX VIVO TISSUES AND IN VIVO MOUSE PANCREATIC TUMORS

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Li, Tong; Chen, Hong; Khokhlova, Tatiana; Wang, Yak-Nam; Kreider, Wayne; He, Xuemei; Hwang, Joo Ha

2014-01-01

Pulsed high-intensity focused ultrasound (pHIFU) has been demonstrated to enhance vascular permeability, disrupt tumor barriers and enhance drug penetration into tumor tissue through acoustic cavitation. Monitoring of cavitation activity during pHIFU treatments and knowing the ultrasound pressure levels sufficient to reliably induce cavitation in a given tissue are therefore very important. Here, three metrics of cavitation activity induced by pHIFU and evaluated by confocal passive cavitation detection were introduced: cavitation probability, cavitation persistence and the level of the broadband acoustic emissions. These metrics were used to characterize cavitation activity in several ex vivo tissue types (bovine tongue and liver and porcine adipose tissue and kidney) and gel phantoms (polyacrylamide and agarose) at varying peak-rarefactional focal pressures (1–12 MPa) during the following pHIFU protocol: frequency 1.1 MHz, pulse duration 1 ms, pulse repetition frequency 1 Hz. To evaluate the relevance of the measurements in ex vivo tissue, cavitation metrics were also investigated and compared in the ex vivo and in vivo murine pancreatic tumors that develop spontaneously in transgenic KPC mice and closely recapitulate human disease in their morphology. The cavitation threshold, defined at 50 % cavitation probability, was found to vary broadly among the investigated tissues (within 2.5–10 MPa), depending mostly on the water-lipid ratio that characterizes the tissue composition. Cavitation persistence and the intensity of broadband emissions depended both on tissue structure and lipid concentration. Both the cavitation threshold and broadband noise emission level were similar between ex vivo and in vivo pancreatic tumor tissue. The largest difference between in vivo and ex vivo settings was found in the pattern of cavitation occurrence throughout pHIFU exposure: it was sporadic in vivo, but ex vivo it decreased rapidly and stopped over the first few pulses

Passive cavitation detection during pulsed HIFU exposures of ex vivo tissues and in vivo mouse pancreatic tumors.

Science.gov (United States)

Li, Tong; Chen, Hong; Khokhlova, Tatiana; Wang, Yak-Nam; Kreider, Wayne; He, Xuemei; Hwang, Joo Ha

2014-07-01

Pulsed high-intensity focused ultrasound (pHIFU) has been shown to enhance vascular permeability, disrupt tumor barriers and enhance drug penetration into tumor tissue through acoustic cavitation. Monitoring of cavitation activity during pHIFU treatments and knowing the ultrasound pressure levels sufficient to reliably induce cavitation in a given tissue are therefore very important. Here, three metrics of cavitation activity induced by pHIFU and evaluated by confocal passive cavitation detection were introduced: cavitation probability, cavitation persistence and the level of the broadband acoustic emissions. These metrics were used to characterize cavitation activity in several ex vivo tissue types (bovine tongue and liver and porcine adipose tissue and kidney) and gel phantoms (polyacrylamide and agarose) at varying peak-rare factional focal pressures (1-12 MPa) during the following pHIFU protocol: frequency 1.1 MHz, pulse duration 1 ms and pulse repetition frequency 1 Hz. To evaluate the relevance of the measurements in ex vivo tissue, cavitation metrics were also investigated and compared in the ex vivo and in vivo murine pancreatic tumors that develop spontaneously in transgenic KrasLSL.G12 D/+; p53 R172 H/+; PdxCretg/+ (KPC) mice and closely re-capitulate human disease in their morphology. The cavitation threshold, defined at 50% cavitation probability, was found to vary broadly among the investigated tissues (within 2.5-10 MPa), depending mostly on the water-lipid ratio that characterizes the tissue composition. Cavitation persistence and the intensity of broadband emissions depended both on tissue structure and lipid concentration. Both the cavitation threshold and broadband noise emission level were similar between ex vivo and in vivo pancreatic tumor tissue. The largest difference between in vivo and ex vivo settings was found in the pattern of cavitation occurrence throughout pHIFU exposure: it was sporadic in vivo, but it decreased rapidly and stopped

Visualisation of distribution of gold nanoparticles in liver tissues ex vivo and in vitro using the method of optical coherence tomography

International Nuclear Information System (INIS)

Genina, Elina A; Terentyuk, G S; Khlebtsov, B N; Bashkatov, A N; Tuchin, Valerii V

2012-01-01

The possibility of visualising the distribution of gold nanoparticles in liver by means of the method of optical coherence tomography is studied experimentally in model samples of beef liver in vitro and rat liver ex vivo. In the experiments we used the gold nanoparticles in the form of nanocages with resonance absorption in the near-IR spectral region. In the model studies the suspension of nanoparticles was applied to the surface of the sample, which then was treated with ultrasound. In the ex vivo studies the suspension of nanoparticles was injected to the laboratory rats intravenously. The image contrast and the optical depth of detection of blood vessels and liver structure components are calculated, as well as the depth of liver optical probing before and after the injection of nanoparticles. It was shown that the administration of the nanoparticle increases significantly the imaging contrast of liver blood vessels owing to the localisation of the nanoparticles therein.

Visualisation of distribution of gold nanoparticles in liver tissues ex vivo and in vitro using the method of optical coherence tomography

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Genina, Elina A; Terentyuk, G S; Khlebtsov, B N; Bashkatov, A N; Tuchin, Valerii V

2012-06-30

The possibility of visualising the distribution of gold nanoparticles in liver by means of the method of optical coherence tomography is studied experimentally in model samples of beef liver in vitro and rat liver ex vivo. In the experiments we used the gold nanoparticles in the form of nanocages with resonance absorption in the near-IR spectral region. In the model studies the suspension of nanoparticles was applied to the surface of the sample, which then was treated with ultrasound. In the ex vivo studies the suspension of nanoparticles was injected to the laboratory rats intravenously. The image contrast and the optical depth of detection of blood vessels and liver structure components are calculated, as well as the depth of liver optical probing before and after the injection of nanoparticles. It was shown that the administration of the nanoparticle increases significantly the imaging contrast of liver blood vessels owing to the localisation of the nanoparticles therein.

Mouse precision-cut liver slices as an ex vivo model to study idiosyncratic drug-induced liver injury.

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Hadi, Mackenzie; Chen, Yixi; Starokozhko, Viktoriia; Merema, Marjolijn T; Groothuis, Geny M M

2012-09-17

Idiosyncratic drug-induced liver injury (IDILI) has been the top reason for withdrawing drugs from the market or for black box warnings. IDILI may arise from the interaction of a drug's reactive metabolite with a mild inflammation that renders the liver more sensitive to injury resulting in increased toxicity (inflammatory stress hypothesis). Aiming to develop a robust ex vivo screening method to study inflammatory stress-related IDILI mechanisms and to find biomarkers that can detect or predict IDILI, mouse precision-cut liver slices (mPCLS) were coincubated for 24 h with IDILI-related drugs and lipopolysaccharide. Lipopolysaccharide exacerbated ketoconazole (15 μM) and clozapine (45 μM) toxicity but not their non-IDILI-related comparators, voriconazole (1500 μM) and olanzapine (45 μM). However, the other IDILI-related drugs tested [diclofenac (200 μM), carbamazepine (400 μM), and troglitazone (30 μM)] did not cause synergistic toxicity with lipopolysaccharide after 24 h of incubation. Lipopolysaccharide further decreased the reduced glutathione levels caused by ketoconazole or clozapine in mPCLS after 24 h of incubation, which was not the case for the other drugs. Lipopolysaccharide significantly increased nitric oxide (NO), cytokine, and chemokine release into the mPCLS media, while the treatment with the drugs alone did not cause any substantial change. All seven drugs drastically reduced lipopolysaccharide-induced NO production. Interestingly, only ketoconazole and clozapine increased the lipopolysaccharide-induced granulocyte colony-stimulating factor (G-CSF) and granulocyte-macrophage colony-stimulating factor (GM-CSF) release. Pilot experiments showed that diclofenac and troglitazone, but not carbamazepine, demonstrated synergistic toxicity with lipopolysaccharide after a longer incubation of 48 h in mPCLS. In conclusion, we have developed an ex vivo model to detect inflammatory stress-related liver toxicity and identified ketoconazole, clozapine

Microwave Ablation Compared with Radiofrequency Ablation for Breast Tissue in an Ex Vivo Bovine Udder Model

International Nuclear Information System (INIS)

Tanaka, Toshihiro; Westphal, Saskia; Isfort, Peter; Braunschweig, Till; Penzkofer, Tobias; Bruners, Philipp; Kichikawa, Kimihiko; Schmitz-Rode, Thomas; Mahnken, Andreas H.

2012-01-01

Purpose: To compare the effectiveness of microwave (MW) ablation with radiofrequency (RF) ablation for treating breast tissue in a nonperfused ex vivo model of healthy bovine udder tissue. Materials and Methods: MW ablations were performed at power outputs of 25W, 35W, and 45W using a 915-MHz frequency generator and a 2-cm active tip antenna. RF ablations were performed with a bipolar RF system with 2- and 3-cm active tip electrodes. Tissue temperatures were continuously monitored during ablation. Results: The mean short-axis diameters of the coagulation zones were 1.34 ± 0.14, 1.45 ± 0.13, and 1.74 ± 0.11 cm for MW ablation at outputs of 25W, 35W, and 45W. For RF ablation, the corresponding values were 1.16 ± 0.09 and 1.26 ± 0.14 cm with electrodes having 2- and 3-cm active tips, respectively. The mean coagulation volumes were 2.27 ± 0.65, 2.85 ± 0.72, and 4.45 ± 0.47 cm 3 for MW ablation at outputs of 25W, 35W, and 45W and 1.18 ± 0.30 and 2.29 ± 0.55 cm 3 got RF ablation with 2- and 3-cm electrodes, respectively. MW ablations at 35W and 45W achieved significantly longer short-axis diameters than RF ablations (P < 0.05). The highest tissue temperature was achieved with MW ablation at 45W (P < 0.05). On histological examination, the extent of the ablation zone in MW ablations was less affected by tissue heterogeneity than that in RF ablations. Conclusion: MW ablation appears to be advantageous with respect to the volume of ablation and the shape of the margin of necrosis compared with RF ablation in an ex vivo bovine udder.

Ex vivo permeation of carprofen from nanoparticles: A comprehensive study through human, porcine and bovine skin as anti-inflammatory agent.

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Parra, Alexander; Clares, Beatriz; Rosselló, Ana; Garduño-Ramírez, María L; Abrego, Guadalupe; García, María L; Calpena, Ana C

2016-03-30

The purpose of this study was the development of poly(d,l-lactide-co-glycolide) acid (PLGA) nanoparticles (NPs) for the dermal delivery of carprofen (CP). The developed nanovehicle was then lyophilized using hydroxypropyl-β-cyclodextrin (HPβCD) as cryoprotectant. The ex vivo permeation profiles were evaluated using Franz diffusion cells using three different types of skin membranes: human, porcine and bovine. Furthermore, biomechanical properties of skin (trans-epidermal water loss and skin hydration) were tested. Finally, the in vivo skin irritation and the anti-inflammatory efficacy were also assayed. Results demonstrated the achievement of NPs 187.32 nm sized with homogeneous distribution, negatively charged surface (-23.39 mV) and high CP entrapment efficiency (75.38%). Permeation studies showed similar diffusion values between human and porcine skins and higher for bovine. No signs of skin irritation were observed in rabbits. Topically applied NPs significantly decreased in vivo inflammation compared to the reference drug in a TPA-induced mouse ear edema model. Thus, it was concluded that NPs containing CP may be a useful tool for the dermal treatment of local inflammation. Copyright © 2016 Elsevier B.V. All rights reserved.

Rat liver contains a limited number of binding sites for hepatic lipase

NARCIS (Netherlands)

G.C. Schoonderwoerd (Kees); A.J.M. Verhoeven (Adrie); H. Jansen (Hans)

1994-01-01

textabstractThe binding of hepatic lipase to rat liver was studied in an ex vivo perfusion model. The livers were perfused with media containing partially purified rat hepatic lipase or bovine milk lipoprotein lipase. The activity of the enzymes was determined in the

Comparison of temperature curve and ablation zone between 915- and 2450-MHz cooled-shaft microwave antenna: Results in ex vivo porcine livers

International Nuclear Information System (INIS)

Sun Yuanyuan; Cheng Zhigang; Dong Lei; Zhang Guoming; Wang Yang; Liang Ping

2012-01-01

Objective: To compare temperature curve and ablation zone between 915- and 2450-MHz cooled-shaft microwave antenna in ex vivo porcine livers. Materials and methods: The 915- and 2450-MHz microwave ablation and thermal monitor system were used in this study. A total of 56 ablation zones and 280 temperature data were obtained in ex vivo porcine livers. The output powers were 50, 60, 70, and 80 W and the setting time was 600 s. The temperature curve of every temperature spot, the short- and long-axis diameters of the coagulation zones were recorded and measured. Results: At all four power output settings, the peak temperatures of every temperature spot had a tendency to increase accordingly as the MW output power was increased, and except for 5 mm away from the antenna, the peak temperatures for the 915 MHz cooled-shaft antenna were significantly higher than those for the 2450 MHz cooled-shaft antenna (p < 0.05). Meanwhile, the short- and long-axis diameters for the 915 MHz cooled-shaft antenna were significantly larger than those for the 2450 MHz cooled-shaft antenna (p < 0.05). Conclusion: The 915 MHz cooled-shaft antenna can yield a significantly larger ablation zone and achieve higher temperature in ablation zone than a 2450 MHz cooled-shaft antenna in ex vivo porcine livers.

EX VIVO STUDY OF QUANTITATIVE ULTRASOUND PARAMETERS IN FATTY RABBIT LIVERS

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Ghoshal, Goutam; Lavarello, Roberto J.; Kemmerer, Jeremy P.; Miller, Rita J.; Oelze, Michael L.

2012-01-01

Nonalcoholic fatty liver disease (NAFLD) affects more than 30% of Americans, and with increasing problems of obesity in the United States, NAFLD is poised to become an even more serious medical concern. At present, accurate classification of steatosis (fatty liver) represents a significant challenge. In this study, the use of high-frequency (8 to 25 MHz) quantitative ultrasound (QUS) imaging to quantify fatty liver was explored. QUS is an imaging technique that can be used to quantify properties of tissue giving rise to scattered ultrasound. The changes in the ultrasound properties of livers in rabbits undergoing atherogenic diets of varying durations were investigated using QUS. Rabbits were placed on a special fatty diet for 0, 3, or 6 weeks. The fattiness of the livers was quantified by estimating the total lipid content of the livers. Ultrasonic properties, such as speed of sound, attenuation, and backscatter coefficients, were estimated in ex vivo rabbit liver samples from animals that had been on the diet for varying periods. Two QUS parameters were estimated based on the backscatter coefficient: effective scatterer diameter (ESD) and effective acoustic concentration (EAC), using a spherical Gaussian scattering model. Two parameters were estimated based on the backscattered envelope statistics (the k parameter and the μ parameter) according to the homodyned K distribution. The speed of sound decreased from 1574 to 1565 m/s and the attenuation coefficient increased from 0.71 to 1.27 dB/cm/MHz, respectively, with increasing fat content in the liver. The ESD decreased from 31 to 17 μm and the EAC increased from 38 to 63 dB/cm3 with increasing fat content in the liver. A significant increase in the μ parameter from 0.18 to 0.93 scatterers/mm3 was observed with increasing fat content in the liver samples. The results of this study indicate that QUS parameters are sensitive to fat content in the liver. PMID:23062376

21 CFR 184.1034 - Catalase (bovine liver).

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2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Catalase (bovine liver). 184.1034 Section 184.1034... Listing of Specific Substances Affirmed as GRAS § 184.1034 Catalase (bovine liver). (a) Catalase (bovine liver) (CAS Reg. No. 81457-95-6) is an enzyme preparation obtained from extracts of bovine liver. It is...

High-powered microwave ablation with a small-gauge, gas-cooled antenna: initial ex vivo and in vivo results.

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Lubner, Meghan G; Hinshaw, J Louis; Andreano, Anita; Sampson, Lisa; Lee, Fred T; Brace, Christopher L

2012-03-01

To evaluate the performance of a gas-cooled, high-powered microwave system. Investigators performed 54 ablations in ex vivo bovine livers using three devices-a single 17-gauge cooled radiofrequency(RF) electrode; a cluster RF electrode; and a single 17-gauge, gas-cooled microwave (MW) antenna-at three time points (n = 6 at 4 minutes, 12 minutes, and 16 minutes). RF power was applied using impedance-based pulsing with maximum 200 W generator output. MW power of 135 W at 2.45 GHz was delivered continuously. An approved in vivo study was performed using 13 domestic pigs. Hepatic ablations were performed using single applicators and the above-mentioned MW and RF generator systems at treatment times of 2 minutes (n = 7 MW, n = 6 RF), 5 minutes (n = 23 MW, n = 8 RF), 7 minutes (n = 11 MW, n = 6 RF), and 10 minutes (n = 7 MW, n = 9 RF). Mean transverse diameter and length of the ablation zones were compared using analysis of variance (ANOVA) with post-hoc t tests and Wilcoxon rank-sum tests. Single ex vivo MW ablations were larger than single RF ablations at all time points (MW mean diameter range 3.5-4.8 cm 4-16 minutes; RF mean diameter range 2.6-3.1 cm 4-16 minutes) (P generation of large ablation zones in short times. Copyright © 2012 SIR. Published by Elsevier Inc. All rights reserved.

Ultrasound-mediated nanoparticle delivery across ex vivo bovine retina after intravitreal injection.

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Huang, Di; Chen, Ying-Shan; Thakur, Sachin S; Rupenthal, Ilva D

2017-10-01

Intravitreal injection is the most common administration route for the treatment of retinal diseases. However, the vitreous and some of the retinal layers themselves act as significant barriers to efficient delivery of drugs administered intravitreally. This study aimed to improve the diffusive mobility of nanoparticles (NPs) in the vitreous and enhance their permeation across the retina after intravitreal injection by application of ultrasound (US). Ex vivo posterior bovine eye cups were used and the vitreous was either left intact or removed gently from the neural retina. Hyaluronic acid coated human serum albumin NPs were administered into the eye cups and continuous US with a frequency of 1MHz, an intensity of 0.5W/cm 2 , and a duration of 30s was applied once or repeatedly via the transscleral route. After pre-determined time points, fluorescence intensities in the vitreous and the retina were analyzed. Short pulses of US significantly improved the diffusive mobility of NPs through the vitreous as well as their penetration across the neural retina into the retinal pigment epithelium and choroid without causing any detectable damage to the ocular tissues. Therefore, transscleral US could be a powerful and safe tool to enhance retinal delivery of intravitreally injected NPs. Copyright © 2017 Elsevier B.V. All rights reserved.

Effects of different hydrostatic pressure on lesions in ex vivo bovine livers induced by high intensity focused ultrasound.

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He, Min; Zhong, Zhiqiang; Li, Xing; Gong, Xiaobo; Wang, Zhibiao; Li, Faqi

2017-05-01

It is well-known that acoustic cavitation associated with the high intensity focused ultrasound (HIFU) treatment often would change the morphology and size of lesions in its treatment. In most studies reported in literature, high ambient hydrostatic pressure was used to suppress the cavitation completely. Investigation of the effects by varying the ambient hydrostatic pressure (P stat ) is still lacking. In this paper, the effects of HIFU on lesions in ex vivo bovine liver specimens under various P stat are systematically investigated. A 1MHz HIFU transducer, with an aperture diameter of 70mm and a focal length of 55mm, was used to generate two groups US exposure of different acoustic intensities and exposure time (6095W/cm 2 ×8s and 9752W/cm 2 ×5s), while keeping the same acoustic energies per unit area (48760J/cm 2 ). The peak acoustic negative pressures (p - ) of the two groups were p 1 - =9.58MPa and p 2 - =10.82MPa, respectively, with the difference p d - =p 2 - -p 1 - =1.24MPa. A passive cavitation detection (PCD) was used to monitor the ultrasonic cavitation signal during exposure of the two groups. The US exposures were done under the following ambient hydrostatic pressures, P stat : atmospheric pressure, 0.5MPa, 1.0MPa, 1.5MPa, 2.0MPa, 2.5MPa and3.0MPa, respectively. The result of PCD showed that there was a statistically significant increase above background noise level in broadband emissions at dose of 9752W/cm 2 ×5s, but not at dose of 6095W/cm 2 ×8s under atmospheric pressure; i.e., the acoustic cavitation took place for p 2 - but not for p 1 - when under atmospheric pressure. The results also showed that there was no statistically difference of the morphology and size of lesions for 6095W/cm 2 ×8s exposure under the aforementioned different ambient hydrostatic pressures. But the lesions generated at 9752W/cm 2 ×5s exposure under P stat =atmospheric pressure, 0.5MPa, 1.0MPa (all of them are less than p d - ), were larger than those under 1.5MPa

Intra-Tissue Pressure Measurement in Ex Vivo Liver Undergoing Laser Ablation with Fiber-Optic Fabry-Perot Probe

Directory of Open Access Journals (Sweden)

Daniele Tosi

2016-04-01

Full Text Available We report the first-ever intra-tissue pressure measurement performed during 1064 nm laser ablation (LA of an ex vivo porcine liver. Pressure detection has been performed with a biocompatible, all-glass, temperature-insensitive Extrinsic Fabry-Perot Interferometry (EFPI miniature probe; the proposed methodology mimics in-vivo treatment. Four experiments have been performed, positioning the probe at different positions from the laser applicator tip (from 0.5 mm to 5 mm. Pressure levels increase during ablation time, and decrease with distance from applicator tip: the recorded peak parenchymal pressure levels range from 1.9 kPa to 71.6 kPa. Different pressure evolutions have been recorded, as pressure rises earlier in proximity of the tip. The present study is the first investigation of parenchymal pressure detection in liver undergoing LA: the successful detection of intra-tissue pressure may be a key asset for improving LA, as pressure levels have been correlated to scattered recurrences of tumors by different studies.

Fiber-optic combined FPI/FBG sensors for monitoring of radiofrequency thermal ablation of liver tumors: ex vivo experiments.

Science.gov (United States)

Tosi, Daniele; Macchi, Edoardo Gino; Braschi, Giovanni; Cigada, Alfredo; Gallati, Mario; Rossi, Sandro; Poeggel, Sven; Leen, Gabriel; Lewis, Elfed

2014-04-01

We present a biocompatible, all-glass, 0.2 mm diameter, fiber-optic probe that combines an extrinsic Fabry-Perot interferometry and a proximal fiber Bragg grating sensor; the probe enables dual pressure and temperature measurement on an active 4 mm length, with 40 Pa and 0.2°C nominal accuracy. The sensing system has been applied to monitor online the radiofrequency thermal ablation of tumors in liver tissue. Preliminary experiments have been performed in a reference chamber with uniform heating; further experiments have been carried out on ex vivo porcine liver, which allowed the measurement of a steep temperature gradient and monitoring of the local pressure increase during the ablation procedure.

Microwave Ablation: Comparison of Simultaneous and Sequential Activation of Multiple Antennas in Liver Model Systems.

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Harari, Colin M; Magagna, Michelle; Bedoya, Mariajose; Lee, Fred T; Lubner, Meghan G; Hinshaw, J Louis; Ziemlewicz, Timothy; Brace, Christopher L

2016-01-01

To compare microwave ablation zones created by using sequential or simultaneous power delivery in ex vivo and in vivo liver tissue. All procedures were approved by the institutional animal care and use committee. Microwave ablations were performed in both ex vivo and in vivo liver models with a 2.45-GHz system capable of powering up to three antennas simultaneously. Two- and three-antenna arrays were evaluated in each model. Sequential and simultaneous ablations were created by delivering power (50 W ex vivo, 65 W in vivo) for 5 minutes per antenna (10 and 15 minutes total ablation time for sequential ablations, 5 minutes for simultaneous ablations). Thirty-two ablations were performed in ex vivo bovine livers (eight per group) and 28 in the livers of eight swine in vivo (seven per group). Ablation zone size and circularity metrics were determined from ablations excised postmortem. Mixed effects modeling was used to evaluate the influence of power delivery, number of antennas, and tissue type. On average, ablations created by using the simultaneous power delivery technique were larger than those with the sequential technique (P Simultaneous ablations were also more circular than sequential ablations (P = .0001). Larger and more circular ablations were achieved with three antennas compared with two antennas (P simultaneous power delivery creates larger, more confluent ablations with greater temperatures than those created with sequential power delivery. © RSNA, 2015.

Magnetic resonance imaging for the exploitation of bubble-enhanced heating by high-intensity focused ultrasound: a feasibility study in ex vivo liver.

Science.gov (United States)

Elbes, Delphine; Denost, Quentin; Robert, Benjamin; Köhler, Max O; Tanter, Mickaël; Bruno, Quesson

2014-05-01

Bubble-enhanced heating (BEH) may be exploited to improve the heating efficiency of high-intensity focused ultrasound in liver and to protect tissues located beyond the focal point. The objectives of this study, performed in ex vivo pig liver, were (i) to develop a method to determine the acoustic power threshold for induction of BEH from displacement images measured by magnetic resonance acoustic radiation force imaging (MR-ARFI), and (ii) to compare temperature distribution with MR thermometry for HIFU protocols with and without BEH. The acoustic threshold for generation of BEH was determined in ex vivo pig liver from MR-ARFI calibration curves of local tissue displacement resulting from sonication at different powers. Temperature distributions (MR thermometry) resulting from "conventional" sonications (20 W, 30 s) were compared with those from "composite" sonications performed at identical parameters, but after a HIFU burst pulse (0.5 s, acoustic power over the threshold for induction of BEH). Displacement images (MR-ARFI) were acquired between sonications to measure potential modifications of local tissue displacement associated with modifications of tissue acoustic characteristics induced by the burst HIFU pulse. The acoustic threshold for induction of BEH corresponded to a displacement amplitude of approximately 50 μm in ex vivo liver. The displacement and temperature images of the composite group exhibited a nearly spherical pattern, shifted approximately 4 mm toward the transducer, in contrast to elliptical shapes centered on the natural focal position for the conventional group. The gains in maximum temperature and displacement values were 1.5 and 2, and the full widths at half-maximum of the displacement data were 1.7 and 2.2 times larger than in the conventional group in directions perpendicular to ultrasound propagation axes. Combination of MR-ARFI and MR thermometry for calibration and exploitation of BEH appears to increase the efficiency and safety

Radiofrequency Ablation with a New Perfused-Cooled Electrode Using a Single Pump: An Experimental Study in Ex Vivo Bovine Liver

International Nuclear Information System (INIS)

Kim, Seung Kwon; Seo, Jung Wook

2005-01-01

The purpose of this study was to assess the efficacy of a new perfused-cooled electrode that uses a single pump for creating a large ablation zone in explanted bovine liver. This was done by comparing with the radiofrequency (RF) ablation zones that were created with a monopolar cooled electrode to the RF ablation zones that were created by the new perfused-cooled electrode. We developed a new perfused-cooled electrode that uses a single pump by modifying a 17-gauge cooled electrode (Radionics) with a 2.5-cm outer metallic sheath (15-gauge) in order to allow use of the internal cooling water (5.85 % hypertonic saline) for the infused saline. Thirty ablation zones were created in explanted bovine livers (12-min ablation cycle; pulsed technique; 2000 mA, maximum) with three different regimens: group A, RF ablation with the 17-gauge cooled electrode; group B, RF ablation with the 15-gauge cooled electrode; group C, RF ablation with the perfused-cooled electrode. T2-weighted magnetic resonance (MR) imaging was obtained immediately after RF ablation for calculating volumes of the ablation zone. Following MR imaging, the ablation zones were excised and measured for transverse diameters and vertical diameters. The transverse diameter, vertical diameter, and the calculated volumes of the ablation zones on MR imaging were compared among the groups. Ablation zones created with the perfused-cooled electrode (group C) were significantly larger than those created with the 17-gauge cooled electrode (group A) and the 15-gauge cooled electrode (group B) according to the transverse diameter and vertical diameter on the gross specimens (p 3 in group A, 28.9 ± 5.7 cm 3 in group B, and 80.0 ± 34 cm 3 in group C, respectively. A new perfused-cooled electrode using a single pump could efficiently increase the size of the ablation zone in liver compared with a monopolar cooled electrode, and this was due to its simultaneous use of internal cooling and saline infusion

Ex vivo MR volumetry of human brain hemispheres.

Science.gov (United States)

Kotrotsou, Aikaterini; Bennett, David A; Schneider, Julie A; Dawe, Robert J; Golak, Tom; Leurgans, Sue E; Yu, Lei; Arfanakis, Konstantinos

2014-01-01

The aims of this work were to (a) develop an approach for ex vivo MR volumetry of human brain hemispheres that does not contaminate the results of histopathological examination, (b) longitudinally assess regional brain volumes postmortem, and (c) investigate the relationship between MR volumetric measurements performed in vivo and ex vivo. An approach for ex vivo MR volumetry of human brain hemispheres was developed. Five hemispheres from elderly subjects were imaged ex vivo longitudinally. All datasets were segmented. The longitudinal behavior of volumes measured ex vivo was assessed. The relationship between in vivo and ex vivo volumetric measurements was investigated in seven elderly subjects imaged both antemortem and postmortem. This approach for ex vivo MR volumetry did not contaminate the results of histopathological examination. For a period of 6 months postmortem, within-subject volume variation across time points was substantially smaller than intersubject volume variation. A close linear correspondence was detected between in vivo and ex vivo volumetric measurements. Regional brain volumes measured with this approach for ex vivo MR volumetry remain relatively unchanged for a period of 6 months postmortem. Furthermore, the linear relationship between in vivo and ex vivo MR volumetric measurements suggests that this approach captures information linked to antemortem macrostructural brain characteristics. Copyright © 2013 Wiley Periodicals, Inc.

Ex-vivo MR Volumetry of Human Brain Hemispheres

Science.gov (United States)

Kotrotsou, Aikaterini; Bennett, David A.; Schneider, Julie A.; Dawe, Robert J.; Golak, Tom; Leurgans, Sue E.; Yu, Lei; Arfanakis, Konstantinos

2013-01-01

Purpose The aims of this work were to: a) develop an approach for ex-vivo MR volumetry of human brain hemispheres that does not contaminate the results of histopathological examination, b) longitudinally assess regional brain volumes postmortem, and c) investigate the relationship between MR volumetric measurements performed in-vivo and ex-vivo. Methods An approach for ex-vivo MR volumetry of human brain hemispheres was developed. Five hemispheres from elderly subjects were imaged ex-vivo longitudinally. All datasets were segmented. The longitudinal behavior of volumes measured ex-vivo was assessed. The relationship between in-vivo and ex-vivo volumetric measurements was investigated in seven elderly subjects imaged both ante-mortem and postmortem. Results The presented approach for ex-vivo MR volumetry did not contaminate the results of histopathological examination. For a period of 6 months postmortem, within-subject volume variation across time points was substantially smaller than inter-subject volume variation. A close linear correspondence was detected between in-vivo and ex-vivo volumetric measurements. Conclusion Regional brain volumes measured with the presented approach for ex-vivo MR volumetry remain relatively unchanged for a period of 6 months postmortem. Furthermore, the linear relationship between in-vivo and ex-vivo MR volumetric measurements suggests that the presented approach captures information linked to ante-mortem macrostructural brain characteristics. PMID:23440751

Extracellular Vesicles from Human Liver Stem Cells Reduce Injury in an Ex Vivo Normothermic Hypoxic Rat Liver Perfusion Model.

Science.gov (United States)

Rigo, Federica; De Stefano, Nicola; Navarro-Tableros, Victor; David, Ezio; Rizza, Giorgia; Catalano, Giorgia; Gilbo, Nicholas; Maione, Francesca; Gonella, Federica; Roggio, Dorotea; Martini, Silvia; Patrono, Damiano; Salizzoni, Mauro; Camussi, Giovanni; Romagnoli, Renato

2018-05-01

The gold standard for organ preservation before transplantation is static cold storage, which is unable to fully protect suboptimal livers from ischemia/reperfusion injury. An emerging alternative is normothermic machine perfusion (NMP), which permits organ reconditioning. Here, we aimed to explore the feasibility of a pharmacological intervention on isolated rat livers by using a combination of NMP and human liver stem cells-derived extracellular vesicles (HLSC-EV). We established an ex vivo murine model of NMP capable to maintain liver function despite an ongoing hypoxic injury induced by hemodilution. Livers were perfused for 4 hours without (control group, n = 10) or with HLSC-EV (treated group, n = 9). Bile production was quantified; perfusate samples were collected hourly to measure metabolic (pH, pO2, pCO2) and cytolysis parameters (AST, alanine aminotransferase, lactate dehydrogenase). At the end of perfusion, we assessed HLSC-EV engraftment by immunofluorescence, tissue injury by histology, apoptosis by terminal deoxynucleotidyl transferase dUTP nick-end labeling assay, tissue hypoxia-inducible factor 1-α, and transforming growth factor-beta 1 RNA expression by quantitative reverse transcription-polymerase chain reaction. During hypoxic NMP, livers were able to maintain homeostasis and produce bile. In the treated group, AST (P = 0.018) and lactate dehydrogenase (P = 0.032) levels were significantly lower than those of the control group at 3 hours of perfusion, and AST levels persisted lower at 4 hours (P = 0.003). By the end of NMP, HLSC-EV had been uptaken by hepatocytes, and EV treatment significantly reduced histological damage (P = 0.030), apoptosis (P = 0.049), and RNA overexpression of hypoxia-inducible factor 1-α (P < 0.0001) and transforming growth factor-beta 1 (P = 0.014). HLSC-EV treatment, even in a short-duration model, was feasible and effectively reduced liver injury during hypoxic NMP.

In-line X-ray phase-contrast imaging of murine liver microvasculature ex vivo

International Nuclear Information System (INIS)

Li Beilei; Xu Min; Shi Hongcheng; Chen Shaoliang; Wu Weizhong; Peng Guanyun; Zhang Xi; Peng Yifeng

2012-01-01

Imaging blood vessels is of importance for determining the vascular distribution of organs and tumors. Phase-contrast X-ray imaging can reveal the vessels in much more detail than conventional X-ray absorption method. Visualizing murine liver microvasculature ex vivo with phase-contrast X-ray imaging was performed at Shanghai Synchrotron Radiation Facility. Barium sulfate and physiological saline were used as contrast agents for the blood vessels. Blood vessels of <Φ20 μm could be detected by replacing resident blood with physiological saline or barium sulfate. An entire branch of the portal vein (from the main axial portal vein to the ninth generation of branching) could be captured in a single phase-contrast image. It is demonstrated that selective angiography based on phase contrast X-ray imaging, with a physiological material of low Z elements (such as saline) being the contrast agent, is a viable imaging strategy. Further efforts will be focused on using the technique to image tumor angiogenesis. (authors)

Effect of nutritional status on oxidative stress in an ex vivo perfused rat liver.

Science.gov (United States)

Stadler, Michaela; Nuyens, Vincent; Seidel, Laurence; Albert, Adelin; Boogaerts, Jean G

2005-11-01

Normothermic ischemia-reperfusion is a determinant in liver injury occurring during surgical procedures, ischemic state, and multiple organ failure. The preexisting nutritional status of the liver might contribute to the extent of tissue injury and primary nonfunction. The aim of this study was to determine the role of starvation on hepatic ischemia-reperfusion injury in normal rat livers. Rats were randomly divided into two groups: one had free access to food, the other was fasted for 16 h. The portal vein was cannulated, and the liver was removed and perfused in a closed ex vivo system. Two modes of perfusion were applied in each series of rats, fed and fasting. In the ischemia-reperfusion mode, the experiment consisted of perfusion for 15 min, warm ischemia for 60 min, and reperfusion during 60 min. In the nonischemia mode, perfusion was maintained during the 135-min study period. Five rats were included in each experimental condition, yielding a total of 20 rats. Liver enzymes, potassium, glucose, lactate, free radicals, i.e., dienes and trienes, and cytochrome c were analyzed in perfusate samples. The proportion of glycogen in hepatocytes was determined in tissue biopsies. Transaminases, lactate dehydrogenase, potassium, and free radical concentrations were systematically higher in fasting rats in both conditions, with and without ischemia. Cytochrome c was higher after reperfusion in the fasting rats. Glucose and lactate concentrations were greater in the fed group. The glycogen content decreased in both groups during the experiment but was markedly lower in the fasting rats. In fed rats, liver injury was moderate, whereas hepatocytes integrity was notably impaired both after continuous perfusion and warm ischemia in fasting animals. Reduced glycogen store in hepatocytes may explain reduced tolerance.

Radiofrequency ablation in the liver using two cooled-wet electrodes in the bipolar mode

International Nuclear Information System (INIS)

Han, Joon Koo; Lee, Jeong Min; Kim, Se Hyung; Lee, Jae Young; Park, Hee Sun; Eo, Hong; Choi, Byung Ihn

2005-01-01

The purpose of this study was to demonstrate the efficacy of bipolar radiofrequency ablation (RFA) using cooled-wet electrodes inducing coagulation in ex vivo bovine livers and in in vivo canine livers. In ex vivo experiments, 20 coagulations were created by monopolar (group A), and bipolar RFA (group B) using a 200 W generator (Valleylab) and one or two cooled-wet electrodes. In in vivo experiments, one coagulation was created by bipolar RFA in each of eight dogs via laparotomy. In ex vivo and in vivo experiments, RF was applied to one or two electrodes at 100 W for 10 min. The dimensions of the coagulations were compared in the two groups. In ex vivo experiments, the mean volumes of the coagulations produced in group B (54.0±16.5 cm 3 ) were greater than those produced in group A (33.9±12.7 cm 3 ) (P=0.007). In in vivo experiments, bipolar RFA produced a coagulation of 39.4±15.6 cm 3 without a major complication. The present study showed that a RF electrode system using two cooled-wet electrodes in the bipolar mode created larger coagulation volumes than the monopolar mode, and this system can be used to create large coagulation without major complications. (orig.)

Comparison of wet radiofrequency ablation with dry radiofrequency ablation and radiofrequency ablation using hypertonic saline preinjection: ex vivo bovine liver

International Nuclear Information System (INIS)

Lee, Jeong Min; Han, Joon Koo; Kim, Se Hyung; Lee, Jae Young; Park, Hee Sun; Hur, Hurn; Choi, Byung Ihn; Shin, Kyung Sook

2004-01-01

We wished to compare the in-vitro efficiency of wet radiofrequency (RF) ablation with the efficiency of dry RF ablation and RF ablation with preinjection of NaCl solutions using excised bovine liver. Radiofrequency was applied to excised bovine livers in a monopolar mode for 10 minutes using a 200 W generator and a perfused-cooled electrode with or without injection or slow infusion of NaCl solutions. After placing the perfused-cooled electrode in the explanted liver, 50 ablation zones were created with five different regimens: group A; standard dry RF ablation, group B; RF ablation with 11 mL of 5% NaCl solution preinjection, group C; RF ablation with infusion of 11 mL of 5% NaCl solution at a rate of 1 mL/min, group D; RFA with 6 mL of 36% NaCl solution preinjection, group E; RF ablation with infusion of 6 mL of 36% NaCl solution at a rate of 0.5 mL/min. In groups C and E, infusion of the NaCl solutions was started 1 min before RF ablation and then maintained during RF ablation (wet RF ablation). During RF ablation, we measured the tissue temperature at 15 mm from the electrode. The dimensions of the ablation zones and changes in impedance, current and liver temperature during RF ablation were then compared between the groups. With injection or infusion of NaCl solutions, the mean initial tissue impedance prior to RF ablation was significantly less in groups B, C, D, and E (43-75 Ω) than for group A (80 Ω) (ρ 3 in group A; 12.4 ± 3.8 cm 3 in group B; 80.9 ± 9.9 cm 3 in group C; 45.3 ± 11.3 cm 3 in group D and 81.6 ± 8.6 cm 3 in group E. The tissue temperature measured at 15 mm from the electrode was higher in groups C, D and E than other groups (ρ < 0.05): 53 ± 12 .deg. C in group A, 42 ± 2 .deg. C in group B, 93 ± 8 .deg. C in group C; 79 ± 12 .deg. C in group D and 83 ± 8 .deg.C in group E. Wet RF ablation with 5% or 36% NaCl solutions shows better efficiency in creating a large ablation zone than does dry RF ablation or RF ablation with

Subnormothermic ex vivo liver perfusion reduces endothelial cell and bile duct injury after donation after cardiac death pig liver transplantation.

Science.gov (United States)

Knaak, Jan M; Spetzler, Vinzent N; Goldaracena, Nicolas; Boehnert, Markus U; Bazerbachi, Fateh; Louis, Kristine S; Adeyi, Oyedele A; Minkovich, Leonid; Yip, Paul M; Keshavjee, Shaf; Levy, Gary A; Grant, David R; Selzner, Nazia; Selzner, Markus

2014-11-01

An ischemic-type biliary stricture (ITBS) is a common feature after liver transplantation using donation after cardiac death (DCD) grafts. We compared sequential subnormothermic ex vivo liver perfusion (SNEVLP; 33°C) with cold storage (CS) for the prevention of ITBS in DCD liver grafts in pig liver transplantation (n = 5 for each group). Liver grafts were stored for 10 hours at 4°C (CS) or preserved with combined 7-hour CS and 3-hour SNEVLP. Parameters of hepatocyte [aspartate aminotransferase (AST), international normalized ratio (INR), factor V, and caspase 3 immunohistochemistry], endothelial cell (EC; CD31 immunohistochemistry and hyaluronic acid), and biliary injury and function [alkaline phosphatase (ALP), total bilirubin, and bile lactate dehydrogenase (LDH)] were determined. Long-term survival (7 days) after transplantation was similar between the SNEVLP and CS groups (60% versus 40%, P = 0.13). No difference was observed between SNEVLP- and CS-treated animals with respect to the peak of serum INR, factor V, or AST levels within 24 hours. CD31 staining 8 hours after transplantation demonstrated intact EC lining in SNEVLP-treated livers (7.3 × 10(-4) ± 2.6 × 10(-4) cells/μm(2)) but not in CS-treated livers (3.7 × 10(-4) ± 1.3 × 10(-4) cells/μm(2) , P = 0.03). Posttransplant SNEVLP animals had decreased serum ALP and serum bilirubin levels in comparison with CS animals. In addition, LDH in bile fluid was lower in SNEVLP pigs versus CS pigs (14 ± 10 versus 60 ± 18 μmol/L, P = 0.02). Bile duct histology revealed severe bile duct necrosis in 3 of 5 animals in the CS group but none in the SNEVLP group (P = 0.03). Sequential SNEVLP preservation of DCD grafts reduces bile duct and EC injury after liver transplantation. © 2014 American Association for the Study of Liver Diseases.

Fourier and non-Fourier bio-heat transfer models to predict ex vivo temperature response to focused ultrasound heating

Science.gov (United States)

Li, Chenghai; Miao, Jiaming; Yang, Kexin; Guo, Xiasheng; Tu, Juan; Huang, Pintong; Zhang, Dong

2018-05-01

Although predicting temperature variation is important for designing treatment plans for thermal therapies, research in this area is yet to investigate the applicability of prevalent thermal conduction models, such as the Pennes equation, the thermal wave model of bio-heat transfer, and the dual phase lag (DPL) model. To address this shortcoming, we heated a tissue phantom and ex vivo bovine liver tissues with focused ultrasound (FU), measured the temperature response, and compared the results with those predicted by these models. The findings show that, for a homogeneous-tissue phantom, the initial temperature increase is accurately predicted by the Pennes equation at the onset of FU irradiation, although the prediction deviates from the measured temperature with increasing FU irradiation time. For heterogeneous liver tissues, the predicted response is closer to the measured temperature for the non-Fourier models, especially the DPL model. Furthermore, the DPL model accurately predicts the temperature response in biological tissues because it increases the phase lag, which characterizes microstructural thermal interactions. These findings should help to establish more precise clinical treatment plans for thermal therapies.

Inducing Hepatitis C Virus Resistance After Pig Liver Transplantation-A Proof of Concept of Liver Graft Modification Using Warm Ex Vivo Perfusion.

Science.gov (United States)

Goldaracena, N; Spetzler, V N; Echeverri, J; Kaths, J M; Cherepanov, V; Persson, R; Hodges, M R; Janssen, H L A; Selzner, N; Grant, D R; Feld, J J; Selzner, M

2017-04-01

Normothermic ex vivo liver perfusion (NEVLP) offers the potential to optimize graft function prior to liver transplantation (LT). Hepatitis C virus (HCV) is dependent on the presence of miRNA(microRNA)-122. Miravirsen, a locked-nucleic acid oligonucleotide, sequesters miR-122 and inhibits HCV replication. The aim of this study was to assess the efficacy of delivering miravirsen during NEVLP to inhibit miR-122 function in a pig LT model. Pig livers were treated with miravirsen during NEVLP or cold storage (CS). Miravirsen absorption, miR-122 sequestration, and miR-122 target gene derepression were determined before and after LT. The effect of miravirsen treatment on HCV infection of hepatoma cells was also assessed. NEVLP improved miravirsen uptake versus CS. Significant miR-122 sequestration and miR-122 target gene derepression were seen with NEVLP but not with CS. In vitro data confirmed miravirsen suppression of HCV replication after established infection and prevented HCV infection with pretreatment of cells, analogous to the pretreatment of grafts in the transplant setting. In conclusion, miravirsen delivery during NEVLP is a potential strategy to prevent HCV reinfection after LT. This is the first large-animal study to provide "proof of concept" for using NEVLP to modify and optimize liver grafts for transplantation. © 2016 The American Society of Transplantation and the American Society of Transplant Surgeons.

The Hepatoprotection Provided by Taurine and Glycine against Antineoplastic Drugs Induced Liver Injury in an Ex Vivo Model of Normothermic Recirculating Isolated Perfused Rat Liver

Directory of Open Access Journals (Sweden)

Reza Heidari

2016-03-01

Full Text Available Taurine (2-aminoethane sulfonic acid is a non-protein amino acid found in high concentration in different tissues. Glycine (Amino acetic acid is the simplest amino acid incorporated in the structure of proteins. Several investigations indicate the hepatoprotective properties of these amino acids. On the other hand, antineoplastic agents-induced serum transaminase elevation and liver injury is a clinical complication. The current investigation was designed to screen the possible hepatoprotective properties of taurine and glycine against antineoplastic drugs-induced hepatic injury in an ex vivo model of isolated perfused rat liver. Rat liver was perfused with different concentration (10 μM, 100 μM and 1000 μM of antineoplastic drugs (Mitoxantrone, Cyclophosphamide, Cisplatin, 5 Fluorouracil, Doxorubicin and Dacarbazine via portal vein. Taurine and glycine were administered to drug-treated livers and liver perfusate samples were collected for biochemical measurements (ALT, LDH, AST, and K+. Markers of oxidative stress (reactive oxygen species formation, lipid peroxidation, total antioxidant capacity and glutathione were also assessed in liver tissue. Antineoplastic drugs caused significant pathological changes in perfusate biochemistry. Furthermore, markers of oxidative stress were significantly elevated in drug treated livers. It was found that taurine (5 and 10 mM and glycine (5 and 10 mM administration significantly mitigated the biomarkers of liver injury and attenuated drug induced oxidative stress. Our data indicate that taurine and glycine supplementation might help as potential therapeutic options to encounter anticancer drugs-induced liver injury.

Comparison of in vivo and ex vivo imaging of the microvasculature with 2-photon fluorescence microscopy

Science.gov (United States)

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.

Avaliação e recondicionamento pulmonar ex vivo Ex vivo lung evaluation and reconditioning

Directory of Open Access Journals (Sweden)

Paulo Manuel Pêgo-Fernandes

2010-12-01

Full Text Available OBJETIVO: Apenas 15% dos pulmões doados são aproveitados para transplante. Um novo método de Perfusão Pulmonar Ex Vivo (PPEV foi desenvolvido e pode ser usado para avaliação e recondicionamento de pulmões "marginais" e rejeitados para o transplante. Esse trabalho relata nossa experiência com a avaliação funcional da PPEV. MÉTODOS: Foram estudados pulmões de 12 doadores considerados inapropriados para transplante pulmonar. Após a captação, os pulmões são perfundidos ex vivo com Steen Solution, uma solução de composição eletrolítica extracelular com alta pressão coloidosmótica. Um oxigenador de membrana ligado ao circuito recebe uma mistura gasosa (nitrogênio e dióxido de carbono e "desoxigena" o perfusato, mantendo uma concentração de gases semelhante a do sangue venoso. Os pulmões são gradualmente aquecidos, perfundidos e ventilados. A avaliação dos órgãos é feita por gasometrias e medidas como a resistência vascular pulmonar (RVP e complacência pulmonar (CP. RESULTADOS: A PaO2 (FiO2 100% passou de um valor médio de 193,3 mmHg no doador para 495,3 mmHg durante a PPEV. Após uma hora de PPEV, a RVP média era de 737,3 dinas/seg/ cm5 e a CP era de 42,2 ml/cmH2O. CONCLUSÕES: O modelo de avaliação pulmonar ex vivo pode melhorar a capacidade de oxigenação de pulmões "marginais" inicialmente rejeitados para transplante. Isso denota um grande potencial do método para aumentar a disponibilidade de pulmões para transplante e, possivelmente, reduzir o tempo de espera nas filas.OBJECTIVE: Only about 15% of the potential candidates for lung donation are considered suitable for transplantation. A new method for ex vivo lung perfusion (EVLP has been developed and can be used for evaluation and reconditioning of "marginal" and unacceptable lungs. This is a report of functional evaluation experience with ex vivo perfusion of twelve donor lungs deemed unacceptable in São Paulo, Brazil. METHODS: After harvesting, the

Evaluation of the in vivo and ex vivo optical properties in a mouse ear model

Energy Technology Data Exchange (ETDEWEB)

Salomatina, E; Yaroslavsky, A N [Wellman Center for Photomedicine, 40 Blossom Street, Boston, MA 02114 (United States)], E-mail: Yaroslav@helix.mgh.harvard.edu

2008-06-07

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, {mu}{sub a}, scattering coefficients, {mu}{sub s}, 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 deg. C wrapped in a gauze moistened with saline solution. Then the specimens were frozen at -25 deg. 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.

Ex-vivo machine perfusion for kidney preservation.

Science.gov (United States)

Hamar, Matyas; Selzner, Markus

2018-06-01

Machine perfusion is a novel strategy to decrease preservation injury, improve graft assessment, and increase organ acceptance for transplantation. This review summarizes the current advances in ex-vivo machine-based kidney preservation technologies over the last year. Ex-vivo perfusion technologies, such as hypothermic and normothermic machine perfusion and controlled oxygenated rewarming, have gained high interest in the field of organ preservation. Keeping kidney grafts functionally and metabolically active during the preservation period offers a unique chance for viability assessment, reconditioning, and organ repair. Normothermic ex-vivo kidney perfusion has been recently translated into clinical practice. Preclinical results suggest that prolonged warm perfusion appears superior than a brief end-ischemic reconditioning in terms of renal function and injury. An established standardized protocol for continuous warm perfusion is still not available for human grafts. Ex-vivo machine perfusion represents a superior organ preservation method over static cold storage. There is still an urgent need for the optimization of the perfusion fluid and machine technology and to identify the optimal indication in kidney transplantation. Recent research is focusing on graft assessment and therapeutic strategies.

3D morphological analysis of the mouse cerebral vasculature: Comparison of in vivo and ex vivo methods.

Directory of Open Access Journals (Sweden)

Joe Steinman

Full Text Available Ex vivo 2-photon fluorescence microscopy (2PFM with optical clearing enables vascular imaging deep into tissue. However, optical clearing may also produce spherical aberrations if the objective lens is not index-matched to the clearing material, while the perfusion, clearing, and fixation procedure may alter vascular morphology. We compared in vivo and ex vivo 2PFM in mice, focusing on apparent differences in microvascular signal and morphology. Following in vivo imaging, the mice (four total were perfused with a fluorescent gel and their brains fructose-cleared. The brain regions imaged in vivo were imaged ex vivo. Vessels were segmented in both images using an automated tracing algorithm that accounts for the spatially varying PSF in the ex vivo images. This spatial variance is induced by spherical aberrations caused by imaging fructose-cleared tissue with a water-immersion objective. Alignment of the ex vivo image to the in vivo image through a non-linear warping algorithm enabled comparison of apparent vessel diameter, as well as differences in signal. Shrinkage varied as a function of diameter, with capillaries rendered smaller ex vivo by 13%, while penetrating vessels shrunk by 34%. The pial vasculature attenuated in vivo microvascular signal by 40% 300 μm below the tissue surface, but this effect was absent ex vivo. On the whole, ex vivo imaging was found to be valuable for studying deep cortical vasculature.

In vivo field dependence of proton relaxation times in human brain, liver and skeletal muscle: a multicenter study

DEFF Research Database (Denmark)

Henriksen, O; de Certaines, J D; Spisni, A

1993-01-01

and MRS, the in vivo field dispersion of T1 and T2 has been measured in order to evaluate whether ex vivo data are representative for the in vivo situation. Brain, skeletal muscle, and liver of healthy human volunteers were studied. Fifteen MR units with a field strength ranging from 0.08 T to 1.5 T took......T1 and T2 relaxation times are fundamental parameters for signal contrast behaviour in MRI. A number of ex vivo relaxometry studies have dealt with the magnetic field dispersion of T1. By means of multicenter study within the frame of the COMAC BME Concerted Action on Tissue Characterization by MRI......, whereas no significant variations were seen for T2. Our in vivo data were generally in reasonable agreement with proposed models based on ex vivo measurements....

A Newly Developed Perfused Umbrella Electrode for Radiofrequency Ablation: An Ex Vivo Evaluation Study in Bovine Liver

International Nuclear Information System (INIS)

Bruners, Philipp; Pfeffer, Jochen; Kazim, Rana M.; Guenther, Rolf W.; Schmitz-Rode, Thomas; Mahnken, Andreas H.

2007-01-01

The purpose of this study was to evaluate the effectiveness of a newly developed perfused monopolar radiofrequency (RF) probe with an umbrella-shaped array. A perfused umbrella-shaped monopolar RF probe based on a LeVeen electrode (Boston Scientific Corp., Natick, MA, USA) with a 3-cm array diameter was developed. Five different configurations of this electrode were tested: (a) perfusion channel/endhole, (b) perfusion channel/endhole + sideholes, (c) 1 cm insulation removed at the tip, (d) 1 cm insulation removed at the tip + perfusion channel/endhole, and (e) 1 cm insulation removed at the tip + perfusion channel/endhole + sideholes. An unmodified LeVeen electrode served as a reference standard. RF ablations were performed in freshly excised bovine liver using a commercial monopolar RF system with a 200-W generator (RF 3000; Boston Scientific Corp.). Each electrode was tested 10 times applying the vendor's recommended ablation protocol combined with the preinjection of 2 ml 0.9% saline. Volumes and shapes of the lesions were compared. Lesions generated with the original LeVeen electrode showed a mean volume of 12.74 ± 0.52 cm 3 . Removing parts of the insulation led to larger coagulation volumes (22.65 ± 2.12 cm 3 ). Depending on the configuration, saline preinjection resulted in a further increase in coagulation volume (25.22 ± 3.37 to 31.28 ± 2.32 cm 3 ). Besides lesion volume, the shape of the ablation zone was influenced by the configuration of the electrode used. We conclude that saline preinjection in combination with increasing the active tip length of the umbrella-shaped LeVeen RF probe allows the reliable ablation of larger volumes in comparison to the originally configured electrode

Comparison between coupled KZK-BHTE numerical simulations and scanned HIFU exposures in excised bovine liver

Science.gov (United States)

Andrew, Marilee A.; Brayman, Andrew A.; Kaczkowski, Peter J.; Kargl, Steven G.

2004-05-01

The use of moving high intensity focused ultrasound (HIFU) treatment protocols is of interest in achieving efficient formation of large-volume lesions in tissue. However, potentially unwanted thermal effects, such as prefocal heating, should be considered. A KZK acoustic model coupled with the BioHeat Transfer Equation has been extended to simulate multiple, moving scans in tissue. Simulation results are compared with experimental data collected over a range of exposure regimes for linear and concentric circular scans with a 3.5-MHz single-element transducer in ex vivo bovine liver. Of particular interest are investigating prefocal thermal buildup and ablating the central core of a circular pattern through conductive heating, that is without direct HIFU exposure. Qualitative agreement is observed between experimental and simulated data; limits of the predictive capability of the model in cavitation regimes will be discussed. [Support provided by the U.S. Army Medical Research Acquisition Activity through The University of Mississippi under terms of Agreement No. DAMD17-02-2-0014. The opinions expressed herein are those of the author(s) and do not necessarily reflect the views of U.S. Army Medical Research Acquisition Activity or The University of Mississippi.

Comparison of ex vivo stability of copeptin and vasopressin

NARCIS (Netherlands)

Heida, Judith E; Boesten, Lianne S M; Ettema, Esmée M; Muller Kobold, Anneke C.; Franssen, Casper F M; Gansevoort, Ron T; Zittema, Debbie

BACKGROUND: Copeptin, part of the vasopressin precursor, is increasingly used as marker for vasopressin and is claimed to have better ex vivo stability. However, no study has directly compared the ex vivo stability of copeptin and vasopressin. METHODS: Blood of ten healthy volunteers was collected

A comparison of microwave ablation and bipolar radiofrequency ablation both with an internally cooled probe: Results in ex vivo and in vivo porcine livers

International Nuclear Information System (INIS)

Yu Jie; Liang Ping; Yu Xiaoling; Liu Fangyi; Chen Lei; Wang Yang

2011-01-01

Purpose: The purpose of this study was to compare the effectiveness of microwave (MW) ablation and radiofrequency (RF) ablation using a single internally cooled probe in a hepatic porcine model. Materials and methods: In the ex vivo experiment, MW ablations (n = 40) were performed with a 2450 MHz and 915 MHz needle antenna, respectively at 60 W, 70 W power settings. Bipolar RF ablations (n = 20) were performed with a 3-cm (T30) and 4-cm (T40) active tip needle electrodes, respectively at a rated power 30 W and 40 W according to automatically systematic power setting. In the in vivo experiment, the 2450 MHz and 915 MHz MW ablation both at 60 W and T30 bipolar RF ablation at 30 W were performed (n = 30). All of the application time were 10 min. Long-axis diameter (Dl), short-axis diameter (Ds), ratio of Ds/Dl, the temperature data 5 mm from the needle and the time of temperature 5 mm from the needle rising to 54 deg. C were measured. Results: Both in ex vivo and in vivo models, Ds and Dl of 915 MHz MW ablations were significantly larger than all the RF ablations (P < 0.05); the Ds for all the 2450 MHz MW ablations were significantly larger than that of T30 RF ablations (P < 0.05). 2450 MHz MW and T30 RF ablation tended to produce more elliptical-shaped ablation zone. Tissue temperatures 5 mm from the needle were considerably higher with MW ablation, meanwhile MW ablation achieved significantly faster rate of temperature rising to 54 deg. C than RF ablation. For in vivo study after 10 min of ablation, the Ds and Dl of 2450 MHz MW, 915 MHz MW and Bipolar RF were 2.35 ± 0.75, 2.95 ± 0.32, 1.61 ± 0.33 and 3.86 ± 0.81, 5.79 ± 1.03, 3.21 ± 0.51, respectively. Highest tissue temperatures 5 mm from the needle were 80.07 ± 12.82 deg. C, 89.07 ± 3.52 deg. C and 65.56 ± 15.31 deg. C and the time of temperature rising to 54 deg. C were respectively 37.50 ± 7.62 s, 24.50 ± 4.09 s and 57.29 ± 23.24 s for three applicators. Conclusion: MW ablation may have higher

ESR signal changes recorded in γ-irradiated bovine livers

International Nuclear Information System (INIS)

Kikuchi, Masahiro; Kobayashi, Yasuhiko

2015-01-01

After fresh raw livers of bovine were exposed to γ-rays on ice, radiation-induced radicals in the livers were measured in liquid nitrogen using electron spin resonance spectroscopy. In the magnetic field of 320.5 to 335.5 mT, the signals responsible to absorbed doses were found. The signal intensity from a main peak was increased up to 5 kGy. The side peaks existing both low and high magnetic field of the main peak showed linear responses as increasing absorbed doses. Radiation-induced radicals were found in a tissue without bones from animals. Without complicated sample preparations, the raw bovine livers can be easily measured on ESR at liquid nitrogen temperature. The ESR method may be applicable to distinguish irradiated fresh raw livers within a few days after irradiation. (author)

Mechanical properties of porcine brain tissue in vivo and ex vivo estimated by MR elastography.

Science.gov (United States)

Guertler, Charlotte A; Okamoto, Ruth J; Schmidt, John L; Badachhape, Andrew A; Johnson, Curtis L; Bayly, Philip V

2018-03-01

The mechanical properties of brain tissue in vivo determine the response of the brain to rapid skull acceleration. These properties are thus of great interest to the developers of mathematical models of traumatic brain injury (TBI) or neurosurgical simulations. Animal models provide valuable insight that can improve TBI modeling. In this study we compare estimates of mechanical properties of the Yucatan mini-pig brain in vivo and ex vivo using magnetic resonance elastography (MRE) at multiple frequencies. MRE allows estimations of properties in soft tissue, either in vivo or ex vivo, by imaging harmonic shear wave propagation. Most direct measurements of brain mechanical properties have been performed using samples of brain tissue ex vivo. It has been observed that direct estimates of brain mechanical properties depend on the frequency and amplitude of loading, as well as the time post-mortem and condition of the sample. Using MRE in the same animals at overlapping frequencies, we observe that porcine brain tissue in vivo appears stiffer than porcine brain tissue samples ex vivo at frequencies of 100 Hz and 125 Hz, but measurements show closer agreement at lower frequencies. Copyright © 2018 Elsevier Ltd. All rights reserved.

Buttressing staples with cholecyst-derived extracellular matrix (CEM) reinforces staple lines in an ex vivo peristaltic inflation model.

LENUS (Irish Health Repository)

Burugapalli, Krishna

2008-11-01

Staple line leakage and bleeding are the most common problems associated with the use of surgical staplers for gastrointestinal resection and anastomotic procedures. These complications can be reduced by reinforcing the staple lines with buttressing materials. The current study reports the potential use of cholecyst-derived extracellular matrix (CEM) in non-crosslinked (NCEM) and crosslinked (XCEM) forms, and compares their mechanical performance with clinically available buttress materials [small intestinal submucosa (SIS) and bovine pericardium (BP)] in an ex vivo small intestine model.

Quantitative characterization of viscoelastic behavior in tissue-mimicking phantoms and ex vivo animal tissues.

Directory of Open Access Journals (Sweden)

Ashkan Maccabi

Full Text Available Viscoelasticity of soft tissue is often related to pathology, and therefore, has become an important diagnostic indicator in the clinical assessment of suspect tissue. Surgeons, particularly within head and neck subsites, typically use palpation techniques for intra-operative tumor detection. This detection method, however, is highly subjective and often fails to detect small or deep abnormalities. Vibroacoustography (VA and similar methods have previously been used to distinguish tissue with high-contrast, but a firm understanding of the main contrast mechanism has yet to be verified. The contributions of tissue mechanical properties in VA images have been difficult to verify given the limited literature on viscoelastic properties of various normal and diseased tissue. This paper aims to investigate viscoelasticity theory and present a detailed description of viscoelastic experimental results obtained in tissue-mimicking phantoms (TMPs and ex vivo tissues to verify the main contrast mechanism in VA and similar imaging modalities. A spherical-tip micro-indentation technique was employed with the Hertzian model to acquire absolute, quantitative, point measurements of the elastic modulus (E, long term shear modulus (η, and time constant (τ in homogeneous TMPs and ex vivo tissue in rat liver and porcine liver and gallbladder. Viscoelastic differences observed between porcine liver and gallbladder tissue suggest that imaging modalities which utilize the mechanical properties of tissue as a primary contrast mechanism can potentially be used to quantitatively differentiate between proximate organs in a clinical setting. These results may facilitate more accurate tissue modeling and add information not currently available to the field of systems characterization and biomedical research.

Quantitative characterization of viscoelastic behavior in tissue-mimicking phantoms and ex vivo animal tissues.

Science.gov (United States)

Maccabi, Ashkan; Shin, Andrew; Namiri, Nikan K; Bajwa, Neha; St John, Maie; Taylor, Zachary D; Grundfest, Warren; Saddik, George N

2018-01-01

Viscoelasticity of soft tissue is often related to pathology, and therefore, has become an important diagnostic indicator in the clinical assessment of suspect tissue. Surgeons, particularly within head and neck subsites, typically use palpation techniques for intra-operative tumor detection. This detection method, however, is highly subjective and often fails to detect small or deep abnormalities. Vibroacoustography (VA) and similar methods have previously been used to distinguish tissue with high-contrast, but a firm understanding of the main contrast mechanism has yet to be verified. The contributions of tissue mechanical properties in VA images have been difficult to verify given the limited literature on viscoelastic properties of various normal and diseased tissue. This paper aims to investigate viscoelasticity theory and present a detailed description of viscoelastic experimental results obtained in tissue-mimicking phantoms (TMPs) and ex vivo tissues to verify the main contrast mechanism in VA and similar imaging modalities. A spherical-tip micro-indentation technique was employed with the Hertzian model to acquire absolute, quantitative, point measurements of the elastic modulus (E), long term shear modulus (η), and time constant (τ) in homogeneous TMPs and ex vivo tissue in rat liver and porcine liver and gallbladder. Viscoelastic differences observed between porcine liver and gallbladder tissue suggest that imaging modalities which utilize the mechanical properties of tissue as a primary contrast mechanism can potentially be used to quantitatively differentiate between proximate organs in a clinical setting. These results may facilitate more accurate tissue modeling and add information not currently available to the field of systems characterization and biomedical research.

In vivo and ex vivo EPR detection of spin-labelled ovalbumin in mice.

Science.gov (United States)

Abramović, Zrinka; Brgles, Marija; Habjanec, Lidija; Tomasić, Jelka; Sentjurc, Marjeta; Frkanec, Ruza

2010-10-01

In this study, spin-labelled ovalbumin (SL-OVA), free or entrapped in liposomes, was administered to mice subcutaneously (s.c.) or intravenously (i.v.) with the aim to determine the conditions for pharmacokinetic studies of spin-labelled proteins by EPR and to measure the time course of SL-OVA distribution in vivo in live mice and ex vivo in isolated organs. Upon s.c. administration, the decay of the EPR signal was followed for 60min at the site of application using an L-band EPR spectrometer. Within this time period, the signal of free SL-OVA was diminished by about 70%. It was estimated with the help of the oxidizing agent K(3)[(FeCN)(6)] that approximately 30% was a consequence of the spin label reduction to EPR non-visible hydroxylamine and about 40% was due to the SL-OVA elimination from the site of measurement. For liposome encapsulated SL-OVA, the intensity diminished only by approx. 40% in the same period, indicating that liposomes successfully protect the protein from reduction. EPR signal could not be detected directly over live mouse organs within 60min after s.c. application of SL-OVA. With the available L-band EPR spectrometer, the measurements at the site of s.c. application are possible if the amount of SL-OVA applied to a mouse is more than 3mg. For the pharmacokinetic studies of the protein distribution in organs after s.c. or i.v. injection the concentration of the spin-labelled protein should be more than 0.5mmol/kg. After i.v. administration, only ex vivo measurements were possible using an X-band EPR spectrometer, since the total amount of SL-OVA was not sufficient for in vivo detection and also because of rapid reduction of nitroxide. After 2min, the protein was preferentially distributed to liver and, to a smaller extent, to spleen.

Quinoid radio-toxin (QRT) induced metabolic changes in mice: An ex vivo and in vivo EPR investigation

Science.gov (United States)

Ibragimova, M.I.; Petukhov, V.Yu.; Zheglov, E.P.; Khan, N.; Hou, H.; Swartz, H.M.; Konjukhov, G.V.; Nizamov, R.N.

2013-01-01

Radio-toxins are toxic metabolites produced by ionizing irradiation and have toxic effects similar to those caused by direct irradiation. We have investigated the effect of a quinoid radio-toxin (QRT) obtained from γ-irradiated potato tuber on various organs in mice using ex vivo and in vivo EPR spectroscopy. Results indicate a decrease in the activity of ribonucleotide reductase enzyme in spleen of mice treated with 0.2 mg QRT. A dose of 2 mg QRT was fatal to mice within 45–60 min of treatment. Nitrosyl hemoglobin complexes α-(Fe2+–NO)α-(Fe2+)β-(Fe2+)2 were detected from spleen, blood, liver, kidney, heart, and lung tissue samples of mice treated with lethal doses of QRT. A significant decrease of pO2 in liver and brain was observed after administration of QRT at the lethal dose. The time of the appearance of the nitrosyl hemoglobin complex and its intensity varied with the dose of QRT and the type of tissue. These results indicate that the effect of the QRT is more prominent in spleen and to a lesser extent in liver and blood. The QRT action at the lethal doses resulted in an increased hypoxia over time with disruption of compensatory adaptive response. The results indicate similar outcome of QRT as observed with γ-irradiation. PMID:18230367

NAA characterization of the new Bovine Liver SRM

International Nuclear Information System (INIS)

Zeisler, R.; Mackey, E.A.; Spatz, R.O.; Greenberg, R.R.; James, W.D.

2008-01-01

The National Institute of Standards and Technology (NIST) is preparing a freeze-dried powdered bovine liver tissue Standard Reference Material (SRM) to replace SRM 1577b Bovine Liver as the stock of this material was exhausted during 2006. Like the original SRM 1577 issued in 1972, this renewal focuses on the key elements for diagnostic, nutritional, and toxicological measurements that are important to medical, veterinary, and environmental sciences investigations. NIST's approach for value assignment included extensive characterization by neutron activation analysis (NAA). Difficulties in the determination of some elements present at very low levels were overcome by use of radiochemical separations. Twentyone elements were characterized in SRM 1577c by NAA. The previous materials, SRM 1577 and 1577b, served as quality control. (author)

A novel method of mouse ex utero transplantation of hepatic progenitor cells into the fetal liver

International Nuclear Information System (INIS)

Shikanai, Mima; Asahina, Kinji; Iseki, Sachiko; Teramoto, Kenichi; Nishida, Tomohiro; Shimizu-Saito, Keiko; Ota, Masato; Eto, Kazuhiro; Teraoka, Hirobumi

2009-01-01

Avoiding the limitations of the adult liver niche, transplantation of hepatic stem/progenitor cells into fetal liver is desirable to analyze immature cells in a hepatic developmental environment. Here, we established a new monitor tool for cell fate of hepatic progenitor cells transplanted into the mouse fetal liver by using ex utero surgery. When embryonic day (ED) 14.5 hepatoblasts were injected into the ED14.5 fetal liver, the transplanted cells expressed albumin abundantly or α-fetoprotein weakly, and contained glycogen in the neonatal liver, indicating that transplanted hepatoblasts can proliferate and differentiate in concord with surrounding recipient parenchymal cells. The transplanted cells became mature in the liver of 6-week-old mice. Furthermore, this method was applicable to transplantation of hepatoblast-like cells derived from mouse embryonic stem cells. These data indicate that this unique technique will provide a new in vivo experimental system for studying cell fate of hepatic stem/progenitor cells and liver organogenesis.

Minimal vascular flows cause strong heat sink effects in hepatic radiofrequency ablation ex vivo.

Science.gov (United States)

Lehmann, Kai S; Poch, Franz G M; Rieder, Christian; Schenk, Andrea; Stroux, Andrea; Frericks, Bernd B; Gemeinhardt, Ole; Holmer, Christoph; Kreis, Martin E; Ritz, Jörg P; Zurbuchen, Urte

2016-08-01

The present paper aims to assess the lower threshold of vascular flow rate on the heat sink effect in bipolar radiofrequency ablation (RFA) ex vivo. Glass tubes (vessels) of 3.4 mm inner diameter were introduced in parallel to bipolar RFA applicators into porcine liver ex vivo. Vessels were perfused with flow rates of 0 to 1,500 ml/min. RFA (30 W power, 15 kJ energy input) was carried out at room temperature and 37°C. Heat sink effects were assessed in RFA cross sections by the decrease in ablation radius, area and by a high-resolution sector planimetry. Flow rates of 1 ml/min already caused a significant cooling effect (P ≤ 0.001). The heat sink effect reached a maximum at 10 ml/min (18.4 mm/s) and remained stable for flow rates up to 1,500 ml/min. Minimal vascular flows of ≥1 ml/min cause a significant heat sink effect in hepatic RFA ex vivo. A lower limit for volumetric flow rate was not found. The maximum of the heat sink effect was reached at a flow rate of 10 ml/min and remained stable for flow rates up to 1,500 ml/min. Hepatic inflow occlusion should be considered in RFA close to hepatic vessels. © 2016 Japanese Society of Hepato-Biliary-Pancreatic Surgery.

Ex vivo sentinel lymph node investigation in colorectal cancer

Directory of Open Access Journals (Sweden)

Antônio Hilário Alves Freitas

2013-01-01

Full Text Available Introduction: In Brazil, about 26,000 cases of colorectal cancer are diagnosed per year. Pa- tients considered at the early stage of disease (without lymph node evolve with tumor relapse or recurrence in up to a quarter of cases, probably due to understaging. Objective: Research on ex vivo sentinel lymph node in patients with colorectal adenocarcinoma. Materials and methods: We studied 37 patients who underwent curative surgical resection. The marker used to identify lymph nodes was patent blue dye injected into the peritu- moral submucosa of the open surgical specimen immediately after its removal from the abdominal cavity. Results: Ex vivo identification of sentinel lymph node with marker occurred in 13 (35.1% patients. The sensitivity was 40% and 60% false negative. The detailed histological examina- tion of sentinel lymph nodes with multilevel section and immunohistochemistry showed metastasis in one (4.3% individual, considered ultra-staging. Conclusion: The ex vivo identification of sentinel lymph node had questionable benefits, and worse results when include patients with rectal cancer. Restaging of one patient was possible after multilevel section and immunohistochemistry of the sentinel lymph node, but more research is needed to evaluate the role of micrometastases in patients with colorectal cancer. Resumo: Introdução: No Brasil, a cada ano são diagnosticados cerca de 26.000 casos de câncer colorre- tal. Pacientes com estadiamento considerado inicial, sem linfonodo metastático, evoluem com recorrência ou recidiva do tumor em até um quarto dos casos, por provável subesta- diamento. Objetivo: pesquisar sobre linfonodo-sentinela ex vivo em pacientes com adeno- carcinoma colorretal. Objetivo: Foram estudados 37 pacientes, submetidos à cirurgia oncológica com ressecção caráter curativo. O marcador de linfonodos utilizado foi o corante azul patente, injetado na submucosa peritumoral da peça cirúrgica aberta imediatamente

Aspects of the regulation of long-chain fatty acid oxidation in bovine liver

International Nuclear Information System (INIS)

Jesse, B.W.; Emery, R.S.; Thomas, J.W.

1986-01-01

Factors involved in regulation of bovine hepatic fatty acid oxidation were examined using liver slices. Fatty acid oxidation was measured as the conversion of l-[ 14 C] palmitate to 14 CO 2 and total [ 14 C] acid-soluble metabolites. Extended (5 to 7 d) fasting of Holstein cows had relatively little effect on palmitate oxidation to acid-soluble metabolites by liver slices, although oxidation to CO 2 was decreased. Feeding a restricted roughage, high concentrate ration to lactating cows resulted in inhibition of palmitate oxidation. Insulin, glucose, and acetate inhibited palmitate oxidation by bovine liver slices. The authors suggest the regulation of bovine hepatic fatty acid oxidation may be less dependent on hormonally induced alterations in enzyme activity as observed in rat liver and more dependent upon action of rumen fermentation products or their metabolites on enzyme systems involved in fatty acid oxidation

In vivo and ex vivo characterization of a novel Er fiber laser system for fractional treatment of soft oral tissues

Science.gov (United States)

Shatilova, Ksenia; Aloian, Georgii; Karabut, Maria; Ryabova, Valentina; Yaroslavsky, Ilya V.; Altshuler, Gregory

2018-02-01

In this work, we present the first histological in vivo and ex vivo study of effects of fractional Er fiber laser (wavelength 1550 nm, peak power 25 W) on keratinized gum and alveolar mucosa for gum regeneration. Biopsy with subsequent NBTC staining was used as primary evaluation technique. Ex vivo, porcine tissue model was used. Effects of pulse energy, beam diameter, and beam divergence were investigated in detail. It has been demonstrated that under optimal conditions columns up to 800 μm in depth could be reliably produced with 130 mJ pulses. Clinically, 2 subjects were treated and 4 punch biopsies were collected. The results were compared with ex vivo data. Both ex vivo and in vivo datasets suggest feasibility of a dental fractional system intended for gum regeneration.

Feasibility of Commercially Available, Fully Automated Hepatic CT Volumetry for Assessing Both Total and Territorial Liver Volumes in Liver Transplantation

Energy Technology Data Exchange (ETDEWEB)

Shin, Cheong Il; Kim, Se Hyung; Rhim, Jung Hyo; Yi, Nam Joon; Suh, Kyung Suk; Lee, Jeong Min; Han, Joon Koo; Choi, Byung Ihn [Seoul National University Hospital, Seoul (Korea, Republic of)

2013-02-15

To assess the feasibility of commercially-available, fully automated hepatic CT volumetry for measuring both total and territorial liver volumes by comparing with interactive manual volumetry and measured ex-vivo liver volume. For the assessment of total and territorial liver volume, portal phase CT images of 77 recipients and 107 donors who donated right hemiliver were used. Liver volume was measured using both the fully automated and interactive manual methods with Advanced Liver Analysis software. The quality of the automated segmentation was graded on a 4-point scale. Grading was performed by two radiologists in consensus. For the cases with excellent-to-good quality, the accuracy of automated volumetry was compared with interactive manual volumetry and measured ex-vivo liver volume which was converted from weight using analysis of variance test and Pearson's or Spearman correlation test. Processing time for both automated and interactive manual methods was also compared. Excellent-to-good quality of automated segmentation for total liver and right hemiliver was achieved in 57.1% (44/77) and 17.8% (19/107), respectively. For both total and right hemiliver volumes, there were no significant differences among automated, manual, and ex-vivo volumes except between automate volume and manual volume of the total liver (p = 0.011). There were good correlations between automate volume and ex-vivo liver volume ({gamma}= 0.637 for total liver and {gamma}= 0.767 for right hemiliver). Both correlation coefficients were higher than those with manual method. Fully automated volumetry required significantly less time than interactive manual method (total liver: 48.6 sec vs. 53.2 sec, right hemiliver: 182 sec vs. 244.5 sec). Fully automated hepatic CT volumetry is feasible and time-efficient for total liver volume measurement. However, its usefulness for territorial liver volumetry needs to be improved.

Feasibility of Commercially Available, Fully Automated Hepatic CT Volumetry for Assessing Both Total and Territorial Liver Volumes in Liver Transplantation

International Nuclear Information System (INIS)

Shin, Cheong Il; Kim, Se Hyung; Rhim, Jung Hyo; Yi, Nam Joon; Suh, Kyung Suk; Lee, Jeong Min; Han, Joon Koo; Choi, Byung Ihn

2013-01-01

To assess the feasibility of commercially-available, fully automated hepatic CT volumetry for measuring both total and territorial liver volumes by comparing with interactive manual volumetry and measured ex-vivo liver volume. For the assessment of total and territorial liver volume, portal phase CT images of 77 recipients and 107 donors who donated right hemiliver were used. Liver volume was measured using both the fully automated and interactive manual methods with Advanced Liver Analysis software. The quality of the automated segmentation was graded on a 4-point scale. Grading was performed by two radiologists in consensus. For the cases with excellent-to-good quality, the accuracy of automated volumetry was compared with interactive manual volumetry and measured ex-vivo liver volume which was converted from weight using analysis of variance test and Pearson's or Spearman correlation test. Processing time for both automated and interactive manual methods was also compared. Excellent-to-good quality of automated segmentation for total liver and right hemiliver was achieved in 57.1% (44/77) and 17.8% (19/107), respectively. For both total and right hemiliver volumes, there were no significant differences among automated, manual, and ex-vivo volumes except between automate volume and manual volume of the total liver (p = 0.011). There were good correlations between automate volume and ex-vivo liver volume (γ= 0.637 for total liver and γ= 0.767 for right hemiliver). Both correlation coefficients were higher than those with manual method. Fully automated volumetry required significantly less time than interactive manual method (total liver: 48.6 sec vs. 53.2 sec, right hemiliver: 182 sec vs. 244.5 sec). Fully automated hepatic CT volumetry is feasible and time-efficient for total liver volume measurement. However, its usefulness for territorial liver volumetry needs to be improved.

Radiofrequency Ablation (RFA): Development of a Flow Model for Bovine Livers for Extensive Bench Testing

International Nuclear Information System (INIS)

Lubienski, Andreas; Bitsch, Rudi G.; Lubienski, Katrin; Kauffmann, Guenter; Duex, Markus

2006-01-01

Purpose. To develop a flow model for bovine livers for extensive bench testing of technical improvements or procedure-related developments of radiofrequency ablation excluding animal experiments. Methods. The perfusion of bovine livers directly from the slaughterhouse was simulated in a liver perfusion tank developed for the experimental work. The liver perfusion medium used was a Tyrode solution prepared in accordance with physiologic criteria (as for liver transplants) which was oxygenated by an oxygenator and heated to 36.5 deg. C. Portal vein circulation was regulated via a flow- and pressure-controlled pump and arterial circulation using a dialysis machine. Flow rate and pressure were adjusted as for the physiology of a human liver converted to bovine liver conditions. The fluid discharged from the liver was returned into the perfusion system through the vena cava. Extendable precision swivel arms with the radiofrequency probe attached were mounted on the liver perfusion tank. RFA was conducted with the RF3000 generator and a 2 cm LeVeen needle (Boston Scientific, Ratingen, Germany) in a three-dimensional grid for precise localization of the generated thermolesions. Results. Four bovine livers weighing 8.4 ± 0.4 kg each were prepared, connected to the perfusion system, and consecutively perfused for the experiments. Mean arterial flow was 569 ± 43 ml/min, arterial pressure 120 mmHg, portovenous flow 1440 ± 305 ml/min, and portal pressure 10 mmHg. Macroscopic evaluation after the experiments revealed no thrombi within the hepatic vessels. A total of 136 RF thermolesions were generated with an average number of 34 per liver. Mean RF duration was 2:59 ± 2:01 min:sec with an average baseline impedance of 28.2 ± 3.4 ohms. The mean diameter of the thermolesions along the puncture channel was 22.98 ± 4.34 mm and perpendicular to the channel was 23.27 ± 4.82 mm. Conclusion. Extracorporeal perfusion of bovine livers with consecutive standardized RF ablation was

Studies on the transverse localization of lysophospholipase II in bovine liver microsomes by immunological techniques

NARCIS (Netherlands)

Moonen, H.; Bosch, H. van den

1979-01-01

1. 1. Lysophospholipase activity solubilized from bovine liver microsomes could be precipitated for more than 80% by antibodies evoked in rabbits against the purified bovine liver lysophospholipase II. 2. 2. After solubilization of the microsomes in 1.5% sodium deoxycholate, an immunoprecipitate

MRI parcellation of ex vivo medial temporal lobe.

Science.gov (United States)

Augustinack, Jean C; Magnain, Caroline; Reuter, Martin; van der Kouwe, André J W; Boas, David; Fischl, Bruce

2014-06-01

Recent advancements in radio frequency coils, field strength and sophisticated pulse sequences have propelled modern brain mapping and have made validation to biological standards - histology and pathology - possible. The medial temporal lobe has long been established as a pivotal brain region for connectivity, function and unique structure in the human brain, and reveals disconnection in mild Alzheimer's disease. Specific brain mapping of mesocortical areas affected with neurofibrillary tangle pathology early in disease progression provides not only an accurate description for location of these areas but also supplies spherical coordinates that allow comparison between other ex vivo cases and larger in vivo datasets. We have identified several cytoarchitectonic features in the medial temporal lobe with high resolution ex vivo MRI, including gray matter structures such as the entorhinal layer II 'islands', perirhinal layer II-III columns, presubicular 'clouds', granule cell layer of the dentate gyrus as well as lamina of the hippocampus. Localization of Brodmann areas 28 and 35 (entorhinal and perirhinal, respectively) demonstrates MRI based area boundaries validated with multiple methods and histological stains. Based on our findings, both myelin and Nissl staining relate to contrast in ex vivo MRI. Precise brain mapping serves to create modern atlases for cortical areas, allowing accurate localization with important applications to detecting early disease processes. Copyright © 2013 Elsevier Inc. All rights reserved.

Endovascular optical coherence tomography ex vivo: venous wall anatomy and tissue alterations after endovenous therapy

International Nuclear Information System (INIS)

Meissner, Oliver A.; Schmedt, Claus-Georg; Steckmeier, Bernd M.; Hunger, Kathrin; Reiser, Maximilian; Mueller-Lisse, Ullrich; Hetterich, Holger; Rieber, Johannes; Sroka, Ronald; Babaryka, Gregor; Siebert, Uwe

2007-01-01

Endovascular optical coherence tomography (OCT) is a new imaging modality providing histology-like information of the venous wall. Radiofrequency ablation (RFA) and laser therapy (ELT) are accepted alternatives to surgery. This study evaluated OCT for qualitative assessment of venous wall anatomy and tissue alterations after RFA and ELT in bovine venous specimens. One hundred and thirty-four venous segments were obtained from ten ex-vivo bovine hind limbs. OCT signal characteristics for different wall layers were assessed in 180/216 (83%) quadrants from 54 normal venous cross-sections. Kappa statistics (κ) were used to calculate intra- and inter-observer agreement. Qualitative changes after RFA (VNUS-Closure) and ELT (diode laser 980 nm, energy densities 15 Joules (J)/cm, 25 J/cm, 35 J/cm) were described in 80 venous cross-sections. Normal veins were characterized by a three-layered appearance. After RFA, loss of three-layered appearance and wall thickening at OCT corresponded with circular destruction of tissue structures at histology. Wall defects after ELT ranged from non-transmural punctiform damage to complete perforation, depending on the energy density applied. Intra- and inter-observer agreement for reading OCT images was very high (0.90 and 0.88, respectively). OCT allows for reproducible evaluation of normal venous wall and alterations after endovenous therapy. OCT could prove to be valuable for optimizing endovenous therapy in vivo. (orig.)

Mice with humanized liver endothelium

NARCIS (Netherlands)

el Filali, E.

2014-01-01

The only curative treatment option for a large proportion of patients suffering from a liver disorder is liver transplantation. The use of ex vivo genetically modified autologous liver cells instead of whole liver transplantation could overcome the problem of donor scarcity. Even though clinical

Digital Radiography for Determination of Primary Tooth Length: In Vivo and Ex Vivo Studies

Directory of Open Access Journals (Sweden)

Maria D. Basso

2015-01-01

Full Text Available Background. Methods for determining the root canal length of the primary tooth should yield accurate and reproducible results. In vitro studies show some limitations, which do not allow their findings to be directly transferred to a clinical situation. Aim. To compare the accuracy of radiographic tooth length obtained from in vivo digital radiograph with that obtained from ex vivo digital radiograph. Method. Direct digital radiographs of 20 upper primary incisors were performed in teeth (2/3 radicular resorption that were radiographed by an intraoral sensor, according to the long-cone technique. Teeth were extracted, measured, and mounted in a resin block, and then radiographic template was used to standardise the sensor-target distance (30 cm. The apparent tooth length (APTL was obtained from the computer screen by means of an electronic ruler accompanying the digital radiography software (CDR 2.0, whereas the actual tooth length (ACTL was obtained by means of a digital calliper following extraction. Data were compared to the ACTL by variance analysis and Pearson’s correlation test. Results. The values for APTL obtained from in vivo radiography were slightly underestimated, whereas those values obtained from ex vivo were slightly overestimated. No significance was observed (P≤0.48 between APTL and ACTL. Conclusion. The length of primary teeth estimated by in vivo and ex vivo comparisons using digital radiography was found to be similar to the actual tooth length.

Electromagnetic tracking for CT-guided spine interventions: phantom, ex-vivo and in-vivo results

International Nuclear Information System (INIS)

Bruners, Philipp; Penzkofer, Tobias; Nagel, Markus; Elfring, Robert; Schmitz-Rode, Thomas; Gronloh, Nina; Guenther, Rolf W.; Mahnken, Andreas H.

2009-01-01

An electromagnetic-based tracking and navigation system was evaluated for interventional radiology. The electromagnetic tracking system (CAPPA IRAD EMT, CASinnovations, Erlangen, Germany) was used for real-time monitoring of punctures of the lumbar facet joints and intervertebral disks in a spine phantom, three pig cadavers and three anaesthesized pigs. Therefore, pre-interventional computed tomography (CT) datasets were transferred to the navigation system and puncture trajectories were planned. A coaxial needle was advanced along the trajectories while the position of the needle tip was monitored in real time. After puncture tracts were marked with pieces of wire another CT examination was performed and distances between wires and anatomical targets were measured. Performing punctures of the facet joints mean needle positioning errors were 0.4 ± 0.8 mm in the spine phantom, 2.8 ± 2.1 mm ex vivo and 3.0 ± 2.0 mm in vivo with mean length of the puncture tract of 54.0 ± 10.4 mm (phantom), 51.6 ± 12.6 mm (ex vivo) and 50.9 ± 17.6 mm (in vivo). At first attempt, intervertebral discs were successfully punctured in 15/15 in the phantom study, in 12/15 in the ex-vivo study and 14/15 in the in-vivo study, respectively. Immobilization of the patient and optimal positioning of the field generator are essential to achieve a high accuracy of needle placement in a clinical CT setting. (orig.)

Feasibility of computed tomography based thermometry during interstitial laser heating in bovine liver

International Nuclear Information System (INIS)

Pandeya, G.D.; Klaessens, J.H.G.M.; Greuter, M.J.W.; Oudkerk, M.; Schmidt, B.; Flohr, T.; Hillegersberg, R. van

2011-01-01

To assess the feasibility of computed tomography (CT) based thermometry during interstitial laser heating in the bovine liver. Four freshly exercised cylindrical blocks of bovine tissue were heated using a continuous laser of Nd:YAG (wavelength: 1064 nm, active length: 30 mm, power: 10-30 W). All tissues were imaged at least once before and 7 times during laser heating using CT and temperatures were simultaneously measured with 5 calibrated thermal sensors. The dependency of the average CT numbers as a function of temperature was analysed with regression analysis and a CT thermal sensitivity was derived. During laser heating, the growing hypodense area was observed around the laser source and that area showed an increase as a function of time. The formation of hypodense area was caused by declining in CT numbers at increasing temperatures. The regression analysis showed an inverse linear dependency between temperature and average CT number with -0.65 ± 0.048 HU/ C (R 2 = 0.75) for the range of 18-85 C in bovine liver. The non-invasive CT based thermometry during interstitial laser heating is feasible in the bovine liver. CT based thermometry could be further developed and may be of potential use during clinical LITT of the liver. (orig.)

Optimized isolation enables Ex vivo analysis of microglia from various central nervous system regions

NARCIS (Netherlands)

De Haas, Alexander H.; Boddeke, Hendricus W. G. M.; Brouwer, Nieske; Biber, Knut

2007-01-01

Ex vivo analysis is an accurate and convenient way to study in vivo microglia phenotype and function. However, current microglia isolation protocols for ex vivo analysis show many differences in isolation steps (perfusion, removal of meninges and blood vessels, mechanical dissociation, enzymatic

Measurement of the coagulation dynamics of bovine liver using the modified microscopic Beer-Lambert law.

Science.gov (United States)

Terenji, Albert; Willmann, Stefan; Osterholz, Jens; Hering, Peter; Schwarzmaier, Hans-Joachim

2005-06-01

During heating, the optical properties of biological tissues change with the coagulation state. In this study, we propose a technique, which uses these changes to monitor the coagulation process during laser-induced interstitial thermotherapy (LITT). Untreated and coagulated (water bath, temperatures between 35 degrees C and 90 degrees C for 20 minutes.) samples of bovine liver tissue were examined using a Nd:YAG (lambda = 1064 nm) frequency-domain reflectance spectrometer. We determined the time integrated intensities (I(DC)) and the phase shifts (Phi) of the photon density waves after migration through the tissue. From these measured quantities, the time of flight (TOF) of the photons and the absorption coefficients of the samples were derived using the modified microscopic Beer-Lambert law. The absorption coefficients of the liver samples decreased significantly with the temperature in the range between 50 degrees C and 70 degrees C. At the same time, the TOF of the investigated photos was found increased indicating an increased scattering. The coagulation dynamics could be well described using the Arrhenius formalism with the activation energy of 106 kJ/mol and the frequency factor of 1.59 x 10(13)/second. Frequency-domain reflectance spectroscopy in combination with the modified microscopic Beer-Lambert (MBL) is suitable to measure heat induced changes in the absorption and scattering properties of bovine liver in vitro. The technique may be used to monitor the coagulation dynamics during local thermo-coagulation in vivo. Copyright 2005 Wiley-Liss, Inc.

Mice with diet-induced obesity demonstrate a relative prothrombotic factor profile and a thicker aorta with reduced ex-vivo function.

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Uner, Aykut G; Unsal, Cengiz; Unsal, Humeyra; Erdogan, Mumin A; Koc, Ece; Ekici, Mehmet; Avci, Hamdi; Balkaya, Muharrem; Belge, Ferda; Tarin, Lokman

2018-04-01

: Classical risk factors such as cholesterol and lipoproteins are currently not sufficient to explain all physiopathological processes of obesity-related vascular dysfunction as well as atherosclerosis and arteriosclerosis. Therefore, the discovery of potential markers involved in vascular dysfunction in the obese state is still needed. Disturbances in hemostatic factors may be involved in the developmental processes associated with obesity-related cardiovascular disorders. We hypothesized that alterations of several hemostatic factors in the obese state could correlate with the function and morphology of the aorta and it could play an important role in the development of vascular dysfunction. To test this, we fed mice with a high-fat diet for 18 weeks and investigated the relationships between selected hemostatic factors (in either plasma or in the liver), metabolic hormones and morphology, and ex-vivo function of the aorta. Here, we show that 18-week exposure to a high-fat diet results in a higher plasma fibrinogen and prolonged prothrombin time in diet-induced obese mice compared to the controls. In addition, liver levels or activities of FII, FX, activated protein C, AT-III, and protein S are significantly different in diet-induced obese mice as compared to the controls. Curiously, FII, FVIII, FX, activated protein C, PTT, and protein S are correlated with both the aorta histology (aortic thickness and diameter) and ex-vivo aortic function. Notably, ex-vivo studies revealed that diet-induced obese mice show a marked attenuation in the functions of the aorta. Taken together, aforementioned hemostatic factors may be considered as critical markers for obesity-related vascular dysfunction and they could play important roles in diagnosing of the dysfunction.

Sex differences in the pro-inflammatory cytokine response to endotoxin unfold in vivo but not ex vivo in healthy humans.

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Wegner, Alexander; Benson, Sven; Rebernik, Laura; Spreitzer, Ingo; Jäger, Marcus; Schedlowski, Manfred; Elsenbruch, Sigrid; Engler, Harald

2017-07-01

Clinical data indicate that inflammatory responses differ across sexes, but the mechanisms remain elusive. Herein, we assessed in vivo and ex vivo cytokine responses to bacterial endotoxin in healthy men and women to elucidate the role of systemic and cellular factors underlying sex differences in inflammatory responses. Participants received an i.v. injection of low-dose endotoxin (0.4 ng/kg body mass), and plasma TNF-α and IL-6 responses were analyzed over a period of 6 h. In parallel, ex vivo cytokine production was measured in endotoxin-stimulated blood samples obtained immediately before in vivo endotoxin administration. As glucocorticoids (GCs) play an important role in the negative feedback regulation of the inflammatory response, we additionally analyzed plasma cortisol concentrations and ex vivo GC sensitivity of cytokine production. Results revealed greater in vivo pro-inflammatory responses in women compared with men, with significantly higher increases in plasma TNF-α and IL-6 concentrations. In addition, the endotoxin-induced rise in plasma cortisol was more pronounced in women. In contrast, no sex differences in ex vivo cytokine production and GC sensitivity were observed. Together, these findings demonstrate major differences in in vivo and ex vivo responses to endotoxin and underscore the importance of systemic factors underlying sex differences in the inflammatory response.

Radiation effects on vitamin A and β-carotene contents in bovine liver and swine pate de foie

International Nuclear Information System (INIS)

Taipina, Magda Sinigalia

2001-01-01

In this study, vitamin A and β-carotene contents and the percent of activity retention were analyzed in 3 kGy and 30 kGy 60 Co γ-irradiated samples of bovine liver and swine liver pate. Six different lots of liver samples weighing 100 g obtained at the meat market were employed. Irradiation was performed with liver samples initially frozen (-15 deg C). Similarly, six different lots of swine liver pate samples weighing about 100 - 125 g , at temperature of 7 deg C were irradiated. Pre-treatments and analysis methods were those described in Instituto Adolfo Lutz Norms for food analyses. Two and five- gram samples in duplicates for bovine liver and five gram-samples for swine liver pate were used. The results showed that there were no losses of either vitamin A or provitamin A activities in the samples of bovine liver and swine liver pate irradiated with a dose of 3 kGy (retention about 100%). On the other hand, for swine liver pate samples of vitamin A were maintained after irradiation with 30 kGy. (author)

Ex vivo tracheomalacia model with 3D-printed external tracheal splint.

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Kaye, Rachel; Goldstein, Todd; Aronowitz, Danielle; Grande, Daniel A; Zeltsman, David; Smith, Lee P

2017-04-01

To design and evaluate an ex vivo model of tracheomalacia with and without a three-dimensional (3D)-printed external tracheal splint. Prospective, ex vivo animal trial. Three groups of ex vivo porcine tracheas were used: 1) control (unmanipulated trachea), 2) tracheomalacia (tracheal rings partially incised and crushed), and 3) splinted tracheomalacia (external custom tracheal splint fitted onto group 2 trachea). Each end of an ex vivo trachea was sealed with a custom-designed and 3D-printed cap; a transducer was placed through one end to measure the pressure inside the trachea. Although the negative pressure was applied to the tracheal lumen, the tracheal wall collapse was measured externally and internally using a bronchoscope. Each group had at least three recorded trials. Tracheal diameter was evaluated using ImageJ software (National Institutes of Health, Bethesda, MD) and was averaged between two raters. Average tracheal occlusion percentage was compared using Student t test. The average occlusion was 31% for group 1, 87.4% for group 2, and 20% for group 3. Significant differences were found between the control and tracheomalacia groups (P tracheomalacia and splinted tracheomalacia groups (P tracheomalacia groups (P = 0.13). Applied pressure was plotted against occlusion and regression line slope differed between the tracheomalacia (0.91) and control (0.12) or splinted tracheomalacia (0.39) groups. We demonstrate the potential for an ex vivo tracheomalacia model to reproduce airway collapse and show that this collapse can be treated successfully with a 3D-printed external splint. These results are promising and justify further studies. N/A. Laryngoscope, 127:950-955, 2017. © 2016 The American Laryngological, Rhinological and Otological Society, Inc.

In-vivo and ex-vivo assessment of the accuracy of the computer-aided volumetry of porcine kidney in CT images

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Cai, W.; Harris, G.; Holalkere, N.; Sahani, D.; Yoshida, H. [Massachusetts General Hospital and Harvard Medical School (United States)

2007-06-15

Measurement of kidney volume by computed tomography (CT), called renal volumetry, is indispensable for diagnosis and treatment of kidney-related diseases. Computer-aided volumetry (CAV) of kidney relies on an efficient and accurate segmentation method of the kidney. The purpose of this study is to assess the accuracy of our CAV of kidney scheme using dynamic-threshold (DT) level set method, based on in-vivo and ex-vivo reference standards. Eight Yorkshire breed anesthetized pigs were scanned on a 64-slice multi-detector CT scanner (Sensation 64, Siemens) after an injection of iodinated (300 mgl/ml) contrast agent through an IV cannula. The kidneys of the pigs were then surgically resected and scanned on CT in the same manner. Both in-vivo and ex-vivo CT images were subjected to our volumetry scheme. The resulting volumes of the kidneys were compared with the in-vivo and ex-vivo reference standards: the former was established by manual contouring of the kidneys on the CT images by an experienced radiologist, and the latter was established as the water displacement volume of the resected kidney. Our CAV of kidney scheme demonstrated accurate in-vivo and ex-vivo measurement of kidney volume, despite a large difference between the two reference standards. (orig.)

In-vivo and ex-vivo assessment of the accuracy of the computer-aided volumetry of porcine kidney in CT images

International Nuclear Information System (INIS)

Cai, W.; Harris, G.; Holalkere, N.; Sahani, D.; Yoshida, H.

2007-01-01

Measurement of kidney volume by computed tomography (CT), called renal volumetry, is indispensable for diagnosis and treatment of kidney-related diseases. Computer-aided volumetry (CAV) of kidney relies on an efficient and accurate segmentation method of the kidney. The purpose of this study is to assess the accuracy of our CAV of kidney scheme using dynamic-threshold (DT) level set method, based on in-vivo and ex-vivo reference standards. Eight Yorkshire breed anesthetized pigs were scanned on a 64-slice multi-detector CT scanner (Sensation 64, Siemens) after an injection of iodinated (300 mgl/ml) contrast agent through an IV cannula. The kidneys of the pigs were then surgically resected and scanned on CT in the same manner. Both in-vivo and ex-vivo CT images were subjected to our volumetry scheme. The resulting volumes of the kidneys were compared with the in-vivo and ex-vivo reference standards: the former was established by manual contouring of the kidneys on the CT images by an experienced radiologist, and the latter was established as the water displacement volume of the resected kidney. Our CAV of kidney scheme demonstrated accurate in-vivo and ex-vivo measurement of kidney volume, despite a large difference between the two reference standards. (orig.)

In vitro and ex vivo effect of hyaluronic acid on erythrocyte flow properties

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Palatnik S

2010-02-01

Full Text Available Abstract Background Hyaluronic acid (HA is present in many tissues; its presence in serum may be related to certain inflammatory conditions, tissue damage, sepsis, liver malfunction and some malignancies. In the present work, our goal was to investigate the significance of hyaluronic acid effect on erythrocyte flow properties. Therefore we performed in vitro experiments incubating red blood cells (RBCs with several HA concentrations. Afterwards, in order to corroborate the pathophysiological significance of the results obtained, we replicated the in vitro experiment with ex vivo RBCs from diagnosed rheumatoid arthritis (RA patients, a serum HA-increasing pathology. Methods Erythrocyte deformability (by filtration through nucleopore membranes and erythrocyte aggregability (EA were tested on blood from healthy donors additioned with purified HA. EA was measured by transmitted light and analyzed with a mathematical model yielding two parameters, the aggregation rate and the size of the aggregates. Conformational changes of cytoskeleton proteins were estimated by electron paramagnetic resonance spectroscopy (EPR. Results In vitro, erythrocytes treated with HA showed increased rigidity index (RI and reduced aggregability, situation strongly related to the rigidization of the membrane cytoskeleton triggered by HA, as shown by EPR results. Also, a significant correlation (r: 0.77, p Conclusions Our results lead us to postulate the hypothesis that HA interacts with the erythrocyte surface leading to modifications in erythrocyte rheological and flow properties, both ex vivo and in vitro.

Testicular cells exhibit similar molecular responses to cigarette smoke condensate ex vivo and in vivo.

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Esakky, Prabagaran; Hansen, Deborah A; Drury, Andrea M; Felder, Paul; Cusumano, Andrew; Moley, Kelle H

2018-01-01

Male exposure to cigarette smoke is associated with seminal defects and with congenital anomalies and childhood cancers in offspring. In mice, paternal exposure to cigarette smoke condensate (CSC) causes molecular defects in germ cells and phenotypic effects in their offspring. Here we used an ex vivo testicular explant model and in vivo exposure to determine the concentration at which CSC impairs spermatogenesis and offspring development. We explanted testis tissue at postnatal day (P)5.5 and cultured it until P11.5. Assessment of growth parameters by analyzing expression of cell-specific markers revealed that the explant system maintained structural and functional integrity. We exposed the P5.5 to -11.5 explants to various concentrations (40-160 µg/ml) of CSC and confirmed that nicotine in the CSC was metabolized to cotinine. We assessed various growth and differentiation parameters, as well as testosterone production, and observed that many spermatogenesis features were impaired at 160 µg/ml CSC. The same parameters were impaired by a similar CSC concentration in vivo Finally, females mated to males that were exposed to 160 µg/ml CSC neonatally had increased rates of pup resorption. We conclude that male exposure to CSC impairs offspring development and that the concentration at which CSC impairs spermatogenesis is similar in vivo and ex vivo. Given that the concentrations of CSC we used contained similar doses of nicotine as human smokers are exposed to, we argue that our model mimics human male reproductive effects of smoking.-Esakky, P., Hansen, D. A., Drury, A. M., Felder, P., Cusumano, A., Moley, K. H. Testicular cells exhibit similar molecular responses to cigarette smoke condensate ex vivo and in vivo . © FASEB.

Population-averaged macaque brain atlas with high-resolution ex vivo DTI integrated into in vivo space.

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Feng, Lei; Jeon, Tina; Yu, Qiaowen; Ouyang, Minhui; Peng, Qinmu; Mishra, Virendra; Pletikos, Mihovil; Sestan, Nenad; Miller, Michael I; Mori, Susumu; Hsiao, Steven; Liu, Shuwei; Huang, Hao

2017-12-01

Animal models of the rhesus macaque (Macaca mulatta), the most widely used nonhuman primate, have been irreplaceable in neurobiological studies. However, a population-averaged macaque brain diffusion tensor imaging (DTI) atlas, including comprehensive gray and white matter labeling as well as bony and facial landmarks guiding invasive experimental procedures, is not available. The macaque white matter tract pathways and microstructures have been rarely recorded. Here, we established a population-averaged macaque brain atlas with high-resolution ex vivo DTI integrated into in vivo space incorporating bony and facial landmarks, and delineated microstructures and three-dimensional pathways of major white matter tracts in vivo MRI/DTI and ex vivo (postmortem) DTI of ten rhesus macaque brains were acquired. Single-subject macaque brain DTI template was obtained by transforming the postmortem high-resolution DTI data into in vivo space. Ex vivo DTI of ten macaque brains was then averaged in the in vivo single-subject template space to generate population-averaged macaque brain DTI atlas. The white matter tracts were traced with DTI-based tractography. One hundred and eighteen neural structures including all cortical gyri, white matter tracts and subcortical nuclei, were labeled manually on population-averaged DTI-derived maps. The in vivo microstructural metrics of fractional anisotropy, axial, radial and mean diffusivity of the traced white matter tracts were measured. Population-averaged digital atlas integrated into in vivo space can be used to label the experimental macaque brain automatically. Bony and facial landmarks will be available for guiding invasive procedures. The DTI metric measurements offer unique insights into heterogeneous microstructural profiles of different white matter tracts.

Ebola Virus Persistence in Semen Ex Vivo.

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Fischer, Robert J; Judson, Seth; Miazgowicz, Kerri; Bushmaker, Trent; Munster, Vincent J

2016-02-01

On March 20, 2015, a case of Ebola virus disease was identified in Liberia that most likely was transmitted through sexual contact. We assessed the efficiency of detecting Ebola virus in semen samples by molecular diagnostics and the stability of Ebola virus in ex vivo semen under simulated tropical conditions.

In vitro and in vivo studies of the endocrine disrupting potency of cadmium in roach (Rutilus rutilus) liver

International Nuclear Information System (INIS)

Gerbron, M.; Geraudie, P.; Xuereb, B.; Marie, S.; Minier, C.

2015-01-01

Highlights: • Ex vivo and in vivo experiments suggest estrogen receptor-driven effects of cadmium. • CdCl 2 is strongly anti-estrogenic when co-exposed to E2. • CdCl 2 + E2 significantly inhibit vtg and erα mRNA expressions. • CdCl 2 compromises the E2-mediated induction of the ar mRNA expression in vivo. - Abstract: Cadmium has been reported to exert estrogenic, antiestrogenic or both effects in vertebrate species. To elucidate the endocrine disrupting action of CdCl 2 , ex vivo and in vivo experiments were performed in roach (Rutilus rutilus). Roach liver explants were exposed to a range of CdCl 2 concentrations alone (0.1–50 μM) or with an effective concentration (100 nM) of 17β-estradiol (E2). In addition, juvenile roach were intraperitoneally injected with CdCl 2 (0.1–2.5 mg/kg) with or without 1 mg E2/kg. Subsequent analysis evaluated the effect of CdCl 2 on vitellogenin (VTG) synthesis both at the mRNA and protein level, on estrogen receptors (erα and erβ1) and on androgen receptor (ar) mRNA expression. Ex vivo and in vivo experiments indicated that CdCl 2 is strongly anti-estrogenic as, when co-exposed to E2, CdCl 2 significantly inhibited VTG production as well as vtg and erα mRNA expressions. Moreover, CdCl 2 compromised the E2-mediated induction of the ar mRNA expression in vivo

Light Emission Requires Exposure to the Atmosphere in Ex Vivo Bioluminescence Imaging

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Yusuke Inoue

2006-04-01

Full Text Available The identification of organs bearing luciferase activity by in vivo bioluminescence imaging (BLI is often difficult, and ex vivo imaging of excised organs plays a complementary role. This study investigated the importance of exposure to the atmosphere in ex vivo BLI. Mice were inoculated with murine pro-B cell line Ba/F3 transduced with firefly luciferase and p190 BCR-ABL. They were killed following in vivo BLI, and whole-body imaging was done after death and then after intraperitoneal air injection. In addition, the right knee was exposed and imaged before and after the adjacent bones were cut. Extensive light signals were seen on in vivo imaging. The luminescence disappeared after the animal was killed, and air injection restored the light emission from the abdomen only, suggesting a critical role of atmospheric oxygen in luminescence after death. Although no substantial light signal at the right knee was seen before bone cutting, light emission was evident after cutting. In conclusion, in ex vivo BLI, light emission requires exposure to the atmosphere. Bone destruction is required to demonstrate luciferase activity in the bone marrow after death.

Nutritional Quality and Physicochemical Characteristics of Defatted Bovine Liver Treated by Supercritical Carbon Dioxide and Organic Solvent

Science.gov (United States)

Kang, Sung-Won; Kim, Hye-Min; Rahman, M. Shafiur; Kim, Ah-Na; Yang, Han-Sul

2017-01-01

Defatted bovine liver (DBL) is a potential source of protein and minerals. Supercritical carbon dioxide (SC-CO2) and a traditional organic solvent method were used to remove lipid from bovine liver, and the quality characteristics of a control bovine liver (CBL), bovine liver defatted by SC-CO2 (DBLSC-CO2) at different pressures, and bovine liver defatted by organic solvent (DBL-OS) were compared. The DBLSC-CO2 samples had significantly higher (pSC-CO2 than the organic solvent method. SDS-PAGE analysis demonstrated that the CBL and DBLSC-CO2 had protein bands of a similar intensity and area, whereas DBL-OS appeared extremely poor bands or no bands due to the degradation of proteins, particularly in the 50 to 75 kDa and 20 to 25 kDa molecular weight ranges. In addition, DBLSC-CO2 was shown to have superior functional properties in terms of total soluble content, water and oil absorption, and foaming and emulsification properties. Therefore, SC-CO2 treatment offers a nutritionally and environmentally friendly approach for the removal of lipid from high protein food sources. In addition, SC-CO2 may be a better substitute of traditional organic solvent extraction for producing more stable and high quality foods with high-protein, fat-free, and low calorie contents. PMID:28316468

Alginate Microencapsulation for Oral Immunisation of Finfish: Release Characteristics, Ex Vivo Intestinal Uptake and In Vivo Administration in Atlantic Salmon, Salmo salar L.

Science.gov (United States)

Ghosh, Bikramjit; Nowak, Barbara F; Bridle, Andrew R

2015-12-01

This study examined the feasibility of alginate microcapsules manufactured using a low-impact technology and reagents to protect orally delivered immunogens for use as immunoprophylactics for fish. Physical characteristics and protein release kinetics of the microcapsules were examined at different pH and temperature levels using a microencapsulated model protein, bovine serum albumin (BSA). Impact of the microencapsulation process on contents was determined by analysing change in bioactivity of microencapsulated lysozyme. Feasibility of the method for oral immunoprophylaxis of finfish was assessed using FITC-labelled microcapsules. These were applied to distal intestinal explants of Atlantic salmon (Salmo salar) to investigate uptake ex vivo. Systemic distribution of microcapsules was investigated by oral administration of FITC-labelled microcapsules to Atlantic salmon fry by incorporating into feed. The microcapsules produced were structurally robust and retained surface integrity, with a modal size distribution of 250-750 nm and a tendency to aggregate. Entrapment efficiency of microencapsulation was 51.2 % for BSA and 43.2 % in the case of lysozyme. Microcapsules demonstrated controlled release of protein, which increased with increasing pH or temperature, and the process had no significant negative effect on bioactivity of lysozyme. Uptake of fluorescent-labelled microcapsules was clearly demonstrated by intestinal explants over a 24-h period. Evidence of microcapsules was found in the intestine, spleen, kidney and liver of fry following oral administration. Amenability of the microcapsules to intestinal uptake and distribution reinforced the strong potential for use of this microencapsulation method in oral immunoprophylaxis of finfish using sensitive immunogenic substances.

Automated Segmentation of in Vivo and Ex Vivo Mouse Brain Magnetic Resonance Images

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Alize E.H. Scheenstra

2009-01-01

Full Text Available Segmentation of magnetic resonance imaging (MRI data is required for many applications, such as the comparison of different structures or time points, and for annotation purposes. Currently, the gold standard for automated image segmentation is nonlinear atlas-based segmentation. However, these methods are either not sufficient or highly time consuming for mouse brains, owing to the low signal to noise ratio and low contrast between structures compared with other applications. We present a novel generic approach to reduce processing time for segmentation of various structures of mouse brains, in vivo and ex vivo. The segmentation consists of a rough affine registration to a template followed by a clustering approach to refine the rough segmentation near the edges. Compared with manual segmentations, the presented segmentation method has an average kappa index of 0.7 for 7 of 12 structures in in vivo MRI and 11 of 12 structures in ex vivo MRI. Furthermore, we found that these results were equal to the performance of a nonlinear segmentation method, but with the advantage of being 8 times faster. The presented automatic segmentation method is quick and intuitive and can be used for image registration, volume quantification of structures, and annotation.

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

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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.

The use of ex vivo human skin tissue for genotoxicity testing

Energy Technology Data Exchange (ETDEWEB)

Reus, Astrid A.; Usta, Mustafa [TNO Triskelion BV, Utrechtseweg 48, 3704 HE, Zeist (Netherlands); Krul, Cyrille A.M., E-mail: cyrille.krul@tno.nl [TNO, Utrechtseweg 48, 3704 HE Zeist (Netherlands)

2012-06-01

As a result of the chemical legislation concerning the registration, evaluation, authorization and restriction of chemicals (REACH), and the Seventh Amendment to the Cosmetics Directive, which prohibits animal testing in Europe for cosmetics, alternative methods for safety evaluation of chemicals are urgently needed. Current in vitro genotoxicity assays are not sufficiently predictive for the in vivo situation, resulting in an unacceptably high number of misleading positives. For many chemicals and ingredients of personal care products the skin is the first site of contact, but there are no in vitro genotoxicity assays available in the skin for additional evaluation of positive or equivocal responses observed in regulatory in vitro genotoxicity assays. In the present study ex vivo human skin tissue obtained from surgery was used for genotoxicity evaluation of chemicals by using the comet assay. Fresh ex vivo human skin tissue was cultured in an air–liquid interface and topically exposed to 20 chemicals, including true positive, misleading positive and true negative genotoxins. Based on the results obtained in the present study, the sensitivity, specificity and accuracy of the ex vivo skin comet assay to predict in vivo genotoxicity were 89%, 90% and 89%, respectively. Donor and experimental variability were mainly reflected in the magnitude of the response and not the difference between the presence and absence of a genotoxic response. The present study indicates that human skin obtained from surgery is a promising and robust model for safety evaluation of chemicals that are in direct contact with the skin. -- Highlights: ► We use human skin obtained from surgery for genotoxicity evaluation of chemicals. ► We use the comet assay as parameter for genotoxicity in ex vivo human skin. ► Sensitivity, specificity and accuracy to predict in vivo genotoxins are determined. ► Sensitivity, specificity and accuracy are 89%, 90% and 90%, respectively. ► The method

The use of ex vivo human skin tissue for genotoxicity testing

International Nuclear Information System (INIS)

Reus, Astrid A.; Usta, Mustafa; Krul, Cyrille A.M.

2012-01-01

As a result of the chemical legislation concerning the registration, evaluation, authorization and restriction of chemicals (REACH), and the Seventh Amendment to the Cosmetics Directive, which prohibits animal testing in Europe for cosmetics, alternative methods for safety evaluation of chemicals are urgently needed. Current in vitro genotoxicity assays are not sufficiently predictive for the in vivo situation, resulting in an unacceptably high number of misleading positives. For many chemicals and ingredients of personal care products the skin is the first site of contact, but there are no in vitro genotoxicity assays available in the skin for additional evaluation of positive or equivocal responses observed in regulatory in vitro genotoxicity assays. In the present study ex vivo human skin tissue obtained from surgery was used for genotoxicity evaluation of chemicals by using the comet assay. Fresh ex vivo human skin tissue was cultured in an air–liquid interface and topically exposed to 20 chemicals, including true positive, misleading positive and true negative genotoxins. Based on the results obtained in the present study, the sensitivity, specificity and accuracy of the ex vivo skin comet assay to predict in vivo genotoxicity were 89%, 90% and 89%, respectively. Donor and experimental variability were mainly reflected in the magnitude of the response and not the difference between the presence and absence of a genotoxic response. The present study indicates that human skin obtained from surgery is a promising and robust model for safety evaluation of chemicals that are in direct contact with the skin. -- Highlights: ► We use human skin obtained from surgery for genotoxicity evaluation of chemicals. ► We use the comet assay as parameter for genotoxicity in ex vivo human skin. ► Sensitivity, specificity and accuracy to predict in vivo genotoxins are determined. ► Sensitivity, specificity and accuracy are 89%, 90% and 90%, respectively. ► The method

Relationship between polychlorinated biphenyl 126 treatment and cytochrome P4501A activity in chickens, as measured by in vivo caffeine and ex vivo ethoxyresorufin metabolism

Energy Technology Data Exchange (ETDEWEB)

Feyk, L.A.; Giesy, J.P.; Lambert, G.H.

1999-09-01

Cytochrome P4501A (CYPIA) activity is often used as a biomarker of exposure of wildlife to polyhalogenated diaromatic hydrocarbons (PHDHs) and is usually measured ex vivo in liver tissue. A caffeine breath test with radiolabeled substrate ({sup 14}C-CBT) has been developed to measure in vivo avian CYPIA activity. Research goals were to develop stable isotope methods ({sup 13}C-CBT), determine dose-response relationships between caffeine N-demethylation (CNDM) and PHDH exposure, and assess the relative utility of the CBT and ex vivo ethoxyresorufin-O-deethylase (EROD) assay. The {sup 13}C-CBT methods were developed with 20 chickens (Gallus domesticus). Chickens received three intraperitoneal injections of 0, 1, 5, or 50 {micro}g 3,3{prime},4,4{prime},5-pentachlorobiphenyl (PCB 126)/kg body weight, and CNDM was quantified by measurement of {sup 13}CO{sub 2}/{sup 12}CO{sub 2} in expired breath. The {sup 13}C-CBT was not as sensitive or specific as the EROD assay as an indicator of PHDH exposure and effect in birds. Constitutive CNDM of great interindividual variability was observed, and the magnitude of induction was greater for EROD activity than for CNDM (approximately 1,000- and 2-fold, respectively). Variability associated with baseline {sup 13}CO{sub 2}/{sup 12}CO{sub 2} ratios in expired breath reduced the sensitivity of the {sup 13}C-CBT method.

Single nucleotide polymorphism discovery in bovine liver using RNA-seq technology.

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Chandra Shekhar Pareek

Full Text Available RNA-seq is a useful next-generation sequencing (NGS technology that has been widely used to understand mammalian transcriptome architecture and function. In this study, a breed-specific RNA-seq experiment was utilized to detect putative single nucleotide polymorphisms (SNPs in liver tissue of young bulls of the Polish Red, Polish Holstein-Friesian (HF and Hereford breeds, and to understand the genomic variation in the three cattle breeds that may reflect differences in production traits.The RNA-seq experiment on bovine liver produced 107,114,4072 raw paired-end reads, with an average of approximately 60 million paired-end reads per library. Breed-wise, a total of 345.06, 290.04 and 436.03 million paired-end reads were obtained from the Polish Red, Polish HF, and Hereford breeds, respectively. Burrows-Wheeler Aligner (BWA read alignments showed that 81.35%, 82.81% and 84.21% of the mapped sequencing reads were properly paired to the Polish Red, Polish HF, and Hereford breeds, respectively. This study identified 5,641,401 SNPs and insertion and deletion (indel positions expressed in the bovine liver with an average of 313,411 SNPs and indel per young bull. Following the removal of the indel mutations, a total of 195,3804, 152,7120 and 205,3184 raw SNPs expressed in bovine liver were identified for the Polish Red, Polish HF, and Hereford breeds, respectively. Breed-wise, three highly reliable breed-specific SNP-databases (SNP-dbs with 31,562, 24,945 and 28,194 SNP records were constructed for the Polish Red, Polish HF, and Hereford breeds, respectively. Using a combination of stringent parameters of a minimum depth of ≥10 mapping reads that support the polymorphic nucleotide base and 100% SNP ratio, 4,368, 3,780 and 3,800 SNP records were detected in the Polish Red, Polish HF, and Hereford breeds, respectively. The SNP detections using RNA-seq data were successfully validated by kompetitive allele-specific PCR (KASPTM SNP genotyping assay. The

Single nucleotide polymorphism discovery in bovine liver using RNA-seq technology.

Science.gov (United States)

Pareek, Chandra Shekhar; Błaszczyk, Paweł; Dziuba, Piotr; Czarnik, Urszula; Fraser, Leyland; Sobiech, Przemysław; Pierzchała, Mariusz; Feng, Yaping; Kadarmideen, Haja N; Kumar, Dibyendu

2017-01-01

RNA-seq is a useful next-generation sequencing (NGS) technology that has been widely used to understand mammalian transcriptome architecture and function. In this study, a breed-specific RNA-seq experiment was utilized to detect putative single nucleotide polymorphisms (SNPs) in liver tissue of young bulls of the Polish Red, Polish Holstein-Friesian (HF) and Hereford breeds, and to understand the genomic variation in the three cattle breeds that may reflect differences in production traits. The RNA-seq experiment on bovine liver produced 107,114,4072 raw paired-end reads, with an average of approximately 60 million paired-end reads per library. Breed-wise, a total of 345.06, 290.04 and 436.03 million paired-end reads were obtained from the Polish Red, Polish HF, and Hereford breeds, respectively. Burrows-Wheeler Aligner (BWA) read alignments showed that 81.35%, 82.81% and 84.21% of the mapped sequencing reads were properly paired to the Polish Red, Polish HF, and Hereford breeds, respectively. This study identified 5,641,401 SNPs and insertion and deletion (indel) positions expressed in the bovine liver with an average of 313,411 SNPs and indel per young bull. Following the removal of the indel mutations, a total of 195,3804, 152,7120 and 205,3184 raw SNPs expressed in bovine liver were identified for the Polish Red, Polish HF, and Hereford breeds, respectively. Breed-wise, three highly reliable breed-specific SNP-databases (SNP-dbs) with 31,562, 24,945 and 28,194 SNP records were constructed for the Polish Red, Polish HF, and Hereford breeds, respectively. Using a combination of stringent parameters of a minimum depth of ≥10 mapping reads that support the polymorphic nucleotide base and 100% SNP ratio, 4,368, 3,780 and 3,800 SNP records were detected in the Polish Red, Polish HF, and Hereford breeds, respectively. The SNP detections using RNA-seq data were successfully validated by kompetitive allele-specific PCR (KASPTM) SNP genotyping assay. The comprehensive

Physiological and Molecular Effects of in vivo and ex vivo Mild Skin Barrier Disruption.

Science.gov (United States)

Pfannes, Eva K B; Weiss, Lina; Hadam, Sabrina; Gonnet, Jessica; Combardière, Béhazine; Blume-Peytavi, Ulrike; Vogt, Annika

2018-01-01

The success of topically applied treatments on skin relies on the efficacy of skin penetration. In order to increase particle or product penetration, mild skin barrier disruption methods can be used. We previously described cyanoacrylate skin surface stripping as an efficient method to open hair follicles, enhance particle penetration, and activate Langerhans cells. We conducted ex vivo and in vivo measurements on human skin to characterize the biological effect and quantify barrier disruption-related inflammation on a molecular level. Despite the known immunostimulatory effects, this barrier disruption and hair follicle opening method was well accepted and did not result in lasting changes of skin physiological parameters, cytokine production, or clinical side effects. Only in ex vivo human skin did we find a discrete increase in IP-10, TGF-β, IL-8, and GM-CSF mRNA. The data underline the safety profile of this method and demonstrate that the procedure per se does not cause substantial inflammation or skin damage, which is also of interest when applied to non-invasive sampling of biomarkers in clinical trials. © 2018 S. Karger AG, Basel.

Imaging of prostate cancer: a platform for 3D co-registration of in-vivo MRI ex-vivo MRI and pathology

Science.gov (United States)

Orczyk, Clément; Mikheev, Artem; Rosenkrantz, Andrew; Melamed, Jonathan; Taneja, Samir S.; Rusinek, Henry

2012-02-01

Objectives: Multi-parametric MRI is emerging as a promising method for prostate cancer diagnosis. prognosis and treatment planning. However, the localization of in-vivo detected lesions and pathologic sites of cancer remains a significant challenge. To overcome this limitation we have developed and tested a system for co-registration of in-vivo MRI, ex-vivo MRI and histology. Materials and Methods: Three men diagnosed with localized prostate cancer (ages 54-72, PSA levels 5.1-7.7 ng/ml) were prospectively enrolled in this study. All patients underwent 3T multi-parametric MRI that included T2W, DCEMRI, and DWI prior to robotic-assisted prostatectomy. Ex-vivo multi-parametric MRI was performed on fresh prostate specimen. Excised prostates were then sliced at regular intervals and photographed both before and after fixation. Slices were perpendicular to the main axis of the posterior capsule, i.e., along the direction of the rectal wall. Guided by the location of the urethra, 2D digital images were assembled into 3D models. Cancer foci, extra-capsular extensions and zonal margins were delineated by the pathologist and included in 3D histology data. A locally-developed software was applied to register in-vivo, ex-vivo and histology using an over-determined set of anatomical landmarks placed in anterior fibro-muscular stroma, central. transition and peripheral zones. The mean root square distance across corresponding control points was used to assess co-registration error. Results: Two specimens were pT3a and one pT2b (negative margin) at pathology. The software successfully fused invivo MRI. ex-vivo MRI fresh specimen and histology using appropriate (rigid and affine) transformation models with mean square error of 1.59 mm. Coregistration accuracy was confirmed by multi-modality viewing using operator-guided variable transparency. Conclusion: The method enables successful co-registration of pre-operative MRI, ex-vivo MRI and pathology and it provides initial evidence

Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos

International Nuclear Information System (INIS)

Souza, D.K. de; Salles, L.P.; Rosa e Silva, A.A.M.

2015-01-01

Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract

Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos

Directory of Open Access Journals (Sweden)

D.K. de Souza

2015-03-01

Full Text Available Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract.

Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos

Energy Technology Data Exchange (ETDEWEB)

Souza, D.K. de [Laboratório de Biotecnologia da Saúde, Faculdade de Medicina, Universidade de Brasília, Brasília, DF (Brazil); Faculdade da Ceilândia, Universidade de Brasília, Brasília, DF (Brazil); Salles, L.P. [Laboratório de Biotecnologia da Saúde, Faculdade de Medicina, Universidade de Brasília, Brasília, DF (Brazil); Departamento de Biologia Molecular, Instituto de Biologia, Universidade de Brasília, Brasília, DF (Brazil); Rosa e Silva, A.A.M. [Laboratório de Biotecnologia da Saúde, Faculdade de Medicina, Universidade de Brasília, Brasília, DF (Brazil)

2015-01-23

Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract.

Descemet Membrane Thickening as a Sign for the Diagnosis of Corneal Graft Rejection: An Ex Vivo Study.

Science.gov (United States)

VanDenBerg, Ryan; Diakonis, Vasilios F; Bozung, Alison; Gameiro, Gustavo Rosa; Fischer, Oliver; El Dakkak, Ahmed; Ulloa-Padilla, Jan Paul; Anagnostopoulos, Apostolos; Dubovy, Sander; Abou Shousha, Mohamed

2017-12-01

To disclose, using an ex vivo study, the histopathological mechanism behind in vivo thickening of the endothelium/Descemet membrane complex (En/DM) observed in rejected corneal grafts (RCGs). Descemet membrane (DM), endothelium, and retrocorneal membranes make up the total En/DM thickness. These layers are not differentiable by high-definition optical coherence tomography; therefore, the source of thickening is unclear from an in vivo perspective. A retrospective ex vivo study (from September 2015 to December 2015) was conducted to measure the thicknesses of DM, endothelium, and retrocorneal membrane in 54 corneal specimens (31 RCGs and 23 controls) using light microscopy. Controls were globes with posterior melanoma without corneal involvement. There were 54 corneas examined ex vivo with mean age 58.1 ± 12.2 in controls and 51.7 ± 27.9 years in RCGs. The ex vivo study uncovered the histopathological mechanism of En/DM thickening to be secondary to significant thickening (P < 0.001) of DM (6.5 ± 2.4 μm) in RCGs compared with controls (3.9 ± 1.5 μm). Our ex vivo study shows that DM is responsible for thickening of the En/DM in RCGs observed in vivo by high-definition optical coherence tomography and not the endothelium or retrocorneal membrane.

Sorbitol increases muscle glucose uptake ex vivo and inhibits intestinal glucose absorption ex vivo and in normal and type 2 diabetic rats.

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Chukwuma, Chika Ifeanyi; Islam, Md Shahidul

2017-04-01

Previous studies have suggested that sorbitol, a known polyol sweetener, possesses glycemic control potentials. However, the effect of sorbitol on intestinal glucose absorption and muscle glucose uptake still remains elusive. The present study investigated the effects of sorbitol on intestinal glucose absorption and muscle glucose uptake as possible anti-hyperglycemic or glycemic control potentials using ex vivo and in vivo experimental models. Sorbitol (2.5% to 20%) inhibited glucose absorption in isolated rat jejuna (IC 50 = 14.6% ± 4.6%) and increased glucose uptake in isolated rat psoas muscle with (GU 50 = 3.5% ± 1.6%) or without insulin (GU 50 = 7.0% ± 0.5%) in a concentration-dependent manner. Furthermore, sorbitol significantly delayed gastric emptying, accelerated digesta transit, inhibited intestinal glucose absorption, and reduced blood glucose increase in both normoglycemic and type 2 diabetic rats after 1 h of coingestion with glucose. Data of this study suggest that sorbitol exhibited anti-hyperglycemic potentials, possibly via increasing muscle glucose uptake ex vivo and reducing intestinal glucose absorption in normal and type 2 diabetic rats. Hence, sorbitol may be further investigated as a possible anti-hyperglycemic sweetener.

Large-Animal Biventricular Working Heart Perfusion System with Low Priming Volume-Comparison between in vivo and ex vivo Cardiac Function.

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Abicht, Jan-Michael; Mayr, Tanja Axinja Jelena; Jauch, Judith; Guethoff, Sonja; Buchholz, Stefan; Reichart, Bruno; Bauer, Andreas

2018-01-01

Existing large-animal, ex vivo, cardiac perfusion models are restricted in their ability to establish an ischemia/reperfusion condition as seen in cardiac surgery or transplantation. Other working heart systems only challenge one ventricle or require a substantially larger priming volume. We describe a novel biventricular cardiac perfusion system with reduced priming volume. Juvenile pig hearts were cardiopleged, explanted, and reperfused ex vivo after 150 minutes of cold ischemia. Autologous whole blood was used as perfusate (minimal priming volume 350 mL). After 15 minutes of Langendorff perfusion (LM), the system was switched into a biventricular working mode (WM) and studied for 3 hours. During reperfusion, complete unloading of both ventricles and constant-pressure coronary perfusion was achieved. During working mode perfusion, the preload and afterload pressure of both ventricles was controlled within the targeted physiologic range. Functional parameters such as left ventricular work index were reduced in ex vivo working mode (in vivo: 787 ± 186 vs. 1 h WM 498 ± 66 mm Hg·mL/g·min; p  hours while functional and blood parameters are easily accessible. Moreover, because of the minimal priming volume, the novel ex vivo cardiac perfusion circuit allows for autologous perfusion, using the limited amount of blood available from the organ donating animal. Georg Thieme Verlag KG Stuttgart · New York.

Liver cell-targeted delivery of therapeutic molecules.

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Kang, Jeong-Hun; Toita, Riki; Murata, Masaharu

2016-01-01

The liver is the largest internal organ in mammals and is involved in metabolism, detoxification, synthesis of proteins and lipids, secretion of cytokines and growth factors and immune/inflammatory responses. Hepatitis, alcoholic or non-alcoholic liver disease, hepatocellular carcinoma, hepatic veno-occlusive disease, and liver fibrosis and cirrhosis are the most common liver diseases. Safe and efficient delivery of therapeutic molecules (drugs, genes or proteins) into the liver is very important to increase the clinical efficacy of these molecules and to reduce their side effects in other organs. Several liver cell-targeted delivery systems have been developed and tested in vivo or ex vivo/in vitro. In this review, we discuss the literature concerning liver cell-targeted delivery systems, with a particular emphasis on the results of in vivo studies.

Stratum corneum damage and ex vivo porcine skin water absorption - a pilot study

DEFF Research Database (Denmark)

Duch Lynggaard, C; Bang Knudsen, D; Jemec, G B E

2009-01-01

A simple ex vivo screening technique would be of interest for mass screening of substances for potential barrier disruptive qualities. Ex vivo water absorption as a marker of skin barrier integrity was studied on pig ear skin. Skin water absorption was quantified by weighing and weight changes were...... found to reflect prehydration barrier damage. It is suggested that this simple model may be elaborated to provide a rapid, economical screening tool for potential skin irritants....

Celiac Disease-Specific TG2-Targeted Autoantibodies Inhibit Angiogenesis Ex Vivo and In Vivo in Mice by Interfering with Endothelial Cell Dynamics.

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Suvi Kalliokoski

Full Text Available A characteristic feature of celiac disease is the presence of circulating autoantibodies targeted against transglutaminase 2 (TG2, reputed to have a function in angiogenesis. In this study we investigated whether TG2-specific autoantibodies derived from celiac patients inhibit angiogenesis in both ex vivo and in vivo models and sought to clarify the mechanism behind this phenomenon. We used the ex vivo murine aorta-ring and the in vivo mouse matrigel-plug assays to address aforementioned issues. We found angiogenesis to be impaired as a result of celiac disease antibody supplementation in both systems. Our results also showed the dynamics of endothelial cells was affected in the presence of celiac antibodies. In the in vivo angiogenesis assays, the vessels formed were able to transport blood despite impairment of functionality after treatment with celiac autoantibodies, as revealed by positron emission tomography. We conclude that celiac autoantibodies inhibit angiogenesis ex vivo and in vivo and impair vascular functionality. Our data suggest that the anti-angiogenic mechanism of the celiac disease-specific autoantibodies involves extracellular TG2 and inhibited endothelial cell mobility.

Celiac Disease–Specific TG2-Targeted Autoantibodies Inhibit Angiogenesis Ex Vivo and In Vivo in Mice by Interfering with Endothelial Cell Dynamics

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Kalliokoski, Suvi; Sulic, Ana-Marija; Korponay-Szabó, Ilma R.; Szondy, Zsuzsa; Frias, Rafael; Perez, Mileidys Alea; Martucciello, Stefania; Roivainen, Anne; Pelliniemi, Lauri J.; Esposito, Carla; Griffin, Martin; Sblattero, Daniele; Mäki, Markku; Kaukinen, Katri; Lindfors, Katri; Caja, Sergio

2013-01-01

A characteristic feature of celiac disease is the presence of circulating autoantibodies targeted against transglutaminase 2 (TG2), reputed to have a function in angiogenesis. In this study we investigated whether TG2-specific autoantibodies derived from celiac patients inhibit angiogenesis in both ex vivo and in vivo models and sought to clarify the mechanism behind this phenomenon. We used the ex vivo murine aorta-ring and the in vivo mouse matrigel-plug assays to address aforementioned issues. We found angiogenesis to be impaired as a result of celiac disease antibody supplementation in both systems. Our results also showed the dynamics of endothelial cells was affected in the presence of celiac antibodies. In the in vivo angiogenesis assays, the vessels formed were able to transport blood despite impairment of functionality after treatment with celiac autoantibodies, as revealed by positron emission tomography. We conclude that celiac autoantibodies inhibit angiogenesis ex vivo and in vivo and impair vascular functionality. Our data suggest that the anti-angiogenic mechanism of the celiac disease-specific autoantibodies involves extracellular TG2 and inhibited endothelial cell mobility. PMID:23824706

Magnetic resonance imaging of trabecular and cortical bone in mice: comparison of high resolution in vivo and ex vivo MR images with corresponding histology

International Nuclear Information System (INIS)

Weber, Michael H.; Sharp, Jonathan C.; Latta, Peter; Sramek, Milos; Hassard, H. Thomas; Orr, F. William

2005-01-01

Measurements of bone morphometry and remodeling have been shown to reflect bone strength and can be used to diagnose degenerative bone disease. In this study, in vivo and ex vivo magnetic resonance imaging (MRI) techniques to assess trabecular and cortical bone properties have been compared to each other and to histology as a novel means for the quantification of bone. Femurs of C57Bl/6 mice were examined both in vivo and ex vivo on an 11.7 T MRI scanner, followed by histologic processing and morphometry. A thresholding analysis technique was applied to the MRI images to generate contour lines and to delineate the boundaries between bone and marrow. Using MRI, an optimal correlation with histology was obtained with an in vivo longitudinal sectioned short echo time gradient-echo versus an in vivo long echo time spin-echo sequence or an ex vivo pulse sequence. Gradient-echo images were acquired with a maximum in-plane resolution of 35 μm. Our results demonstrated that in both the in vivo and ex vivo data sets, the percent area of marrow increases and percent area of trabecular bone and cortical bone thickness decreases moving from the epiphyseal growth plate to the diaphysis. These changes, observed with MRI, correlate with the histological data. Investigations using in vivo MRI gradient-echo sequences consistently gave the best correlation with histology. Our quantitative evaluation using both ex vivo and in vivo MRI was found to be an effective means to visualize non-invasively the normal variation in trabecular and cortical bone as compared to a histological 'gold standard' The experiments validated in vivo MRI as a potential high resolution technique for investigating both soft tissue, such as marrow, and bone without radiation exposure

Ex Vivo Liver Experiment of Hydrochloric Acid-Infused and Saline-Infused Monopolar Radiofrequency Ablation: Better Outcomes in Temperature, Energy, and Coagulation

Energy Technology Data Exchange (ETDEWEB)

Jiang, Xiong-ying; Gu, Yang-kui; Huang, Jin-hua, E-mail: huangjh@sysucc.org.cn; Gao, Fei; Zou, Ru-hai; Zhang, Tian-qi [Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, State Key Laboratory of Oncology in South China (China)

2016-04-15

ObjectiveTo compare temperature, energy, and coagulation between hydrochloric acid-infused radiofrequency ablation (HAIRFA) and normal saline-infused radiofrequency ablation (NSIRFA) in ex vivo porcine liver model.Materials and Methods30 fresh porcine livers were excised in 60 lesions, 30 with HAIRFA and the other 30 with NSIRFA. Both modalities used monopolar perfusion electrode connected to a RF generator set at 103 °C and 30 W. In each group, ablation time was set at 10, 20, or 30 min (10 lesions from each group at each time). We compared tissue temperatures (at 0.5, 1.0, 1.5, 2.0, 2.5, and 3.0 cm away from the electrode tip), average power, deposited energy, deposited energy per coagulation volume (DEV), coagulation diameters, coagulative volume, and spherical ratio between the two groups.ResultsTemperature–time curves showed that HAIRFA provided progressively greater heating than that of NSIRFA. At 30 min, mean average power, deposited energy, coagulation volumes (113.67 vs. 12.28 cm{sup 3}) and diameters, and increasing in tissue temperature were much greater with HAIRFA (P < 0.001 for all), except DEV was lower (456 vs. 1396 J/cm{sup 3}, P < 0.001). The spherical ratio was closer to 1 with HAIRFA (1.23 vs. 1.46). Coagulation diameters, volume, and average power of HAIRFA increased significantly with longer ablation times. While with NSIRFA, these characteristics were stable till later 20 min, except the power decreased with longer ablation times.ConclusionsHAIRFA creates much larger and more spherical lesions by increasing overall energy deposition, modulating thermal conductivity, and transferring heat during ablation.

High-resolution ex vivo magnetic resonance angiography: a feasibility study on biological and medical tissues

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Boel Lene WT

2010-03-01

Full Text Available Abstract Background In biomedical sciences, ex vivo angiography is a practical mean to elucidate vascular structures three-dimensionally with simultaneous estimation of intravascular volume. The objectives of this study were to develop a magnetic resonance (MR method for ex vivo angiography and to compare the findings with computed tomography (CT. To demonstrate the usefulness of this method, examples are provided from four different tissues and species: the human placenta, a rice field eel, a porcine heart and a turtle. Results The optimal solution for ex vivo MR angiography (MRA was a compound containing gelatine (0.05 g/mL, the CT contrast agent barium sulphate (0.43 mol/L and the MR contrast agent gadoteric acid (2.5 mmol/L. It was possible to perform angiography on all specimens. We found that ex vivo MRA could only be performed on fresh tissue because formalin fixation makes the blood vessels permeable to the MR contrast agent. Conclusions Ex vivo MRA provides high-resolution images of fresh tissue and delineates fine structures that we were unable to visualise by CT. We found that MRA provided detailed information similar to or better than conventional CTA in its ability to visualize vessel configuration while avoiding interfering signals from adjacent bones. Interestingly, we found that vascular tissue becomes leaky when formalin-fixed, leading to increased permeability and extravascular leakage of MR contrast agent.

Ex vivo confocal microscopy: a new diagnostic technique for mucormycosis.

Science.gov (United States)

Leclercq, A; Cinotti, E; Labeille, B; Perrot, J L; Cambazard, F

2016-05-01

Skin-dedicated ex vivo confocal microscopy (EVCM) has so far mainly been employed to identify cutaneous tumours on freshly excised samples. We present two cases where EVCM has been used to diagnose cutaneous mucormycosis. The skin biopsies were evaluated by the skin-dedicated ex vivo confocal microscope VivaScope 2500(®) (MAVIG GmbH, Munich Germany) under both reflectance and fluorescence mode. Conventional direct optical examination on skin scraping and histological examination were later performed. Mucormycetes observed by EVCM presented as hyper-reflective elongated 20 μm in diameter structures with perpendicular ramifications. Fungi were found both under reflectance and fluorescence mode and were better visible with acridine orange under fluorescence EVCM. Conventional direct optical examination on skin scraping and histological examination found the same elongated and branching structures confirming the presence of Mucormycetes. Ex vivo confocal microscopy has both the advantages of being fast as the direct optical examination, and to be able to show the localisation of the fungi in the tissue like the histological examination. In our cases, EVCM allowed to rapidly confirm the clinical diagnosis of mucormycosis, which is essential for the treatment of this fungal infection. Further studies are needed to compare the performance of EVCM with the findings of conventional histological and mycological examinations. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Graft downsizing during ex vivo lung perfusion: case report and technical notes.

Science.gov (United States)

Nosotti, M; Rosso, L; Mendogni, P; Tosi, D; Palleschi, A; Righi, I; Froio, S; Valenza, F; Santambrogio, L

2014-09-01

Among patients with respiratory insufficiency awaiting lung transplantation, small adult patients have a lower opportunity of receiving size-matched pulmonary grafts, because of the shortage of donors, particularly those of small size. Reducing the size of an oversized graft is one of the methods to increase the donor pool; similarly, ex vivo lung perfusion is an emerging technique aimed toward the same purpose. We describe how we combined the 2 techniques (lobar transplantation plus contralateral nonanatomic graft reduction during ex vivo lung perfusion) to overcome graft shortage in a clinical case. For the 1st time, this case report demonstrates that surgical manipulation during ex vivo lung perfusion does not affect the functional improvement in a lung previously judged to be not suitable for transplantation. The 6-month follow-up results are similar to those of standard bilateral lung transplantation. Copyright © 2014 Elsevier Inc. All rights reserved.

STUDY OF INTRA TESTICULAR REGULATIONS OF SPERMATOGENESIS DIFFERENTIATION BY EX-VIVO APPROACH

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A. Adaika

2010-12-01

Full Text Available The aim of this work is to study the regulation of intratesticular during spermatogenesis ex vivo. To highlight the progress of spermatogenesis ex vivo, we developed two cell culture systems of seminiferous tubules to study the role of local factors that control the proliferation and differentiation of male germ cells. Our studies are based on two main techniques: RT-PCR and RNA extraction to examine changes in the expression of some growth factors in the culture of seminiferous tubules as the SCF, c- Kit and TGFß. The results show, using RT-PCR, that expression of SCF, c-Kit and TGFb is probably not involved in the alterations of spermatogenesis ex vivo. Indeed, their expressions are not modified during three weeks of culture, and their expressions depend on the proportion of cells where they are expressed. Our results also show that clusterin is a marker of Sertoli cells in the culture of seminiferous tubules and its expression is not altered by the presence of germ cells.

Ex Vivo (Fluorescence) Confocal Microscopy in Surgical Pathology: State of the Art.

Science.gov (United States)

Ragazzi, Moira; Longo, Caterina; Piana, Simonetta

2016-05-01

First developed in 1957, confocal microscopy is a powerful imaging tool that can be used to obtain near real-time reflected light images of untreated human tissue with nearly histologic resolution. Besides its research applications, in the last decades, confocal microscopy technology has been proposed as a useful device to improve clinical diagnosis, especially in ophthalmology, dermatology, and endomicroscopy settings, thanks to advances in instrument development. Compared with the wider use of the in vivo tissue assessment, ex vivo applications of confocal microscopy are not fully explored. A comprehensive review of the current literature was performed here, focusing on the reliable applications of ex vivo confocal microscopy in surgical pathology and on some potential evolutions of this new technique from pathologists' viewpoint.

Carlecortemcel-l: an ex vivo expanded umbilical cord blood cell graft for allogeneic transplantation.

Science.gov (United States)

Petropoulos, Demetrios; Chan, Ka Wah

2009-11-01

Success of umbilical cord blood transplantation (UCBT) is mostly affected by the cell dose infused and its application is limited by the size of the recipient. For most adults and older children it is not possible to find a single UCB unit large enough for reliable engraftment. One strategy to increase the number of progenitor cells available is ex vivo expansion of the unit. The main challenge of ex vivo expansion systems is how not to deplete the self-renewing cell population by driving them into differentiation into committed progenitors. Copper modulates basic cell functions, such as survival, proliferation, and differentiation. Reduction of cellular copper in ex vivo culture conditions enabled preferential proliferation of early progenitors and increased engraftment capabilities. The result of a Phase I study of carlecortemcel-l, a product derived from ex vivo expansion of UCB progenitors in the presence of a copper chelator and early-acting cytokines, and the study design for the current pivotal study are presented. A literature review using PubMed and the investigator's brochure from the manufacturer. Early results suggest that carlecortemcel-l infusion is safe and may be associated with favorable non-relapse mortality rates. A pivotal global study is currently being conducted to evaluate safety and efficacy of this product from centralized manufacturing facilities.

Vaginal Lactobacillus Inhibits HIV-1 Replication in Human Tissues Ex Vivo

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Rogers A. Ñahui Palomino

2017-05-01

Full Text Available Lactobacillus species, which dominate vaginal microbiota of healthy reproductive-age women, lower the risks of sexually transmitted infections, including the risk of human immunodeficiency virus (HIV acquisition. The exact mechanisms of this protection remain to be understood. Here, we investigated these mechanisms in the context of human cervico-vaginal and lymphoid tissues ex vivo. We found that all six Lactobacillus strains tested in these systems significantly suppressed HIV type-1 (HIV-1 infection. We identified at least three factors that mediated this suppression: (i Acidification of the medium. The pH of the undiluted medium conditioned by lactobacilli was between 3.8 and 4.6. Acidification of the culture medium with hydrochloric acid (HCl to this pH in control experiments was sufficient to abrogate HIV-1 replication. However, the pH of the Lactobacillus-conditioned medium (CM diluted fivefold, which reached ∼6.9, was also suppressive for HIV-1 infection, while in control experiments HIV-1 infection was not abrogated when the pH of the medium was brought to 6.9 through the use of HCl. This suggested the existence of other factors responsible for HIV-1 inhibition by lactobacilli. (ii Lactic acid. There was a correlation between the concentration of lactic acid in the Lactobacillus-CM and its ability to suppress HIV-1 infection in human tissues ex vivo. Addition of lactic acid isomers D and L to tissue culture medium at the concentration that corresponded to their amount released by lactobacilli resulted in HIV-1 inhibition. Isomer L was produced in higher quantities than isomer D and was mostly responsible for HIV-1 inhibition. These results indicate that lactic acid, in particular its L-isomer, inhibits HIV-1 independently of lowering of the pH. (iii Virucidal effect. Incubation of HIV-1 in Lactobacillus-CM significantly suppressed viral infectivity for human tissues ex vivo. Finally, lactobacilli adsorb HIV-1, serving as a sink

Measuring in-vivo and in-situ ex-vivo the 3D deformation of the lamina cribrosa microstructure under elevated intraocular pressure

Science.gov (United States)

Wei, Junchao; Yang, Bin; Voorhees, Andrew P.; Tran, Huong; Brazile, Bryn; Wang, Bo; Schuman, Joel; Smith, Matthew A.; Wollstein, Gadi; Sigal, Ian A.

2018-02-01

Elevated intraocular pressure (IOP) deforms the lamina cribrosa (LC), a structure within the optic nerve head (ONH) in the back of the eye. Evidence suggests that these deformations trigger events that eventually cause irreversible blindness, and have therefore been studied in-vivo using optical coherence tomography (OCT), and ex-vivo using OCT and a diversity of techniques. To the best of our knowledge, there have been no in-situ ex-vivo studies of LC mechanics. Our goal was two-fold: to introduce a technique for measuring 3D LC deformations from OCT, and to determine whether deformations of the LC induced by elevated IOP differ between in-vivo and in-situ ex-vivo conditions. A healthy adult rhesus macaque monkey was anesthetized and IOP was controlled by inserting a 27- gauge needle into the anterior chamber of the eye. Spectral domain OCT was used to obtain volumetric scans of the ONH at normal and elevated IOPs. To improve the visibility of the LC microstructure the scans were first processed using a novel denoising technique. Zero-normalized cross-correlation was used to find paired corresponding locations between images. For each location pair, the components of the 3D strain tensor were determined using non-rigid image registration. A mild IOP elevation from 10 to 15mmHg caused LC effective strains as large as 3%, and about 50% larger in-vivo than in-situ ex-vivo. The deformations were highly heterogeneous, with substantial 3D components, suggesting that accurate measurement of LC microstructure deformation requires high-resolution volumes. This technique will help improve understanding of LC biomechanics and how IOP contributes to glaucoma.

Clinical application of sentinel lymph node mapping in colon cancer: in vivo vs. ex vivo techniques.

Science.gov (United States)

Oh, Seung Yeop; Kim, Do Yoon; Kim, Young Bae; Suh, Kwang Wook

2014-09-01

Clinical usefulness of sentinel lymph node (SLN) mapping in colorectal cancer remains controversial. The aim of this study is to evaluate the accuracy of the SLN mapping technique using serial sectioning, and to compare the results between ex vivo and in vivo techniques. From February 2011 to October 2012, 34 colon cancer patients underwent SLN mapping during surgical resection. Eleven patients were analyzed with the in vivo method, and 23 patients with the ex vivo method. Patient characteristics and results of SLN mapping were evaluated. The SLN mapping was performed in 34 patients. Mean age was 67.3 years (range, 44-81 years). Primary tumors were located in the following sites: 13 in the right colon (38.2%) and 21 in the left colon (61.8%). SLN mapping was performed successfully in 88.2% of the patients. There was no significant difference in the identification rate between the two methods (90.9% vs. 87.0%, P = 1.000). Both the mapping methods showed a low sensitivity and high rate of skip metastasis. This study showed that SLN evaluation using serial sectioning could not predict the nodal status with clinically acceptable accuracy despite the high detection rate.

The correlation of in vivo and ex vivo tissue dielectric properties to validate electromagnetic breast imaging: initial clinical experience

International Nuclear Information System (INIS)

Halter, Ryan J; Zhou, Tian; Meaney, Paul M; Hartov, Alex; Barth, Richard J Jr; Rosenkranz, Kari M; Wells, Wendy A; Kogel, Christine A; Borsic, Andrea; Rizzo, Elizabeth J; Paulsen, Keith D

2009-01-01

Electromagnetic (EM) breast imaging provides low-cost, safe and potentially a more specific modality for cancer detection than conventional imaging systems. A primary difficulty in validating these EM imaging modalities is that the true dielectric property values of the particular breast being imaged are not readily available on an individual subject basis. Here, we describe our initial experience in seeking to correlate tomographic EM imaging studies with discrete point spectroscopy measurements of the dielectric properties of breast tissue. The protocol we have developed involves measurement of in vivo tissue properties during partial and full mastectomy procedures in the operating room (OR) followed by ex vivo tissue property recordings in the same locations in the excised tissue specimens in the pathology laboratory immediately after resection. We have successfully applied all of the elements of this validation protocol in a series of six women with cancer diagnoses. Conductivity and permittivity gauged from ex vivo samples over the frequency range 100 Hz–8.5 GHz are found to be similar to those reported in the literature. A decrease in both conductivity and permittivity is observed when these properties are gauged from ex vivo samples instead of in vivo. We present these results in addition to a case study demonstrating how discrete point spectroscopy measurements of the tissue can be correlated and used to validate EM imaging studies

DNA damage in lens epithelium of cataract patients in vivo and ex vivo.

Science.gov (United States)

Øsnes-Ringen, Oyvind; Azqueta, Amaia O; Moe, Morten C; Zetterström, Charlotta; Røger, Magnus; Nicolaissen, Bjørn; Collins, Andrew R

2013-11-01

DNA damage has been described in the human cataractous lens epithelium, and oxidative stress generated by UV radiation and endogenous metabolic processes has been suggested to play a significant role in the pathogenesis of cataract. In this study, the aim was to explore the quality and relative quantity of DNA damage in lens epithelium of cataract patients in vivo and after incubation in a cell culture system. Capsulotomy specimens were analysed, before and after 1 week of ex vivo cultivation, using the comet assay to measure DNA strand breaks, oxidized purine and pyrimidine bases and UV-induced cyclobutane pyrimidine dimers. DNA strand breaks were barely detectable, oxidized pyrimidines and pyrimidine dimers were present at low levels, whereas there was a relatively high level of oxidized purines, which further increased after cultivation. The observed levels of oxidized purines in cataractous lens epithelium may support a theory consistent with light damage and oxidative stress as mediators of molecular damage to the human lens epithelium. Damage commonly associated with UV-B irradiation was relatively low. The levels of oxidized purines increased further in a commonly used culture system. This is of interest considering the importance and versatility of ex vivo systems in studies exploring the pathogenesis of cataract. © 2012 The Authors. Acta Ophthalmologica © 2012 Acta Ophthalmologica Scandinavica Foundation.

Combined in vivo and ex vivo analysis of mesh mechanics in a porcine hernia model.

Science.gov (United States)

Kahan, Lindsey G; Lake, Spencer P; McAllister, Jared M; Tan, Wen Hui; Yu, Jennifer; Thompson, Dominic; Brunt, L Michael; Blatnik, Jeffrey A

2018-02-01

Hernia meshes exhibit variability in mechanical properties, and their mechanical match to tissue has not been comprehensively studied. We used an innovative imaging model of in vivo strain tracking and ex vivo mechanical analysis to assess effects of mesh properties on repaired abdominal walls in a porcine model. We hypothesized that meshes with dissimilar mechanical properties compared to native tissue would alter abdominal wall mechanics more than better-matched meshes. Seven mini-pigs underwent ventral hernia creation and subsequent open repair with one of two heavyweight polypropylene meshes. Following mesh implantation with attached radio-opaque beads, fluoroscopic images were taken at insufflation pressures from 5 to 30 mmHg on postoperative days 0, 7, and 28. At 28 days, animals were euthanized and ex vivo mechanical testing performed on full-thickness samples across repaired abdominal walls. Testing was conducted on 13 mini-pig controls, and on meshes separately. Stiffness and anisotropy (the ratio of stiffness in the transverse versus craniocaudal directions) were assessed. 3D reconstructions of repaired abdominal walls showed stretch patterns. As pressure increased, both meshes expanded, with no differences between groups. Over time, meshes contracted 17.65% (Mesh A) and 0.12% (Mesh B; p = 0.06). Mesh mechanics showed that Mesh A deviated from anisotropic native tissue more than Mesh B. Compared to native tissue, Mesh A was stiffer both transversely and craniocaudally. Explanted repaired abdominal walls of both treatment groups were stiffer than native tissue. Repaired tissue became less anisotropic over time, as mesh properties prevailed over native abdominal wall properties. This technique assessed 3D stretch at the mesh level in vivo in a porcine model. While the abdominal wall expanded, mesh-ingrown areas contracted, potentially indicating stresses at mesh edges. Ex vivo mechanics demonstrate that repaired tissue adopts mesh properties, suggesting

Valproic acid modulates platelet and coagulation function ex vivo

DEFF Research Database (Denmark)

Bambakidis, Ted; Dekker, Simone E; Halaweish, Ihab

2017-01-01

of coagulopathy, it remains unknown whether this is a direct effect of the drug, or the establishment of an overall prosurvival phenotype. We thus conducted an ex-vivo experiment to determine if VPA has an effect on coagulation and platelet function. Ten swine were subjected to traumatic brain injury (TBI...

Design and development of vitamin C-encapsulated proliposome with improved in-vitro and ex-vivo antioxidant efficacy.

Science.gov (United States)

Parhizkar, Elahehnaz; Rashedinia, Marzieh; Karimi, Maryam; Alipour, Shohreh

2018-06-06

Vitamin C, as an antioxidant additive in pharmaceutical and food products, is susceptible to environmental conditions, and new design strategies are needed to enhance its stability. The aim of this study is to prepare vitamin C proliposome using film deposition on the carrier by applying different factors, and optimise the characteristics of the obtained powder using the design expert ® software. The optimised formulation demonstrated acceptable flowability with 20% vitamin C loading. This formulation released about 90% vitamin C within 2 h and showed higher (1.7-fold) in-vitro antioxidant activity. Ex-vivo antioxidant activity was 1.9 and 1.6 times higher in brain and liver cells, respectively. A 27% reduction in malondialdehyde (MDA) level of liver cell was obtained comparing free vitamin C. Therefore, this study results suggest that the vitamin C-encapsulated proliposome powder might be an appropriate carrier for oral drug delivery of vitamin C with better antioxidant efficacy.

LukMF′ is the major secreted leukocidin of bovine Staphylococcus aureus and is produced in vivo during bovine mastitis

Science.gov (United States)

Vrieling, Manouk; Boerhout, Eveline M.; van Wigcheren, Glenn F.; Koymans, Kirsten J.; Mols-Vorstermans, Tanja G.; de Haas, Carla J. C.; Aerts, Piet C.; Daemen, Ineke J. J. M.; van Kessel, Kok P. M.; Koets, Ad P.; Rutten, Victor P. M. G.; Nuijten, Piet J.M.; van Strijp, Jos A. G.; Benedictus, Lindert

2016-01-01

Staphylococcus aureus is a major human and animal pathogen and a common cause of mastitis in cattle. S. aureus secretes several leukocidins that target bovine neutrophils, crucial effector cells in the defence against bacterial pathogens. In this study, we investigated the role of staphylococcal leukocidins in the pathogenesis of bovine S. aureus disease. We show that LukAB, in contrast to the γ-hemolysins, LukED, and LukMF′, was unable to kill bovine neutrophils, and identified CXCR2 as a bovine receptor for HlgAB and LukED. Furthermore, we assessed functional leukocidin secretion by bovine mastitis isolates and observed that, although leukocidin production was strain dependent, LukMF′ was most abundantly secreted and the major toxin killing bovine neutrophils. To determine the role of LukMF′ in bovine mastitis, cattle were challenged with high (S1444) or intermediate (S1449, S1463) LukMF′-producing isolates. Only animals infected with S1444 developed severe clinical symptoms. Importantly, LukM was produced in vivo during the course of infection and levels in milk were associated with the severity of mastitis. Altogether, these findings underline the importance of LukMF′ as a virulence factor and support the development of therapeutic approaches targeting LukMF′ to control S. aureus mastitis in cattle. PMID:27886237

Analysis of antigen-specific B-cell memory directly ex vivo.

Science.gov (United States)

McHeyzer-Williams, Louise J; McHeyzer-Williams, Michael G

2004-01-01

Helper T-cell-regulated B-cell memory develops in response to initial antigen priming as a cellular product of the germinal center (GC) reaction. On antigen recall, memory response precursors expand rapidly with exaggerated differentiation into plasma cells to produce the high-titer, high-affinity antibody(Ab) that typifies the memory B-cell response in vivo. We have devised a high-resolution flow cytometric strategy to quantify the emergence and maintenance of antigen-specific memory B cells directly ex vivo. Extended cell surface phenotype establishes a level of cellular diversity not previously appreciated for the memory B-cell compartment. Using an "exclusion transfer" strategy, we ascertain the capacity of two distinct memory B-cell populations to transfer antigen-specific memory into naive adoptive hosts. Finally, we sequence expressed messenger ribonucleic acid (mRNA) from single cells within the population to estimate the level of somatic hypermutation as the best molecular indicator of B-cell memory. In this chapter, we describe the methods used in each of these four sections that serve to provide high-resolution quantification of antigen-specific B-cell memory responses directly ex vivo.

Nigrosome-1 on Susceptibility Weighted Imaging to Differentiate Parkinson’s Disease From Atypical Parkinsonism: An In Vivo and Ex Vivo Pilot Study

International Nuclear Information System (INIS)

Meijer, Frederick J.A.; Steens, Stefan C.; Rumund, Anouke van; Cappellen van Walsum, Anne-Marie van; Küsters, Benno; Esselink, Rianne A.J.; Verbeek, Marcel M.; Bloem, Bastiaan R.; Goraj, Bożena

2016-01-01

Previous case-control studies have suggested that the absence of a swallow-tail appearance in the substantia nigra on high-resolution SWI, representing nigrosome-1, has high accuracy to identify Parkinson’s disease (PD). The first goal of our study was to evaluate nigrosome-1 ex vivo using optimized high-resolution susceptibility sensitive MRI. Our second goal was to evaluate its diagnostic value in vivo using a clinical 3T SWI sequence to differentiate between PD and atypical parkinsonism (AP) in a cohort of patients with early-stage parkinsonism. Case-control pilot study to evaluate nigrosome-1 ex vivo (2 PD, 2 controls), using high-resolution susceptibility sensitive sequences at 11.7 T MRI. Next, evaluation of nigrosome-1 in vivo using a clinical 3 T SWI sequence in a prospective cohort of 60 patients with early-stage parkinsonism (39 PD, 21 AP). Moreover, 12 control subjects were scanned. The bilateral substantia nigra was evaluated by two neuroradiologists for the presence, absence or indecisive presence of nigrosome-1. The discriminative power was evaluated by Receiver-Operating Characteristic. We identified nigrosome-1 in ex vivo control subjects. Nigrosome-1 was not identified in the ex vivo PD cases. In our prospective clinical cohort study, the AUC for the swallow-tail sign to discriminate between PD and AP was 0.56 (0.41–0.71) for reader 1 and 0.68 (0.55–0.82) for reader 2. The diagnostic accuracy of the swallow-tail sign was marginal to discriminate between PD and AP using our clinical 3 T SWI sequence

PEDIATRIC LIVER TRANSPLANTATION WITH EX-SITU LIVER TRANSECTION AND THE APPLICATION OF THE HUMAN FIBRINOGEN AND THROMBIN SPONGE IN THE WOUND AREA.

Science.gov (United States)

Vicentine, Fernando Pompeu Piza; Gonzalez, Adriano Miziara; Azevedo, Ramiro Anthero de; Benini, Barbara Burza; Linhares, Marcelo Moura; Lopes-Filho, Gaspar de Jesus; Martins, Jose Luiz; Salzedas-Netto, Alcides Augusto

2016-01-01

Surgical strategy to increase the number of liver transplants in the pediatric population is the ex-situ liver transection (reduction or split). However, it is associated with complications such as hemorrhage and leaks. The human fibrinogen and thrombin sponge is useful for improving hemostasis in liver surgery. Compare pediatric liver transplants with ex-situ liver transection (reduction or split) with or without the human fibrinogen and thrombin sponge. Was performed a prospective analysis of 21 patients submitted to liver transplantation with ex-situ liver transection with the application of the human fibrinogen and thrombin sponge in the wound area (group A) and retrospective analysis of 59 patients without the sponge (group B). The characteristics of recipients and donors were similar. There were fewer reoperations due to bleeding in the wound area in group A (14.2%) compared to group B (41.7%, p=0.029). There was no difference in relation to the biliary leak (group A: 17.6%, group B: 5.1%, p=0.14). There was a lower number of reoperations due to bleeding of the wound area of ​​the hepatic graft when the human fibrinogen and thrombin sponge were used. Estratégia cirúrgica para aumentar o número de transplantes hepáticos na população pediátrica é a transecção hepática ex-situ (redução ou split). No entanto, ela está associada com complicações, tais como hemorragia e fístulas. A esponja de fibrinogênio e trombina humana é útil para melhorar a hemostasia nas operações hepáticas. Comparar transplantes hepáticos pediátricos com transecção hepática ex-situ (redução ou split) com ou sem a esponja de fibrinogênio e trombina humana. Foi realizada análise prospectiva de 21 pacientes submetidos ao transplante de fígado com transecção hepática ex-situ com a aplicação da esponja de fibrinogênio e trombina humana na área cruenta (grupo A) e análise retrospectiva de 59 pacientes sem a esponja (grupo B). As características dos

In and ex vivo breast disease study by Raman spectroscopy

DEFF Research Database (Denmark)

Raniero, L.; Canevari, R. A.; Ramalho, L. N. Z.

2011-01-01

ex vivo measurements gave the highest specificity and sensitivity: 96 and 97%, respectively, as well as a largest percentage for correct discrimination: 94%. Now that the important bands have been experimentally determined in this and other works, what remains is for first principles molecular...

3D thoracoscopic ultrasound volume measurement validation in an ex vivo and in vivo porcine model of lung tumours

International Nuclear Information System (INIS)

Hornblower, V D M; Yu, E; Fenster, A; Battista, J J; Malthaner, R A

2007-01-01

The purpose of this study was to validate the accuracy and reliability of volume measurements obtained using three-dimensional (3D) thoracoscopic ultrasound (US) imaging. Artificial 'tumours' were created by injecting a liquid agar mixture into spherical moulds of known volume. Once solidified, the 'tumours' were implanted into the lung tissue in both a porcine lung sample ex vivo and a surgical porcine model in vivo. 3D US images were created by mechanically rotating the thoracoscopic ultrasound probe about its long axis while the transducer was maintained in close contact with the tissue. Volume measurements were made by one observer using the ultrasound images and a manual-radial segmentation technique and these were compared with the known volumes of the agar. In vitro measurements had average accuracy and precision of 4.76% and 1.77%, respectively; in vivo measurements had average accuracy and precision of 8.18% and 1.75%, respectively. The 3D thoracoscopic ultrasound can be used to accurately and reproducibly measure 'tumour' volumes both in vivo and ex vivo

3D thoracoscopic ultrasound volume measurement validation in an ex vivo and in vivo porcine model of lung tumours

Energy Technology Data Exchange (ETDEWEB)

Hornblower, V D M [Canadian Surgical Technologies and Advanced Robotics, London, Ontario (Canada); Yu, E [Canadian Surgical Technologies and Advanced Robotics, London, Ontario (Canada); Fenster, A [Canadian Surgical Technologies and Advanced Robotics, London, Ontario (Canada); Battista, J J [Canadian Surgical Technologies and Advanced Robotics, London, Ontario (Canada); Malthaner, R A [Canadian Surgical Technologies and Advanced Robotics, London, Ontario (Canada)

2007-01-07

The purpose of this study was to validate the accuracy and reliability of volume measurements obtained using three-dimensional (3D) thoracoscopic ultrasound (US) imaging. Artificial 'tumours' were created by injecting a liquid agar mixture into spherical moulds of known volume. Once solidified, the 'tumours' were implanted into the lung tissue in both a porcine lung sample ex vivo and a surgical porcine model in vivo. 3D US images were created by mechanically rotating the thoracoscopic ultrasound probe about its long axis while the transducer was maintained in close contact with the tissue. Volume measurements were made by one observer using the ultrasound images and a manual-radial segmentation technique and these were compared with the known volumes of the agar. In vitro measurements had average accuracy and precision of 4.76% and 1.77%, respectively; in vivo measurements had average accuracy and precision of 8.18% and 1.75%, respectively. The 3D thoracoscopic ultrasound can be used to accurately and reproducibly measure 'tumour' volumes both in vivo and ex vivo.

Some studies on liver and rumen flukes of bovines in Sri Lanka

African Journals Online (AJOL)

nkpc001

Keywords: liver and rumen flukes, bovines, influence of climate, Sri Lanka. INTRODUCTION .... they were given two changes of absolute alcohol, each longing ..... inland and where fresh water ditches, streams and large water ... India. Res. ull. Panjab Univ., New series, 18: 369-337. sen J, Perry B. 1994. The epidemiology,.

Robust methods to create ex vivo minimum deformation atlases for brain mapping.

Science.gov (United States)

Janke, Andrew L; Ullmann, Jeremy F P

2015-02-01

Highly detailed ex vivo 3D atlases of average structure are of critical importance to neuroscience and its current push to understanding the global microstructure of the brain. Multiple single slice histology sections can no longer provide sufficient detail of inter-slice microstructure and lack out of plane resolution. Two ex vivo methods have emerged that can create such detailed models. High-field micro MRI with the addition of contrast media has allowed intact whole brain microstructure imaging with an isotropic resolution of 15 μm in mouse. Blockface imaging has similarly evolved to a point where it is now possible to image an entire brain in a rigorous fashion with an out of plane resolution of 10 μm. Despite the destruction of the tissue as part of this process it allows a reconstructed model that is free from cutting artifacts. Both of these methods have been utilised to create minimum deformation atlases that are representative of the respective populations. The MDA atlases allow us unprecedented insight into the commonality and differences in microstructure in cortical structures in specific taxa. In this paper we provide an overview of how to create such MDA models from ex vivo data. Copyright © 2015 Elsevier Inc. All rights reserved.

Ex vivo culture of patient tissue & examination of gene delivery.

LENUS (Irish Health Repository)

Rajendran, Simon

2012-01-31

This video describes the use of patient tissue as an ex vivo model for the study of gene delivery. Fresh patient tissue obtained at the time of surgery is sliced and maintained in culture. The ex vivo model system allows for the physical delivery of genes into intact patient tissue and gene expression is analysed by bioluminescence imaging using the IVIS detection system. The bioluminescent detection system demonstrates rapid and accurate quantification of gene expression within individual slices without the need for tissue sacrifice. This slice tissue culture system may be used in a variety of tissue types including normal and malignant tissue and allows us to study the effects of the heterogeneous nature of intact tissue and the high degree of variability between individual patients. This model system could be used in certain situations as an alternative to animal models and as a complementary preclinical mode prior to entering clinical trial.

Evaluation of the In Vivo and Ex Vivo Binding of Novel BC1 Cannabinoid Receptor Radiotracers

Energy Technology Data Exchange (ETDEWEB)

Miller, A.; Gatley, J.; Gifford, A.

2002-01-01

The primary active ingredient of marijuana, 9-tetrahydrocannabinol, exerts its psychoactive effects by binding to cannabinoid CB1 receptors. These receptors are found throughout the brain with high concentrations in the hippocampus and cerebellum. The current study was conducted to evaluate the binding of a newly developed putative cannabinoid antagonist, AM630, and a classical cannabinoid 8-tetrahydrocannabinol as potential PET and/or SPECT imaging agents for brain CB1 receptors. For both of these ligands in vivo and ex vivo studies in mice were conducted. AM630 showed good overall brain uptake (as measure by %IA/g) and a moderately rapid clearance from the brain with a half-clearance time of approximately 30 minutes. However, AM630 did not show selective binding to CB1 cannabinoid receptors. Ex vivo autoradiography supported the lack of selective binding seen in the in vivo study. Similar to AM630, 8-tetrahydrocanibol also failed to show selective binding to CB1 receptor rich brain areas. The 8-tetrahydrocanibol showed moderate overall brain uptake and relatively slow brain clearance as compared to AM630. Further studies were done with AM2233, a cannabinoid ligand with a similar structure as AM630. These studies were done to develop an ex vivo binding assay to quantify the displacement of [131I]AM2233 binding by other ligands in Swiss-Webster and CB1 receptor knockout mice. By developing this assay we hoped to determine the identity of an unknown binding site for AM2233 present in the hippocampus of CB1 knockout mice. Using an approach based on incubation of brain slices prepared from mice given intravenous [131I]AM2233 in either the presence or absence of AM2233 (unlabelled) it was possible to demonstrate a significant AM2233-displacable binding in the Swiss-Webster mice. Future studies will determine if this assay is appropriate for identifying the unknown binding site for AM2233 in the CB1 knockout mice.

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

Science.gov (United States)

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 culture...

[A new technique for ensuring negative surgical margins during partial nephrectomy: the ex vivo ultrasound control].

Science.gov (United States)

Desmonts, A; Tillou, X; Le Gal, S; Secco, M; Orczyk, C; Bensadoun, H; Doerfler, A

2013-10-01

To evaluate the feasibility and the efficiency of intraoperative ex vivo ultrasound of resection margins in patients undergoing partial nephrectomy by urologist. Patients undergoing partial nephrectomy from July 2010 to November 2012 for T1-T2 renal tumors were included in analysis. Tumor margin status was immediately determined by ex vivo ultrasound done by the surgeon himself. Results were compared with margin status on definitive pathological evaluation. A total of 26 men and 15 women with a median age of 61 (30-82) years old were included in analysis. Intraoperative ex vivo ultrasound revealed negative surgical margins in 38 cases and positive margins in two. Final pathological results revealed negative margins in all except one case. Ultrasound sensitivity and specificity were 100% and 97%, respectively. Mean ultrasound duration was 1minute±1. Mean tumor and margin sizes were 3.4±1.8cm and 2.38±1.76mm, respectively. Intraoperative ex vivo ultrasound of resection margins in patients undergoing partial nephrectomy by a urologist seemed to be feasible, efficient and easy. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Correlation between the dielectric properties and biological activities of human ex vivo hepatic tissue

International Nuclear Information System (INIS)

Wang, Hang; You, Fusheng; Fu, Feng; Dong, Xiuzhen; Shi, Xuetao; He, Yong; Yang, Min; Yan, Qingguo

2015-01-01

Dielectric properties are vital biophysical features of biological tissues, and biological activity is an index to ascertain the active state of tissues. This study investigated the potential correlation between the dielectric properties and biological activities of human hepatic tissue with prolonged ex vivo time through correlation and regression analyses. The dielectric properties of 26 cases of normal human hepatic tissue at 10 Hz to 100 MHz were measured from 15 min after isolation to 24 h at 37 °C with 90% humidity. Cell morphologies, including nucleus area (NA) and alteration rate of intercellular area (ICAR), were analyzed as indicators of biological activities. Conductivity, complex resistivity, and NA exhibited opposing changes 1 h after isolation. Relative permittivity and ex vivo time were not closely correlated (p > 0.05). The dielectric properties measured at low frequencies (i.e. <1 MHz) were more sensitive than those measured at high frequencies in reflecting the biological activity of ex vivo tissue. Highly significant correlations were found between conductivity, resistivity and the ex vivo time (p < 0.05) as well as conductivity and the cell morphology (p < 0.05). The findings indicated that establishing the correlation between the dielectric properties and biological activities of human hepatic tissue is of great significance for promoting the role of dielectric properties in biological science, particularly in human biology. (paper)

Intraoperative diagnosis of nonpigmented nail tumours with ex vivo fluorescence confocal microscopy: 10 cases.

Science.gov (United States)

Debarbieux, S; Gaspar, R; Depaepe, L; Dalle, S; Balme, B; Thomas, L

2015-04-01

Ex vivo fluorescence confocal microscopy (FCM) permits real-time imaging of freshly excised skin tissues. Its usefulness as a time-sparing alternative to frozen sections in Mohs surgery of basal cell carcinoma is well documented. The purpose of this study was to describe the ex vivo FCM features of a series of benign and malignant nonpigmented tumours of the nail unit, and to correlate them with conventional histopathology. Nail apparatus tumours from 10 patients were imaged during surgical exploration using ex vivo FCM after immersion in acridine orange. Confocal mosaics of the freshly performed biopsies were evaluated in real time and retrospectively compared with haematoxylin and eosin sections. Our series included two invasive epithelial tumours (Group 1), four in situ or minimally invasive squamous cell carcinomas (SCC) (Group 2), three benign epithelial tumours (Group 3) and one nodular melanoma (Group 4). The correlation was excellent for malignant epithelial tumours exhibiting marked cytological and architectural atypias (Bowen disease, invasive SCC and onycholemmal carcinoma). Onychomatricomas exhibited a very peculiar aspect with densely cellular papillae. The correlation was less favourable for minimally invasive well-differentiated SCCs with slight cytological atypias. The correlation was poor for our case of amelanotic invasive subungual melanoma. Ex vivo FCM could be a useful tool to shorten management of nonpigmented nail tumours: in the case of a malignant tumour, it could indeed lead to performing wide excision during the same surgical procedure and possibly assessing the surgical margins. © 2014 British Association of Dermatologists.

Ex vivo generation of a functional and regenerative wound epithelium from axolotl (Ambystoma mexicanum) skin.

Science.gov (United States)

Ferris, Donald R; Satoh, Akira; Mandefro, Berhan; Cummings, Gillian M; Gardiner, David M; Rugg, Elizabeth L

2010-10-01

Urodele amphibians (salamanders) are unique among adult vertebrates in their ability to regenerate structurally complete and fully functional limbs. Regeneration is a stepwise process that requires interactions between keratinocytes, nerves and fibroblasts. The formation of a wound epithelium covering the amputation site is an early and necessary event in the process but the molecular mechanisms that underlie the role of the wound epithelium in regeneration remain unclear. We have developed an ex vivo model that recapitulates many features of in vivo wound healing. The model comprises a circular explant of axolotl (Ambystoma mexicanum) limb skin with a central circular, full thickness wound. Re-epithelialization of the wound area is rapid (typically <11 h) and is dependent on metalloproteinase activity. The ex vivo wound epithelium is viable, responds to neuronal signals and is able to participate in ectopic blastema formation and limb regeneration. This ex vivo model provides a reproducible and tractable system in which to study the cellular and molecular events that underlie wound healing and regeneration. © 2010 The Authors. Journal compilation © 2010 Japanese Society of Developmental Biologists.

Evaluation of an ex vivo murine local lymph node assay: multiple endpoint comparison.

Science.gov (United States)

Piccotti, Joseph R; Knight, Stephanie A; Gillhouse, Kimberly; Lagattuta, Mark S; Bleavins, Michael R

2006-01-01

The local lymph node assay (LLNA) is used to assess the skin sensitization potential of chemicals. In the standard assay, mice are treated topically on the dorsum of both ears with test substance for 3 days. Following 2 days of rest, the initiation of the hypersensitivity response is evaluated by injecting (3)H-thymidine into a tail vein, and then measuring the levels of radioisotope incorporated into the DNA of lymph node cells draining the ears. In the current study, BALB/c mice were treated with the contact sensitizers hexylcinnamic aldehyde (HCA) and oxazolone, and the nonsensitizer methyl salicylate. The proliferative response of lymph node cells was evaluated in an ex vivo assay, in which isolated cells were cultured in vitro with (3)H-thymidine. Treatment of mice with HCA at 5-50% resulted in concentration-related increases in (3)H-thymidine incorporation, with stimulation indices ranging from 3 to 14. Low animal-to-animal variability was seen in three replicate assays testing HCA at 25%. As anticipated, the proliferative response induced by the potent sensitizer oxazolone at 0.25% was greater than HCA at all concentrations tested. Stimulation indices of 1.5 and 3 were seen in two independent experiments with methyl salicylate. These equivocal findings were likely due to the irritancy properties of the compound. Importantly, measuring ex vivo (3)H-thymidine incorporation was more sensitive than evaluating lymph node weight and cellularity, and in vitro bromodeoxyuridine incorporation. Furthermore, the results of the ex vivo LLNA were comparable to the standard assay. This study provided evidence that supports the use of an ex vivo LLNA for hazard assessment of contact hypersensitivity. Copyright 2006 John Wiley & Sons, Ltd.

Ex vivo lung perfusion with adenosine A2A receptor agonist allows prolonged cold preservation of lungs donated after cardiac death.

Science.gov (United States)

Wagner, Cynthia E; Pope, Nicolas H; Charles, Eric J; Huerter, Mary E; Sharma, Ashish K; Salmon, Morgan D; Carter, Benjamin T; Stoler, Mark H; Lau, Christine L; Laubach, Victor E; Kron, Irving L

2016-02-01

Ex vivo lung perfusion has been successful in the assessment of marginal donor lungs, including donation after cardiac death (DCD) donor lungs. Ex vivo lung perfusion also represents a unique platform for targeted drug delivery. We sought to determine whether ischemia-reperfusion injury would be decreased after transplantation of DCD donor lungs subjected to prolonged cold preservation and treated with an adenosine A2A receptor agonist during ex vivo lung perfusion. Porcine DCD donor lungs were preserved at 4°C for 12 hours and underwent ex vivo lung perfusion for 4 hours. Left lungs were then transplanted and reperfused for 4 hours. Three groups (n = 4/group) were randomized according to treatment with the adenosine A2A receptor agonist ATL-1223 or the dimethyl sulfoxide vehicle: Infusion of dimethyl sulfoxide during ex vivo lung perfusion and reperfusion (DMSO), infusion of ATL-1223 during ex vivo lung perfusion and dimethyl sulfoxide during reperfusion (ATL-E), and infusion of ATL-1223 during ex vivo lung perfusion and reperfusion (ATL-E/R). Final Pao2/Fio2 ratios (arterial oxygen partial pressure/fraction of inspired oxygen) were determined from samples obtained from the left superior and inferior pulmonary veins. Final Pao2/Fio2 ratios in the ATL-E/R group (430.1 ± 26.4 mm Hg) were similar to final Pao2/Fio2 ratios in the ATL-E group (413.6 ± 18.8 mm Hg), but both treated groups had significantly higher final Pao2/Fio2 ratios compared with the dimethyl sulfoxide group (84.8 ± 17.7 mm Hg). Low oxygenation gradients during ex vivo lung perfusion did not preclude superior oxygenation capacity during reperfusion. After prolonged cold preservation, treatment of DCD donor lungs with an adenosine A2A receptor agonist during ex vivo lung perfusion enabled Pao2/Fio2 ratios greater than 400 mm Hg after transplantation in a preclinical porcine model. Pulmonary function during ex vivo lung perfusion was not predictive of outcomes after transplantation. Copyright �

Formulation Optimization and Ex Vivo and In Vivo Evaluation of Celecoxib Microemulsion-Based Gel for Transdermal Delivery.

Science.gov (United States)

Cao, Mengyuan; Ren, Lili; Chen, Guoguang

2017-08-01

Celecoxib (CXB) is a poorly aqueous solubility sulfonamide non-steroidal anti-inflammatory drug (NSAID). Hence, the formulation of CXB was selected for solubilization and bioavailability. To find out suitable formulation for microemulsion, the solubility of CXB in triacetin (oil phase), Tween 80 (surfactant), and Transcutol-P (co-surfactant) was screened respectively and optimized by using orthogonal experimental design. The Km value and concentration of oil, S mix , and water were confirmed by pseudo-ternary phase diagram studies and central composite design. One percent carbopol 934 was added to form CXB microemulsion-based gel. The final formulation was evaluated for its appearance, pH, viscosity, stability, drug content determination, globule size, and zeta potential. Its ex vivo drug permeation and the in vivo pharmacokinetic was investigated. Further research was performed to ensure the safety and validity by skin irritation study and in vivo anti-inflammatory activity study. Ex vivo permeation study in mice was designed to compare permeation and transdermal ability between microemulsion formulation and conventional gel. The results revealed that optimized microemulsion-based gel gained higher permeation based on smaller globule size and high drug loading of microemulsion. Transdermal ability was also greatly improved. Bioavailability was compared to market Celebrex® by the in vivo pharmacokinetic study in rabbits. The results indicated that CXB microemulsion-based gel had better bioavailability than Celebrex®.

PGE2 suppresses NK activity in vivo directly and through adrenal hormones: Effects that cannot be reflected by ex-vivo assessment of NK cytotoxicity

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Meron, G.; Tishler, Y.; Shaashua, L.; Rosenne, E.; Levi, B.; Melamed, R.; Gotlieb, N.; Matzner, P.; Sorski, L.; Ben-Eliyahu, S.

2013-01-01

Surgery can suppress in vivo levels of NK cell cytotoxicity (NKCC) through various mechanisms, including catecholamine-, glucocorticoid (CORT)-, and prostaglandin (PG)-mediated responses. However, PGs are synthesized locally following tissue damage, driving proinflammatory and CORT responses, while their systemic levels are often unaffected. Thus, we herein studied the role of adrenal factors in mediating in vivo effects of PGs on NKCC, using adrenalectomized and sham-operated F344 rats subjected to surgery or PGE2 administration. In vivo and ex-vivo approaches were employed, based on intravenous administration of the NK-sensitive MADB106 tumor line, and based on ex-vivo assessment of YAC-1 and MADB106 target-line lysis. Additionally, in vitro studies assessed the kinetics of the impact of epinephrine, CORT, and PGE2 on NKCC. The results indicated that suppression of NKCC by epinephrine and PGE2 are short lasting, and cannot be evident when these compounds are removed from the in vitro assay milieu, or in the context of ex-vivo assessment of NKCC. In contrast, the effects of CORT are long-lasting and are reflected in both conditions even after its removal. Marginating-pulmonary NKCC was less susceptible to suppression than circulating NKCC, when tested against the xenogeneic YAC-1 target line, but not against the syngeneic MADB106 line, which seems to involve different cytotoxicity mechanisms. Overall, these findings indicate that elevated systemic PG levels can directly suppress NKCC in vivo, but following laparotomy adrenal hormones mediate most of the effects of endogenously-released PGs. Additionally, the ex-vivo approach seems limited in reflecting the short-lasting NK-suppressive effects of catecholamines and PGs. PMID:23153554

PGE2 suppresses NK activity in vivo directly and through adrenal hormones: effects that cannot be reflected by ex vivo assessment of NK cytotoxicity.

Science.gov (United States)

Meron, G; Tishler, Y; Shaashua, L; Rosenne, E; Levi, B; Melamed, R; Gotlieb, N; Matzner, P; Sorski, L; Ben-Eliyahu, S

2013-02-01

Surgery can suppress in vivo levels of NK cell cytotoxicity (NKCC) through various mechanisms, including catecholamine-, glucocorticoid (CORT)-, and prostaglandin (PG)-mediated responses. However, PGs are synthesized locally following tissue damage, driving proinflammatory and CORT responses, while their systemic levels are often unaffected. Thus, we herein studied the role of adrenal factors in mediating in vivo effects of PGs on NKCC, using adrenalectomized and sham-operated F344 rats subjected to surgery or PGE(2) administration. In vivo and ex vivo approaches were employed, based on intravenous administration of the NK-sensitive MADB106 tumor line, and based on ex vivo assessment of YAC-1 and MADB106 target-line lysis. Additionally, in vitro studies assessed the kinetics of the impact of epinephrine, CORT, and PGE(2) on NKCC. The results indicated that suppression of NKCC by epinephrine and PGE(2) are short lasting, and cannot be evident when these compounds are removed from the in vitro assay milieu, or in the context of ex vivo assessment of NKCC. In contrast, the effects of CORT are long-lasting and are reflected in both conditions even after its removal. Marginating-pulmonary NKCC was less susceptible to suppression than circulating NKCC, when tested against the xenogeneic YAC-1 target line, but not against the syngeneic MADB106 line, which seems to involve different cytotoxicity mechanisms. Overall, these findings indicate that elevated systemic PG levels can directly suppress NKCC in vivo, but following laparotomy adrenal hormones mediate most of the effects of endogenously-released PGs. Additionally, the ex vivo approach seems limited in reflecting the short-lasting NK-suppressive effects of catecholamines and PGs. Copyright © 2012 Elsevier Inc. All rights reserved.

Formulation and in Vitro, ex Vivo and in Vivo Evaluation of Elastic Liposomes for Transdermal Delivery of Ketorolac Tromethamine

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Néstor Mendoza

2011-12-01

Full Text Available The objective of the current study was to formulate ketorolac tromethamine-loaded elastic liposomes and evaluate their in vitro drug release and their ex vivo and in vivo transdermal delivery. Ketorolac tromethamine (KT, which is a potent analgesic, was formulated in elastic liposomes using Tween 80 as an edge activator. The elastic vesicles were prepared by film hydration after optimizing the sonication time and number of extrusions. The vesicles exhibited an entrapment efficiency of 73 ± 11%, vesicle size of 127.8 ± 3.4 nm and a zeta potential of −12 mV. In vitro drug release was analyzed from liposomes and an aqueous solution, using Franz diffusion cells and a cellophane dialysis membrane with molecular weight cut-off of 8000 Da. Ex vivo permeation of KT across pig ear skin was studied using a Franz diffusion cell, with phosphate buffer (pH 7.4 at 32 °C as receptor solution. An in vivo drug permeation study was conducted on healthy human volunteers using a tape-stripping technique. The in vitro results showed (i a delayed release when KT was included in elastic liposomes, compared to an aqueous solution of the drug; (ii a flux of 0.278 mg/cm2h and a lag time of about 10 h for ex vivo permeation studies, which may indicate that KT remains in the skin (with the possibility of exerting a local effect before reaching the receptor medium; (iii a good correlation between the total amount permeated, the penetration distance (both determined by tape stripping and transepidermal water loss (TEWL measured during the in vivo permeation studies. Elastic liposomes have the potential to transport the drug through the skin, keep their size and drug charge, and release the drug into deep skin layers. Therefore, elastic liposomes hold promise for the effective topical delivery of KT.

"Resuscitation" of marginal liver allografts for transplantation with machine perfusion technology.

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Graham, Jay A; Guarrera, James V

2014-08-01

As the rate of medically suitable donors remains relatively static worldwide, clinicians have looked to novel methods to meet the ever-growing demand of the liver transplant waiting lists worldwide. Accordingly, the transplant community has explored many strategies to offset this deficit. Advances in technology that target the ex vivo "preservation" period may help increase the donor pool by augmenting the utilization and improving the outcomes of marginal livers. Novel ex vivo techniques such as hypothermic, normothermic, and subnormothermic machine perfusion may be useful to "resuscitate" marginal organs by reducing ischemia/reperfusion injury. Moreover, other preservation techniques such as oxygen persufflation are explored as they may also have a role in improving function of "marginal" liver allografts. Currently, marginal livers are frequently discarded or can relegate the patient to early allograft dysfunction and primary non-function. Bench to bedside advances are rapidly emerging and hold promise for expanding liver transplantation access and improving outcomes. Copyright © 2014 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Paediatric laparoscopic hernia repair: Ex vivo skills in the reduced training era

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Chris Parsons

2013-01-01

Full Text Available Introduction: Changes to surgical working hours have resulted in shorter training times and fewer learning opportunities. Tools that develop surgical skills ex-vivo are of particular interest in this era. Laparoscopic skills are regarded as essential by many for modern paediatric surgery practice. Several generic skills models have been reported and validated. However, there is limited evidence regarding the role of procedure specific models. Here, a laparoscopic paediatric hernia repair model is trialled with surgical trainees and their competence compared with consultant colleagues. Patients and Methods: An ex-vivo paediatric inguinal hernia repair model was devised. Surgical trainees from 5 specialist centres were recruited and performed multiple standardised repairs. Results: 23 trainees performed 192 repairs. Experts performed 10 repairs for comparison. Trainees were timed performing the repair and their accuracy measured. With repeated attempts trainee′s timings and accuracy improved until by the 10 th repair they were no different from benchmark consultant scores. Conclusion: A simple, procedure specific ex-vivo training model has been evaluated for laparoscopic hernia training in paediatric surgery. The results suggest improvements in competence with repetition. Trainee and benchmark consultant scores are no different by the 10 th trainee attempt. We conclude that this model may have a valuable role in the training and assessment of future paediatric surgeons.

Flat panel computed tomography of human ex vivo heart and bone specimens: initial experience

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Nikolaou, Konstantin; Becker, Christoph R.; Reiser, Maximilian F. [Ludwig-Maximilians-University, Department of Clinical Radiology, Munich (Germany); Flohr, Thomas; Stierstorfer, Karl [CT Division, Siemens Medical Solutions, Forchheim (Germany)

2005-02-01

The aim of this technical investigation was the detailed description of a prototype flat panel detector computed tomography system (FPCT) and its initial evaluation in an ex vivo setting. The prototype FPCT scanner consists of a conventional radiographic flat panel detector, mounted on a multi-slice CT scanner gantry. Explanted human ex vivo heart and foot specimens were examined. Images were reformatted with various reconstruction algorithms and were evaluated for high-resolution anatomic information. For comparison purposes, the ex vivo specimens were also scanned with a conventional 16-detector-row CT scanner (Sensation 16, Siemens Medical Solutions, Forchheim, Germany). With the FPCT prototype used, a 1,024 x 768 resolution matrix can be obtained, resulting in an isotropic voxel size of 0.25 x 0.25 x 0.25 mm at the iso-center. Due to the high spatial resolution, very small structures such as trabecular bone or third-degree, distal branches of coronary arteries could be visualized. This first evaluation showed that flat panel detector systems can be used in a cone-beam computed tomography scanner and that very high spatial resolutions can be achieved. However, there are limitations for in vivo use due to constraints in low contrast resolution and slow scan speed. (orig.)

Detecting hepatic steatosis using ultrasound-induced thermal strain imaging: an ex vivo animal study

International Nuclear Information System (INIS)

Mahmoud, Ahmed M; Ding, Xuan; Dutta, Debaditya; Kim, Kang; Singh, Vijay P

2014-01-01

�± 0.037%). Using histology as a gold standard to classify mouse livers, US-TSI had a sensitivity and specificity of 70% and 90%, respectively. The area under the receiver operating characteristic curve was 0.775. This ex vivo study demonstrates the feasibility of using US-TSI to detect fatty livers and warrants further investigation of US-TSI as a diagnostic tool for hepatic steatosis. (paper)

Preparation of bovine muscle, bovine liver and pig kidney reference materials and the certification of the contents of nine elements of toxicological and nutritional interest

International Nuclear Information System (INIS)

Wagstaffe, P.J.; Muntau, H.

1987-01-01

The preparation of 3 meat reference materials (bovine muscle, bovine liver and pig kidney) and the steps taken to confirm their homogeneity and stability are described. Details are presented of a preliminary intercomparison and of the final collaborative exercise which led to the certification of the contents of 9 elements of toxicological and nutritional importance. Indicative values are given for the contents of a further 9 elements in these materials. (orig.)

The Response of RIF-1 Fibrosarcomas to the Vascular-Disrupting Agent ZD6126 Assessed by In Vivo and Ex Vivo1H Magnetic Resonance Spectroscopy

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Basetti Madhu

2006-07-01

Full Text Available The response of radiation-induced fibrosarcoma1 (RIF-1 tumors treated with the vascular-disrupting agent (VDA ZD6126 was assessed by in vivo and ex vivo1H magnetic resonance spectroscopy (MRS methods. Tumors treated with 200 mg/kg ZD6126 showed a significant reduction in total choline (tCho in vivo 24 hours after treatment, whereas control tumors showed a significant increase in tCho. This response was investigated further within both ex vivo unprocessed tumor tissues and tumor tissue metabolite extracts. Ex vivo high-resolution magic angle spinning (HRMAS and 1H MRS of metabolite extracts revealed a significant reduction in phosphocholine and glycerophosphocholine in biopsies of ZD6126-treated tumors, confirming in vivo tCho response. ZD6126-induced reduction in choline compounds is consistent with a reduction in cell membrane turnover associated with necrosis and cell death following disruption of the tumor vasculature. In vivo tumor tissue water diffusion and lactate measurements showed no significant changes in response to ZD6126. Spin-spin relaxation times (T2 of water and metabolites also remained unchanged. Noninvasive 1H MRS measurement of tCho in vivo provides a potential biomarker of tumor response to VDAs in RIF-1 tumors.

Spinal cord dopamine D2/D3 receptors: in vivo and ex vivo imaging in the rat using 18F/11C-fallypride

International Nuclear Information System (INIS)

Kaur, Jasmeet; Khararjian, Armen; Coleman, Robert A.; Constantinescu, Cristian C.; Pan, Min-Liang; Mukherjee, Jogeshwar

2014-01-01

Objectives: The spinal cord is known to be innervated with dopaminergic cells with catecholaminergic projections arising from the medulla and pons and dopaminergic transmission in the spinal cord is vital for sensory and motor function. Our goal was to evaluate and compare the imaging capability of dopamine D2/D3 receptors in the rat spinal cord using PET ligands 18 F-fallypride and 11 C-fallypride. Methods: Male Sprague–Dawley rats were used in all in vitro and in vivo studies. Spinal cord and brain sections were used for in vitro autoradiography and ex vivo autoradiography. For in vivo studies animals received a 18 F-fallypride scan or a 11 C-fallypride PET scan. The spinal cord and the brain were then harvested, flash-frozen and imaged ex vivo. For in vivo analysis Logan plots with cerebellum as a reference was used to evaluate binding potentials (BP). Tissue ratios were used for ex vivo analysis. Drug effects were evaluated using clozapine, haloperidol and dopamine were evaluated on spinal cord sections in vitro. Results: In vitro studies showed 18 F-fallypride binding to superficial dorsal horn (SDH), dorsal horn (DH), ventral horn (VH) and the pars centralis (PC). In the cervical section, the greatest amount of binding appeared to be in the SDH. Ex vivo studies showed approximately 6% of 18 F-fallypride in SDH compared to that observed in the striatum. In vivo analysis of both 18 F-fallypride and 11 C-fallypride in the spinal cord were comparable to that in the extrastriatal regions. Haloperidol and clozapine displaced more than 75% of the 18 F-fallypride in spinal cord sections. Conclusions: Our studies showed 18 F-fallypride and 11 C-fallypride binding in the spinal cord in vitro and in vivo. The binding pattern correlates well with the known distribution of dopamine D2/D3 receptors in the spinal cord

A high pulse repetition frequency ultrasound system for the ex vivo measurement of mechanical properties of crystalline lenses with laser-induced microbubbles interrogated by acoustic radiation force

International Nuclear Information System (INIS)

Yoon, Sangpil; Emelianov, Stanislav; Aglyamov, Salavat; Karpiouk, Andrei

2012-01-01

A high pulse repetition frequency ultrasound system for an ex vivo measurement of mechanical properties of an animal crystalline lens was developed and validated. We measured the bulk displacement of laser-induced microbubbles created at different positions within the lens using nanosecond laser pulses. An impulsive acoustic radiation force was applied to the microbubble, and spatio-temporal measurements of the microbubble displacement were assessed using a custom-made high pulse repetition frequency ultrasound system consisting of two 25 MHz focused ultrasound transducers. One of these transducers was used to emit a train of ultrasound pulses and another transducer was used to receive the ultrasound echoes reflected from the microbubble. The developed system was operating at 1 MHz pulse repetition frequency. Based on the measured motion of the microbubble, Young’s moduli of surrounding tissue were reconstructed and the values were compared with those measured using the indentation test. Measured values of Young’s moduli of four bovine lenses ranged from 2.6 ± 0.1 to 26 ± 1.4 kPa, and there was good agreement between the two methods. Therefore, our studies, utilizing the high pulse repetition frequency ultrasound system, suggest that the developed approach can be used to assess the mechanical properties of ex vivo crystalline lenses. Furthermore, the potential of the presented approach for in vivo measurements is discussed. (paper)

Effective in vivo and ex vivo gene transfer to intestinal mucosa by VSV-G-pseudotyped lentiviral vectors

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Kasahara Noriyuki

2010-05-01

Full Text Available Abstract Background Gene transfer to the gastrointestinal (GI mucosa is a therapeutic strategy which could prove particularly advantageous for treatment of various hereditary and acquired intestinal disorders, including inflammatory bowel disease (IBD, GI infections, and cancer. Methods We evaluated vesicular stomatitis virus glycoprotein envelope (VSV-G-pseudotyped lentiviral vectors (LV for efficacy of gene transfer to both murine rectosigmoid colon in vivo and human colon explants ex vivo. LV encoding beta-galactosidase (LV-β-Gal or firefly-luciferase (LV-fLuc reporter genes were administered by intrarectal instillation in mice, or applied topically for ex vivo transduction of human colorectal explant tissues from normal individuals. Macroscopic and histological evaluations were performed to assess any tissue damage or inflammation. Transduction efficiency and systemic biodistribution were evaluated by real-time quantitative PCR. LV-fLuc expression was evaluated by ex vivo bioluminescence imaging. LV-β-Gal expression and identity of transduced cell types were examined by histochemical and immunofluorescence staining. Results Imaging studies showed positive fLuc signals in murine distal colon; β-Gal-positive cells were found in both murine and human intestinal tissue. In the murine model, β-Gal-positive epithelial and lamina propria cells were found to express cytokeratin, CD45, and CD4. LV-transduced β-Gal-positive cells were also seen in human colorectal explants, consisting mainly of CD45, CD4, and CD11c-positive cells confined to the LP. Conclusions We have demonstrated the feasibility of LV-mediated gene transfer into colonic mucosa. We also identified differential patterns of mucosal gene transfer dependent on whether murine or human tissue was used. Within the limitations of the study, the LV did not appear to induce mucosal damage and were not distributed beyond the distal colon.

Adhesion of Plasmodium falciparum infected erythrocytes in ex vivo perfused placental tissue

DEFF Research Database (Denmark)

Pehrson, Caroline; Mathiesen, Line; Heno, Kristine K

2016-01-01

placental tissue. RESULTS: The ex vivo placental perfusion model was modified to study adhesion of infected erythrocytes binding to CSA, endothelial protein C receptor (EPCR) or a transgenic parasite where P. falciparum erythrocyte membrane protein 1 expression had been shut down. Infected erythrocytes......, such as binding to immunoglobulins. Furthermore, other parasite antigens have been associated with placental malaria. These findings have important implications for placental malaria vaccine design. The objective of this study was to adapt and describe a biologically relevant model of parasite adhesion in intact...... expressing VAR2CSA accumulated in perfused placental tissue whereas the EPCR binding and the transgenic parasite did not. Soluble CSA and antibodies specific against VAR2CSA inhibited binding of infected erythrocytes. CONCLUSION: The ex vivo model provides a novel way of studying receptor-ligand interactions...

Ex vivo analysis identifies effective HIV-1 latency–reversing drug combinations

Science.gov (United States)

Laird, Gregory M.; Bullen, C. Korin; Rosenbloom, Daniel I.S.; Martin, Alyssa R.; Hill, Alison L.; Durand, Christine M.; Siliciano, Janet D.; Siliciano, Robert F.

2015-01-01

Reversal of HIV-1 latency by small molecules is a potential cure strategy. This approach will likely require effective drug combinations to achieve high levels of latency reversal. Using resting CD4+ T cells (rCD4s) from infected individuals, we developed an experimental and theoretical framework to identify effective latency-reversing agent (LRA) combinations. Utilizing ex vivo assays for intracellular HIV-1 mRNA and virion production, we compared 2-drug combinations of leading candidate LRAs and identified multiple combinations that effectively reverse latency. We showed that protein kinase C agonists in combination with bromodomain inhibitor JQ1 or histone deacetylase inhibitors robustly induce HIV-1 transcription and virus production when directly compared with maximum reactivation by T cell activation. Using the Bliss independence model to quantitate combined drug effects, we demonstrated that these combinations synergize to induce HIV-1 transcription. This robust latency reversal occurred without release of proinflammatory cytokines by rCD4s. To extend the clinical utility of our findings, we applied a mathematical model that estimates in vivo changes in plasma HIV-1 RNA from ex vivo measurements of virus production. Our study reconciles diverse findings from previous studies, establishes a quantitative experimental approach to evaluate combinatorial LRA efficacy, and presents a model to predict in vivo responses to LRAs. PMID:25822022

Bovine liver sample preparation and micro-homogeneity study for Cu and Zn determination by solid sampling electrothermal atomic absorption spectrometry

International Nuclear Information System (INIS)

Nomura, Cassiana S.; Silva, Cintia S.; Nogueira, Ana R.A.; Oliveira, Pedro V.

2005-01-01

This work describes a systematic study for the bovine liver sample preparation for Cu and Zn determination by solid sampling electrothermal atomic absorption spectrometry. The main parameters investigated were sample drying, grinding process, particle size, sample size, microsample homogeneity, and their relationship with the precision and accuracy of the method. A bovine liver sample was prepared using different drying procedures: (1) freeze drying, and (2) drying in a household microwave oven followed by drying in a stove at 60 deg. C until constant mass. Ball and cryogenic mills were used for grinding. Less sensitive wavelengths for Cu (216.5 nm) and Zn (307.6 nm), and Zeeman-based three-field background correction for Cu were used to diminish the sensitivities. The pyrolysis and atomization temperatures adopted were 1000 deg. C and 2300 deg. C for Cu, and 700 deg. C and 1700 deg. C for Zn, respectively. For both elements, it was possible to calibrate the spectrometer with aqueous solutions. The use of 250 μg of W + 200 μg of Rh as permanent chemical modifier was imperative for Zn. Under these conditions, the characteristic mass and detection limit were 1.4 ng and 1.6 ng for Cu, and 2.8 ng and 1.3 ng for Zn, respectively. The results showed good agreement (95% confidence level) for homogeneity of the entire material (> 200 mg) when the sample was dried in microwave/stove and ground in a cryogenic mill. The microsample homogeneity study showed that Zn is more dependent on the sample pretreatment than Cu. The bovine liver sample prepared in microwave/stove and ground in a cryogenic mill presented results with the lowest relative standard deviation for Cu than Zn. Good accuracy and precision were observed for bovine liver masses higher than 40 μg for Cu and 30 μg for Zn. The concentrations of Cu and Zn in the prepared bovine liver sample were 223 mg kg - 1 and 128 mg kg - 1 , respectively. The relative standard deviations were lower than 6% (n = 5). The

Recovery following Thyroxine Treatment Withdrawal, but Not Propylthiouracil, Averts In Vivo and Ex Vivo Thyroxine-Provoked Cardiac Complications in Adult FVB/N Mice

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Nancy S. Saad

2017-01-01

Full Text Available Persistent cardiovascular pathology has been described in hyperthyroid patients even with effective antithyroid treatment. Here, we studied the effect of a well-known antithyroid drug, propylthiouracil (PTU; 20 mg/kg/day, on thyroxine (T4; 500 µg/kg/day-induced increase in blood pressure (BP, cardiac hypertrophy, and altered responses of the contractile myocardium both in vivo and ex vivo after 2 weeks of treatment. Furthermore, the potential recovery through 2 weeks of T4 treatment discontinuation was also investigated. PTU and T4 recovery partially reduced the T4-prompted increase in BP. Alternatively, PTU significantly improved the in vivo left ventricular (LV function with no considerable effects on cardiac hypertrophy or ex vivo right ventricular (RV contractile alterations subsequent to T4 treatment. Conversely, T4 recovery considerably enhanced the T4-provoked cardiac changes both in vivo and ex vivo. Altogether, our data is in agreement with the proposal that hyperthyroidism-induced cardiovascular pathology could persevere even with antithyroid treatments, such as PTU. However, this cannot be generalized and further investigation with different antithyroid treatments should be executed. Moreover, we reveal that recovery following experimental hyperthyroidism could potentially ameliorate cardiac function and decrease the risk for additional cardiac complications, yet, this appears to be model-dependent and should be cautiously construed.

CALCIUM-DRIVEN TRANSCRIPTION OF CARDIAC SPECIFYING GENE PROGRAM IN LIVER STEM CELLS

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We have previously shown that a cloned liver stem cell line (WB F344) acquires a cardiac phenotype when seeded in a cardiac microenvironment in vivo and ex vivo. Here we investigated the mechanisms of this transdifferentiation in early (cell, rat neonatal ventricu...

Ex vivo instability of glycated albumin: A role for autoxidative glycation.

Science.gov (United States)

Jeffs, Joshua W; Ferdosi, Shadi; Yassine, Hussein N; Borges, Chad R

2017-09-01

Ex vivo protein modifications occur within plasma and serum (P/S) samples due to prolonged exposure to the thawed state-which includes temperatures above -30 °C. Herein, the ex vivo glycation of human serum albumin from healthy and diabetic subjects was monitored in P/S samples stored for hours to months at -80 °C, -20 °C, and room temperature, as well as in samples subjected to multiple freeze-thaw cycles, incubated at different surface area-to-volume ratios or under different atmospheric compositions. A simple dilute-and-shoot method utilizing trap-and-elute LC-ESI-MS was employed to determine the relative abundances of the glycated forms of albumin-including forms of albumin bearing more than one glucose molecule. Significant increases in glycated albumin were found to occur within hours at room temperature, and within days at -20 °C. These increases continued over a period of 1-2 weeks at room temperature and over 200 days at -20 °C, ultimately resulting in a doubling of glycated albumin in both healthy and diabetic patients. It was also shown that samples stored at lower surface area-to-volume ratios or incubated under a nitrogen atmosphere experienced less rapid glucose adduction of albumin-suggesting a role for oxidative glycation in the ex vivo glycation of albumin. Copyright © 2017 Elsevier Inc. All rights reserved.

Quantification of fibronectin as a method to assess ex vivo extracellular matrix remodeling

DEFF Research Database (Denmark)

Bager, Cecilie Liv; Gudmann, N.; Willumsen, N.

2016-01-01

Altered architecture, composition and quality of the extracellular matrix (ECM) are pathological hallmarks of several inflammatory and fibro-proliferative pathological processes such as osteoarthritis (OA), rheumatoid arthritis (RA), fibrosis and cancer. One of the most important components...... of the ECM is fibronectin. Fibronectin serves as an adhesion molecule anchoring cells to the underlying basement membrane through direct interaction with integrin receptors. Fibronectin hereby modulates the properties of the ECM and affects cellular processes. Quantification of fibronectin remodeling could...... therefore be used to assess the changes in the ECM that occur during progression of fibro-proliferative pathologies. Ex vivo models are becoming state-of-the-art tools to study ECM remodeling as the cellular composition and the organization of the ECM are preserved. Ex vivo models may therefore...

Carcinoma-associated perisinusoidal laminin may signal tumour cell metastasis to the liver

DEFF Research Database (Denmark)

Wewer, U M; Albrechtsen, R

1992-01-01

using chain-specific monoclonal antibodies against B2 laminin. In an ex vivo assay, viable tumour cells (Panc-1 and clone A) were found to bind with remarkable specificity to frozen sections of liver tissue containing perisinusoidal laminin as opposed to liver tissues without laminin. We suggest...

An ex vivo human skin model for studying skin barrier repair.

Science.gov (United States)

Danso, Mogbekeloluwa O; Berkers, Tineke; Mieremet, Arnout; Hausil, Farzia; Bouwstra, Joke A

2015-01-01

In the studies described in this study, we introduce a novel ex vivo human skin barrier repair model. To develop this, we removed the upper layer of the skin, the stratum corneum (SC) by a reproducible cyanoacrylate stripping technique. After stripping the explants, they were cultured in vitro to allow the regeneration of the SC. We selected two culture temperatures 32 °C and 37 °C and a period of either 4 or 8 days. After 8 days of culture, the explant generated SC at a similar thickness compared to native human SC. At 37 °C, the early and late epidermal differentiation programmes were executed comparably to native human skin with the exception of the barrier protein involucrin. At 32 °C, early differentiation was delayed, but the terminal differentiation proteins were expressed as in stripped explants cultured at 37 °C. Regarding the barrier properties, the SC lateral lipid organization was mainly hexagonal in the regenerated SC, whereas the lipids in native human SC adopt a more dense orthorhombic organization. In addition, the ceramide levels were higher in the cultured explants at 32 °C and 37 °C than in native human SC. In conclusion, we selected the stripped ex vivo skin model cultured at 37 °C as a candidate model to study skin barrier repair because epidermal and SC characteristics mimic more closely the native human skin than the ex vivo skin model cultured at 32 °C. Potentially, this model can be used for testing formulations for skin barrier repair. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Efficient ex vivo delivery of chemically modified messenger RNA using lipofection and magnetofection.

Science.gov (United States)

Badieyan, Zohreh Sadat; Pasewald, Tamara; Mykhaylyk, Olga; Rudolph, Carsten; Plank, Christian

2017-01-22

Recently, chemically modified mRNA (cmRNA) therapeutics have been the subject of extensive application-oriented research in both academia and industry as a safer alternative for gene and recombinant protein therapies. However, the lack of an efficient delivery system hinders widespread application. Here we used ∼100-nm lipoplexes and magnetic lipoplexes that can protect cmRNA from RNases and efficiently deliver it into muscle and fat tissues as well as to the endothelium of the carotid artery. Establishing magnetofection for ex vivo cmRNA delivery for the first time, we suggest this method for potential enhanced and targeted delivery of cmRNA. This study introduces optimal cmRNA complexes with high ex vivo efficiency as good candidates for further in vivo cmRNA delivery. Copyright © 2016 Elsevier Inc. All rights reserved.

An elegant technique for ex vivo imaging in experimental research—Optical coherence tomography (OCT)

DEFF Research Database (Denmark)

Tschernig, T.; Thrane, Lars; Jørgensen, Thomas Martini

2013-01-01

Optical coherence tomography (OCT) is an elegant technology for imaging of tissues and organs and has been established for clinical use for around a decade. Thus, it is used in vivo but can also serve as a valuable ex vivo imaging tool in experimental research. Here, a brief overview is given...

Ex Vivo Activity of Endoperoxide Antimalarials, Including Artemisone and Arterolane, against Multidrug-Resistant Plasmodium falciparum Isolates from Cambodia

Science.gov (United States)

2014-10-01

OCT 2014 2. REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Ex Vivo Activity of Endoperoxide Antimalarials , Including Artemisone...Prescribed by ANSI Std Z39-18 Ex Vivo Activity of Endoperoxide Antimalarials , Including Artemisone and Arterolane, against Multidrug-Resistant...potent antimalarial activity (2, 3). Despite having a rapid mecha- nism of action, artemisinin resistance eventually emerged and was first detected

Chick embryo partial ischemia model: a new approach to study ischemia ex vivo.

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Syamantak Majumder

Full Text Available BACKGROUND: Ischemia is a pathophysiological condition due to blockade in blood supply to a specific tissue thus damaging the physiological activity of the tissue. Different in vivo models are presently available to study ischemia in heart and other tissues. However, no ex vivo ischemia model has been available to date for routine ischemia research and for faster screening of anti-ischemia drugs. In the present study, we took the opportunity to develop an ex vivo model of partial ischemia using the vascular bed of 4(th day incubated chick embryo. METHODOLOGY/PRINCIPAL FINDINGS: Ischemia was created in chick embryo by ligating the right vitelline artery using sterile surgical suture. Hypoxia inducible factor- 1 alpha (HIF-1alpha, creatine phospho kinase-MB and reactive oxygen species in animal tissues and cells were measured to confirm ischemia in chick embryo. Additionally, ranolazine, N-acetyl cysteine and trimetazidine were administered as an anti-ischemic drug to validate the present model. Results from the present study depicted that blocking blood flow elevates HIF-1alpha, lipid peroxidation, peroxynitrite level in ischemic vessels while ranolazine administration partially attenuates ischemia driven HIF-1alpha expression. Endothelial cell incubated on ischemic blood vessels elucidated a higher level of HIF-1alpha expression with time while ranolazine treatment reduced HIF-1alpha in ischemic cells. Incubation of caprine heart strip on chick embryo ischemia model depicted an elevated creatine phospho kinase-MB activity under ischemic condition while histology of the treated heart sections evoked edema and disruption of myofibril structures. CONCLUSIONS/SIGNIFICANCE: The present study concluded that chick embryo partial ischemia model can be used as a novel ex vivo model of ischemia. Therefore, the present model can be used parallel with the known in vivo ischemia models in understanding the mechanistic insight of ischemia development and in

Identification of factors promoting ex vivo maintenance of mouse hematopoietic stem cells by long-term single-cell quantification.

Science.gov (United States)

Kokkaliaris, Konstantinos D; Drew, Erin; Endele, Max; Loeffler, Dirk; Hoppe, Philipp S; Hilsenbeck, Oliver; Schauberger, Bernhard; Hinzen, Christoph; Skylaki, Stavroula; Theodorou, Marina; Kieslinger, Matthias; Lemischka, Ihor; Moore, Kateri; Schroeder, Timm

2016-09-01

The maintenance of hematopoietic stem cells (HSCs) during ex vivo culture is an important prerequisite for their therapeutic manipulation. However, despite intense research, culture conditions for robust maintenance of HSCs are still missing. Cultured HSCs are quickly lost, preventing their improved analysis and manipulation. Identification of novel factors supporting HSC ex vivo maintenance is therefore necessary. Coculture with the AFT024 stroma cell line is capable of maintaining HSCs ex vivo long-term, but the responsible molecular players remain unknown. Here, we use continuous long-term single-cell observation to identify the HSC behavioral signature under supportive or nonsupportive stroma cocultures. We report early HSC survival as a major characteristic of HSC-maintaining conditions. Behavioral screening after manipulation of candidate molecules revealed that the extracellular matrix protein dermatopontin (Dpt) is involved in HSC maintenance. DPT knockdown in supportive stroma impaired HSC survival, whereas ectopic expression of the Dpt gene or protein in nonsupportive conditions restored HSC survival. Supplementing defined stroma- and serum-free culture conditions with recombinant DPT protein improved HSC clonogenicity. These findings illustrate a previously uncharacterized role of Dpt in maintaining HSCs ex vivo. © 2016 by The American Society of Hematology.

'En face' ex vivo reflectance confocal microscopy to help the surgery of basal cell carcinoma of the eyelid.

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Espinasse, Marine; Cinotti, Elisa; Grivet, Damien; Labeille, Bruno; Prade, Virginie; Douchet, Catherine; Cambazard, Frédéric; Thuret, Gilles; Gain, Philippe; Perrot, Jean Luc

2017-07-01

Ex vivo confocal microscopy is a recent imaging technique for the perioperative control of skin tumour margins. Up to date, it has been used in the fluorescence mode and with vertical sections of the specimen margins. The aim of this study was to evaluate its use in the reflectance mode and with a horizontal ('en face') scanning of the surgical specimen in a series of basal cell carcinoma of the eyelid. Prospective consecutive cohort study was performed at the University Hospital of Saint-Etienne, France. Forty-one patients with 42 basal cell carcinoma of the eyelid participated in this study. Basal cell carcinomas were excised with a 2-mm-wide clinically safe margin. The surgical specimens were analysed under ex vivo confocal microscopy in the reflectance mode and with an en face scanning in order to control at a microscopic level if the margins were free from tumour invasion. Histopathogical examination was later performed in order to compare the results. Sensitivity and specificity of ex vivo confocal microscopy for the presence of tumour-free margins. Ex vivo confocal microscopy results were consistent with histopathology in all cases (tumour-free margins in 40 out of 42 samples; sensitivity and specificity of 100%). Ex vivo confocal microscopy in the reflectance mode with an 'en face' scanning can control tumour margins of eyelid basal cell carcinomas and optimize their surgical management. This procedure has the advantage on the fluorescent mode of not needing any contrast agent to examine the samples. © 2016 Royal Australian and New Zealand College of Ophthalmologists.

Ex-vivo evaluation of crab shell chitosan as absorption enhancer in ...

African Journals Online (AJOL)

This study was aimed at evaluating crab shell chitosan as absorption enhancer in ciprofloxacin tablet formulation using the ex-vivo model. Six batches of ciprofloxacin tablets containing varying concentrations of crab shell-derived chitosan ranging from 0 to 5% w/w at 1% w/w intervals were produced. Batch CTS-0 ...

Ex-vivo assessment and non-invasive in vivo imaging of internal hemorrhages in Aga2/+ mutant mice

Energy Technology Data Exchange (ETDEWEB)

Ermolayev, Vladimir [Institute for Biological and Medical Imaging, Helmholtz Zentrum München, Building 56, Ingolstädter Landstraße 1, D-85764 Neuherberg (Germany); Cohrs, Christian M. [Institute for Experimental Genetics, Helmholtz Zentrum München, Ingolstädter Landstraße 1, D-85764 Neuherberg (Germany); Mohajerani, Pouyan; Ale, Angelique [Institute for Biological and Medical Imaging, Helmholtz Zentrum München, Building 56, Ingolstädter Landstraße 1, D-85764 Neuherberg (Germany); Hrabé de Angelis, Martin [Institute for Experimental Genetics, Helmholtz Zentrum München, Ingolstädter Landstraße 1, D-85764 Neuherberg (Germany); Ntziachristos, Vasilis, E-mail: v.ntziachristos@tum.de [Institute for Biological and Medical Imaging, Helmholtz Zentrum München, Building 56, Ingolstädter Landstraße 1, D-85764 Neuherberg (Germany)

2013-03-08

Highlights: ► Aga2/+ mice, model for Osteogenesis imperfecta, have type I collagen mutation. ► Aga2/+ mice display both moderate and severe phenotypes lethal 6–11th postnatal. ► Internal hemorrhages studied in Aga2/+ vs. control mice at 6 and 9 days postnatal. ► Anatomical and functional findings in-vivo contrasted to the ex-vivo appearance. -- Abstract: Mutations in type I collagen genes (COL1A1/2) typically lead to Osteogenesis imperfecta, the most common heritable cause of skeletal fractures and bone deformation in humans. Heterozygous Col1a1{sup Aga2/+}, animals with a dominant mutation in the terminal C-propeptide domain of type I collagen develop typical skeletal hallmarks and internal hemorrhages starting from 6 day after birth. The disease progression for Aga2/+ mice, however, is not uniform differing between severe phenotype lethal at the 6–11th day of life, and moderate-to-severe one with survival to adulthood. Herein we investigated whether a new modality that combines X-ray computer tomography with fluorescence tomography in one hybrid system can be employed to study internal bleedings in relation to bone fractures and obtain insights into disease progression. The disease phenotype was characterized on Aga2/+ vs. wild type mice between 6 and 9 days postnatal. Anatomical and functional findings obtained in-vivo were contrasted to the ex-vivo appearance of the same tissues under cryo-slicing.

Ex-vivo assessment and non-invasive in vivo imaging of internal hemorrhages in Aga2/+ mutant mice

International Nuclear Information System (INIS)

Ermolayev, Vladimir; Cohrs, Christian M.; Mohajerani, Pouyan; Ale, Angelique; Hrabé de Angelis, Martin; Ntziachristos, Vasilis

2013-01-01

Highlights: ► Aga2/+ mice, model for Osteogenesis imperfecta, have type I collagen mutation. ► Aga2/+ mice display both moderate and severe phenotypes lethal 6–11th postnatal. ► Internal hemorrhages studied in Aga2/+ vs. control mice at 6 and 9 days postnatal. ► Anatomical and functional findings in-vivo contrasted to the ex-vivo appearance. -- Abstract: Mutations in type I collagen genes (COL1A1/2) typically lead to Osteogenesis imperfecta, the most common heritable cause of skeletal fractures and bone deformation in humans. Heterozygous Col1a1 Aga2/+ , animals with a dominant mutation in the terminal C-propeptide domain of type I collagen develop typical skeletal hallmarks and internal hemorrhages starting from 6 day after birth. The disease progression for Aga2/+ mice, however, is not uniform differing between severe phenotype lethal at the 6–11th day of life, and moderate-to-severe one with survival to adulthood. Herein we investigated whether a new modality that combines X-ray computer tomography with fluorescence tomography in one hybrid system can be employed to study internal bleedings in relation to bone fractures and obtain insights into disease progression. The disease phenotype was characterized on Aga2/+ vs. wild type mice between 6 and 9 days postnatal. Anatomical and functional findings obtained in-vivo were contrasted to the ex-vivo appearance of the same tissues under cryo-slicing

Fusing in vivo and ex vivo NMR sources of information for brain tumor classification

International Nuclear Information System (INIS)

Croitor-Sava, A R; Laudadio, T; Sima, D M; Van Huffel, S; Martinez-Bisbal, M C; Celda, B; Piquer, J; Heerschap, A

2011-01-01

In this study we classify short echo-time brain magnetic resonance spectroscopic imaging (MRSI) data by applying a model-based canonical correlation analyses algorithm and by using, as prior knowledge, multimodal sources of information coming from high-resolution magic angle spinning (HR-MAS), MRSI and magnetic resonance imaging. The potential and limitations of fusing in vivo and ex vivo nuclear magnetic resonance sources to detect brain tumors is investigated. We present various modalities for multimodal data fusion, study the effect and the impact of using multimodal information for classifying MRSI brain glial tumors data and analyze which parameters influence the classification results by means of extensive simulation and in vivo studies. Special attention is drawn to the possibility of considering HR-MAS data as a complementary dataset when dealing with a lack of MRSI data needed to build a classifier. Results show that HR-MAS information can have added value in the process of classifying MRSI data

Bacterial transmission from contact lenses to porcine corneas : An ex vivo study

NARCIS (Netherlands)

Vermeltfoort, Pit B. J.; van Kooten, Theo G.; Bruinsma, Gerda .; Hooymans, Anneke M. M.; van der Mei, Henny C.; Busscher, Henk J.

PURPOSE. To quantify the transmission to ex vivo porcine eyes of Staphylococcus aureus 835 and Pseudomonas aeruginosa 3 from three types of contact lenses - one daily wear and two extended wear - differing in hydrophobicity and roughness. METHODS. One daily wear lens (etafilcon) and two

Tailored nanostructured platforms for boosting transcorneal permeation: Box-Behnken statistical optimization, comprehensive in vitro, ex vivo and in vivo characterization.

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Elsayed, Ibrahim; Sayed, Sinar

2017-01-01

Ocular drug delivery systems suffer from rapid drainage, intractable corneal permeation and short dosing intervals. Transcorneal drug permeation could increase the drug availability and efficiency in the aqueous humor. The aim of this study was to develop and optimize nanostructured formulations to provide accurate doses, long contact time and enhanced drug permeation. Nanovesicles were designed based on Box-Behnken model and prepared using the thin film hydration technique. The formed nanodispersions were evaluated by measuring the particle size, polydispersity index, zeta potential, entrapment efficiency and gelation temperature. The obtained desirability values were utilized to develop an optimized nanostructured in situ gel and insert. The optimized formulations were imaged by transmission and scanning electron microscopes. In addition, rheological characters, in vitro drug diffusion, ex vivo and in vivo permeation and safety of the optimized formulation were investigated. The optimized insert formulation was found to have a relatively lower viscosity, higher diffusion, ex vivo and in vivo permeation, when compared to the optimized in situ gel. So, the lyophilized nanostructured insert could be considered as a promising carrier and transporter for drugs across the cornea with high biocompatibility and effectiveness.

Tailored nanostructured platforms for boosting transcorneal permeation: Box–Behnken statistical optimization, comprehensive in vitro, ex vivo and in vivo characterization

Science.gov (United States)

Elsayed, Ibrahim; Sayed, Sinar

2017-01-01

Ocular drug delivery systems suffer from rapid drainage, intractable corneal permeation and short dosing intervals. Transcorneal drug permeation could increase the drug availability and efficiency in the aqueous humor. The aim of this study was to develop and optimize nanostructured formulations to provide accurate doses, long contact time and enhanced drug permeation. Nanovesicles were designed based on Box–Behnken model and prepared using the thin film hydration technique. The formed nanodispersions were evaluated by measuring the particle size, polydispersity index, zeta potential, entrapment efficiency and gelation temperature. The obtained desirability values were utilized to develop an optimized nanostructured in situ gel and insert. The optimized formulations were imaged by transmission and scanning electron microscopes. In addition, rheological characters, in vitro drug diffusion, ex vivo and in vivo permeation and safety of the optimized formulation were investigated. The optimized insert formulation was found to have a relatively lower viscosity, higher diffusion, ex vivo and in vivo permeation, when compared to the optimized in situ gel. So, the lyophilized nanostructured insert could be considered as a promising carrier and transporter for drugs across the cornea with high biocompatibility and effectiveness. PMID:29133980

Novel In Vitro/Ex Vivo Animal Modeling for Filovirus Aerosol Infection

Science.gov (United States)

2014-09-01

Infection PRINCIPAL INVESTIGATOR: Ayesha Mahmood, Ph.D. CONTRACTING ORGANIZATION: Sanofi Pasteur VaxDesign Corporation...ORGANIZATION NAME(S) AND ADDRESS(ES) 8. PERFORMING ORGANIZATION REPORT Sanofi Pasteur VaxDesign Corporation Orlando, Florida, 32826 9...a collaborative research effort between the USAMRIID Labs and Sanofi Pasteur VaxDesign to develop in vitro and ex vivo viral disease model systems

Sistemas osso-implante ex vivo utilizando haste intramedular polimérica para imobilização de fraturas femorais em bovinos jovens Ex vivo bone-implant systems using polymeric intramedullary nails for fixation of femoral fractures in young calves

Directory of Open Access Journals (Sweden)

Odael Spadeto Junior

2011-02-01

Full Text Available O desenvolvimento de novos aparatos usando materiais disponíveis e de baixo custo pode ser uma alternativa viável para o tratamento cirúrgico de fraturas em ossos longos de bovinos. O objetivo deste estudo foi avaliar a resistência mecânica de fêmures de bovinos jovens com fratura diafiseal, imobilizados com hastes intramedulares bloqueadas, compostas por diferentes polímeros. Para tanto, testes físicos de compressão e flexão, por meio de uma máquina universal de ensaios foram realizados em quatro grupos distintos de seis fêmures obtidos de bovinos jovens. Em um dos grupos, os ossos foram mantidos íntegros (grupo controle, enquanto que os outros os ossos, foram fraturados e imobilizados com uma haste intramedular bloqueada, composta por polipropileno, poliacetal ou poliamida (uma para cada grupo. Independente do polímero utilizado, nenhuma das hastes estudadas ofereceu aos fêmures fraturados resistência comparável ao osso íntegro, quando consideradas em conjunto as forças de flexão e compressão. A concordância desses achados com resultados in vivo previamente publicados, demonstra que a metodologia utilizada para testes ex vivo pode ser útil na seleção de materiais mais resistentes para confecção de novos modelos.The development of new devices using available and low cost materials may be an useful alternative for the treatment of long bone fractures in large animals. The aim of this study was to evaluate the mechanical strength of young bovine femur with diaphyseal fracture fixed with different polymeric intramedullary nails. Bending and compression tests using a universal machine were carried out in 4 distinct groups of 6 femurs from young calves. In one of the groups bones were intact while in the other three fractured bones were fixed using an intramedullary nail made of polypropylene, polyacetal or polyamide (one for each group. Considering bending and compression tests together, none of the used polymers offered

Measurement of histamine release from human lung tissue ex vivo by microdialysis technique

DEFF Research Database (Denmark)

Nissen, Dan; Petersen, Lars Jelstrup; Nolte, H

1998-01-01

OBJECTIVE AND DESIGN: Currently no method is available for measurement of mediator release from intact human lung. In this study, a microdialysis technique was used to measure histamine release from mast cells in human lung tissue ex vivo. MATERIAL: Microdialysis fibers of 216 microm were inserted...... responses were observed but data could be reproduced within individual donors. Monocyte chemoattractant protein-1, a potent basophil secretagogue, did not induce histamine release in lung tissue which indicated mast cells to be the histamine source. Substance P did not release histamine in the lung tissue....... CONCLUSIONS: The microdialysis technique allowed measurements of histamine release from mast cells in intact lung ex vivo. The method may prove useful since a number of experiments can be performed in a few hours in intact lung tissue without any dispersion or enzymatic treatment....

Anti-Inflammatory Inhibitors Targeting Jak and Ikk Have An Anabolic Effect on Type II Collagen Turnover ex Vivo

DEFF Research Database (Denmark)

Kjelgaard-Petersen, Cecilie Freja; Bay-Jensen, Anne-Christine; Karsdal, M.A.

2016-01-01

be beneficial for the selection of novel anti-inflammatory treatments for RA and iOA. Objectives The aim of this study was to investigate the direct effect of the anti-inflammatory inhibitors R406 (the active metabolite of Fostamatinib), Tofacitinib, TPCA-1 and SB203580 on the cartilage ECM turnover. Methods...... Full depth bovine cartilage ex vivo cultures were cultured for 3 weeks with OSM [10 ng/mL] and TNFα [2 ng/mL] (O+T) or together with R406, Tofacitinib or TPCA-1 at 10 μM and a two-fold dilution to 0.16 μM. SB203580 was tested at 3 μM, 1 μM and 0.3 μM. As negative control, untreated explants were...... R406, the Jak inhibitor Tofacitinib, and the IKK inhibitor TPCA-1 inhibited the release of ARGS or AGNx1, while the p38 inhibitor, SB203580, had no effect. The turnover of type II collagen was measured by the formation of type II collagen (ProC2) and MMP-mediated degradation of type II collagen (C2M...

Mouse Precision-Cut Liver Slices as an ex Vivo Model To Study Idiosyncratic Drug-Induced Liver Injury

NARCIS (Netherlands)

Hadi, Mackenzie; Chen, Yixi; Starokozhko, Viktoriia; Groothuis, Geny M. M.; Merema, M.T.

Idiosyncratic drug-induced liver injury (IDILI) has been the top reason for withdrawing drugs from the market or for black box warnings. IDILI may arise from the interaction of a drug's reactive metabolite with a mild inflammation that renders the liver more sensitive to injury resulting in

Micron-sized and submicron-sized aerosol deposition in a new ex vivo preclinical model.

Science.gov (United States)

Perinel, Sophie; Leclerc, Lara; Prévôt, Nathalie; Deville, Agathe; Cottier, Michèle; Durand, Marc; Vergnon, Jean-Michel; Pourchez, Jérémie

2016-07-07

The knowledge of where particles deposit in the respiratory tract is crucial for understanding the health effects associated with inhaled drug particles. An ex vivo study was conducted to assess regional deposition patterns (thoracic vs. extrathoracic) of radioactive polydisperse aerosols with different size ranges [0.15 μm-0.5 μm], [0.25 μm-1 μm] and [1 μm-9 μm]. SPECT/CT analyses were performed complementary in order to assess more precisely the regional deposition of aerosols within the pulmonary tract. Experiments were set using an original respiratory tract model composed of a human plastinated head connected to an ex vivo porcine pulmonary tract. The model was ventilated by passive expansion, simulating pleural depressions. Aerosol was administered during nasal breathing. Planar scintigraphies allowed to calculate the deposited aerosol fractions for particles in the three size ranges from sub-micron to micron The deposited fractions obtained, for thoracic vs. extra-thoracic regions respectively, were 89 ± 4 % vs. 11 ± 4 % for [0.15 μm-0.5 μm], 78 ± 5 % vs. 22 ± 5 % for [0.25 μm-1 μm] and 35 ± 11 % vs.65 ± 11 % for [1 μm-9 μm]. Results obtained with this new ex vivo respiratory tract model are in good agreement with the in vivo data obtained in studies with baboons and humans.

[Infecting glial cells with antimony resistant Leishmania tropica: A new ex-vivo model].

Science.gov (United States)

Zorbozan, Orçun; Harman, Mehmet; Evren, Vedat; Erdoğan, Mümin Alper; Kılavuz, Aslı; Tunalı, Varol; Çavuş, İbrahim; Yılmaz, Özlem; Özbilgin, Ahmet; Turgay, Nevin

2018-01-01

Leishmaniasis is a vector-borne zoonotic disease that shows different clinical features like cutaneous, mucocutaneous, visceral and viscerotropic forms. The protocols used in the treatment of leishmaniasis are toxic and have many limitations during administration. One of the limitations of treatment is the resistance against the protocols in practice. There is also a need to define new treatment options especially for resistant patients. Ex-vivo models using primary cell cultures may be a good source for evaluating new drug options in patients with antimony resistance, in addition to in-vitro and in-vivo studies. In this study, it was aimed to define a new ex-vivo culture model to evaluate treatment options in patients with cutaneous leishmaniasis who did not respond to treatment. In our experimental model of ex-vivo infection, Leishmania tropica promastigotes isolated from a case previously diagnosed with cutaneous leishmaniasis were used. The primary astroglial cell culture used for the ex-vivo model was prepared from 2-3 days old neonatal Sprague Dawley rat brains under sterile conditions by the modification McCarthy's method. The astroglia cells, which reached sufficient density, were infected with antimony resistant L.tropica promastigotes. After 24 hours of incubation, the supernatant on the cells were collected, the cell culture plate was dried at room temperature, then fixed with methyl alcohol and stained with Giemsa to search for L.tropica amastigotes. Amastigotes were intensely observed in glia cells in primary cell cultures infected with L.tropica promastigotes. No promastigotes were seen on Giemsa stained preparations of the precipitates prepared from the bottom sediment after the centrifugation of the liquid medium removed from the infected plates. In this study, promastigotes from a cutaneous leishmaniasis patient unable to respond to pentavalent antimony therapy were shown to infect rat glia cells and converted to amastigote form. This amastigote

High resolution SAW elastography for ex-vivo porcine skin specimen

Science.gov (United States)

Zhou, Kanheng; Feng, Kairui; Wang, Mingkai; Jamera, Tanatswa; Li, Chunhui; Huang, Zhihong

2018-02-01

Surface acoustic wave (SAW) elastography has been proven to be a non-invasive, non-destructive method for accurately characterizing tissue elastic properties. Current SAW elastography technique tracks generated surface acoustic wave impulse point by point which are a few millimeters away. Thus, reconstructed elastography has low lateral resolution. To improve the lateral resolution of current SAW elastography, a new method was proposed in this research. A M-B scan mode, high spatial resolution phase sensitive optical coherence tomography (PhS-OCT) system was employed to track the ultrasonically induced SAW impulse. Ex-vivo porcine skin specimen was tested using this proposed method. A 2D fast Fourier transform based algorithm was applied to process the acquired data for estimating the surface acoustic wave dispersion curve and its corresponding penetration depth. Then, the ex-vivo porcine skin elastogram was established by relating the surface acoustic wave dispersion curve and its corresponding penetration depth. The result from the proposed method shows higher lateral resolution than that from current SAW elastography technique, and the approximated skin elastogram could also distinguish the different layers in the skin specimen, i.e. epidermis, dermis and fat layer. This proposed SAW elastography technique may have a large potential to be widely applied in clinical use for skin disease diagnosis and treatment monitoring.

Effects of ex-vivo and in-vivo treatment with probiotics on the inflammasome in dogs with chronic enteropathy.

Directory of Open Access Journals (Sweden)

Silke Schmitz

Full Text Available Inflammasomes coordinate the maturation of IL-1β and IL-18 in response to danger signals. They are vital for maintenance of intestinal homeostasis and have been linked to chronic intestinal inflammation in humans. Probiotics have been advocated as treatment in intestinal inflammation. So far, no study has investigated the role of the inflammasome in canine chronic enteropathy (CE. In this study the intestinal expression of inflammasome components was assessed in CE dogs compared to controls, when treated with probiotic Enterococcus faecium (EF ex-vivo and in-vivo. RNA extraction from endoscopic biopsies and reverse-transcriptase quantitative PCR was performed for NLRP3, casp-1, IL-1β and IL-18. Immunohistochemistry was performed to investigate protein expression in tissues. Gene expression of casp-1 and NLRP3 was lower in CE samples than controls. Ex-vivo treatment with EF reduced NLRP3 expression in control samples. Treatment of CE dogs with EF alongside dietary intervention had no effect on gene expression. In contrast, IL-1β protein expression in CE decreased with dietary treatment (but not with probiotics. The results of this study suggest that the inflammasome or its components may be partially involved in the inflammatory process seen in CE, but distinct from intestinal inflammation in humans.

Rapid ex vivo imaging of PAIII prostate to bone tumor with SWIFT-MRI.

Science.gov (United States)

Luhach, Ihor; Idiyatullin, Djaudat; Lynch, Conor C; Corum, Curt; Martinez, Gary V; Garwood, Michael; Gillies, Robert J

2014-09-01

The limiting factor for MRI of skeletal/mineralized tissue is fast transverse relaxation. A recent advancement in MRI technology, SWIFT (Sweep Imaging with Fourier Transform), is emerging as a new approach to overcome this difficulty. Among other techniques like UTE, ZTE, and WASPI, the application of SWIFT technology has the strong potential to impact preclinical and clinical imaging, particularly in the context of primary or metastatic bone cancers because it has the added advantage of imaging water in mineralized tissues of bone allowing MRI images to be obtained of tissues previously visible only with modalities such as computed tomography (CT). The goal of the current study is to examine the feasibility of SWIFT for the assessment of the prostate cancer induced changes in bone formation (osteogenesis) and destruction (osteolysis) in ex vivo specimens. A luciferase expressing prostate cancer cell line (PAIII) or saline control was inoculated directly into the tibia of 6-week-old immunocompromised male mice. Tumor growth was assessed weekly for 3 weeks before euthanasia and dissection of the tumor bearing and sham tibias. The ex vivo mouse tibia specimens were imaged with a 9.4 Tesla (T) and 7T MRI systems. SWIFT images are compared with traditional gradient-echo and spin-echo MRI images as well as CT and histological sections. SWIFT images with nominal resolution of 78 μm are obtained with the tumor and different bone structures identified. Prostate cancer induced changes in the bone microstructure are visible in SWIFT images, which is supported by spin-echo, high resolution CT and histological analysis. SWIFT MRI is capable of high-quality high-resolution ex vivo imaging of bone tumor and surrounding bone and soft tissues. Furthermore, SWIFT MRI shows promise for in vivo bone tumor imaging, with the added benefits of nonexposure to ionizing radiation, quietness, and speed. Copyright © 2013 Wiley Periodicals, Inc.

Ex-vivo release of Pipeline Embolization Device polytetrafluoroethylene (PTFE) sleeves for improved distal landing zone accuracy in-vivo: A technical note.

Science.gov (United States)

Griessenauer, Christoph J; Gupta, Raghav; Moore, Justin; Thomas, Ajith J; Ogilvy, Christopher S

2016-12-01

Distal landing zone accuracy is critical in some intracranial aneurysms treated with the Pipeline Embolization Device (PED), and delayed opening of the distal end of the device can complicate the procedure. Here, we report a technical nuance that facilitates accurate placement of the distal end of the PED by ex-vivo, pre-implantation release of the PED Flex polytetrafluoroethylene (PTFE) sleeves. The PED Flex is partially pushed out of the introducer sheath ex-vivo, pre-implantation until the distal PED opens entirely and the PTFE sleeves are located distal to the device. Without inverting the PTFE sleeves, the PED is carefully pulled back into the introducer sheath placing the PTFE sleeves inside the device. The PED is loaded into the microcatheter and advanced toward the site of implantation. When the PED is initially deployed and pushed out of the microcatheter, it opens immediately and provides an anchor for the remainder of the deployment process. We present a video (supplementary material) that illustrates the technique along with an illustrative case. Ex-vivo, pre-implantation release of the PTFE sleeves is an option in aneurysm treatment where distal landing accuracy is critical. Even without the protection of the PTFE sleeves, our clinical observation shows that the PED can be advanced safely through the microcatheter in selected cases. © The Author(s) 2016.

An ex vivo approach to botanical-drug interactions: a proof of concept study.

Science.gov (United States)

Wang, Xinwen; Zhu, Hao-Jie; Munoz, Juliana; Gurley, Bill J; Markowitz, John S

2015-04-02

Botanical medicines are frequently used in combination with therapeutic drugs, imposing a risk for harmful botanical-drug interactions (BDIs). Among the existing BDI evaluation methods, clinical studies are the most desirable, but due to their expense and protracted time-line for completion, conventional in vitro methodologies remain the most frequently used BDI assessment tools. However, many predictions generated from in vitro studies are inconsistent with clinical findings. Accordingly, the present study aimed to develop a novel ex vivo approach for BDI assessment and expand the safety evaluation methodology in applied ethnopharmacological research. This approach differs from conventional in vitro methods in that rather than botanical extracts or individual phytochemicals being prepared in artificial buffers, human plasma/serum collected from a limited number of subjects administered botanical supplements was utilized to assess BDIs. To validate the methodology, human plasma/serum samples collected from healthy subjects administered either milk thistle or goldenseal extracts were utilized in incubation studies to determine their potential inhibitory effects on CYP2C9 and CYP3A4/5, respectively. Silybin A and B, two principal milk thistle phytochemicals, and hydrastine and berberine, the purported active constituents in goldenseal, were evaluated in both phosphate buffer and human plasma based in vitro incubation systems. Ex vivo study results were consistent with formal clinical study findings for the effect of milk thistle on the disposition of tolbutamide, a CYP2C9 substrate, and for goldenseal׳s influence on the pharmacokinetics of midazolam, a widely accepted CYP3A4/5 substrate. Compared to conventional in vitro BDI methodologies of assessment, the introduction of human plasma into the in vitro study model changed the observed inhibitory effect of silybin A, silybin B and hydrastine and berberine on CYP2C9 and CYP3A4/5, respectively, results which more

Ex vivo and in vivo evaluation of microemulsion based transdermal delivery of E. coli specific T4 bacteriophage: A rationale approach to treat bacterial infection.

Science.gov (United States)

Rastogi, Vaibhav; Yadav, Pragya; Verma, Anurag; Pandit, Jayanta K

2017-09-30

This study is focused on the development and evaluation of transdermal delivery of E. coli-specific T4 bacteriophages both ex-vivo and in-vivo using microemulsion as delivery carrier in eradicating the infection caused by E. coli. Microemulsions were prepared by mixing selected oil, surfactants and aqueous phase containing bacteriophages. The formulations were subjected to physicochemical characterization, ex-vivo and in-vivo permeation, stability studies, histological and immunofluorescence examination. The colloidal system exhibits a uniform size distribution, of finite size (150-320nm). Transmission electron microscopy revealed the encapsulation of bacteriophage in the aqueous globule. Ex-vivo permeation across skin was successfully achieved as 6×10 6 PFU/mL and 6.7×10 6 PFU/mL of T4 permeated from ME 6% and 10%, respectively. ME 6% was found to be thermodynamically stable and in-vivo permeation resulted in 5.49×10 5 PFU/mL of bacteriophages in the blood of the E. coli challenged rats, while 2.48×10 5 PFU/mL was detected in germ free rats, at the end of the study. Infected rats that were treated with bacteriophage were survived while significant mortality was observed in others. Histological and IL-6 immunofluorescence examination of the tissues revealed the efficacy/safety of the therapy. The microemulsion-based transdermal delivery of bacteriophage could be a promising approach to treat the infections caused by antibiotic-resistant bacteria. Copyright © 2017 Elsevier B.V. All rights reserved.

Implementing an innovated preservation technology: The American Society of Transplant Surgeons' (ASTS) Standards Committee White Paper on Ex Situ Liver Machine Perfusion.

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Quintini, Cristiano; Martins, Paulo N; Shah, Shimul; Killackey, Mary; Reed, Alan; Guarrera, James; Axelrod, David A

2018-05-23

The pervasive shortage of deceased donor liver allografts contributes to significant waitlist mortality despite efforts to increase organ donation. Ex vivo liver perfusion appears to enhance preservation of donor organs, extending viability and potentially evaluating function in organs previously considered too high risk for transplant. These devices pose novel challenges for organ allocation, safety, training, and finances. This white paper describes the American Society of Transplant Surgeons' belief that organ preservation technology is a vital advance, but its use should not change fundamental aspects of organ allocation. Additional data elements need to be collected, made available for organ assessment by transplant professionals to allow determination of organ suitability in the case of reallocation and incorporated into risk adjustment methodology. Finally, further work is needed to determine the optimal strategy for management and oversight of perfused organs prior to transplantation. © 2018 The American Society of Transplantation and the American Society of Transplant Surgeons.

A randomized comparison of laparoscopic, flexible endoscopic, and wired and wireless magnetic cameras on ex vivo and in vivo NOTES surgical performance.

Science.gov (United States)

Chang, Victoria C; Tang, Shou-Jiang; Swain, C Paul; Bergs, Richard; Paramo, Juan; Hogg, Deborah C; Fernandez, Raul; Cadeddu, Jeffrey A; Scott, Daniel J

2013-08-01

The influence of endoscopic video camera (VC) image quality on surgical performance has not been studied. Flexible endoscopes are used as substitutes for laparoscopes in natural orifice translumenal endoscopic surgery (NOTES), but their optics are originally designed for intralumenal use. Manipulable wired or wireless independent VCs might offer advantages for NOTES but are still under development. To measure the optical characteristics of 4 VC systems and to compare their impact on the performance of surgical suturing tasks. VC systems included a laparoscope (Storz 10 mm), a flexible endoscope (Olympus GIF 160), and 2 prototype deployable cameras (magnetic anchoring and guidance system [MAGS] Camera and PillCam). In a randomized fashion, the 4 systems were evaluated regarding standardized optical characteristics and surgical manipulations of previously validated ex vivo (fundamentals of laparoscopic surgery model) and in vivo (live porcine Nissen model) tasks; objective metrics (time and errors/precision) and combined surgeon (n = 2) performance were recorded. Subtle differences were detected for color tests, and field of view was variable (65°-115°). Suitable resolution was detected up to 10 cm for the laparoscope and MAGS camera but only at closer distances for the endoscope and PillCam. Compared with the laparoscope, surgical suturing performances were modestly lower for the MAGS camera and significantly lower for the endoscope (ex vivo) and PillCam (ex vivo and in vivo). This study documented distinct differences in VC systems that may be used for NOTES in terms of both optical characteristics and surgical performance. Additional work is warranted to optimize cameras for NOTES. Deployable systems may be especially well suited for this purpose.

Platelet lysate as a substitute for animal serum for the ex-vivo expansion of mesenchymal stem/stromal cells: present and future.

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Astori, Giuseppe; Amati, Eliana; Bambi, Franco; Bernardi, Martina; Chieregato, Katia; Schäfer, Richard; Sella, Sabrina; Rodeghiero, Francesco

2016-07-13

The use of fetal bovine serum (FBS) as a cell culture supplement is discouraged by regulatory authorities to limit the risk of zoonoses and xenogeneic immune reactions in the transplanted host. Additionally, FBS production came under scrutiny due to animal welfare concerns. Platelet derivatives have been proposed as FBS substitutes for the ex-vivo expansion of mesenchymal stem/stromal cells (MSCs) since platelet-derived growth factors can promote MSC ex-vivo expansion. Platelet-derived growth factors are present in platelet lysate (PL) obtained after repeated freezing-thawing cycles of the platelet-rich plasma or by applying physiological stimuli such as thrombin or CaCl2.PL-expanded MSCs have been used already in the clinic, taking advantage of their faster proliferation compared with FBS-expanded preparations. Should PL be applied to other biopharmaceutical products, its demand is likely to increase dramatically. The use of fresh platelet units for the production of PL raises concerns due to limited availability of platelet donors. Expired units might represent an alternative, but further data are needed to define safety, including pathogen reduction, and functionality of the obtained PL. In addition, relevant questions concerning the definition of PL release criteria, including concentration ranges of specific growth factors in PL batches for various clinical indications, also need to be addressed. We are still far from a common definition of PL and standardized PL manufacture due to our limited knowledge of the mechanisms that mediate PL-promoting cell growth. Here, we concisely discuss aspects of PL as MSC culture supplement as a preliminary step towards an agreed definition of the required characteristics of PL for the requirements of manufacturers and users.

Ex Vivo Expansion of Hematopoietic Stem Cells to Improve Engraftment in Stem Cell Transplantation.

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Ko, Kap-Hyoun; Nordon, Robert; O'Brien, Tracey A; Symonds, Geoff; Dolnikov, Alla

2017-01-01

The efficient use of hematopoietic stem cells (HSC) for transplantation is often limited by the relatively low numbers of HSC collected. The ex vivo expansion of HSC for clinical use is a potentially valuable and safe approach to increase HSC numbers thereby increasing engraftment and reducing the risk of morbidity from infection. Here, we describe a protocol for the robust ex vivo expansion of human CD34(+) HSC isolated from umbilical cord blood. The protocol described can efficiently generate large numbers of HSC. We also describe a flow cytometry-based method using high-resolution division tracking to characterize the kinetics of HSC growth and differentiation. Utilizing the guidelines discussed, it is possible for investigators to use this protocol as presented or to modify it for their specific needs.

Association between in vivo bone formation and ex vivo migratory capacity of human bone marrow stromal cells

DEFF Research Database (Denmark)

Andersen, Rikke K.; Zaher, Walid; Larsen, Kenneth Hauberg

2015-01-01

INTRODUCTION: There is a clinical need for developing systemic transplantation protocols for use of human skeletal stem cells (also known bone marrow stromal stem cells) (hBMSC) in tissue regeneration. In systemic transplantation studies, only a limited number of hBMSC home to injured tissues...... populations derived from telomerized hBMSC (hBMSC-TERT) with variable ability to form heterotopic bone when implanted subcutaneously in immune deficient mice. In vitro transwell migration assay was used and the in vivo homing ability of transplanted hBMSC to bone fractures in mice was visualized...... suggesting that only a subpopulation of hBMSC possesses "homing" capacity. Thus, we tested the hypothesis that a subpopulation of hBMSC defined by ability to form heterotopic bone in vivo, is capable of homing to injured bone. METHODS: We tested ex vivo and in vivo homing capacity of a number of clonal cell...

White blood cell-based detection of asymptomatic scrapie infection by ex vivo assays.

Directory of Open Access Journals (Sweden)

Sophie Halliez

Full Text Available Prion transmission can occur by blood transfusion in human variant Creutzfeldt-Jakob disease and in experimental animal models, including sheep. Screening of blood and its derivatives for the presence of prions became therefore a major public health issue. As infectious titer in blood is reportedly low, highly sensitive and robust methods are required to detect prions in blood and blood derived products. The objectives of this study were to compare different methods--in vitro, ex vivo and in vivo assays--to detect prion infectivity in cells prepared from blood samples obtained from scrapie infected sheep at different time points of the disease. Protein misfolding cyclic amplification (PMCA and bioassays in transgenic mice expressing the ovine prion protein were the most efficient methods to identify infected animals at any time of the disease (asymptomatic to terminally-ill stages. However scrapie cell and cerebellar organotypic slice culture assays designed to replicate ovine prions in culture also allowed detection of prion infectivity in blood cells from asymptomatic sheep. These findings confirm that white blood cells are appropriate targets for preclinical detection and introduce ex vivo tools to detect blood infectivity during the asymptomatic stage of the disease.

Helicobacter pylori-induced gastric pathology: insights from in vivo and ex vivo models

Directory of Open Access Journals (Sweden)

Michael D. Burkitt

2017-02-01

Full Text Available Gastric colonization with Helicobacter pylori induces diverse human pathological conditions, including superficial gastritis, peptic ulcer disease, mucosa-associated lymphoid tissue (MALT lymphoma, and gastric adenocarcinoma and its precursors. The treatment of these conditions often relies on the eradication of H. pylori, an intervention that is increasingly difficult to achieve and that does not prevent disease progression in some contexts. There is, therefore, a pressing need to develop new experimental models of H. pylori-associated gastric pathology to support novel drug development in this field. Here, we review the current status of in vivo and ex vivo models of gastric H. pylori colonization, and of Helicobacter-induced gastric pathology, focusing on models of gastric pathology induced by H. pylori, Helicobacter felis and Helicobacter suis in rodents and large animals. We also discuss the more recent development of gastric organoid cultures from murine and human gastric tissue, as well as from human pluripotent stem cells, and the outcomes of H. pylori infection in these systems.

Suppression of MMP activity in bovine cartilage explants cultures has little if any effect on the release of aggrecanase-derived aggrecan fragments

DEFF Research Database (Denmark)

Wang, Bijue; Chen, Pingping; Jensen, Anne-Christine Bay

2009-01-01

BACKGROUND: Progressive loss of articular cartilage is a central hallmark in many joint disease, however, the relative importance of individual proteolytic pathways leading to cartilage erosion is at present unknown. We therefore investigated the time-dependant release ex vivo of MMP- and aggreca......BACKGROUND: Progressive loss of articular cartilage is a central hallmark in many joint disease, however, the relative importance of individual proteolytic pathways leading to cartilage erosion is at present unknown. We therefore investigated the time-dependant release ex vivo of MMP......- and aggrecanase-derived fragments of aggrecan and type II collagen into the supernatant of bovine cartilage explants cultures using neo-epitope specific immunoassays, and to associate the release of these fragments with the activity of proteolytic enzymes using inhibitors. FINDINGS: Bovine cartilage explants were...... cultured in the presence or absence of the catabolic cytokines oncostatin M (OSM) and tumor necrosis factor alpha (TNFalpha). In parallel, explants were co-cultured with protease inhibitors such as GM6001, TIMP1, TIMP2 and TIMP3. Fragments released into the supernatant were determined using a range of neo...

Preservation of the liver for transplantation: Machine perfusion-based strategies for extended preservation and recovery

NARCIS (Netherlands)

Bruinsma, B.G.

2015-01-01

The field of liver transplantation remains burdened by a severe shortage of donor organs. An important limitation to the use of many livers is the ex vivo preservation of the organ, as maintaining the viability of the donor organ while outside the body is essential for successful transplantation.

Chest drainage teaching and training for medical students. Use of a surgical ex vivo pig model.

Science.gov (United States)

Tube, Milton Ignacio Carvalho; Netto, Fernando Antonio Campelo Spencer; Costa, Elaine; Lafayette, Daniell de Siqueira Araújo; Lima, George Augusto da Fonseca Carvalho Antunes; Menezes, Jamile Isabela Santos de; Aires, Vinicius Gueiros Buenos; Ferraz, Álvaro Antônio Bandeira; Campos, Josemberg Marins; Moraes, Fernando Ribeiro de

2016-05-01

Implement a constructivist approach in thoracic drainage training in surgical ex vivo pig models, to compare the acquisition of homogeneous surgical skills between medical students. Experimental study, prospective, transversal, analytical, controlled, three steps. Selection, training, evaluation. a) students without training in thoracic drainage; b) without exposure to constructivist methodology. 2) EXCLUSION CRITERIA: a) students developed surgical skills; b) a history of allergy. (N = 312). Two groups participated in the study: A and B. Lecture equal for both groups. Differentiated teaching: group A, descriptive and informative method; group B, learning method based on problems. A surgical ex vivo pig model for training the chest drain was created. Were applied pre and post-test, test goal-discursive and OSATS scale. Theoretical averages: Group A = 9.5 ± 0.5; Group B = 8.8 ± 1.1 (p = 0.006). Medium Practices: Group A = 22.8 ± 1.8; Group B = 23.0 ± 2.8 (p <0.001). Through the constructivist methodology implemented in the thoracic drainage training in surgical ex vivo pig models, has proven the acquisition of surgical skills homogeneous compared among medical students.

First identification of the herpes simplex virus by skin-dedicated ex vivo fluorescence confocal microscopy during herpetic skin infections.

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Cinotti, E; Perrot, J L; Labeille, B; Campolmi, N; Thuret, G; Naigeon, N; Bourlet, T; Pillet, S; Cambazard, F

2015-06-01

Skin-dedicated ex vivo fluorescence confocal microscopy (FCM) has so far been used to identify cutaneous tumours on freshly excised samples using acridine orange as fluorochrome. To use FCM for a new indication, namely, the identification of the herpes simplex virus (HSV) in skin lesions, using fluorescent antibodies. Six roof samples from skin vesicles suspicious for HSV lesions were incubated with anti-HSV-1 and anti-HSV-2 antibodies coupled with fluorescein isothiocyanate, and examined under skin-dedicated ex vivo FCM. The positive controls were swabs taken from the floor of each vesicle and observed under conventional direct fluorescence assay (DFA) and by viral cultures. Roof samples from three bullae of bullous pemphigoid were the negative controls. Using ex vivo FCM, the samples from the lesions clinically suspicious for HSV infection were seen to be fluorescent after incubation with anti-HSV-1, and were negative after incubation with anti-HSV-2 antibodies. Conventional DFA with an optical microscope and cultures confirmed the presence of HSV-1 infection. By using fluorescent antibodies to identify precise structures, ex vivo FCM can be used for indications other than tumour identification. More specifically, it can be an additional diagnostic tool for HSV infection. © 2014 British Association of Dermatologists.

The development of a three-dimensional scaffold for ex vivo biomimicry of human acute myeloid leukaemia.

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Blanco, Teresa Mortera; Mantalaris, Athanasios; Bismarck, Alexander; Panoskaltsis, Nicki

2010-03-01

Acute myeloid leukaemia (AML) is a cancer of haematopoietic cells that develops in three-dimensional (3-D) bone marrow niches in vivo. The study of AML has been hampered by lack of appropriate ex vivo models that mimic this microenvironment. We hypothesised that fabrication and optimisation of suitable biomimetic scaffolds for culturing leukaemic cells ex vivo might facilitate the study of AML in its native 3-D niche. We evaluated the growth of three leukaemia subtype-specific cell lines, K-562, HL60 and Kasumi-6, on highly porous scaffolds fabricated from biodegradable and non-biodegradable polymeric materials, such as poly (L-lactic-co-glycolic acid) (PLGA), polyurethane (PU), poly (methyl-methacrylate), poly (D, L-lactade), poly (caprolactone), and polystyrene. Our results show that PLGA and PU supported the best seeding efficiency and leukaemic growth. Furthermore, the PLGA and PU scaffolds were coated with extracellular matrix (ECM) proteins, collagen type I (62.5 or 125 microg/ml) and fibronectin (25 or 50 microg/ml) to provide biorecognition signals. The 3 leukaemia subtype-specific lines grew best on PU scaffolds coated with 62.5 microg/ml collagen type I over 6 weeks in the absence of exogenous growth factors. In conclusion, PU-collagen scaffolds may provide a practical model to study the biology and treatment of primary AML in an ex vivo mimicry. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

Surgical model pig ex vivo for venous dissection teaching in medical schools.

Science.gov (United States)

Tube, Milton Ignacio Carvalho; Spencer-Netto, Fernando Antonio Campelo; Oliveira, Anderson Igor Pereira de; Holanda, Arthur Cesário de; Barros, Bruno Leão Dos Santos; Rezende, Caio Cezar Gomes; Cavalcanti, João Pedro Guerra; Batista, Marília Apolinário; Campos, Josemberg Marins

2017-02-01

To investigate a method for development of surgical skills in medical students simulating venous dissection in surgical ex vivo pig model. Prospective, analytical, experimental, controlled study with four stages: selection, theoretical teaching, training and assessment. Sample of 312 students was divided into two groups: Group A - 2nd semester students; Group B - students of 8th semester. The groups were divided into five groups of 12 students, trained two hours per week in the semester. They set up four models to three students in each skill station assisted by a monitor. Teaching protocol emergency procedures training were applied to venous dissection, test goal-discursive and OSATS scale. The pre-test confirmed that the methodology has not been previously applied to the students. The averages obtained in the theoretical evaluation reached satisfactory parameters in both groups. The results of applying OSATS scale showed the best performance in group A compared to group B, however, both groups had satisfactory medium. The method was enough to raise a satisfactory level of skill both groups in venous dissection running on surgical swine ex vivo models.

Ex vivo rabbit cornea diffusion studies with a soluble insert of moxifloxacin.

Science.gov (United States)

Sebastián-Morelló, María; Calatayud-Pascual, María Aracely; Rodilla, Vicent; Balaguer-Fernández, Cristina; López-Castellano, Alicia

2018-02-01

The objective of this research was to develop and evaluate an ocular insert for the controlled drug delivery of moxifloxacin which could perhaps be used in the treatment of corneal keratitis or even bacterial endophthalmitis. We have evaluated the ex vivo ocular diffusion of moxifloxacin through rabbit cornea, both fresh and preserved under different conditions. Histological studies were also carried out. Subsequently, drug matrix inserts were prepared using bioadhesive polymers. The inserts were evaluated for their physicochemical parameters. Ophthalmic ex vivo permeation of moxifloxacin was carried out with the most promising insert. The formulate insert was thin and provided higher ocular diffusion than commercial formulations. Ocular diffusion studies revealed significant differences between fresh and frozen corneas. Histological examinations also showed differences in the thickness of stroma between fresh and frozen corneas. The ophthalmic insert we have developed allows a larger quantity of moxifloxacin to permeate through the cornea than existing commercial formulations of the drug. Ocular delivery of moxifloxacin with this insert could be a new approach for the treatment of eye diseases.

Rat brain sagittal organotypic slice cultures as an ex vivo dopamine cell loss system.

Science.gov (United States)

McCaughey-Chapman, Amy; Connor, Bronwen

2017-02-01

Organotypic brain slice cultures are a useful tool to study neurological function as they provide a more complex, 3-dimensional system than standard 2-dimensional in vitro cell cultures. Building on a previously developed mouse brain slice culture protocol, we have developed a rat sagittal brain slice culture system as an ex vivo model of dopamine cell loss. We show that rat brain organotypic slice cultures remain viable for up to 6 weeks in culture. Using Fluoro-Gold axonal tracing, we demonstrate that the slice 3-dimensional cytoarchitecture is maintained over a 4 week culturing period, with particular focus on the nigrostriatal pathway. Treatment of the cultures with 6-hydroxydopamine and desipramine induces a progressive loss of Fluoro-Gold-positive nigral cells with a sustained loss of tyrosine hydroxylase-positive nigral cells. This recapitulates the pattern of dopaminergic degeneration observed in the rat partial 6-hydroxydopamine lesion model and, most importantly, the progressive pathology of Parkinson's disease. Our slice culture platform provides an advance over other systems, as we demonstrate for the first time 3-dimensional cytoarchitecture maintenance of rat nigrostriatal sagittal slices for up to 6 weeks. Our ex vivo organotypic slice culture system provides a long term cellular platform to model Parkinson's disease, allowing for the elucidation of mechanisms involved in dopaminergic neuron degeneration and the capability to study cellular integration and plasticity ex vivo. Copyright © 2017 Elsevier B.V. All rights reserved.

Ex vivo fluorescence confocal microscopy for fast evaluation of tumour margins during Mohs surgery.

Science.gov (United States)

Bennàssar, A; Vilata, A; Puig, S; Malvehy, J

2014-02-01

Ex vivo fluorescence confocal microscopy (FCM) enables real-time imaging of skin morphology directly in freshly excised tissue. FCM displays wide field-of-view mosaics with cellular resolution, thus enabling a rapid bedside pathology. An application of interest is rapid detection of residual basal cell carcinoma (BCC) in skin excisions during Mohs surgery. We sought to evaluate the sensitivity and specificity of ex vivo imaging with FCM for the detection of residual BCC in Mohs tissue excisions, and to calculate the time invested up to the diagnosis for both FCM and frozen sections. Eighty consecutive BCCs were prospectively collected and the margins scanned with ex vivo FCM, including excisions with and without residual BCC of all major subtypes. Each mosaic was divided into two or four, resulting in 480 submosaics for study. Every confocal submosaic was assessed for the presence or absence of BCC and compared with standard frozen sections as the gold standard. Furthermore, the time spent for each technique was calculated and compared. The overall sensitivity and specificity of detecting residual BCC were 88% and 99%, respectively. Moreover, the new technique reduced by almost two-thirds the time invested when compared with the processing of a frozen section (P confocal mosaicing microscopy in fresh tissue for rapid surgical pathology, potentially to expedite and guide Mohs surgery with high accuracy. This observation is an important step towards the goal of using real-time surgical pathology for skin tumours. © 2013 British Association of Dermatologists.

Linarin Inhibits the Acetylcholinesterase Activity In-vitro and Ex-vivo

Science.gov (United States)

Feng, Xinchi; Wang, Xin; Liu, Youping; Di, Xin

2015-01-01

Linarin is a flavone glycoside in the plants Flos chrysanthemi indici, Buddleja officinalis, Cirsium setosum, Mentha arvensis and Buddleja davidii, and has been reported to possess analgesic, antipyretic, anti-inflammatory and neuroprotective activities. In this paper, linarin was investigated for its AChE inhibitory potential both in-vitro and ex-vivo. Ellman’s colorimetric method was used for the determination of AChE inhibitory activity in mouse brain. In-vitro assays revealed that linarin inhibited AChE activity with an IC50 of 3.801 ± 1.149 μM. Ex-vivo study showed that the AChE activity was significantly reduced in both the cortex and hippocampus of mice treated intraperitoneally with various doses of linarin (35, 70 and 140 mg/Kg). The inhibition effects produced by high dose of linarin were the same as that obtained after huperzine A treatment (0.5 mg/Kg). Molecular docking study revealed that both 4’-methoxyl group and 7-O-sugar moiety of linarin played important roles in ligand-receptor binding and thus they are mainly responsible for AChE inhibitory activity. In view of its potent AChE inhibitory activity, linarin may be a promising therapeutic agent for the treatment of some diseases associated with AChE, such as glaucoma, myasthenia gravis, gastric motility and Alzheimer’s disease. PMID:26330885

Ex-vivo expansion of red blood cells: how real for transfusion in humans?

Science.gov (United States)

Migliaccio, Anna Rita; Masselli, Elena; Varricchio, Lilian; Whitsett, Carolyn

2012-03-01

Blood transfusion is indispensable for modern medicine. In developed countries, the blood supply is adequate and safe but blood for alloimmunized patients is often unavailable. Concerns are increasing that donations may become inadequate in the future as the population ages prompting a search for alternative transfusion products. Improvements in culture conditions and proof-of-principle studies in animal models have suggested that ex-vivo expanded red cells may represent such a product. Compared to other cell therapies transfusion poses the unique challenge of requiring great cell doses (2.5×10(12) cells vs 10(7) cells). Although production of such cell numbers is theoretically possible, current technologies generate red cells in numbers sufficient only for safety studies. It is conceived that by the time these studies will be completed, technical barriers to mass cell production will have been eliminated making transfusion with ex-vivo generated red cells a reality. Copyright © 2011 Elsevier Ltd. All rights reserved.

Comparison of iodinated contrast media for the assessment of atherosclerotic plaque attenuation values by CT coronary angiography: Observations in an ex vivo model

NARCIS (Netherlands)

L. la Grutta (Ludovico); M. Galia (Massimo); G. Gentile; G. Lo Re (G.); E. Grassedonio (Emanuele); F. Coppolino; E. Maffei (Erica); E. Maresi (E.); A. Lo Casto (A.); F. Cademartiri (Filippo); M. Midiri (Massimo)

2013-01-01

textabstractObjective: To compare the influence of different iodinated contrast media with several dilutions on plaque attenuation in an ex vivo coronary model studied by multislice CT coronary angiography. Methods: In six ex vivo left anterior descending coronary arteries immersed in oil, CT

Comparison of Different Cytokine Conditions Reveals Resveratrol as a New Molecule for Ex Vivo Cultivation of Cord Blood-Derived Hematopoietic Stem Cells.

Science.gov (United States)

Heinz, Niels; Ehrnström, Birgitta; Schambach, Axel; Schwarzer, Adrian; Modlich, Ute; Schiedlmeier, Bernhard

2015-09-01

Human cord blood (CB)-derived hematopoietic stem cells (HSCs) are an interesting source for HSC transplantation. However, the number of collected CB-HSCs is often too low for one transplantation; therefore, ex vivo expansion of CB-HSCs is desirable. Current expansion protocols are based on the use of cytokine combinations, including insulin-like growth factor-binding protein 2 (IGFBP2) and angiopoietin-like proteins, or combinations with "small molecules" such as stemregenin-1. The aim of our project was to compare the potential of different CB-HSC expansion strategies side-by-side by phenotypical analysis in vitro and serial engraftment properties in NOD/SCID/IL2rg-/- (NSG) immunodeficient mice. We further identified resveratrol, a naturally occurring polyphenol, as a new, alternative small molecule combined with cytokines to facilitate serum-free ex vivo expansion of human CB-HSCs. The cultivation in resveratrol preserved the CB-HSC phenotype in vitro most efficiently and was ∼2 times more potent than commonly used cytokine conditions (including stem cell factor, thrombopoietin, Fms-related tyrosine kinase 3 ligand, interleukin-6) and the recently established serum-free culture, including IGFBP2 and angiopoietin-like 5. Serial transplantation studies further confirmed resveratrol to support robust multilineage engraftment in primary and secondary NSG recipients. Therefore, our work proposes resveratrol as a new small molecule for improved ex vivo culture and modification of human HSCs based on an efficient ex vivo propagation of the HSC fate. Human cord blood (CB)-derived hematopoietic stem cells (HSCs) are an important source for HSC transplantations but restricted in their usage because of their low numbers. In gene therapy, modifications of HSCs relies on their ex vivo modification without losing their stemness properties. Therefore, ex vivo cultivation and expansion of CB-HSCs is important for their effective application in HSC transplantation and gene

Hydralazine inhibits compression and acrolein-mediated injuries in ex vivo spinal cord.

Science.gov (United States)

Hamann, Kristin; Nehrt, Genevieve; Ouyang, Hui; Duerstock, Brad; Shi, Riyi

2008-02-01

We have previously shown that acrolein, a lipid peroxidation byproduct, is significantly increased following spinal cord injury in vivo, and that exposure to neuronal cells results in oxidative stress, mitochondrial dysfunction, increased membrane permeability, impaired axonal conductivity, and eventually cell death. Acrolein thus may be a key player in the pathogenesis of spinal cord injury, where lipid peroxidation is known to be involved. The current study demonstrates that the acrolein scavenger hydralazine protects against not only acrolein-mediated injury, but also compression in guinea pig spinal cord ex vivo. Specifically, hydralazine (500 mumol/L to 1 mmol/L) can significantly alleviate acrolein (100-500 mumol/L)-induced superoxide production, glutathione depletion, mitochondrial dysfunction, loss of membrane integrity, and reduced compound action potential conduction. Additionally, 500 mumol/L hydralazine significantly attenuated compression-mediated membrane disruptions at 2 and 3 h following injury. This was consistent with our findings that acrolein-lys adducts were increased following compression injury ex vivo, an effect that was prevented by hydralazine treatment. These findings provide further evidence for the role of acrolein in spinal cord injury, and suggest that acrolein-scavenging drugs such as hydralazine may represent a novel therapy to effectively reduce oxidative stress in disorders such as spinal cord injury and neurodegenerative diseases, where oxidative stress is known to play a role.

Investigation of allergenicity of some cosmetic mixtures by using ex vivo local lymph node assay-BrdU endpoints.

Science.gov (United States)

Ulker, Ozge Cemiloglu; Kaymak, Yesim; Karakaya, Asuman

2014-01-01

Balsam of Peru and fragrance mix are commonly used in cosmetic products. Allergy to fragrance is the most common cause of cosmetic contact dermatitis. In the present study, ex vivo local lymph node assay-5-bromo-2'-deoxyuridine (LLNA-BrdU) was used to evaluate the dermal sensitization potential of these cosmetic mixtures. The stimulation index values and estimated concentration (EC3) values were calculated and the potency classification was found for each mixture. At the same time, in order to measure the irritant effect without having to use additional animals, a combination of ex vivo LLNA-BrdU and the irritancy assay was conducted. Th1 [interleukin (IL)-2, interferon-γ] and Th2 cytokine (IL-4, IL-5) releases from lymph node cell culture were investigated as non-radioactive endpoints. According to the results of ex vivo LLNA-BrdU assays, EC3 values were found to be 3.09% (moderate) for balsam of Peru and 4.44% (moderate) for fragrance mix. Cytokine analysis results indicate that both Th1 and Th2 cytokines are involved in the regulation of murine contact allergy and can be considered as useful endpoints. In conclusion, according to our results, fragrance mix and balsam of Peru can be considered as moderate sensitizers; however, in high concentrations, both of them have irritation properties. The cytokines investigated can be considered as the endpoints of the ex vivo LLNA-BrdU assay. © 2014 S. Karger AG, Basel.

Anti-inflammatory activity of Punica granatum L. (Pomegranate) rind extracts applied topically to ex vivo skin.

Science.gov (United States)

Houston, David M J; Bugert, Joachim; Denyer, Stephen P; Heard, Charles M

2017-03-01

Coadministered pomegranate rind extract (PRE) and zinc (II) produces a potent virucidal activity against Herpes simplex virus (HSV); however, HSV infections are also associated with localised inflammation and pain. Here, the objective was to determine the anti-inflammatory activity and relative depth penetration of PRE, total pomegranate tannins (TPT) and zinc (II) in skin, ex vivo. PRE, TPT and ZnSO 4 were dosed onto freshly excised ex vivo porcine skin mounted in Franz diffusion cells and analysed for COX-2, as a marker for modulation of the arachidonic acid inflammation pathway, by Western blotting and immunohistochemistry. Tape stripping was carried out to construct relative depth profiles. Topical application of PRE to ex vivo skin downregulated expression of COX-2, which was significant after just 6h, and maintained for up to 24h. This was achieved with intact stratum corneum, proving that punicalagin penetrated skin, further supported by the depth profiling data. When PRE and ZnSO 4 were applied together, statistically equal downregulation of COX-2 was observed when compared to the application of PRE alone; no effect followed the application of ZnSO 4 alone. TPT downregulated COX-2 less than PRE, indicating that tannins alone may not be entirely responsible for the anti-inflammatory activity of PRE. Punicalagin was found throughout the skin, in particular the lower regions, indicating appendageal delivery as a significant route to the viable epidermis. Topical application of TPT and PRE had significant anti-inflammatory effects in ex vivo skin, confirming that PRE penetrates the skin and modulates COX-2 regulation in the viable epidermis. Pomegranates have potential as a novel approach in ameliorating the inflammation and pain associated with a range of skin conditions, including cold sores and herpetic stromal keratitis. Copyright © 2016 Elsevier B.V. All rights reserved.

Analysis of the ligand binding properties of recombinant bovine liver-type fatty acid binding protein

DEFF Research Database (Denmark)

Rolf, B; Oudenampsen-Krüger, E; Börchers, T

1995-01-01

The coding part of the cDNA for bovine liver-type fatty acid binding protein (L-FABP) has been amplified by RT-PCR, cloned and used for the construction of an Escherichia coli (E. coli) expression system. The recombinant protein made up to 25% of the soluble E. coli proteins and could be isolated...

Time- and temperature-dependent autolysis of urinary bladder epithelium during ex vivo preservation.

Science.gov (United States)

Erman, Andreja; Veranič, Peter

2011-07-01

Morphological and functional preservation of urinary bladder epithelium-urothelium after extirpation from an organism enables physiological studies of that tissue and provides the basis for successful organ transplantations. The aim of this study was to determine the optimal temperature for maintaining urothelium in ex vivo conditions. Mouse urinary bladders were kept at the three temperatures usually used for maintaining tissue during transportation: at the temperature of melting ice (1°C), at room temperature (22-24°C), and at the body temperature of most mammals (37°C). Autolytic structural changes were followed with electron microscopy, while destruction of cytoskeleton and intercellular junctions was observed by immunolabeling. The first ultrastructural changes, swelling of mitochondria and necrosis of individual cells, became evident 30 min after extirpation if the tissue was kept at 1°C. After 60 and 120 min in ex vivo conditions, the most severe changes with increasing plasma membrane ruptures were detected at 1°C, while at room temperature only mild changes were detected. At 37°C, the extent of ultrastructural changes was between those of the other two experimental temperatures. Autolytic destruction of cytoskeleton and intercellular junctions was not observed before 2 h after extirpation. After 4 h, severe degradation of cytokeratin 20 and microtubules were found at 1°C and 37°C, while being almost undisturbed at room temperature. On the other hand, the reduction of desmoplakin and ZO-1 labeling was more evident at 37°C than at 1°C and room temperature. These findings provide evidence that room temperature is most appropriate for short ex vivo preservation of urothelial tissue.

Parsley extract inhibits in vitro and ex vivo platelet aggregation and prolongs bleeding time in rats.

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Gadi, Dounia; Bnouham, Mohamed; Aziz, Mohammed; Ziyyat, Abderrahim; Legssyer, Abdelkhaleq; Legrand, Chantal; Lafeve, Françoise Fauvel; Mekhfi, Hassane

2009-08-17

Many cardiovascular diseases are associated with an increase in blood platelet activity. In Morocco, parsley (Petroselinum crispum, Apiaceae) is one of the medicinal herbs used to treat cardiovascular diseases such as arterial hypertension. In this study, crude aqueous extract (CAE) of parsley was evaluated for its anti-platelet activity in experimental animals on platelet aggregation in vitro and ex vivo; and on bleeding time in vivo. The in vitro aggregation was monitored after pre-incubation of platelets with CAE. The bleeding time and ex vivo aggregation were performed after oral treatment. CAE inhibited dose dependently platelet aggregation in vitro induced by thrombin, ADP, collagen and epinephrine. The oral administration of CAE (3g/kg) inhibited significantly (pparsley may be benefit in the normalization of platelet hyperactivation, in the nutritional prevention of cardiovascular diseases and are potentially interesting in the development of new prevention strategies.

Current Understanding of Acute Bovine Liver Disease in Australia

Directory of Open Access Journals (Sweden)

Elizabeth Read

2016-12-01

Full Text Available Acute bovine liver disease (ABLD is a hepatotoxicity principally of cattle which occurs in southern regions of Australia. Severely affected animals undergo rapid clinical progression with mortalities often occurring prior to the recognition of clinical signs. Less severely affected animals develop photosensitization and a proportion can develop liver failure. The characteristic histopathological lesion in acute fatal cases is severe, with acute necrosis of periportal hepatocytes with hemorrhage into the necrotic areas. Currently there are a small number of toxins that are known to cause periportal necrosis in cattle, although none of these have so far been linked to ABLD. Furthermore, ABLD has frequently been associated with the presence of rough dog’s tail grass (Cynosurus echinatus and Drechslera spp. fungi in the pasture system, but it is currently unknown if these are etiological factors. Much of the knowledge about ABLD is contained within case reports, with very little experimental research investigating the specific cause(s. This review provides an overview of the current and most recently published knowledge of ABLD. It also draws on wider research and unpublished reports to suggest possible fungi and mycotoxins that may give rise to ABLD.

Gravitational physiology of human immune cells: a review of in vivo, ex vivo and in vitro studies

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Cogoli, A.

1996-01-01

The study of the function of immune cells in microgravity has been studied for more than 20 years in several laboratories. It is clear today that the immune system is depressed in more than 50% of the astronauts during and after space flight and that the activation of T lymphocytes by mitogens in vitro changes dramatically. This article gives an overview of the gravitational studies conducted by our laboratory in Spacelab, in MIR station, in sounding rockets and on the ground in the clinostat and the centrifuge. Three experimental approaches are followed in our work: (i) Ex vivo studies are performed with blood samples drawn from astronauts; (ii) in vivo studies are based on the application of seven antigens to the skin of the astronauts; (iii) in vitro studies are carried out with immune cells purified from the blood of healthy donors (not astronauts). The data from our in vivo and ex vivo studies are in agreement with those of other laboratories and show that the immunological function is depressed in the majority of astronauts as a consequence of the stress of space flight rather than by a direct influence of gravity on the cell. Immune depression may become a critical hazard on long duration flights on space stations or to other planets. In vitro experiments show that cultures of free-floating lymphocytes and monocytes undergo a dramatic depression of activation by the mitogen concanavalin A, while activation is more than doubled when the cells are attached to microcarrier beads. Such effects may be attributed to both direct and indirect effects of gravitational unloading on basic biological mechanisms of the cell. While the in vitro data are very important to clarify certain aspects of the biological mechanism of T cells activation, they are not descriptive of the changes of the immunological function of the astronauts.

Ex vivo changes in blood glucose levels seldom change blood glucose control algorithm recommendations

NARCIS (Netherlands)

de Groene, L.; Harmsen, R. E.; Binnekade, J. M.; Spronk, P. E.; Schultz, M. J.

2010-01-01

Background. Hyperglycemia and glycemic variabilities are associated with adverse outcomes in critically ill patients. Blood glucose control with insulin mandates an adequate and precise assessment of blood glucose levels. Blood glucose levels, however, can change ex vivo after sampling. The aim of

Control of bovine hepatic fatty acid oxidation

International Nuclear Information System (INIS)

Jesse, B.W.; Emery, R.S.; Thomas, J.W.

1986-01-01

Fatty acid oxidation by bovine liver slices and mitochondria was examined to determine potential regulatory sites of fatty acid oxidation. Conversion of 1-[ 14 C]palmitate to 14 CO 2 and total [ 14 C]acid-soluble metabolites was used to measure fatty acid oxidation. Oxidation of palmitate (1 mM) was linear in both liver slice weight and incubation time. Carnitine stimulated palmitate oxidation; 2 mM dl-carnitine produced maximal stimulation of palmitate oxidation to both CO 2 and acid-soluble metabolites. Propionate (10 mM) inhibited palmitate oxidation by bovine liver slices. Propionate (.5 to 10 mM) had no effect on palmitate oxidation by mitochondria, but malonyl Coenzyme A, the first committed intermediate of fatty acid synthesis, inhibited mitochondrial palmitate oxidation (inhibition constant = .3 μM). Liver mitochonndrial carnitine palmitoyltransferase exhibited Michaelis constants for palmitoyl Coenzyme A and l-carnitine of 11.5 μM and .59 mM, respectively. Long-chain fatty acid oxidation in bovine liver is regulated by mechanisms similar to those in rats but adapted to the unique digestive physiology of the bovine

A non-covalent peptide-based strategy for ex vivo and in vivo oligonucleotide delivery.

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Crombez, Laurence; Morris, May C; Heitz, Frederic; Divita, Gilles

2011-01-01

The dramatic acceleration in identification of new nucleic acid-based therapeutic molecules such as short interfering RNA (siRNA) and peptide-nucleic acid (PNA) analogues has provided new perspectives for therapeutic targeting of specific genes responsible for pathological disorders. However, the poor cellular uptake of nucleic acids together with the low permeability of the cell membrane to negatively charged molecules remain major obstacles to their clinical development. Several non-viral strategies have been proposed to improve the delivery of synthetic short oligonucleotides both in cultured cells and in vivo. Cell-penetrating peptides constitute very promising tools for non-invasive cellular import of oligonucleotides and analogs. We recently described a non-covalent strategy based on short amphiphatic peptides (MPG8/PEP3) that have been successfully applied ex vivo and in vivo for the delivery of therapeutic siRNA and PNA molecules. PEP3 and MPG8 form stable nanoparticles with PNA analogues and siRNA, respectively, and promote their efficient cellular uptake, independently of the endosomal pathway, into a wide variety of cell lines, including primary and suspension lines, without any associated cytotoxicity. This chapter describes easy-to-handle protocols for the use of MPG-8 or PEP-3-nanoparticle technologies for PNA and siRNA delivery into adherent and suspension cell lines as well as in vivo into cancer mouse models.

Ex Vivo Methods for Informing Computational Models of the Mitral Valve

OpenAIRE

Bloodworth, Charles H.; Pierce, Eric L.; Easley, Thomas F.; Drach, Andrew; Khalighi, Amir H.; Toma, Milan; Jensen, Morten O.; Sacks, Michael S.; Yoganathan, Ajit P.

2016-01-01

Computational modeling of the mitral valve (MV) has potential applications for determining optimal MV repair techniques and risk of recurrent mitral regurgitation. Two key concerns for informing these models are (1) sensitivity of model performance to the accuracy of the input geometry, and, (2) acquisition of comprehensive data sets against which the simulation can be validated across clinically relevant geometries. Addressing the first concern, ex vivo micro-computed tomography (microCT) wa...

Novel ex vivo culture method for human monocytes uses shear flow to prevent total loss of transendothelial diapedesis function.

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Tsubota, Yoshiaki; Frey, Jeremy M; Raines, Elaine W

2014-01-01

Monocyte recruitment to inflammatory sites and their transendothelial migration into tissues are critical to homeostasis and pathogenesis of chronic inflammatory diseases. However, even short-term suspension culture of primary human monocytes leads to phenotypic changes. In this study, we characterize the functional effects of ex vivo monocyte culture on the steps involved in monocyte transendothelial migration. Our data demonstrate that monocyte diapedesis is impaired by as little as 4 h culture, and the locomotion step is subsequently compromised. After 16 h in culture, monocyte diapedesis is irreversibly reduced by ∼90%. However, maintenance of monocytes under conditions mimicking physiological flow (5-7.5 dyn/cm²) is sufficient to reduce diapedesis impairment significantly. Thus, through the application of shear during ex vivo culture of monocytes, our study establishes a novel protocol, allowing functional analyses of monocytes not currently possible under static culture conditions. These data further suggest that monocyte-based therapeutic applications may be measurably improved by alteration of ex vivo conditions before their use in patients.

Fluorescence Confocal Microscopy for Ex Vivo Diagnosis of Conjunctival Tumors: A Pilot Study.

Science.gov (United States)

Iovieno, Alfonso; Longo, Caterina; De Luca, Mariacarla; Piana, Simonetta; Fontana, Luigi; Ragazzi, Moira

2016-08-01

To evaluate the potential use of fluorescence confocal microscopy (FCM) for ex vivo diagnosis and excision margin assessment of conjunctival neoplasms. Validity study. setting: Single institution. Consecutive patients with clinically suspicious conjunctival lesions. Conjunctival lesions were excised in toto using a standard "no-touch technique" by a single surgeon (A.I.). Collected specimens were examined with a commercially available laser scanning fluorescence confocal microscope after immersion in a 0.6 mM solution of acridine orange dye for 10-20 seconds. Specimens were subsequently processed with standard histologic analysis. FCM diagnosis of the nature and extension of conjunctival lesions. Sixteen consecutive patients were included in the study (11 male, 5 female; mean age 58.1 ± 26.1 years, range 10-90 years). The median time needed to process and analyze a sample with FCM was 15 minutes. Eleven of 16 lesions were identified by FCM as squamous (2 benign papillomas, 2 grade 2 conjunctival intraepithelial neoplasias, 7 in situ squamous carcinomas) and 5 as nonsquamous (1 pingueculum, 1 dermolipoma, 2 melanocytic nevi, 1 melanoma). In all cases FCM was able to detect horizontal and vertical extension of the lesion. All FCM findings were confirmed by corresponding subsequent histologic examination. FCM provides a fast ex vivo preliminary diagnosis of suspicious conjunctival lesions with good histologic details and margin assessment, and may represent a novel tool for intraoperative and postsurgical management of conjunctival tumors. This is the first study to investigate ex vivo FCM application in ophthalmology. Copyright © 2016 Elsevier Inc. All rights reserved.

A Novel Ex Vivo Model to Investigate the Underlying Mechanisms in Alzheimer’s Disease

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Emanuele Brai

2017-09-01

Full Text Available Currently there is no widely accepted animal model reproducing the full pathological profile of Alzheimer’s disease (AD, since the basic mechanisms of neurodegeneration are still poorly understood. We have proposed that the interaction between the α7 nicotinic acetylcholine receptor (α7-nAChR and a recently discovered toxic peptide, cleaved from the acetylcholinesterase (AChE C-terminus, could account for the aberrant processes occurring in AD. In this article we describe a new application on ex vivo model procedure, which combines the advantages of both in vivo and in vitro preparations, to study the effects of the AChE-derived peptide on the rat basal forebrain (BF. Western blot analysis showed that the levels of α7-nAChR, p-Tau and Aβ are differentially expressed upon the AChE-peptide administration, in a selective site-dependent manner. In conclusion, this methodology demonstrates the action of a novel peptide in triggering an AD-like phenotype and proposes a new ex vivo approach for manipulating and monitoring neurochemical processes contributing to neurodegeneration, in a time-dependent and site-specific manner.

A marginal anticancer effect of regorafenib on pancreatic carcinoma cells in vitro, ex vivo, and in vivo.

Science.gov (United States)

Mayer, Barbara; Karakhanova, Svetlana; Bauer, Nathalie; Liu, Li; Zhu, Yifan; Philippov, Pavel P; Werner, Jens; Bazhin, Alexandr V

2017-11-01

Activation of receptor tyrosine kinases is recognized as a hallmark of cancer. Vascular endothelial growth factor (VEGF) and its receptor VEGFR are the prominent players in the induction of tumor neoangiogenesis. Strategies to inhibit VEGF and VEGFR are under intensive investigation in preclinical and clinical settings. Regorafenib is a multikinase inhibitor targeting some VEGFR and other receptor kinases. Preclinical results led to the FDA approval of regorafenib for treatment of metastatic colorectal cancer patients. Effects of this drug in pancreatic ductal adenocarcinoma (PDAC) have not been investigated yet. Gene expression was assessed with real-time PCR analysis. In vitro cell viability, proliferation, apoptosis, necrosis, migration, and invasion of the PDAC cells were assessed after regorafenib treatment. Ex vivo anti-tumor effects of regorafenib were investigated in a spheroid model of PDAC. In vivo anti-tumor effects of the drug were evaluated in a fertilized chicken egg model. In this work, we have demonstrated only a marginal anticancer effect of regorafenib in PDAC in vitro and ex vivo. However, in the egg model of PDAC, this drug reduced tumor volume. Besides, regorafenib is capable of modulating the expression of cancer stem cell (CSC) markers and epithelial-to-mesenchymal transition (EMT) markers on PDAC cells. We found out that effects of regorafenib on the expression of CSC and EMT markers are very heterogeneous and depend obviously on original expression of these markers. We concluded that regorafenib might be a potential drug for PDAC and it should be investigated in future clinical trials.

In vitro-ex vivo correlations between a cell-laden hydrogel and mucosal tissue for screening composite delivery systems.

Science.gov (United States)

Blakney, Anna K; Little, Adam B; Jiang, Yonghou; Woodrow, Kim A

2016-11-01

Composite delivery systems where drugs are electrospun in different layers and vary the drug stacking-order are posited to affect bioavailability. We evaluated how the formulation characteristics of both burst- and sustained-release electrospun fibers containing three physicochemically diverse drugs: dapivirine (DPV), maraviroc (MVC) and tenofovir (TFV) affect in vitro and ex vivo release. We developed a poly(hydroxyethyl methacrylate) (pHEMA) hydrogel release platform for the rapid, inexpensive in vitro evaluation of burst- and sustained-release topical or dermal drug delivery systems with varying microarchitecture. We investigated properties of the hydrogel that could recapitulate ex vivo release into nonhuman primate vaginal tissue. Using a dimethyl sulfoxide extraction protocol and high-performance liquid chromatography analysis, we achieved >93% recovery from the hydrogels and >88% recovery from tissue explants for all three drugs. We found that DPV loading, but not stacking order (layers of fiber containing a single drug) or microarchitecture (layers with isolated drug compared to all drugs in the same layer) impacted the burst release in vitro and ex vivo. Our burst-release formulations showed a correlation for DPV accumulation between the hydrogel and tissue (R 2 =   0.80), but the correlation was not significant for MVC or TFV. For the sustained-release formulations, the PLGA/PCL content did not affect TFV release in vitro or ex vivo. Incorporation of cells into the hydrogel matrix improved the correlation between hydrogel and tissue explant release for TFV. We expect that this hydrogel-tissue mimic may be a promising preclinical model to evaluate topical or transdermal drug delivery systems with complex microarchitectures.

Ex vivo expansion of hematopoietic stem cell by fusion protein TAT-Zfx

International Nuclear Information System (INIS)

Xu Chong; Zhang Yanbing; Jiang Hua

2009-01-01

The relative inability of hemopoietic stem cells (HSCs) to reproduce themselves (self-renew) ex vivo imposes substantial limitations on the current use of HSC transplantation. Recently, the transcription factor Zfx has been demonstrated that played an important in controlling the self-renewal of hematopoietic stem cells. Here, we reported that Zfx could enable high-level expansion of HSCs in vitro, by combination of protein transduction domain, TAT. Furthermore, we also demonstrated that expanded HSCs population retains their normal in vivo potential of pluripotency. It is thus that TAT-Zfx has the potential to expand HSCs significantly in vitro, and will have enormous clinical potentials.

Pulmonary ultrasound elastography: a feasibility study with phantoms and ex-vivo tissue

Science.gov (United States)

Nguyen, Man Minh; Xie, Hua; Paluch, Kamila; Stanton, Douglas; Ramachandran, Bharat

2013-03-01

Elastography has become widely used for minimally invasive diagnosis in many tumors as seen with breast, liver and prostate. Among different modalities, ultrasound-based elastography stands out due to its advantages including being safe, real-time, and relatively low-cost. While lung cancer is the leading cause of cancer mortality among both men and women, the use of ultrasound elastography for lung cancer diagnosis has hardly been investigated due to the limitations of ultrasound in air. In this work, we investigate the use of static-compression based endobronchial ultrasound elastography by a 3D trans-oesophageal echocardiography (TEE) transducer for lung cancer diagnosis. A water-filled balloon was designed to 1) improve the visualization of endobronchial ultrasound and 2) to induce compression via pumping motion inside the trachea and bronchiole. In a phantom study, we have successfully generated strain images indicating the stiffness difference between the gelatin background and agar inclusion. A similar strain ratio was confirmed with Philips ultrasound strain-based elastography product. For ex-vivo porcine lung study, different tissue ablation methods including chemical injection, Radio Frequency (RF) ablation, and direct heating were implemented to achieve tumor-mimicking tissue. Stiff ablated lung tissues were obtained and detected with our proposed method. These results suggest the feasibility of pulmonary elastography to differentiate stiff tumor tissue from normal tissue.

Ex vivo expansion of human umbilical cord blood hematopoietic stem and progenitor cells

NARCIS (Netherlands)

N. Kusadasi (Nuray)

2002-01-01

textabstractThe goal of this thesis research is to establish ex vivo expansion conditions for HSCs derived from UCB. To realize the expansion of HSCs, CD34+ or ACJ33+ UCB cells were cultured in the absence or presence of various cocktails of early acting cytokines including Flt3-L, Tpo, SCF or IL6

Development and utilization of an ex vivo bromodeoxyuridine local lymph node assay protocol for assessing potential chemical sensitizers.

Science.gov (United States)

Williams, W C; Copeland, C; Boykin, E; Quell, S J; Lehmann, D M

2015-01-01

The murine local lymph node assay (LLNA) is widely used to identify chemicals that may cause allergic contact dermatitis. Exposure to a dermal sensitizer results in proliferation of local lymph node T cells, which has traditionally been measured by in vivo incorporation of [(3) H]methyl thymidine. A more recent non-isotopic variation of the assay utilizes bromodeoxyuridine (BrdU) incorporation in vivo. To further improve the utility of this assay, we developed an ex vivo BrdU labeling procedure eliminating the need for in vivo injections. The results of this assay correctly identified a strong sensitizer (i.e., trimellitic anhydride) as well as weak/moderate sensitizers (i.e., eugenol, cinnamaldehyde and hexylcinnaminic aldehyde). As anticipated, neither non-sensitizers isopropanol and lactic acid nor the false negative chemical nickel II sulfate hexahydrate induced a positive threshold response in the assay. The results of this assay are in close agreement with those of the in vivo LLNA:BrdU-enzyme-linked immunosorbent assay labeling procedure. We also used the ex vivo BrdU LLNA procedure to evaluate ammonium hexachloroplatinate, ammonium tetrachloroplatinate and cis-diamminedichloroplatinum(II) and the assay correctly identified them as sensitizers based on the calculation of EC2 values. We conclude that this ex vivo BrdU labeling method offers predictive capacity comparable to previously established LLNA protocols while eliminating animal injections and the use of radioisotope. Published 2014. This article is a U.S. Government work and is in the public domain in the USA. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

Ex vivo MRI evaluation of prostate cancer: Localization and margin status prediction of prostate cancer in fresh radical prostatectomy specimens.

Science.gov (United States)

Heidkamp, Jan; Hoogenboom, Martijn; Kovacs, Iringo E; Veltien, Andor; Maat, Arie; Sedelaar, J P Michiel; Hulsbergen-van de Kaa, Christina A; Fütterer, Jurgen J

2018-02-01

To investigate the ability of high field ex vivo magnetic resonance imaging (MRI) to localize prostate cancer (PCa) and to predict the margin status in fresh radical prostatectomy (RP) specimens using histology as the reference standard. This Institutional Review Board (IRB)-approved study had written informed consent. Patients with biopsy-proved PCa and a diagnostic multiparametric 3T MRI examination of the prostate prior to undergoing RP were prospectively included. A custom-made container provided reference between the 7T ex vivo MRI obtained from fresh RP specimens and histological slicing. On ex vivo MRI, PCa was localized and the presence of positive surgical margins was determined in a double-reading session. These findings were compared with histological findings obtained from completely cut, whole-mount embedded, prostate specimens. In 12 RP specimens, histopathology revealed 36 PCa lesions, of which 17 (47%) and 20 (56%) were correlated with the ex vivo MRI in the first and second reading session, respectively. Nine of 12 (75%) index lesions were localized in the first session, in the second 10 of 12 (83%). Seven and 8 lesions of 11 lesions with Gleason score >6 and >0.5 cc were localized in the first and second session, respectively. In the first session none of the four histologically positive surgical margins (sensitivity 0%) and 9 of 13 negative margins (specificity 69%) were detected. In second session the sensitivity and specificity were 25% and 88%, respectively. Ex vivo MRI enabled accurate localization of PCa in fresh RP specimens, and the technique provided information on the margin status with high specificity. 1 Technical Efficacy: Stage 1 J. Magn. Reson. Imaging 2018;47:439-448. © 2017 International Society for Magnetic Resonance in Medicine.

Evaluation of non-radioactive endpoints of ex vivo local lymph node assay-BrdU to investigate select contact sensitizers.

Science.gov (United States)

Ulker, Ozge Cemiloglu; Ates, Ilker; Atak, Aysegul; Karakaya, Asuman

2013-01-01

The present study sought to verify the utility of the non-radioactive endpoints LLNA BrdU (5-bromo-2'-deoxyuridine) ex vivo incorporation and cytokine release using auricular lymph node cells isolated from BALB/c mice topically treated with a strong (formaldehyde or p-phenylene-diamine [PPD]), moderate sensitizer (cinnamal), or weak sensitizer (eugenol). Stimulation index (SI) and EC₃ values were calculated for each agent. Based on the results of ex vivo LLNA-BrdU assays, EC₃ values were calculated to be 0.29, 0.09, 1.91, and 16.60% for formaldehyde, PPD, cinnamal, and eugenol, respectively. These results were in good agreement with data from previous standard radioactive LLNA. Cytokine analyses indicated T(H)1 and T(H)2 cytokine involvement in the regulation of murine contact allergy and these could be utilized as endpoints in assessments of contact allergy in mice. In conclusion, the current study provided evidence that the non-radioactive endpoint LLNA BrdU ex vivo incorporation could be of use as a viable alternative approach to assess the skin sensitization potential of test compound with respect to improving animal welfare. This is of particular importance in the case of any laboratory where it might be difficult to handle and/or readily employ radioisotopes. Further studies will be required to confirm--across test agents--the reproducibility as well as the limits of utility of this new ex vivo BrdU method.

Laser-induced cartilage damage: an ex-vivo model using confocal microscopy

Science.gov (United States)

Frenz, Martin; Zueger, Benno J.; Monin, D.; Weiler, C.; Mainil-Varlet, P. M.; Weber, Heinz P.; Schaffner, Thomas

1999-06-01

Although there is an increasing popularity of lasers in orthopedic surgery, there is a growing concern about negative side effects of this therapy e.g. prolonged restitution time, radiation damage to adjacent cartilage or depth effects like bone necrosis. Despite case reports and experimental investigations over the last few years little is known about the extent of acute cartilage damage induced by different lasers types and energies. Histological examination offers only limited insights in cell viability and metabolism. Ho:YAG and Er:YAG lasers emitting at 2.1 micrometer and 2.94 micrometer, respectively, are ideally suited for tissue treatment because these wavelengths are strongly absorbed in water. The Purpose of the present study is to evaluate the effect of laser type and energy on chondrocyte viability in an ex vivo model. Free running Er:YAG (E equals 100 and 150 mJ) and Ho:YAG (E equals 500 and 800 mJ) lasers were used at different energy levels using a fixed pulse length of 400 microseconds. The energy was delivered at 8 Hz through optical fibers. Fresh bovine hyaline cartilage samples were mounted in a water bath at room temperature and the fiber was positioned at 30 degree and 180 degree angles relative to the tissue surface. After laser irradiation the samples were assessed by a life-dead cell viability test using a confocal microscope and by standard histology. Thermal damage was much deeper with Ho:YAG (up to 1800 micrometer) than with the Er:YAG laser (up to 70 micrometer). The cell viability test revealed a damage zone about twice the one determined by standard histology. Confocal microscopy is a powerful tool for assessing changes in tissue structure after laser treatment. In addition this technique allows to quantify these alterations without necessitating time consuming and expensive animal experiments.

Freesurfer cortical normative data for adults using Desikan-Killiany-Tourville and ex vivo protocols.

Science.gov (United States)

Potvin, Olivier; Dieumegarde, Louis; Duchesne, Simon

2017-08-01

We recently built normative data for FreeSurfer morphometric estimates of cortical regions using its default atlas parcellation (Desikan-Killiany or DK) according to individual and scanner characteristics. We aimed to produced similar normative values for Desikan-Killianny-Tourville (DKT) and ex vivo-based labeling protocols, as well as examine the differences between these three atlases. Surfaces, thicknesses, and volumes of cortical regions were produced using cross-sectional magnetic resonance scans from the same 2713 healthy individuals aged 18-94 years as used in the reported DK norms. Models predicting regional cortical estimates of each hemisphere were produced using age, sex, estimated intracranial volume (eTIV), scanner manufacturer and magnetic field strength (MFS) as predictors. The DKT and DK models generally included the same predictors and produced similar R 2 . Comparison between DK, DKT, ex vivo atlases normative cortical measures showed that the three protocols generally produced similar normative values. Copyright © 2017 Elsevier Inc. All rights reserved.

Optically Sectioned Imaging of Microvasculature of In-Vivo and Ex-Vivo Thick Tissue Models with Speckle-illumination HiLo Microscopy and HiLo Image Processing Implementation in MATLAB Architecture

Science.gov (United States)

Suen, Ricky Wai

The work described in this thesis covers the conversion of HiLo image processing into MATLAB architecture and the use of speckle-illumination HiLo microscopy for use of ex-vivo and in-vivo imaging of thick tissue models. HiLo microscopy is a wide-field fluorescence imaging technique and has been demonstrated to produce optically sectioned images comparable to confocal in thin samples. The imaging technique was developed by Jerome Mertz and the Boston University Biomicroscopy Lab and has been implemented in our lab as a stand-alone optical setup and a modification to a conventional fluorescence microscope. Speckle-illumination HiLo microscopy combines two images taken under speckle-illumination and standard uniform-illumination to generate an optically sectioned image that reject out-of-focus fluorescence. The evaluated speckle contrast in the images is used as a weighting function where elements that move out-of-focus have a speckle contrast that decays to zero. The experiments shown here demonstrate the capability of our HiLo microscopes to produce optically-sectioned images of the microvasculature of ex-vivo and in-vivo thick tissue models. The HiLo microscope were used to image the microvasculature of ex-vivo mouse heart sections prepared for optical histology and the microvasculature of in-vivo rodent dorsal window chamber models. Studies in label-free surface profiling with HiLo microscopy is also presented.

Development, standardization and testing of a bacterial wound infection model based on ex vivo human skin.

Directory of Open Access Journals (Sweden)

Christoph Schaudinn

Full Text Available Current research on wound infections is primarily conducted on animal models, which limits direct transferability of these studies to humans. Some of these limitations can be overcome by using-otherwise discarded-skin from cosmetic surgeries. Superficial wounds are induced in fresh ex vivo skin, followed by intradermal injection of Pseudomonas aeruginosa under the wound. Subsequently, the infected skin is incubated for 20 hours at 37°C and the CFU/wound are determined. Within 20 hours, the bacteria count increased from 107 to 109 bacteria per wound, while microscopy revealed a dense bacterial community in the collagen network of the upper wound layers as well as numerous bacteria scattered in the dermis. At the same time, IL-1alpha and IL-1beta amounts increased in all infected wounds, while-due to bacteria-induced cell lysis-the IL-6 and IL-8 concentrations rose only in the uninfected samples. High-dosage ciprofloxacin treatment resulted in a decisive decrease in bacteria, but consistently failed to eradicate all bacteria. The main benefits of the ex vivo wound model are the use of healthy human skin, a quantifiable bacterial infection, a measureable donor-dependent immune response and a good repeatability of the results. These properties turn the ex vivo wound model into a valuable tool to examine the mechanisms of host-pathogen interactions and to test antimicrobial agents.

Comparison of In-Vitro and Ex-Vivo Wound Healing Assays for the Investigation of Diabetic Wound Healing and Demonstration of a Beneficial Effect of a Triterpene Extract.

Directory of Open Access Journals (Sweden)

Christopher Ueck

Full Text Available Diabetes mellitus is a frequent cause for chronic, difficult-to-treat wounds. New therapies for diabetic wounds are urgently needed and in-vitro or ex-vivo test systems are essential for the initial identification of new active molecules. The aim of this study is to compare in-vitro and ex-vivo test systems for their usability for early drug screening and to investigate the efficacy of a birch bark triterpene extract (TE that has been proven ex-vivo and clinically to accelerate non-diabetic wound healing (WH, in a diabetic context. We investigated in-vitro models for diabetic WH, i.e. scratch assays with human keratinocytes from diabetic donors or cultured under hyperglycaemic conditions and a newly developed porcine ex-vivo hyperglycaemic WH model for their potential to mimic delayed diabetic WH and for the influence of TE in these test systems. We show that keratinocytes from diabetic donors often fail to exhibit significantly delayed WH. For cells under hyperglycaemic conditions significant decrease is observed but is influenced by choice of medium and presence of supplements. Also, donor age plays a role. Interestingly, hyperglycaemic effects are mainly hyperosmolaric effects in scratch assays. Ex-vivo models under hyperglycaemic conditions show a clear and substantial decrease of WH, and here both glucose and hyperosmolarity effects are involved. Finally, we provide evidence that TE is also beneficial for ex-vivo hyperglycaemic WH, resulting in significantly increased length of regenerated epidermis to 188±16% and 183±11% (SEM; p<0.05 compared to controls when using two different TE formulations. In conclusion, our results suggest that microenvironmental influences are important in WH test systems and that therefore the more complex hyperglycaemic ex-vivo model is more suitable for early drug screening. Limitations of the in-vitro and ex-vivo models are discussed. Furthermore our data recommend TE as a promising candidate for in-vivo

Soft tissue influence on ex vivo mobility in the hip of Iguana: comparison with in vivo movement and its bearing on joint motion of fossil sprawling tetrapods.

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Arnold, Patrick; Fischer, Martin S; Nyakatura, John A

2014-07-01

The reconstruction of a joint's maximum range of mobility (ROM) often is a first step when trying to understand the locomotion of fossil tetrapods. But previous studies suggest that the ROM of a joint is restricted by soft tissues surrounding the joint. To expand the limited informative value of ROM studies for the reconstruction of a fossil species' locomotor characteristics, it is moreover necessary to better understand the relationship of ex vivo ROM with the actual in vivo joint movement. To gain insight into the relationship between ex vivo mobility and in vivo movement, we systematically tested for the influence of soft tissues on joint ROM in the hip of the modern lizard Iguana iguana. Then, we compared the ex vivo mobility to in vivo kinematics of the hip joint in the same specimens using X-ray sequences of steady-state treadmill locomotion previously recorded. With stepwise removal of soft tissues and a repeated-measurement protocol, we show that soft tissues surrounding the hip joint considerably limit ROM, highlighting the problems when joint ROM is deduced from bare bones only. We found the integument to have the largest effect on the range of long-axis rotation, pro- and retraction. Importantly, during locomotion the iguana used only a fragment of the ROM that was measured in our least restrictive dissection situation (i.e. pelvis and femur only conjoined by ligaments), demonstrating the discrepancy between hip joint ROM and actual in vivo movement. Our study emphasizes the necessity for caution when attempting to reconstruct joint ROM or even locomotor kinematics from fossil bones only, as actual in vivo movement cannot be deduced directly from any condition of cadaver mobility in Iguana and likely in other tetrapods. © 2014 Anatomical Society.

Exploring sex differences in the adult zebra finch brain: In vivo diffusion tensor imaging and ex vivo super-resolution track density imaging.

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Hamaide, Julie; De Groof, Geert; Van Steenkiste, Gwendolyn; Jeurissen, Ben; Van Audekerke, Johan; Naeyaert, Maarten; Van Ruijssevelt, Lisbeth; Cornil, Charlotte; Sijbers, Jan; Verhoye, Marleen; Van der Linden, Annemie

2017-02-01

Zebra finches are an excellent model to study the process of vocal learning, a complex socially-learned tool of communication that forms the basis of spoken human language. So far, structural investigation of the zebra finch brain has been performed ex vivo using invasive methods such as histology. These methods are highly specific, however, they strongly interfere with performing whole-brain analyses and exclude longitudinal studies aimed at establishing causal correlations between neuroplastic events and specific behavioral performances. Therefore, the aim of the current study was to implement an in vivo Diffusion Tensor Imaging (DTI) protocol sensitive enough to detect structural sex differences in the adult zebra finch brain. Voxel-wise comparison of male and female DTI parameter maps shows clear differences in several components of the song control system (i.e. Area X surroundings, the high vocal center (HVC) and the lateral magnocellular nucleus of the anterior nidopallium (LMAN)), which corroborate previous findings and are in line with the clear behavioral difference as only males sing. Furthermore, to obtain additional insights into the 3-dimensional organization of the zebra finch brain and clarify findings obtained by the in vivo study, ex vivo DTI data of the male and female brain were acquired as well, using a recently established super-resolution reconstruction (SRR) imaging strategy. Interestingly, the SRR-DTI approach led to a marked reduction in acquisition time without interfering with the (spatial and angular) resolution and SNR which enabled to acquire a data set characterized by a 78μm isotropic resolution including 90 diffusion gradient directions within 44h of scanning time. Based on the reconstructed SRR-DTI maps, whole brain probabilistic Track Density Imaging (TDI) was performed for the purpose of super resolved track density imaging, further pushing the resolution up to 40μm isotropic. The DTI and TDI maps realized atlas

Integrity and stability of oral liposomes containing bile salts studied in simulated and ex vivo gastrointestinal media.

Science.gov (United States)

Hu, Shunwen; Niu, Mengmeng; Hu, Fuqiang; Lu, Yi; Qi, Jianping; Yin, Zongning; Wu, Wei

2013-01-30

The objective of this study was to investigate the integrtity and stability of oral liposomes containing glycocholate (SGC-Lip) in simulated gastrointestinal (GI) media and ex vivo GI media from rats in comparison with conventional liposomes (CH-Lip) composed of soybean phosphatidylcholine and cholesterol. Membrane integrity of liposomes was evaluated by monitoring calcein release, particle size and distribution in different simulated GI media. The stability of liposomes encapsulating insulin was investigated in simulated GI fluids containing pepsin or pancreatin and ex vivo GI enzyme fluids. Simulated GI media with low pH or physiological bile salts resulted in significant increase in calcein release, but dynamic laser scattering data showed that the size and distribution were generally stable. SGC-Lip retained the major amount of the initially encapsulated insulin as compared with CH-Lip in simulated GI fluids (SGF, FaSSGF, SIF and FeSSIF-V2). SGC-Lip retained respectively 17.1% and 20.5% of the initially encapsulated insulin in ex vivo GI fluid, which were also significantly more than CH-Lip. These results suggested that SGC-Lip could protect insulin from degradation to some degree during their transit through the gastrointestinal tract and contributed to enhanced oral absorption. Copyright © 2012 Elsevier B.V. All rights reserved.

Liver Effects of Clinical Drugs Differentiated in Human Liver Slices

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Alison E. M. Vickers

2017-03-01

Full Text Available Drugs with clinical adverse effects are compared in an ex vivo 3-dimensional multi-cellular human liver slice model. Functional markers of oxidative stress and mitochondrial function, glutathione GSH and ATP levels, were affected by acetaminophen (APAP, 1 mM, diclofenac (DCF, 1 mM and etomoxir (ETM, 100 μM. Drugs targeting mitochondria more than GSH were dantrolene (DTL, 10 μM and cyclosporin A (CSA, 10 μM, while GSH was affected more than ATP by methimazole (MMI, 500 μM, terbinafine (TBF, 100 μM, and carbamazepine (CBZ 100 μM. Oxidative stress genes were affected by TBF (18%, CBZ, APAP, and ETM (12%–11%, and mitochondrial genes were altered by CBZ, APAP, MMI, and ETM (8%–6%. Apoptosis genes were affected by DCF (14%, while apoptosis plus necrosis were altered by APAP and ETM (15%. Activation of oxidative stress, mitochondrial energy, heat shock, ER stress, apoptosis, necrosis, DNA damage, immune and inflammation genes ranked CSA (75%, ETM (66%, DCF, TBF, MMI (61%–60%, APAP, CBZ (57%–56%, and DTL (48%. Gene changes in fatty acid metabolism, cholestasis, immune and inflammation were affected by DTL (51%, CBZ and ETM (44%–43%, APAP and DCF (40%–38%, MMI, TBF and CSA (37%–35%. This model advances multiple dosing in a human ex vivo model, plus functional markers and gene profile markers of drug induced human liver side-effects.

Ex-vivo α-galactosylceramide activation of NKT cells in humans and macaques.

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Fernandez, Caroline S; Cameron, Garth; Godfrey, Dale I; Kent, Stephen J

2012-08-31

NKT cells are key mediators of antiviral and anticancer immunity. Experiments in mice have demonstrated that activation of NKT cells in vivo induces the expression of multiple effector molecules critical to successful immunity. Human clinical trials have shown similar responses, although in vivo activation of NKT cells in humans or primate models are far more limited in number and scope. Measuring ex vivo activation of NKT cells by the CD1d-restricted glycolipid ligand α-Galactosylceramide (α-GalCer) through cytokine expression profiles is a useful marker of NKT cell function, but for reasons that are unclear, this approach does not appear to work as well in humans and non-human primate macaque models in comparison to mice. We performed a series of experiments on human and macaque (Macaca nemestrina) fresh whole blood samples to define optimal conditions to detect NKT cell cytokine (TNF, IFNγ, IL-2) and degranulation marker (CD107a) expression by flow cytometry. We found that conditions previously described for mouse splenocyte NKT cell activation were suboptimal on human or macaque blood NKT cells. In contrast, a 6h incubation with brefeldin A added for the last 4h, in a 96-well plate based assay, and using an α-GalCer concentration of 1 μg/ml were optimal methods to stimulate NKT cells in fresh blood from both humans and macaques. Unexpectedly, we noted that blood NKT cells from macaques infected with SIV were more readily activated by α-GalCer than NKT cells from uninfected macaques, suggesting that SIV infection may have primed the NKT cells. In conclusion, we describe optimized methods for the ex vivo antigen-specific activation of human and macaque blood NKT cells. These assays should be useful in monitoring NKT cells in disease and in immunotherapy studies. Copyright © 2012 Elsevier B.V. All rights reserved.

Interleukin-7 facilitates HIV-1 transmission to cervico-vaginal tissue ex vivo.

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Andrea Introini

2013-02-01

Full Text Available The majority of HIV-1 infections in women occur through vaginal intercourse, in which virus-containing semen is deposited on the cervico-vaginal mucosa. Semen is more than a mere carrier of HIV-1, since it contains many biological factors, in particular cytokines, that may affect HIV-1 transmission. The concentration of interleukin (IL-7, one of the most prominent cytokines in semen of healthy individuals, is further increased in semen of HIV-1-infected men. Here, we investigated the potential role of IL-7 in HIV-1 vaginal transmission in an ex vivo system of human cervico-vaginal tissue. We simulated an in vivo situation by depositing HIV-1 on cervico-vaginal tissue in combination with IL-7 at concentrations comparable with those measured in semen of HIV-1-infected individuals. We found that IL-7 significantly enhanced virus replication in ex vivo infected cervico-vaginal tissue. Similarly, we observed an enhancement of HIV-1 replication in lymphoid tissue explants. Analysis of T cells isolated from infected tissues showed that IL-7 reduced CD4⁺ T cell depletion preventing apoptosis, as shown by the decrease in the number of cells expressing the apoptotic marker APO2.7 and the increase in the expression of the anti-apoptotic protein B-cell lymphoma (Bcl-2. Also, IL-7 increased the fraction of cycling CD4⁺ T cells, as evidenced by staining for the nuclear factor Ki-67. High levels of seminal IL-7 in vivo may be relevant to the survival of the founder pool of HIV-1-infected cells in the cervico-vaginal mucosa at the initial stage of infection, promoting local expansion and dissemination of HIV infection.

Persistence of DNA studied in different ex vivo and in vivo rat models simulating the human gut situation

DEFF Research Database (Denmark)

Wilcks, Andrea; van Hoek, A.H.A.M.; Joosten, R.G.

2004-01-01

This study aimed to evaluate the possibility of DNA sequences from genetically modified plants to persist in the gastrointestinal (GI) tract. PCR analysis and transformation assays were used to study DNA persistence and integrity in various ex vivo and in vivo systems using gnotobiotic rats. DNA......, plasmid DNA could be recovered throughout the GI tract when intestinal samples were taken up to 5 h after feeding rats with plasmid. Furthermore, DNA isolated from these intestinal samples was able to transform electro-competent Escherichia coli, showing that the plasmid was still biologically active....... The results indicate that ingested DNA may persist in the GI tract and consequently may be present for uptake by intestinal bacteria....

A novel monoclonal antibody of human stem cell factor inhibits umbilical cord blood stem cell ex vivo expansion

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Fan Jie

2012-12-01

Full Text Available Abstract Stem cell factor (SCF activates hematopoietic stem cell (HSC self-renewal and is being used to stimulate the ex vivo expansion of HSCs. The mechanism by which SCF supports expansion of HSCs remains poorly understood. In cord blood ex vivo expansion assays, a newly produced anti-SCF monoclonal antibody (clone 23C8 was found to significantly inhibit the expansion of CD34+ cells. This antibody appears to bind directly to a part of SCF that is critical for biological activity toward expansion of CD34+ cells, which is located in the first 104 amino acids from the NH2-terminus.

Comprehensive Antiretroviral Restriction Factor Profiling Reveals the Evolutionary Imprint of the ex Vivo and in Vivo IFN-β Response in HTLV-1-Associated Neuroinflammation.

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Leal, Fabio E; Menezes, Soraya Maria; Costa, Emanuela A S; Brailey, Phillip M; Gama, Lucio; Segurado, Aluisio C; Kallas, Esper G; Nixon, Douglas F; Dierckx, Tim; Khouri, Ricardo; Vercauteren, Jurgen; Galvão-Castro, Bernardo; Saraiva Raposo, Rui Andre; Van Weyenbergh, Johan

2018-01-01

HTLV-1-Associated Myelopathy (HAM/TSP) is a progressive neuroinflammatory disorder for which no disease-modifying treatment exists. Modest clinical benefit from type I interferons (IFN-α/β) in HAM/TSP contrasts with its recently identified IFN-inducible gene signature. In addition, IFN-α treatment in vivo decreases proviral load and immune activation in HAM/TSP, whereas IFN-β therapy decreases tax mRNA and lymphoproliferation. We hypothesize this "IFN paradox" in HAM/TSP might be explained by both cell type- and gene-specific effects of type I IFN in HTLV-1-associated pathogenesis. Therefore, we analyzed ex vivo transcriptomes of CD4 + T cells, PBMCs and whole blood in healthy controls, HTLV-1-infected individuals, and HAM/TSP patients. First, we used a targeted approach, simultaneously quantifying HTLV-1 mRNA (HBZ, Tax), proviral load and 42 host genes with known antiretroviral (anti-HIV) activity in purified CD4 + T cells. This revealed two major clusters ("antiviral/protective" vs. "proviral/deleterious"), as evidenced by significant negative (TRIM5/TRIM22/BST2) vs. positive correlation (ISG15/PAF1/CDKN1A) with HTLV-1 viral markers and clinical status. Surprisingly, we found a significant inversion of antiretroviral activity of host restriction factors, as evidenced by opposite correlation to in vivo HIV-1 vs. HTLV-1 RNA levels. The anti-HTLV-1 effect of antiviral cluster genes was significantly correlated to their adaptive chimp/human evolution score, for both Tax mRNA and PVL. Six genes of the proposed antiviral cluster underwent lentivirus-driven purifying selection during primate evolution (TRIM5/TRIM22/BST2/APOBEC3F-G-H), underscoring the cross-retroviral evolutionary imprint. Secondly, we examined the genome-wide type I IFN response in HAM/TSP patients, following short-term ex vivo culture of PBMCs with either IFN-α or IFN-β. Microarray analysis evidenced 12 antiretroviral genes (including TRIM5α/TRIM22/BST2) were significantly up-regulated by IFN

PVA matches human liver in needle-tissue interaction.

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de Jong, Tonke L; Pluymen, Loes H; van Gerwen, Dennis J; Kleinrensink, Gert-Jan; Dankelman, Jenny; van den Dobbelsteen, John J

2017-05-01

Medical phantoms can be used to study needle-tissue interaction and to train medical residents. The purpose of this research is to study the suitability of polyvinyl alcohol (PVA) as a liver tissue mimicking material in terms of needle-tissue interaction. Insertions into ex-vivo human livers were used for reference. Six PVA samples were created by varying the mass percentage of PVA to water (4m% and 7m%) and the number of freeze-thaw cycles (1, 2 and 3 cycles, 16hours of freezing at -19°C, 8hours of thawing). The inner needle of an 18 Gauge trocar needle with triangular tip was inserted 13 times into each of the samples, using an insertion velocity of 5 mm/s. In addition, 39 insertions were performed in two ex-vivo human livers. Axial forces on the needle were captured during insertion and retraction and characterized by friction along the needle shaft, peak forces, and number of peak forces per unit length. The concentration of PVA and the number of freeze-thaw cycles both influenced the mechanical interaction between needle and specimen. Insertions into 4m% PVA phantoms with 2 freeze-thaw cycles were comparable to human liver in terms of estimated friction along the needle shaft and the number of peak forces. Therefore, these phantoms are considered to be suitable liver mimicking materials for image-guided needle interventions. The mechanical properties of PVA hydrogels can be influenced in a controlled manner by varying the concentration of PVA and the number of freeze-thaw cycles, to mimic liver tissue characteristics. Copyright © 2017 Elsevier Ltd. All rights reserved.

Lyophilized insulin nanoparticles prepared from quaternized N-aryl derivatives of chitosan as a new strategy for oral delivery of insulin: in vitro, ex vivo and in vivo characterizations.

Science.gov (United States)

Mahjub, Reza; Radmehr, Moojan; Dorkoosh, Farid Abedin; Ostad, Seyed Naser; Rafiee-Tehrani, Morteza

2014-12-01

The purpose of this research was the development, in vitro, ex vivo and in vivo characterization of lyophilized insulin nanoparticles prepared from quaternized N-aryl derivatives of chitosan. Insulin nanoparticles were prepared from methylated N-(4-N,N-dimethylaminobenzyl), methylated N-(4 pyridinyl) and methylated N-(benzyl). Insulin nanoparticles containing non-modified chitosan and also trimethyl chiotsan (TMC) were also prepared as control. The effects of the freeze-drying process on physico-chemical properties of nanoparticles were investigated. The release of insulin from the nanoparticles was studied in vitro. The mechanism of the release of insulin from different types of nanoparticles was determined using curve fitting. The secondary structure of the insulin released from the nanoparticles was analyzed using circular dichroism and the cell cytotoxicity of nanoparticles on a Caco-2 cell line was determined. Ex vivo studies were performed on excised rat jejunum using Frantz diffusion cells. In vivo studies were performed on diabetic male Wistar rats and blood glucose level and insulin serum concentration were determined. Optimized nanoparticles with proper physico-chemical properties were obtained. The lyophilization process was found to cause a decrease in zeta potential and an increase in PdI as well as and a decrease in entrapment efficiency (EE%) and loading efficiency (LE%) but conservation in size of nanoparticles. Atomic force microscopy (AFM) images showed non-aggregated, stable and spherical to sub-spherical nanoparticles. The in vitro release study revealed higher release rates for lyophilized compared to non-lyophilized nanoparticles. Cytotoxicity studies on Caco-2 cells revealed no significant cytotoxicity for prepared nanoparticles after 3-h post-incubation but did show the concentration-dependent cytotoxicity after 24 h. The percentage of cumulative insulin determined from ex vivo studies was significantly higher in nanoparticles prepared

Pleiotropic functions of magnetic nanoparticles for ex vivo gene transfer.

Science.gov (United States)

Kami, Daisuke; Kitani, Tomoya; Kishida, Tsunao; Mazda, Osam; Toyoda, Masashi; Tomitaka, Asahi; Ota, Satoshi; Ishii, Ryuga; Takemura, Yasushi; Watanabe, Masatoshi; Umezawa, Akihiro; Gojo, Satoshi

2014-08-01

Gene transfer technique has various applications, ranging from cellular biology to medical treatments for diseases. Although nonviral vectors, such as episomal vectors, have been developed, it is necessary to improve their gene transfer efficacy. Therefore, we attempted to develop a highly efficient gene delivery system combining an episomal vector with magnetic nanoparticles (MNPs). In comparison with the conventional method using transfection reagents, polyethylenimine-coated MNPs introduced episomal vectors more efficiently under a magnetic field and could express the gene in mammalian cells with higher efficiency and for longer periods. This novel in vitro separation method of gene-introduced cells utilizing the magnetic property of MNPs significantly facilitated the separation of cells of interest. Transplanted cells in vivo were detected using magnetic resonance. These results suggest that MNPs play multifunctional roles in ex vivo gene transfer, such as improvement of gene transfer efficacy, separation of cells, and detection of transplanted cells. This study convincingly demonstrates enhanced efficiency of gene transfer via magnetic nanoparticles. The method also enables magnetic sorting of cells positive for the transferred gene, and in vivo monitoring of the process with MRI. Copyright © 2014 Elsevier Inc. All rights reserved.

In vivo migration of labeled autologous natural killer cells to liver metastases in patients with colon carcinoma

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Satolli Maria A

2006-11-01

Full Text Available Abstract Background Besides being the effectors of native anti-tumor cytotoxicity, NK cells participate in T-lymphocyte responses by promoting the maturation of dendritic cells (DC. Adherent NK (A-NK cells constitute a subset of IL-2-stimulated NK cells which show increased expression of integrins and the ability to adhere to solid surface and to migrate, infiltrate, and destroy cancer. A critical issue in therapy of metastatic disease is the optimization of NK cell migration to tumor tissues and their persistence therein. This study compares localization to liver metastases of autologous A-NK cells administered via the systemic (intravenous, i.v. versus locoregional (intraarterial, i.a. routes. Patients and methods A-NK cells expanded ex-vivo with IL-2 and labeled with 111In-oxine were injected i.a. in the liver of three colon carcinoma patients. After 30 days, each patient had a new preparation of 111In-A-NK cells injected i.v. Migration of these cells to various organs was evaluated by SPET and their differential localization to normal and neoplastic liver was demonstrated after i.v. injection of 99mTc-phytate. Results A-NK cells expressed a donor-dependent CD56+CD16+CD3- (NK or CD56+CD16+CD3+ (NKT phenotype. When injected i.v., these cells localized to the lung before being visible in the spleen and liver. By contrast, localization of i.a. injected A-NK cells was virtually confined to the spleen and liver. Binding of A-NK cells to liver neoplastic tissues was observed only after i.a. injections. Conclusion This unique study design demonstrates that A-NK cells adoptively transferred to the liver via the intraarterial route have preferential access and substantial accumulation to the tumor site.

Gene expression and apoptosis in bovine embryos during in vitro culture and in vivo development

NARCIS (Netherlands)

Knijn, H.W.

2004-01-01

The first attempts to fertilise in vitro bovine oocytes were done in the late sixties but only in 1982 the first calf was born after transplantation of a complete in vitro produced embryo. Since then the in vitro production system improved a lot but it is still impossible to mimic the in vivo

In vivo time-harmonic multifrequency elastography of the human liver

International Nuclear Information System (INIS)

Tzschätzsch, Heiko; Guo, Jing; Streitberger, Kaspar-Josche; Fischer, Thomas; Sack, Ingolf; Ipek-Ugay, Selcan; Braun, Jürgen; Gentz, Enno; Klaua, Robert; Schultz, Michael

2014-01-01

Elastography is capable of noninvasively detecting hepatic fibrosis by imposing mechanical stress and measuring the viscoelastic response in the liver. Magnetic resonance elastography (MRE) relies on time-harmonic vibrations, while most dynamic ultrasound elastography methods employ transient stimulation methods. This study attempts to benefit from the advantages of time-harmonic tissue stimulation, i.e. relative insensitivity to obesity and ascites and mechanical approachability of the entire liver, and the advantages of ultrasound, i.e. time efficiency, low costs, and wide availability, by introducing in vivo time-harmonic elastography (THE) of the human liver using ultrasound and a broad range of harmonic stimulation frequencies. THE employs continuous harmonic shear vibrations at 7 frequencies from 30 to 60 Hz in a single examination and determines the elasticity and the viscosity of the liver from the dispersion of the shear wave speed within the applied frequency range. The feasibility of the method is demonstrated in the livers of eight healthy volunteers and a patient with cirrhosis. Multifrequency MRE at the same drive frequencies was used as elastographic reference method. Similar values of shear modulus and shear viscosity according the Kelvin–Voigt model were obtained by MRE and THE, indicating that the new method is suitable for in vivo quantification of the shear viscoelastic properties of the liver, however, in real-time and at a fraction of the costs of MRE. In conclusion, THE may provide a useful tool for fast assessment of the viscoelastic properties of the liver at low costs and without limitations in obesity, ascites or hemochromatosis. (paper)

Anti-Bovine Programmed Death-1 Rat–Bovine Chimeric Antibody for Immunotherapy of Bovine Leukemia Virus Infection in Cattle

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Tomohiro Okagawa

2017-06-01

Full Text Available Blockade of immunoinhibitory molecules, such as programmed death-1 (PD-1/PD-ligand 1 (PD-L1, is a promising strategy for reinvigorating exhausted T cells and preventing disease progression in a variety of chronic infections. Application of this therapeutic strategy to cattle requires bovinized chimeric antibody targeting immunoinhibitory molecules. In this study, anti-bovine PD-1 rat–bovine chimeric monoclonal antibody 5D2 (Boch5D2 was constructed with mammalian expression systems, and its biochemical function and antiviral effect were characterized in vitro and in vivo using cattle infected with bovine leukemia virus (BLV. Purified Boch5D2 was capable of detecting bovine PD-1 molecules expressed on cell membranes in flow cytometric analysis. In particular, Biacore analysis determined that the binding affinity of Boch5D2 to bovine PD-1 protein was similar to that of the original anti-bovine PD-1 rat monoclonal antibody 5D2. Boch5D2 was also capable of blocking PD-1/PD-L1 binding at the same level as 5D2. The immunomodulatory and therapeutic effects of Boch5D2 were evaluated by in vivo administration of the antibody to a BLV-infected calf. Inoculated Boch5D2 was sustained in the serum for a longer period. Boch5D2 inoculation resulted in activation of the proliferation of BLV-specific CD4+ T cells and decrease in the proviral load of BLV in the peripheral blood. This study demonstrates that Boch5D2 retains an equivalent biochemical function to that of the original antibody 5D2 and is a candidate therapeutic agent for regulating antiviral immune response in vivo. Clinical efficacy of PD-1/PD-L1 blockade awaits further experimentation with a large number of animals.

In vitro, ex vivo and in vivo examination of buccal absorption of metoprolol with varying pH in TR146 cell culture, porcine buccal mucosa and Göttingen minipigs

DEFF Research Database (Denmark)

Holm, René; Meng-Lund, Emil; Andersen, Morten B.

2013-01-01

This work studied the buccal absorption of metoprolol in vitro, ex vivo and in vivo as a function of buffered pH at 7.4, 8.5, 9.0 and 9.5. Permeability studies showed a correlation (r(2)=0.92) between in vitro TR146 cell culture and ex vivo porcine buccal mucosa in a modified Ussing chamber...... was obtained after buccal dosing (58-107%) compared to oral (3%) administration, ranging 58-107% and 3%, respectively. Macroscopically, no local toxic effects were observed by visual inspection of mini-pig cheeks. A very clear level C in vitro in vivo correlation (r(2)=0.98) was obtained between the observed....... A higher apparent permeability was observed at higher pH values, i.e. the more compound that was unionised the higher the permeability. In vivo studies were conducted in anaesthetised Göttingen mini-pigs. A clear influence of pH on the absorption was seen and a significant higher absolute bioavailability...

Radiation effects on vitamin A and {beta}-carotene contents in bovine liver and swine pate de foie; Efeito da radiacao gama sobre a vitamina A e o {beta}-caroteno de figado bovino e suino

Energy Technology Data Exchange (ETDEWEB)

Taipina, Magda Sinigalia

2001-07-01

In this study, vitamin A and {beta}-carotene contents and the percent of activity retention were analyzed in 3 kGy and 30 kGy {sup 60}Co {gamma}-irradiated samples of bovine liver and swine liver pate. Six different lots of liver samples weighing 100 g obtained at the meat market were employed. Irradiation was performed with liver samples initially frozen (-15 deg C). Similarly, six different lots of swine liver pate samples weighing about 100 - 125 g , at temperature of 7 deg C were irradiated. Pre-treatments and analysis methods were those described in Instituto Adolfo Lutz Norms for food analyses. Two and five- gram samples in duplicates for bovine liver and five gram-samples for swine liver pate were used. The results showed that there were no losses of either vitamin A or provitamin A activities in the samples of bovine liver and swine liver pate irradiated with a dose of 3 kGy (retention about 100%). On the other hand, for swine liver pate samples of vitamin A were maintained after irradiation with 30 kGy. (author)

Imaging of oxygen gradients in giant umbrella cells: an ex vivo PLIM study.

Science.gov (United States)

Zhdanov, A V; Golubeva, A V; Okkelman, I A; Cryan, J F; Papkovsky, D B

2015-10-01

O2 plays a pivotal role in aerobic metabolism and regulation of cell and tissue function. Local differences and fluctuations in tissue O2 levels are well documented; however, the physiological significance of O2 microgradients, particularly at the subcellular level, remains poorly understood. Using the cell-penetrating phosphorescent O2 probe Pt-Glc and confocal fluorescence microscopy, we visualized O2 distribution in individual giant (>100-μm) umbrella cells located superficially in the urinary bladder epithelium. We optimized conditions for in vivo phosphorescent staining of the inner surface of the mouse bladder and subsequent ex vivo analysis of excised live tissue. Imaging experiments revealed significant (≤85 μM) and heterogeneous deoxygenation within respiring umbrella cells, with radial O2 gradients of up to 40 μM across the cell, or ∼0.6 μM/μm. Deeply deoxygenated (5-15 μM O2) regions were seen to correspond to the areas enriched with polarized mitochondria. Pharmacological activation of mitochondrial respiration decreased oxygenation and O2 gradients in umbrella cells, while inhibition with antimycin A dissipated the gradients and caused gradual reoxygenation of the tissue to ambient levels. Detailed three-dimensional maps of O2 distribution potentially can be used for the modeling of intracellular O2-dependent enzymatic reactions and downstream processes, such as hypoxia-inducible factor signaling. Further ex vivo and in vivo studies on intracellular and tissue O2 gradients using confocal imaging can shed light on the molecular mechanisms regulating O2-dependent (patho)physiological processes in the bladder and other tissues. Copyright © 2015 the American Physiological Society.

Effect of different polymers on in vitro and ex vivo permeability of Ofloxacin from its mucoadhesive suspensions

OpenAIRE

Chakraborti, Chandra Kanti; Sahoo, Subhashree; Behera, Pradipta Kumar

2014-01-01

Considering the importance of drug permeation from formulations, in vitro and ex vivo drug permeation characteristics of three oral mucoadhesive suspensions of Ofloxacin were designed and compared. Three suspensions of Ofloxacin were prepared by taking two grades of Carbopol polymer such as Carbopol 934 (C934) and Carbopol 940 (C940); and Hydroxypropyl methylcellulose. The permeability study was performed by using a Franz diffusion cell through both synthetic cellulose acetate membrane and ex...

Ex vivo investigation of ocular tissue distribution following intravitreal administration of connexin43 mimetic peptide using the microdialysis technique and LC-MS/MS.

Science.gov (United States)

Bisht, Rohit; Mandal, Abhirup; Rupenthal, Ilva D; Mitra, Ashim K

2016-12-01

This study aimed to develop and evaluate an ex vivo eye model for intravitreal drug sampling and tissue distribution of connexin43 mimetic peptide (Cx43MP) following intravitreal injection using the microdialysis technique and LC-MS/MS. An LC-MS/MS method was developed, validated, and applied for quantification of Cx43MP in ocular tissues. Microdialysis probes were calibrated for in vitro recovery studies. Bovine eyes were fixed in a customized eye holder and after intravitreal injection of Cx43MP, microdialysis probes were implanted in the vitreous body. Vitreous samples were collected at particular time intervals over 24 h. Moreover, 24 and 48 h after intravitreal injection ocular tissues were collected, processed, and analyzed for Cx43MP concentrations using LC-MS/MS. The LC-MS/MS method showed good linearity (r 2  = 0.9991). The mean percent recovery for lower (LQC), medium (MQC), and higher quality control (HQC) (0.244, 3.906, and 125 μg/mL) was found to be 83.83, 84.92, and 94.52, respectively, with accuracy ranges between 96 and 99 % and limits of detection (LOD) and quantification (LOQ) of 0.122 and 0.412 μg/mL. The in vitro recovery of the probes was found to be over 80 %. As per microdialysis sample analysis, the Cx43MP concentration was found to increase slowly in the vitreous body up to 16 h and thereafter declined. After 48 h, the Cx43MP concentration was higher in vitreous, cornea, and retina compared to lens, iris, and aqueous humor. This ex vivo model may therefore be a useful tool to investigate intravitreal kinetics and ocular disposition of therapeutic molecules after intravitreal injection.

The cryoablation of lung tissue using liquid nitrogen in gel and in the ex vivo pig lung.

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Nomori, Hiroaki; Yamazaki, Ikuo; Kondo, Toshiya; Kanno, Masaya

2017-02-01

To examine the efficiency of cryoablation using liquid nitrogen in lung tissue, we measured the size and temperature distribution of the frozen area (iceball) in gel and in the ex vivo pig lungs. Cryoprobes with diameters of 2.4 and 3.4 mm (2.4D and 3.4D, respectively) were used. Three temperature sensors were positioned at the surface of the cryoprobe and at distances of 0.5 and 1.5 cm from the cryoprobe. The ex vivo pig lungs were perfused with 37 °C saline and inflated using ventilator to simulate in vivo lung conditions. In gel, the 2.4D and 3.4D probes made iceballs of 3.9 ± 0.1 and 4.8 ± 0.3 cm in diameter, respectively, and the temperature at 1.5 cm from those probes reached -32 ± 8 and -53 ± 5 °C, respectively. In the pig lung, the 2.4D and 3.4D probes made iceballs of 5.2 ± 0.1 and 5.5 ± 0.4 cm in diameter, respectively, and the temperature at 1.5 cm from these probes reached -49 ± 5 and -58 ± 3 °C, respectively. Liquid nitrogen cryoablation using both 2.4D and 3.4D probes made iceballs that were of sufficient size, and effective temperatures were reached in both gel and the ex vivo pig lung.

Ex Vivo Model of Human Penile Transplantation and Rejection: Implications for Erectile Tissue Physiology.

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Sopko, Nikolai A; Matsui, Hotaka; Lough, Denver M; Miller, Devin; Harris, Kelly; Kates, Max; Liu, Xiaopu; Billups, Kevin; Redett, Richard; Burnett, Arthur L; Brandacher, Gerald; Bivalacqua, Trinity J

2017-04-01

Penile transplantation is a potential treatment option for severe penile tissue loss. Models of human penile rejection are lacking. Evaluate effects of rejection and immunosuppression on cavernous tissue using a novel ex vivo mixed lymphocyte reaction (MLR) model. Cavernous tissue and peripheral blood mononuclear cells (PBMCs) from 10 patients undergoing penile prosthesis operations and PBMCs from a healthy volunteer were obtained. Ex vivo MLRs were prepared by culturing cavernous tissue for 48h in media alone, in media with autologous PBMCs, or in media with allogenic PBMCs to simulate control, autotransplant, and allogenic transplant conditions with or without 1μM cyclosporine A (CsA) or 20nM tacrolimus (FK506) treatment. Rejection was characterized by PBMC flow cytometry and gene expression transplant array. Cavernous tissues were evaluated by histomorphology and myography to assess contraction and relaxation. Data were analyzed using two-way analysis of variance and unpaired Student t test. Flow cytometry and tissue array demonstrated allogenic PBMC activation consistent with rejection. Rejection impaired cavernous tissue physiology and was associated with cellular infiltration and apoptosis. CsA prevented rejection but did not improve tissue relaxation. CsA treatment impaired relaxation in tissues cultured without PBMCs compared with media and FK506. Study limitations included the use of penile tissue with erectile dysfunction and lack of cross-matching data. This model could be used to investigate the effects of penile rejection and immunosuppression. Additional studies are needed to optimize immunosuppression to prevent rejection and maximize corporal tissue physiology. This report describes a novel ex vivo model of human penile transplantation rejection. Tissue rejection impaired erectile tissue physiology. This report suggests that cyclosporin A might hinder corporal physiology and that other immunosuppressant agents, such as FK506, might be better suited

Ex vivo expansion of alveolar macrophages with Mycobacterium tuberculosis from the resected lungs of patients with pulmonary tuberculosis

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Petrunina, Ekaterina; Umpeleva, Tatiana; Karskanova, Svetlana; Bayborodin, Sergey; Vakhrusheva, Diana; Kravchenko, Marionella; Skornyakov, Sergey

2018-01-01

Tuberculosis (TB), with the Mycobacterium tuberculosis (Mtb) as the causative agent, remains to be a serious world health problem. Traditional methods used for the study of Mtb in the lungs of TB patients do not provide information about the number and functional status of Mtb, especially if Mtb are located in alveolar macrophages. We have developed a technique to produce ex vivo cultures of cells from different parts of lung tissues surgically removed from patients with pulmonary TB and compared data on the number of cells with Mtb inferred by the proposed technique to the results of bacteriological and histological analyses used for examination of the resected lungs. The ex vivo cultures of cells obtained from the resected lungs of all patients were largely composed of CD14-positive alveolar macrophages, foamy or not, with or without Mtb. Lymphocytes, fibroblasts, neutrophils, and multinucleate Langhans giant cells were also observed. We found alveolar macrophages with Mtb in the ex vivo cultures of cells from the resected lungs of even those TB patients, whose sputum smears and lung tissues did not contain acid-fast Mtb or reveal growing Mtb colonies on dense medium. The detection of alveolar macrophages with Mtb in ex vivo culture as soon as 16–18 h after isolation of cells from the resected lungs of all TB patients suggests that the technique proposed for assessing the level of infection in alveolar macrophages of TB patients has higher sensitivity than do prolonged bacteriological or pathomorphological methods. The proposed technique allowed us to rapidly (in two days after surgery) determine the level of infection with Mtb in the cells of the resected lungs of TB patients and, by the presence or absence of Mtb colonies, including those with cording morphology, the functional status of the TB agent at the time of surgery. PMID:29401466

Equine dendritic cells generated with horse serum have enhanced functionality in comparison to dendritic cells generated with fetal bovine serum

OpenAIRE

Ziegler, Anja; Everett, Helen; Hamza, Eman; Garbani, Mattia; Gerber, Vinzenz; Marti, Eliane; Steinbach, Falko

2016-01-01

BACKGROUND: Dendritic cells are professional antigen-presenting cells that play an essential role in the initiation and modulation of T cell responses. They have been studied widely for their potential clinical applications, but for clinical use to be successful, alternatives to xenogeneic substances like fetal bovine serum (FBS) in cell culture need to be found. Protocols for the generation of dendritic cells ex vivo from monocytes are well established for several species, including hor...

Chitosan and Kappa-Carrageenan Vaginal Acyclovir Formulations for Prevention of Genital Herpes. In Vitro and Ex Vivo Evaluation

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María-Pilar Sánchez-Sánchez

2015-09-01

Full Text Available Vaginal formulations for the prevention of sexually transmitted infections are currently gaining importance in drug development. Polysaccharides, such as chitosan and carrageenan, which have good binding capacity with mucosal tissues, are now included in vaginal delivery systems. Marine polymer-based vaginal mucoadhesive solid formulations have been developed for the controlled release of acyclovir, which may prevent the sexual transmission of the herpes simplex virus. Drug release studies were carried out in two media: simulated vaginal fluid and simulated vaginal fluid/simulated seminal fluid mixture. The bioadhesive capacity and permanence time of the bioadhesion, the prepared compacts, and compacted granules were determined ex vivo using bovine vaginal mucosa as substrate. Swelling processes were quantified to confirm the release data. Biocompatibility was evaluated through in vitro cellular toxicity assays, and the results showed that acyclovir and the rest of the materials had no cytotoxicity at the maximum concentration tested. The mixture of hydroxyl-propyl-methyl-cellulose with chitosan- or kappa-carrageenan-originated mucoadhesive systems that presented a complete and sustained release of acyclovir for a period of 8–9 days in both media. Swelling data revealed the formation of optimal mixed chitosan/hydroxyl-propyl-methyl-cellulose gels which could be appropriated for the prevention of sexual transmission of HSV.

Biobanking of human pancreas cancer tissue: impact of ex-vivo procurement times on RNA quality.

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Rudloff, Udo; Bhanot, Umesh; Gerald, William; Klimstra, David S; Jarnagin, William R; Brennan, Murray F; Allen, Peter J

2010-08-01

Tissue banking has become a major initiative at many oncology centers. The influence of warm ex-vivo ischemia times, storage times, and biobanking protocols on RNA integrity and subsequent microarray data is not well documented. A prospective institutional review board-approved protocol for the banking of abdominal neoplasms was initiated at Memorial Sloan-Kettering Cancer Center in 2001. Sixty-four representative pancreas cancer specimens snap-frozen at various ex-vivo procurement times (1 h) and banked during three time periods (2001-2004, 2004-2006, 2006-2008) were processed. RNA integrity was determined by microcapillary electrophoresis using the RNA integrity number (RIN) algorithm and by results of laser-capture microdissection (LCM). Overall, 42% of human pancreas cancer specimens banked under a dedicated protocol yielded RNA with a RIN of > or =7. Limited warm ex-vivo ischemia times did not negatively impact RNA quality (percentage of tissue with total RNA with RIN of > or =7 for 60 min, 42%), and long-term storage of banked pancreas cancer biospecimens did not negatively influence RNA quality (total RNA with RIN of > or =7 banked 2001-2004, 44%; 2004-2006, 38%; 2006-2008, 50%). RNA retrieved from pancreatic cancer samples with RIN of > or =7 subject to LCM yielded RNA suitable for further downstream applications. Fresh-frozen pancreas tissue banked within a standardized research protocol yields high-quality RNA in approximately 50% of specimens and can be used for enrichment by LCM. Quality of tissues of the biobank were not adversely impacted by limited variations of warm ischemia times or different storage periods. This study shows the challenges and investments required to initiate and maintain high-quality tissue repositories.

Comprehensive Antiretroviral Restriction Factor Profiling Reveals the Evolutionary Imprint of the ex Vivo and in Vivo IFN-β Response in HTLV-1-Associated Neuroinflammation

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Fabio E. Leal

2018-05-01

Full Text Available HTLV-1-Associated Myelopathy (HAM/TSP is a progressive neuroinflammatory disorder for which no disease-modifying treatment exists. Modest clinical benefit from type I interferons (IFN-α/β in HAM/TSP contrasts with its recently identified IFN-inducible gene signature. In addition, IFN-α treatment in vivo decreases proviral load and immune activation in HAM/TSP, whereas IFN-β therapy decreases tax mRNA and lymphoproliferation. We hypothesize this “IFN paradox” in HAM/TSP might be explained by both cell type- and gene-specific effects of type I IFN in HTLV-1-associated pathogenesis. Therefore, we analyzed ex vivo transcriptomes of CD4+ T cells, PBMCs and whole blood in healthy controls, HTLV-1-infected individuals, and HAM/TSP patients. First, we used a targeted approach, simultaneously quantifying HTLV-1 mRNA (HBZ, Tax, proviral load and 42 host genes with known antiretroviral (anti-HIV activity in purified CD4+ T cells. This revealed two major clusters (“antiviral/protective” vs. “proviral/deleterious”, as evidenced by significant negative (TRIM5/TRIM22/BST2 vs. positive correlation (ISG15/PAF1/CDKN1A with HTLV-1 viral markers and clinical status. Surprisingly, we found a significant inversion of antiretroviral activity of host restriction factors, as evidenced by opposite correlation to in vivo HIV-1 vs. HTLV-1 RNA levels. The anti-HTLV-1 effect of antiviral cluster genes was significantly correlated to their adaptive chimp/human evolution score, for both Tax mRNA and PVL. Six genes of the proposed antiviral cluster underwent lentivirus-driven purifying selection during primate evolution (TRIM5/TRIM22/BST2/APOBEC3F-G-H, underscoring the cross-retroviral evolutionary imprint. Secondly, we examined the genome-wide type I IFN response in HAM/TSP patients, following short-term ex vivo culture of PBMCs with either IFN-α or IFN-β. Microarray analysis evidenced 12 antiretroviral genes (including TRIM5α/TRIM22/BST2 were significantly

RNA-Seq analysis uncovers transcriptomic variations between morphologically similar in vivo- and in vitro-derived bovine blastocysts

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Driver Ashley M

2012-03-01

Full Text Available Abstract Background A valuable tool for both research and industry, in vitro fertilization (IVF has applications range from gamete selection and preservation of traits to cloning. Although IVF has achieved worldwide use, with approximately 339,685 bovine embryos transferred in 2010 alone, there are still continuing difficulties with efficiency. It is rare to have more than 40% of fertilized in vitro cattle oocytes reach blastocyst stage by day 8 of culture, and pregnancy rates are reported as less than 45% for in vitro produced embryos. To investigate potential influences in-vitro fertilization (IVF has on embryonic development, this study compares in vivo- and in vitro-derived bovine blastocysts at a similar stage and quality grade (expanded, excellent quality to determine the degree of transcriptomic variation beyond morphology using RNA-Seq. Results A total of 26,906,451 and 38,184,547 fragments were sequenced for in vitro and in vivo embryo pools, respectively. We detected expression for a total of 17,634 genes, with 793 genes showing differential expression between the two embryo populations with false discovery rate (FDR Conclusions Thus, our results support that IVF may influence at the transcriptomic level and that morphology is limited in full characterization of bovine preimplantation embryos.

Neutron activation analysis technique and X-ray fluorescence in bovine liver sample

International Nuclear Information System (INIS)

Maihara, V.A.; Favaro, D.I.T.; Vasconcellos, M.B.A.; Sato, I.M.; Salvador, V.L.

2002-01-01

Many analytical techniques have been used in food and diet analysis in order to determine a great number of nutritional elements, ranging from percentage to ng g -1 , with high sensitivity and accuracy. Instrumental Neutron activation Analysis (INAA) has been employed to certificate many trace elements in biological reference materials. More recently, the X-Ray Fluorescence (FRX-WD) has been also used to determine some essential elements in food samples. The INAA has been applied in nutrition studies in our laboratory at IPEN since the 80 s. For the development of analytical methodologies the use of the reference materials with the same characteristics of the sample analyzed is essential. Several Brazilian laboratories do not have conditions to use these materials due their high cost.In this paper preliminary results of commercial bovine liver sample analyses obtained by INAA and WD-XRF methods are presented. This sample was prepared to be a Brazilian candidate of reference material for a group of laboratories participating in a research project sponsored by FAPESP. The concentrations of some elements like Cl, K, Na, P, S and trace elements Br, Ca, Co, Cu, Fe, Mg, Mn, Mo, Rb, Se and Zn were determined by INAA and WD-XFR. For validation methodology of both techniques, NIST SRM 1577b Bovine Liver reference material was analyzed and the detection limits were calculated. The concentrations of elements determined by both analytical techniques were compared by using the Student's t-test and for Cl, Cu, Fe, K, Mg, Na, Rn and Zn the results do show no statistical difference for 95% significance level. (author)

Feasibility of a liver transcriptomics approach to assess bovine treatment with the prohormone dehydroepiandrosterone (DHEA

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Rijk Jeroen CW

2010-09-01

Full Text Available Abstract Background Within the European Union the use of growth promoting agents in animal production is prohibited. Illegal use of natural prohormones like dehydroepiandrosterone (DHEA is hard to prove since prohormones are strongly metabolized in vivo. In the present study, we investigated the feasibility of a novel effect-based approach for monitoring abuse of DHEA. Changes in gene expression profiles were studied in livers of bull calves treated orally (PO or intramuscularly (IM with 1000 mg DHEA versus two control groups, using bovine 44K DNA microarrays. In contrast to controlled genomics studies, this work involved bovines purchased at the local market on three different occasions with ages ranging from 6 to 14 months, thereby reflecting the real life inter-animal variability due to differences in age, individual physiology, season and diet. Results As determined by principal component analysis (PCA, large differences in liver gene expression profiles were observed between treated and control animals as well as between the two control groups. When comparing the gene expression profiles of PO and IM treated animals to that of all control animals, the number of significantly regulated genes (p-value 1.5 was 23 and 37 respectively. For IM and PO treated calves, gene sets were generated of genes that were significantly regulated compared to one control group and validated versus the other control group using Gene Set Enrichment Analysis (GSEA. This cross validation, showed that 6 out of the 8 gene sets were significantly enriched in DHEA treated animals when compared to an 'independent' control group. Conclusions This study showed that identification and application of genomic biomarkers for screening of (prohormone abuse in livestock production is substantially hampered by biological variation. On the other hand, it is demonstrated that comparison of pre-defined gene sets versus the whole genome expression profile of an animal allows to

Reprogrammed chondrocytes engineered to produce IL-12 provide novel ex vivo immune-gene therapy for cancer.

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Tada, Hiroyuki; Kishida, Tsunao; Fujiwara, Hitoshi; Kosuga, Toshiyuki; Konishi, Hirotaka; Komatsu, Shuhei; Shiozaki, Atsushi; Ichikawa, Daisuke; Okamoto, Kazuma; Otsuji, Eigo; Mazda, Osam

2017-03-01

The somatic cell reprogramming technology was applied to a novel and promising ex vivo immune-gene therapy strategy for cancer. To establish a novel ex vivo cytokine gene therapy of cancer using the somatic cell reprogramming procedures. Mouse fibroblasts were converted into chondrocytes and subsequently transduced with IL-12 gene. The resultant IL-12 induced chondrogenic cells were irradiated with x-ray and inoculated into mice bearing CT26 colon cancer. The irradiation at 20 Gy or higher totally eliminated the proliferative potential of the cells, while less significantly influencing the IL-12 production from the cells. An inoculation of the irradiated IL-12 induced chondrogenic cells significantly suppressed tumor by inducing tumor-specific cytotoxic T lymphocytes, enhancing natural killer tumoricidal activity and inhibiting tumor neoangiogenesis in the mice. The somatic cell reprogramming procedures may provide a novel and effective means to treat malignancies.

Astaxanthin, a Carotenoid, Stimulates Immune Responses by Enhancing IFN-γ and IL-2 Secretion in Primary Cultured Lymphocytes in Vitro and ex Vivo

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Lin, Kuan-Hung; Lin, Kao-Chang; Lu, Wan-Jung; Thomas, Philip-Aloysius; Jayakumar, Thanasekaran; Sheu, Joen-Rong

2015-01-01

Astaxanthin, a potent antioxidant carotenoid, plays a major role in modulating the immune response. In this study, we examined the immunomodulatory effects of astaxanthin on cytokine production in primary cultured lymphocytes both in vitro and ex vivo. Direct administration of astaxanthin (70–300 nM) did not produce cytotoxicity in lipopolysaccharide (LPS, 100 µg/ mL)- or concanavalin A (Con A, 10 µg/ mL)-activated lymphocytes, whereas astaxanthin alone at 300 nM induced proliferation of splenic lymphocytes (p < 0.05) in vitro. Although astaxanthin, alone or with Con A, had no apparent effect on interferon (INF-γ) and interleukin (IL-2) production in primary cultured lymphocytes, it enhanced LPS-induced INF-γ production. In an ex vivo experiment, oral administration of astaxanthin (0.28, 1.4 and 7 mg/kg/day) for 14 days did not cause alterations in the body or spleen weights of mice and also was not toxic to lymphocyte cells derived from the mice. Moreover, treatment with astaxanthin significantly increased LPS-induced lymphocyte proliferation ex vivo but not Con A-stimulated lymphocyte proliferation ex vivo. Enzyme linked immunosorbent assay (ELISA) analysis revealed that administration of astaxanthin significantly enhanced INF-γ production in response to both LPS and Con A stimulation, whereas IL-2 production increased only in response to Con A stimulation. Also, astaxanthin treatment alone significantly increased IL-2 production in lymphocytes derived from mice, but did not significantly change production of INF-γ. These findings suggest that astaxanthin modulates lymphocytic immune responses in vitro, and that it partly exerts its ex vivo immunomodulatory effects by increasing INF-γ and IL-2 production without inducing cytotoxicity. PMID:26729100

Phosphatidylcholine contributes to in vivo 31P MRS signal from the human liver

International Nuclear Information System (INIS)

Chmelik, Marek; Bogner, Wolfgang; Gajdosik, Martin; Gruber, Stephan; Trattnig, Siegfried; Valkovic, Ladislav; Wolf, Peter; Krebs, Michael; Halilbasic, Emina; Trauner, Michael; Krssak, Martin

2015-01-01

To demonstrate the overlap of the hepatic and bile phosphorus ( 31 P) magnetic resonance (MR) spectra and provide evidence of phosphatidylcholine (PtdC) contribution to the in vivo hepatic 31 P MRS phosphodiester (PDE) signal, suggested in previous reports to be phosphoenolpyruvate (PEP). Phantom measurements to assess the chemical shifts of PEP and PtdC signals were performed at 7 T. A retrospective analysis of hepatic 3D 31 P MR spectroscopic imaging (MRSI) data from 18 and five volunteers at 3 T and 7 T, respectively, was performed. Axial images were inspected for the presence of gallbladder, and PDE signals in representative spectra were quantified. Phantom experiments demonstrated the strong pH-dependence of the PEP chemical shift and proved the overlap of PtdC and PEP (∝2 ppm relative to phosphocreatine) at hepatic pH. Gallbladder was covered in seven of 23 in vivo 3D-MRSI datasets. The PDE gall /γ-ATP liver ratio was 4.8-fold higher (p = 0.001) in the gallbladder (PDE gall /γ-ATP liver = 3.61 ± 0.79) than in the liver (PDE liver /γ-ATP liver = 0.75 ± 0.15). In vivo 7 T 31 P MRSI allowed good separation of PDE components. The gallbladder is a strong source of contamination in adjacent 31 P MR hepatic spectra due to biliary phosphatidylcholine. In vivo 31 P MR hepatic signal at 2.06 ppm may represent both phosphatidylcholine and phosphoenolpyruvate, with a higher phosphatidylcholine contribution due to its higher concentration. (orig.)

Ex vivo and in vivo coherent Raman imaging of the peripheral and central nervous system

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Huff, Terry Brandon

A hallmark of nervous system disorders is damage or degradation of the myelin sheath. Unraveling the mechanisms underlying myelin degeneration and repair represent one of the great challenges in medicine. This thesis work details the development and utilization of advanced optical imaging methods to gain insight into the structure and function of myelin in both healthy and diseased states in the in vivo environment. This first part of this thesis discusses ex vivo studies of the effects of high-frequency stimulation of spinal tissues on the structure of the node of Ranvier as investigated by coherent anti-Stokes Raman scattering (CARS) imaging (manuscript submitted to Journal of Neurosciece). Reversible paranodal myelin retraction at the nodes of Ranvier was observed during 200 Hz electrical stimulation, beginning minutes after the onset and continuing for up to 10 min after stimulation was ceased. A mechanistic study revealed a Ca2+ dependent pathway: high-frequency stimulation induced paranodal myelin retraction via pathologic calcium influx into axons, calpain activation, and cytoskeleton degradation through spectrin break-down. Also, the construction of dual-scanning CARS microscope for large area mapping of CNS tissues is detailed (Optics Express, 2008, 16:19396-193409). A confocal scanning head equipped with a rotating polygon mirror provides high speed, high resolution imaging and is coupled with a motorized sample stage to generate high-resolution large-area images of mouse brain coronal section and guinea pig spinal cord cross section. The polygon mirror decreases the mosaic acquisition time significantly without reducing the resolution of individual images. The ex vivo studies are then extended to in vivo imaging of mouse sciatic nerve tissue by CARS and second harmonic generation (SHG) imaging (Journal of Microscopy, 2007, 225: 175-182). Following a minimally invasive surgery to open the skin, CARS imaging of myelinated axons and SHG imaging of the

Ex Vivo Expansion of Functional Human UCB-HSCs/HPCs by Coculture with AFT024-hkirre Cells

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Muti ur Rehman Khan

2014-01-01

Full Text Available Kiaa1867 (human Kirre, hKirre has a critical role in brain development and/or maintenance of the glomerular slit diaphragm in kidneys. Murine homolog of this gene, mKirre expressed in OP9 and AFT024 cells could support hematopoietic stem cells/hematopoietic progenitor cells (HSC/HPC expansion in vitro. HKirre is also expressed in human FBMOB-hTERT cell line and fetal liver fibroblast-like cells but its function has remained unclear. In this paper, we cloned a hKirre gene from human fetal liver fibroblast-like cells and established a stably overexpressing hKirre-AFT024 cell line. Resultant cells could promote self-renewal and ex vivo expansion of HSCs/HPCs significantly higher than AFT024-control cells transformed with mock plasmid. The Expanded human umbilical cord blood (hUCB CD34+ cells retained the capacity of multipotent differentiation as long as 8 weeks and successfully repopulated the bone marrow of sublethally irradiated NOD/SCID mice, which demonstrated the expansion of long-term primitive transplantable HSCs/HPCs. Importantly, hkirre could upregulate the expressions of Wnt-5A, BMP4, and SDF-1 and downregulate TGF-β with other hematopoietic growth factors. By SDS-PAGE and Western Blot analysis, a ~89 kDa protein in total lysate of AFT024-hKirre was identified. Supernatants from AFT024-hkirre could also support CD34+CD38− cells expansion. These results demonstrated that the AFT024-hKirre cells have the ability to efficiently expand HSCs/HPCs.

Ex Vivo Gene Therapy Using Human Mesenchymal Stem Cells to Deliver Growth Factors in the Skeletal Muscle of a Familial ALS Rat Model.

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Suzuki, Masatoshi; Svendsen, Clive N

2016-01-01

Therapeutic protein and molecule delivery to target sites by transplanted human stem cells holds great promise for ex vivo gene therapy. Our group has demonstrated the therapeutic benefits of ex vivo gene therapy targeting the skeletal muscles in a transgenic rat model of familial amyotrophic lateral sclerosis (ALS). We used human mesenchymal stem cells (hMSCs) and genetically modified them to release neuroprotective growth factors such as glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor (VEGF). Intramuscular growth factor delivery via hMSCs can enhance neuromuscular innervation and motor neuron survival in a rat model of ALS (SOD1(G93A) transgenic rats). Here, we describe the protocol of ex vivo delivery of growth factors via lentiviral vector-mediated genetic modification of hMSCs and hMSC transplantation into the skeletal muscle of a familial ALS rat model.

Effects of zinc ex vivo on taurine uptake in goldfish retinal cells

OpenAIRE

Nusetti, Sonia; Urbina, Mary; Lima, Lucimey

2010-01-01

Background Taurine and zinc exert neurotrophic effects in the central nervous system. Current studies demonstrate that Na+/Cl- dependent neurotransmitter transporters, similar to that of taurine, are modulated by micromolar concentrations of zinc. This study examined the effect of zinc sulfate ex vivo on [3H]taurine transport in goldfish retina. Methods Isolated cells were incubated in Ringer with zinc (0.1?100 ?M). Taurine transport was done with 50 nM [3H]taurine or by isotopic dilution wit...

Monitoring UV-induced signalling pathways in an ex vivo skin organ culture model using phospho-antibody array.

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Lenain, Christelle; Gamboa, Bastien; Perrin, Agnes; Séraïdaris, Alexia; Bertino, Béatrice; Rival, Yves; Bernardi, Mathieu; Piwnica, David; Méhul, Bruno

2018-05-01

We investigated UV-induced signalling in an ex vivo skin organ culture model using phospho-antibody array. Phosphorylation modulations were analysed in time-course experiments following exposure to solar-simulated UV and validated by Western blot analyses. We found that UV induced P-p38 and its substrates, P-ERK1/2 and P-AKT, which were previously shown to be upregulated by UV in cultured keratinocytes and in vivo human skin. This indicates that phospho-antibody array applied to ex vivo skin organ culture is a relevant experimental system to investigate signalling events following perturbations. As the identified proteins are components of pathways implicated in skin tumorigenesis, UV-exposed skin organ culture model could be used to investigate the effect on these pathways of NMSC cancer drug candidates. In addition, we found that phospho-HCK is induced upon UV exposure, producing a new candidate for future studies investigating its role in the skin response to UV and UV-induced carcinogenesis. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Epidermal protection with cryogen spray cooling during high fluence pulsed dye laser irradiation: an ex vivo study.

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Tunnell, J W; Nelson, J S; Torres, J H; Anvari, B

2000-01-01

Higher laser fluences than currently used in therapy (5-10 J/cm(2)) are expected to result in more effective treatment of port wine stain (PWS) birthmarks. However, higher incident fluences increase the risk of epidermal damage caused by absorption of light by melanin. Cryogen spray cooling offers an effective method to reduce epidermal injury during laser irradiation. The objective of this study was to determine whether high laser incident fluences (15-30 J/cm(2)) could be used while still protecting the epidermis in ex vivo human skin samples. Non-PWS skin from a human cadaver was irradiated with a Candela ScleroPlus Laser (lambda = 585 nm; pulse duration = 1.5 msec) by using various incident fluences (8-30 J/cm(2)) without and with cryogen spray cooling (refrigerant R-134a; spurt durations: 40-250 msec). Assessment of epidermal damage was based on histologic analysis. Relatively short spurt durations (40-100 msec) protected the epidermis for laser incident fluences comparable to current therapeutic levels (8-10 J/cm(2)). However, longer spurt durations (100-250 msec) increased the fluence threshold for epidermal damage by a factor of three (up to 30 J/cm(2)) in these ex vivo samples. Results of this ex vivo study show that epidermal protection from high laser incident fluences can be achieved by increasing the cryogen spurt duration immediately before pulsed laser exposure. Copyright 2000 Wiley-Liss, Inc.

Numerical study and ex vivo assessment of HIFU treatment time reduction through optimization of focal point trajectory

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Grisey, A.; Yon, S.; Pechoux, T.; Letort, V.; Lafitte, P.

2017-03-01

Treatment time reduction is a key issue to expand the use of high intensity focused ultrasound (HIFU) surgery, especially for benign pathologies. This study aims at quantitatively assessing the potential reduction of the treatment time arising from moving the focal point during long pulses. In this context, the optimization of the focal point trajectory is crucial to achieve a uniform thermal dose repartition and avoid boiling. At first, a numerical optimization algorithm was used to generate efficient trajectories. Thermal conduction was simulated in 3D with a finite difference code and damages to the tissue were modeled using the thermal dose formula. Given an initial trajectory, the thermal dose field was first computed, then, making use of Pontryagin's maximum principle, the trajectory was iteratively refined. Several initial trajectories were tested. Then, an ex vivo study was conducted in order to validate the efficicency of the resulting optimized strategies. Single pulses were performed at 3MHz on fresh veal liver samples with an Echopulse and the size of each unitary lesion was assessed by cutting each sample along three orthogonal planes and measuring the dimension of the whitened area based on photographs. We propose a promising approach to significantly shorten HIFU treatment time: the numerical optimization algorithm was shown to provide a reliable insight on trajectories that can improve treatment strategies. The model must now be improved in order to take in vivo conditions into account and extensively validated.

Endocytosis via galactose receptors in vivo. Ligand size directs uptake by hepatocytes and/or liver macrophages

International Nuclear Information System (INIS)

Schlepper-Schaefer, J.; Huelsmann, D.; Djovkar, A.; Meyer, H.E.; Herbertz, L.; Kolb, H.; Kolb-Bachofen, V.

1986-01-01

The intrahepatic binding and uptake of variously sized ligands with terminal galactosyl residues is rat liver was followed. The ligands were administered to prefixed livers in binding studies and in vivo and in situ (serum-free perfused livers) in uptake studies. Gold sols with different particle diameters were prepared: 5 nm (Au 5 ), 17 nm (Au 17 ), 50 nm (Au 50 ) and coated with galactose exposing glycoproteins (asialofetuin (ASF) or lactosylated BSA (LacBSA)). Electron microscopy of mildly prefixed livers perfused with LacBSA-Au 5 in serum-free medium showed ligand binding to liver macrophages, hepatocytes and endothelial cells. Ligands bound to prefixed cell surfaces reflect the initial distribution of receptor activity: pre-aggregated clusters of ligands are found on liver macrophages, single particles statistically distributed on hepatocytes and pre-aggregated clusters of particles restricted to coated pits on endothelial cells. Ligand binding is prevented in the presence of 80 mM N-acetylgalactosamine (GalNAc), while N-acetylglucosamine (GlcNAc) is without effect. Electron microscopy of livers after ligand injection into the tail vein shows that in vivo uptake of electron-dense galactose particles by liver cells is size-dependent. In vivo uptake by liver macrophages is mediated by galactose-specific recognition as shown by inhibition with GalNAc

Ex vivo testing of immune responses in precision-cut lung slices

International Nuclear Information System (INIS)

Henjakovic, M.; Sewald, K.; Switalla, S.; Kaiser, D.; Mueller, M.; Veres, T.Z.; Martin, C.; Uhlig, S.; Krug, N.; Braun, A.

2008-01-01

The aim of this study was the establishment of precision-cut lung slices (PCLS) as a suitable ex vivo alternative approach to animal experiments for investigation of immunomodulatory effects. For this purpose we characterized the changes of cytokine production and the expression of cell surface markers after incubation of PCLS with immunoactive substances lipopolysaccharide (LPS), macrophage-activating lipopeptide-2 (MALP-2), interferon γ (IFNγ), and dexamethasone. Viability of PCLS from wild-type and CD11c-enhanced yellow fluorescent protein (CD11-EYFP)-transgenic mice was controlled by measurement of lactate dehydrogenase (LDH) enzyme activity and live/dead fluorescence staining using confocal microscopy. Cytokines and chemokines were detected with Luminex technology and ELISA. Antigen presenting cell (APC) markers were investigated in living mouse PCLS in situ using confocal microscopy. LPS triggered profound pro-inflammatory effects in PCLS. Dexamethasone prevented LPS-induced production of cytokines/chemokines such as interleukin (IL)-5, IL-1α, TNFα, IL-12(p40), and RANTES in PCLS. Surface expression of MHC class II, CD40, and CD11c, but not CD86 was present in APCs of naive PCLS. Incubation with LPS enhanced specifically the expression of MHC class II on diverse cells. MALP-2 only failed to alter cytokine or chemokine levels, but was highly effective in combination with IFNγ resulting in increased levels of TNFα, IL-12(p40), RANTES, and IL-1α. PCLS showed characteristic responses to typical pro-inflammatory stimuli and may thus provide a suitable ex vivo technique to predict the immunomodulatory potency of inhaled substances

Single-Shot-RARE for rapid 3D hyperpolarized metabolic ex vivo tissue imaging: RF-pulse design for semi-dense spectra

DEFF Research Database (Denmark)

Magnusson, P.O.; Jensen, Pernille Rose; Dyrby, Tim Bjørn

MRS of hyperpolarized (HP) 13C-enriched compounds is a promising method for in vivo cancer diagnosis . Sentinel lymph node ex vivo tissue sample histology used in clinical routine for breast cancer metastasis diagnosis requires time consuming sample analysis. 3D-HP-MRSI can potentially speed up...

An ionic liquid-in-water microemulsion as a potential carrier for topical delivery of poorly water soluble drug: Development, ex-vivo and in-vivo evaluation.

Science.gov (United States)

Goindi, Shishu; Kaur, Ramanpreet; Kaur, Randeep

2015-11-30

In this paper, we report an ionic liquid-in-water (IL/w) microemulsion (ME) formulation which is able to solubilize etodolac (ETO), a poorly water soluble drug for topical delivery using BMIMPF6 (1-butyl-3-methylimidazolium hexafluorophosphate) as IL, Tween 80 as surfactant and ethanol as co-surfactant. The prepared ME was characterized for physicochemical parameters, subjected to ex-vivo permeation studies as well as in-vivo pharmacodynamic evaluation. The ex-vivo drug permeation studies through rat skin was performed using Franz-diffusion cell and the IL/w based ME showed maximum mean cumulative percent permeation of 99.030±0.921% in comparison to oil-in-water (o/w) ME (61.548±1.875%) and oily solution (48.830±2.488%) of ETO. In-vivo anti-arthritic and anti-inflammatory activities of the prepared formulations were evaluated using different rodent models and the results revealed that ETO loaded IL/w based ME was found to be more effective in controlling inflammation than oily solution, o/w ME and marketed formulation of ETO. Histopathological studies also demonstrated that IL/w based ME caused no anatomical and pathological changes in the skin. Copyright © 2015 Elsevier B.V. All rights reserved.

Human Precision-Cut Liver Slices as an ex Vivo Model to Study Idiosyncratic Drug-Induced Liver Injury

NARCIS (Netherlands)

Hadi, Mackenzie; Westra, Inge M.; Starokozhko, Viktoriia; Dragovic, Sanja; Merema, M.T.; Groothuis, Geny M. M.

Idiosyncratic drug-induced liver injury (IDILI) is a major problem during drug development and has caused drug withdrawal and black-box warnings. Because of the low concordance of the hepatotoxicity of drugs in animals and humans, robust screening methods using human tissue are needed to predict

Hilar Renal Artery Aneurysm - Ex-vivo Reconstruction and Autotransplantation.

Science.gov (United States)

Pinto Sousa, Pedro; Veiga, Carlos; Matos, Arlindo; Sá Pinto, Pedro; Almeida, Rui

2017-01-01

Renal artery aneurysm (RAA) is a rare clinical entity with an estimated prevalence of 0.15% to 0.1%in the general population. The majority of patients present asymptomatically and the diagnosis is made incidentally during a hypertension study test, and more rarely, fortuitously after backache. Indications to treat have been subject of intense debate, nevertheless there seems to be some consensus that RAAs greater than 2 cm in diameter, expanding RAA, with thrombus or in pregnant women should be treated. Treatment options vary between surgical or endovascular approach. The complex (hilar) RAA constitute a subset of RAA that present a therapeutic dilemma because of their anatomic location and may require extracorporeal arterial reconstruction and auto-transplantation. We describe a 71-year-old woman with a personal history of hypertension for more than twenty years but normal renal function. Following the study for an abdominal discomfort a complex RAA was incidentally diagnosed. Computed tomographic angiography with three-dimensional reconstruction revealed a 13mm, saccular aneurysm located at the right renal hilum. We performed hand-assisted laparoscopic nephrectomy with ex vivo repair of the RAA. The aneurysm was resected and a polar renal artery was implanted over the resected area with a latero-terminal anastomosis. Complementarily, the renal vein was augmented with a spiral great saphenous vein graft and finally the kidney was implanted into the right iliac fossa. The intervention and postoperative course were uneventful and the patient submitted to ultrasound evaluation on the day after procedure. It revealed normal renal perfusion with normal flow indices. In the last follow-up realized, two months after surgery the patient was alive with a well-functioning auto-transplant. RAA may be nowadays more frequently diagnosed due to the increasing use of imaging techniques. While renal artery trunk aneurysms are most often treated using an endovascular procedure it

Ex vivo modulation of the Foxo1 phosphorylation state does not lead to dysfunction of T regulatory cells.

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Kristen Kelley Penberthy

Full Text Available Peripheral regulatory CD4+ T cells (Treg cells prevent maladaptive inflammatory responses to innocuous foreign antigens. Treg cell dysfunction has been linked to many inflammatory diseases, including allergic airway inflammation. Glucocorticoids that are used to treat allergic airway inflammation and asthma are thought to work in part by promoting Treg cell differentiation; patients who are refractory to these drugs have defective induction of anti-inflammatory Treg cells. Previous observations suggest that Treg cells deficient in the transcription factor FoxO1 are pro-inflammatory, and that FoxO1 activity is regulated by its phosphorylation status and nuclear localization. Here, we asked whether altering the phosphorylation state of FoxO1 through modulation of a regulatory phosphatase might affect Treg cell function. In a mouse model of house dust mite-induced allergic airway inflammation, we observed robust recruitment of Treg cells to the lungs and lymph nodes of diseased mice, without an apparent increase in the Treg cytokine interleukin-10 in the airways. Intriguingly, expression of PP2A, a serine/threonine phosphatase linked to the regulation of FoxO1 phosphorylation, was decreased in the mediastinal lymph nodes of HDM-treated mice, mirroring the decreased PP2A expression seen in peripheral blood monocytes of glucocorticoid-resistant asthmatic patients. When we asked whether modulation of PP2A activity alters Treg cell function via treatment with the PP2A inhibitor okadaic acid, we observed increased phosphorylation of FoxO1 and decreased nuclear localization. However, dysregulation of FoxO1 did not impair Treg cell differentiation ex vivo or cause Treg cells to adopt a pro-inflammatory phenotype. Moreover, inhibition of PP2A activity did not affect the suppressive function of Treg cells ex vivo. Collectively, these data suggest that modulation of the phosphorylation state of FoxO1 via PP2A inhibition does not modify Treg cell function ex

Ex Vivo Produced Oral Mucosa Equivalent by Using the Direct Explant Cell Culture Technique

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Kamile Öztürk

2012-09-01

Full Text Available Objective: The aim of this study is the histological and immunohistochemical evaluation of ex vivo produced oral mucosal equivalents using keratinocytes cultured by direct explant technique.Material and Methods: Oral mucosa tissue samples were obtained from the keratinized gingival tissues of 14 healthy human subjects. Human oral mucosa keratinocytes from an oral mucosa biopsy specimen were dissociated by the explant technique. Once a sufficient population of keratinocytes was reached, they were seeded onto the type IV collagen coated “AlloDerm” and taken for histological and immunohistochemical examinations at 11 days postseeding of the keratinocytes on the cadaveric human dermal matrix.Results: Histopathologically and immunohistochemically, 12 out of 14 successful ex vivo produced oral mucosa equivalents (EVPOME that consisted of a stratified epidermis on a dermal matrix have been developed with keratinocytes cultured by the explant technique.Conclusion: The technical handling involved in the direct explant method at the beginning of the process has fewer steps than the enzymatic method and use of the direct explant technique protocol for culturing of human oral mucosa keratinocyte may be more adequate for EVPOME production.

Characterization of micro-invasive trabecular bypass stents by ex vivo perfusion and computational flow modeling

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Hunter KS

2014-03-01

Full Text Available Kendall S Hunter,1 Todd Fjield,2 Hal Heitzmann,2 Robin Shandas,1 Malik Y Kahook3 1Department of Bioengineering, University of Colorado Denver, Aurora, CO, USA; 2Glaukos Corporation, Laguna Hills, CA, USA; 3University of Colorado Hospital Eye Center, Aurora, CO, USA Abstract: Micro-invasive glaucoma surgery with the Glaukos iStent® or iStent inject® (Glaukos Corporation, Laguna Hills, CA, USA is intended to create a bypass through the trabecular meshwork to Schlemm's canal to improve aqueous outflow through the natural physiologic pathway. While the iStent devices have been evaluated in ex vivo anterior segment models, they have not previously been evaluated in whole eye perfusion models nor characterized by computational fluid dynamics. Intraocular pressure (IOP reduction with the iStent was evaluated in an ex vivo whole human eye perfusion model. Numerical modeling, including computational fluid dynamics, was used to evaluate the flow through the stents over physiologically relevant boundary conditions. In the ex vivo model, a single iStent reduced IOP by 6.0 mmHg from baseline, and addition of a second iStent further lowered IOP by 2.9 mmHg, for a total IOP reduction of 8.9 mmHg. Computational modeling showed that simulated flow through the iStent or iStent inject is smooth and laminar at physiological flow rates. Each stent was computed to have a negligible flow resistance consistent with an expected significant decrease in IOP. The present perfusion results agree with prior clinical and laboratory studies to show that both iStent and iStent inject therapies are potentially titratable, providing clinicians with the opportunity to achieve lower target IOPs by implanting additional stents. Keywords: glaucoma, iStent, trabecular bypass, intraocular pressure, ab-interno, CFD

Ex-Vivo Cow Skin Viscoelastic Effect for Tribological Aspects in Endoprosthesis

Science.gov (United States)

Subhi, K. A.; Tudor, A.; Hussein, E. K.; Wahad, H.; Chisiu, G.

2018-01-01

The viscoelastic behavior of ex-vivo cow skin was experimentally studied by applied load from different indenter types (circle, square and triangle, all types have the same area) for different times (10 sec, 30 sec, and 60 sec). The viscoelastic tests were carried out using a UMT series (UMT-II, CETR Corporation). The experimental results collected at different operating conditions showed that the cow skin has a higher reaction against the triangle indenter compared to the other shapes. Whereas the hysteresis of cow skin was lower at low applied load time and it's increased when the time increased.

[Formulation aspects and ex-vivo examination of buccal drug delivery systems].

Science.gov (United States)

Szabó, Barnabás; Hetényi, Gergely; Majoros, Klaudia; Miszori, Veronika; Kállai, Nikolett; Zelkó, Romána

2011-01-01

Application of buccal dosage forms has several advantages. Buccal route can be used for systemic delivery because the mucosa has a rich blood supply and it is relatively permeable. This route of drug delivery is of special advantages, including the bypass of first pass effect and the avoidance of presystemic elimination within the GIT. Buccal delivery systems enable the systemic delivery of peptides and proteins. In our previous study the physiological background of this application and the excipients of the possible formulations were reviewed. In the present work the formulation and ex vivo examination aspects of buccal drug delivery systems are summarized.

Human precision-cut liver slices as an ex vivo model to study idiosyncratic drug-induced liver injury

NARCIS (Netherlands)

Hadi, Mackenzie; Westra, Inge; Starokozhko, Viktoriia; Dragovic, Sanja; Merema, Maja; Groothuis, Genoveva

2013-01-01

Idiosyncratic drug-induced liver injury (IDILI) is a major problem during drug development and has caused drug withdrawal and black-box warnings. Due to the low concordance of the hepatotoxicity of drugs in animals and humans, robust screening methods using human tissue are needed to predict and to

Ex Vivo Reconstruction and Autotransplantation for Hilar Renal Artery Aneurysms in Patients with Congenital Anomalies.

Science.gov (United States)

Adeyemi, Jaiyeola; Johnson, Jacob; Rits, Yevgeniy; Akingba, A George; Rubin, Jeffrey

2018-02-01

Renal artery aneurysms (RAAs) are an uncommon finding but are more often associated with other congenital disorders. The complex (hilar) RAAs constitute a subset of RAAs that present a therapeutic dilemma for the vascular surgeon because of their anatomic location. This dilemma worsens when hilar RAAs occur with a solitary kidney where organ preservation is vital. Ex vivo reconstruction with autotransplantation is especially suitable for hilar RAAs, even when they are associated with a solitary kidney. We report 2 of such cases of RAAs with a solitary kidney in patients with pertinent congenital anomalies. In 1 case, the hilar RAA was associated with a significant accessory renal artery, whereas in the other case, the hilar RAA was associated with a significant connective tissue disorder. Ex vivo reconstruction and autotransplantation was successful in both cases; however, treatment modalities had to be adapted to the patient's unique conditions. Copyright © 2017 Elsevier Inc. All rights reserved.

Ex vivo confocal microscopy: an emerging technique in dermatology

Science.gov (United States)

Perrot, Jean Luc; Labeille, Bruno; Cambazard, Frédéric; Rubegni, Pietro

2018-01-01

This review aims to give an overview of the current available applications of ex vivo confocal microscopy (EVCM) in dermatology. EVCM is a relatively new imaging technique that allows microscopic examination of freshly excised unfixed tissue. It enables a rapid examination of the skin sample directly in the surgery room and thus represents an alternative to the intraoperative micrographic control of the surgical margins of cutaneous tumors by standard microscopic examination on cryopreserved sections during Mohs surgery. Although this technique has mainly been developed for the margin’s control of basal cell carcinoma, many other skin tumors have been studied, including melanoma. Use of EVCM is continuing to evolve, and many possible applications are under investigation, such as the study of nails and hair diseases and the diagnosis of skin infections. PMID:29785327

Modelo de conductancia hidráulica de la dentina humana ex vivo

OpenAIRE

Hevia, J.; Fresno, C.; Martín, J.; Moncada, G.; Letelier, C.; Oliveira Junior, O.B.; Fernández, E.

2013-01-01

El objetivo principal de este trabajo fue montar y probar un modelo experimental para medir la conductancia hidráulica de la dentina ex vivo. Diecisiete terceros molares sanos, con indicación de exodoncia, de donantes sanos de edades entre 15 y 30 años fueron obtenidos mediante consentimiento informado. Luego de limpiarlos, desinfectarlos, incluirlos en resina epóxica y cortarlos se obtuvieron 17 muestras de dentina, correspondiente a un disco de resina con un corte coronal de diente que pres...

HBV-Derived Synthetic Long Peptide Can Boost CD4+ and CD8+ T-Cell Responses in Chronic HBV Patients Ex Vivo.

Science.gov (United States)

Dou, Yingying; van Montfoort, Nadine; van den Bosch, Aniek; de Man, Robert A; Zom, Gijs G; Krebber, Willem-Jan; Melief, Cornelis J M; Buschow, Sonja I; Woltman, Andrea M

2018-02-14

Vaccination with synthetic long peptides (SLP) is a promising new treatment strategy for chronic hepatitis B virus (CHB). SLP can induce broad T-cell responses for all HLA types. Here we investigated the ability of a prototype HBV-core (HBc)-sequence-derived SLP to boost HBV-specific T cells in CHB patients ex vivo. HBc-SLP was used to assess cross-presentation by monocyte-derived dendritic cells (moDC) and BDCA1+ blood myeloid DC (mDC) to engineered HBV-specific CD8+ T cells. Autologous SLP-loaded and toll-like receptor (TLR)-stimulated DC were used to activate patient HBc-specific CD8+ and CD4+ T cells. HBV-SLP was cross-presented by moDC, which was further enhanced by adjuvants. Patient-derived SLP-loaded moDC significantly increased autologous HBcAg18-27-specific CD8+ T cells and CD4+ T cells ex vivo. HBV-specific T cells were functional as they synthesized tumor necrosis factor-alpha and interferon-gamma. In 6/7 of patients blockade of PD-L1 further increased SLP effects. Also, importantly, patient-derived BDCA1+ mDC cross-presented and activated autologous T-cell responses ex vivo. As a proof of concept, we showed a prototype HBc-SLP can boost T-cell responses in patients ex vivo. These results pave the way for the development of a therapeutic SLP-based vaccine to induce effective HBV-specific adaptive immune responses in CHB patients. © The Author(s) 2017. Published by Oxford University Press for the Infectious Diseases Society of America.

Arg354 in the catalytic centre of bovine liver catalase is protected from methylglyoxal-mediated glycation.

Science.gov (United States)

Scheckhuber, Christian Q

2015-12-30

In addition to controlled post-translational modifications proteins can be modified with highly reactive compounds. Usually this leads to a compromised functionality of the protein. Methylglyoxal is one of the most common agents that attack arginine residues. Methylglyoxal is also regarded as a pro-oxidant that affects cellular redox homeostasis by contributing to the formation of reactive oxygen species. Antioxidant enzymes like catalase are required to protect the cell from oxidative damage. These enzymes are also targets for methylglyoxal-mediated modification which could severely affect their catalytic activity in breaking down reactive oxygen species to less reactive or inert compounds. Here, bovine liver catalase was incubated with high levels of methylglyoxal to induce its glycation. This treatment did not lead to a pronounced reduction of enzymatic activity. Subsequently methylglyoxal-mediated arginine modifications (hydroimidazolone and dihydroxyimidazolidine) were quantitatively analysed by sensitive nano high performance liquid chromatography/electron spray ionisation/tandem mass spectrometry. Whereas several arginine residues displayed low to moderate levels of glycation (e.g., Arg93, Arg365, Arg444) Arg354 in the active centre of catalase was never found to be modified. Bovine liver catalase is able to tolerate very high levels of the modifying α-oxoaldehyde methylglyoxal so that its essential enzymatic function is not impaired.

Use of an ex vivo local lymph node assay to assess contact hypersensitivity potential.

Science.gov (United States)

Piccotti, Joseph R; Kawabata, Thomas T

2008-07-01

The local lymph node assay (LLNA) is used to assess the contact hypersensitivity potential of compounds. In the standard assay, mice are treated topically with test compound to the dorsum of both ears on Days 1-3. The induction of a hypersensitivity response is assessed on Day 6 by injecting [(3)H]-thymidine into a tail vein and measuring thymidine incorporation into DNA of lymph node cells draining the ears. The ex vivo LLNA is conducted similarly except lymphocyte proliferation is assessed after in vitro incubation of lymph node cells with [(3)H]-thymidine, which significantly reduces the amount of radioactive waste. The current study tested the use of this approach for hazard assessment of contact hypersensitivity and to estimate allergenic potency. Female BALB/c mice were treated on Days 1-3 with two nonsensitizers (4' -methoxyacetophenone, diethyl phthalate), three weak sensitizers (hydroxycitronellal, eugenol, citral), one weak-to-moderate sensitizer (hexylcinnamic aldehyde), two moderate sensitizers (isoeugenol, phenyl benzoate), and one strong sensitizer (dinitrochlorobenzene). On Day 6, isolated lymph node cells were incubated overnight with [(3)H]-thymidine and thymidine incorporation was measured by liquid scintillation spectrophotometry. The ex vivo LLNA accurately distinguished the contact sensitizers from the nonsensitizing chemicals, and correctly ranked the relative potency of the compounds tested. The EC3 values, i.e., the effective concentration of test substance needed to induce a stimulation index of 3, were as follows: 4' -methoxyacetophenone (> 50%), diethyl phthalate (> 50%), hydroxycitronellal (20.4%), eugenol (13.6%), citral (8.9%), isoeugenol (3.8%), hexylcinnamic aldehyde (2.7%), phenyl benzoate (2%), and dinitrochlorobenzene (0.02%). In addition, low inter-animal and inter-experiment variability was seen with 25% hexyl-cinnamic aldehyde (assay positive control). The results of the ex vivo LLNA in the current study were consistent with

In vitro and in vivo antivirus activity of an anti-programmed death-ligand 1 (PD-L1 rat-bovine chimeric antibody against bovine leukemia virus infection.

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Asami Nishimori

Full Text Available Programmed death-1 (PD-1, an immunoinhibitory receptor on T cells, is known to be involved in immune evasion through its binding to PD-ligand 1 (PD-L1 in many chronic diseases. We previously found that PD-L1 expression was upregulated in cattle infected with bovine leukemia virus (BLV and that an antibody that blocked the PD-1/PD-L1 interaction reactivated T-cell function in vitro. Therefore, this study assessed its antivirus activities in vivo. First, we inoculated the anti-bovine PD-L1 rat monoclonal antibody 4G12 into a BLV-infected cow. However, this did not induce T-cell proliferation or reduction of BLV provirus loads during the test period, and only bound to circulating IgM+ B cells until one week post-inoculation. We hypothesized that this lack of in vivo effects was due to its lower stability in cattle and so established an anti-PD-L1 rat-bovine chimeric antibody (Boch4G12. Boch4G12 was able to bind specifically with bovine PD-L1, interrupt the PD-1/PD-L1 interaction, and activate the immune response in both healthy and BLV-infected cattle in vitro. Therefore, we experimentally infected a healthy calf with BLV and inoculated it intravenously with 1 mg/kg of Boch4G12 once it reached the aleukemic (AL stage. Cultivation of peripheral blood mononuclear cells (PBMCs isolated from the tested calf indicated that the proliferation of CD4+ T cells was increased by Boch4G12 inoculation, while BLV provirus loads were significantly reduced, clearly demonstrating that this treatment induced antivirus activities. Therefore, further studies using a large number of animals are required to support its efficacy for clinical application.

Rac1 and AMPK Account for the Majority of Muscle Glucose Uptake Stimulated by Ex Vivo Contraction but Not In Vivo Exercise

DEFF Research Database (Denmark)

Sylow, Lykke; Møller, Lisbeth; Kleinert, Maximilian

2017-01-01

, but whether those two signaling pathways jointly account for the entire signal to glucose transport is unknown. We therefore studied the ability of contraction and exercise to stimulate glucose transport in isolated muscles with AMPK loss of function combined with either pharmacological inhibition or genetic...... uptake in vivo was only partially reduced by Rac1 mKO with no additive effect of a2KD. It is concluded that Rac1 and AMPK together account for almost the entire ex vivo contraction response in muscle glucose transport, whereas only Rac1, but not a2 AMPK, regulates muscle glucose uptake during submaximal...

Somatostatin receptor expression in the human spleen - Answer to an enigma by ex-vivo and in-vitro autoradiography after 177Lu-DOTA-octreotate administration

International Nuclear Information System (INIS)

Melis, M.; Swart, J. de; Groen, H.C.; Konijnenberg, M.W.; Van der Graaf, L.M.; Kaemmerer, D.; Kulkami, H.R.; Baum, R.P.; Lupp, A.; Saenger, J.; Jong, M. de

2015-01-01

Full text of publication follows. Aim: radiolabelled somatostatin analogues are being used for diagnostic and therapeutic (PRRT) purposes in patients with somatostatin receptor (SSTR) expressing tumours. During PRRT a significant spleen uptake may lead to radiation doses of > 20 Gy. Yet, the threshold dose for spleen radiation induced toxicity is currently unknown. Based on previous 68 Ga-DOTATOC PET/CT studies, demonstrating higher uptake in spleen than in splenosis, white pulp (WP) localization of radioactivity was suggested. This hypothesis was investigated in the current pilot study using the longer lived 177 Lu-DOTA-octreotate. Methods: a patient diagnosed with neuroendocrine neoplasm of the pancreatic tail (SUV max on 68 Ga-DOTATOC PET/CT 100.4) with liver metastasis (SUV 47.3, normal liver SUV 12.5) and uptake in the spleen (SUV 41.0) received 1 GBq 177 Lu-DOTA-octreotate. 2 h after administration whole-body planar scintigraphy and SPECT/CT of the upper abdomen was performed, followed by laparoscopic resection of the pancreatic tumour and splenectomy the next day. After spleen transport from Bad Berka to Rotterdam ex-vivo micro-SPECT of the removed spleen was acquired for 73 min using 2.5 mm diameter pinholes. Spleen fragments (∼10 * 10 * 5 mm) were either snap-frozen in liquid nitrogen or fixed in 10% formalin and paraffin embedded. Ex-vivo autoradiography of 10 μm cryo-sections was performed and serial sections were used for 111 In-DOTA-octreotate in-vitro autoradiography after decay of 177 Lu. FFPE sections were used for HE- and immunostaining for SSTR2A and cell subsets CD4 (Th-cell), CD8 (Ts-cell), CD20 (B-cell) and CD68 (macrophage). Results: 177 Lu-DOTA-octreotate scintigraphy and SPECT/CT demonstrated high uptake in the pancreatic tumor, hepatic metastasis and homogeneously in the normal spleen. High resolution micro-SPECT imaging of the isolated spleen also revealed a relatively homogeneous uptake (calculated rest activity 60 MBq 177 Lu). The

Real-Time Amperometric Recording of Extracellular H2O2 in the Brain of Immunocompromised Mice: An In Vitro, Ex Vivo and In Vivo Characterisation Study

Science.gov (United States)

Reid, Caroline H.; Finnerty, Niall J.

2017-01-01

We detail an extensive characterisation study on a previously described dual amperometric H2O2 biosensor consisting of H2O2 detection (blank) and degradation (catalase) electrodes. In vitro investigations demonstrated excellent H2O2 sensitivity and selectivity against the interferent, ascorbic acid. Ex vivo studies were performed to mimic physiological conditions prior to in vivo deployment. Exposure to brain tissue homogenate identified reliable sensitivity and selectivity recordings up to seven days for both blank and catalase electrodes. Furthermore, there was no compromise in pre- and post-implanted catalase electrode sensitivity in ex vivo mouse brain. In vivo investigations performed in anaesthetised mice confirmed the ability of the H2O2 biosensor to detect increases in amperometric current following locally perfused/infused H2O2 and antioxidant inhibitors mercaptosuccinic acid and sodium azide. Subsequent recordings in freely moving mice identified negligible effects of control saline and sodium ascorbate interference injections on amperometric H2O2 current. Furthermore, the stability of the amperometric current was confirmed over a five-day period and analysis of 24-h signal recordings identified the absence of diurnal variations in amperometric current. Collectively, these findings confirm the biosensor current responds in vivo to increasing exogenous and endogenous H2O2 and tentatively supports measurement of H2O2 dynamics in freely moving NOD SCID mice. PMID:28698470

Cloning and sequencing of the bovine gastrin gene

DEFF Research Database (Denmark)

Lund, T; Rehfeld, J F; Olsen, Jørgen

1989-01-01

In order to deduce the primary structure of bovine preprogastrin we therefore sequenced a gastrin DNA clone isolated from a bovine liver cosmid library. Bovine preprogastrin comprises 104 amino acids and consists of a signal peptide, a 37 amino acid spacer-sequence, the gastrin-34 sequence followed...

High-resolution ex vivo imaging of coronary artery stents using 64-slice computed tomography - initial experience

International Nuclear Information System (INIS)

Rist, Carsten; Nikolaou, Konstantin; Wintersperger, Bernd J.; Reiser, Maximilian F.; Becker, Christoph R.; Flohr, Thomas

2006-01-01

The aim of the study was to evaluate the potential of new-generation multi-slice computed tomography (CT) scanner technology for the delineation of coronary artery stents in an ex vivo setting. Nine stents of various diameters (seven stents 3 mm, two stents 2.5 mm) were implanted into the coronary arteries of ex vivo porcine hearts and filled with a mixture of an iodine-containing contrast agent. Specimens were scanned with a 16-slice CT (16SCT) machine; (Somatom Sensation 16, Siemens Medical Solutions), slice thickness 0.75 mm, and a 64-slice CT (64SCT, Somatom Sensation 64), slice-thickness 0.6 mm. Stent diameters as well as contrast densities were measured, on both the 16SCT and 64SCT images. No significant differences of CT densities were observed between the 16SCT and 64SCT images outside the stent lumen: 265±25HU and 254±16HU (P=0.33), respectively. CT densities derived from the 64SCT images and 16SCT images within the stent lumen were 367±36HU versus 402±28HU, P<0.05, respectively. Inner and outer stent diameters as measured from 16SCT and 64SCT images were 2.68±0.08 mm versus 2.81±0.07 mm and 3.29±0.06 mm versus 3.18±0.07 mm (P<0.05), respectively. The new 64SCT scanner proved to be superior in the ex vivo assessment of coronary artery stents to the conventional 16SCT machine. Increased spatial resolution allows for improved assessment of the coronary artery stent lumen. (orig.)

Time dependence of electrical bioimpedance on porcine liver and kidney under a 50 Hz ac current

International Nuclear Information System (INIS)

Spottorno, J; Rivero, G; Venta, J de la; Multigner, M; Alvarez, L; Santos, M

2008-01-01

The purpose of this work is to study the changes of the bioimpedance from its 'in vivo' value to the values measured in a few hours after the excision from the body. The evolution of electrical impedance with time after surgical extraction has been studied on two porcine organs: the liver and the kidney. Both in vivo and ex vivo measurements of electrical impedance, measuring its real and imaginary components, have been performed. The in vivo measurements have been carried out with the animal anaesthetized. The ex vivo measurements have been made more than 2 h after the extraction of the organ. The latter experiment has been carried out at two different stabilized temperatures: at normal body temperature and at the standard preservation temperature for transplant surgery. The measurements show a correlation between the biological evolution and the electrical bioimpedance of the organs, which increases from its in vivo value immediately after excision, multiplying its value by 2 in a few hours

Time dependence of electrical bioimpedance on porcine liver and kidney under a 50 Hz ac current

Energy Technology Data Exchange (ETDEWEB)

Spottorno, J; Rivero, G; Venta, J de la [Instituto de Magnetismo Aplicado (ADIF-UCM-CSIC), PO Box 155, Las Rozas, Madrid 28230 (Spain); Multigner, M [Departamento de Fisica de Materiales, UCM, Ciudad Universitaria, 28040 Madrid (Spain); Alvarez, L; Santos, M [Centro de Investigacion Biomedica en Red en BioingenierIa, Biomateriales y Nanomedicina (CIBER-BBN), Madrid (Spain)

2008-03-21

The purpose of this work is to study the changes of the bioimpedance from its 'in vivo' value to the values measured in a few hours after the excision from the body. The evolution of electrical impedance with time after surgical extraction has been studied on two porcine organs: the liver and the kidney. Both in vivo and ex vivo measurements of electrical impedance, measuring its real and imaginary components, have been performed. The in vivo measurements have been carried out with the animal anaesthetized. The ex vivo measurements have been made more than 2 h after the extraction of the organ. The latter experiment has been carried out at two different stabilized temperatures: at normal body temperature and at the standard preservation temperature for transplant surgery. The measurements show a correlation between the biological evolution and the electrical bioimpedance of the organs, which increases from its in vivo value immediately after excision, multiplying its value by 2 in a few hours.

The role of metabolism in Diclofenac-induced intestinal toxicity in human ex vivo

NARCIS (Netherlands)

Niu, Xiaoyu; Makkinje, Miriam; de Graaf, Inge; Groothuis, Genoveva

2012-01-01

The use of Diclofenac (DCF: 2-(2,6-dichloranilino) phenyl acetic acid ), a non-steroidal anti-inflammatory drug is associated with severe gastro-intestinal side-effects. In vivo rat studies suggest that reactive metabolites of DCF, produced by the liver, play an important role in the intestinal

Liver-derived systemic factors drive β-cell hyperplasia in insulin resistant states

Energy Technology Data Exchange (ETDEWEB)

El Ouaamari, Abdelfattah; Kawamori, Dan; Dirice, Ercument; Liew, Chong Wee; Shadrach, Jennifer L.; Hu, Jiang; Katsuta, Hitoshi; Hollister-Lock, Jennifer; Qian, Weijun; Wagers, Amy J.; Kulkarni, Rohit N.

2013-02-21

Integrative organ cross-talk regulates key aspects of energy homeostasis and its dysregulation may underlie metabolic disorders such as obesity and diabetes. To test the hypothesis that cross-talk between the liver and pancreatic islets modulates β-cell growth in response to insulin resistance, we used the Liver-specific Insulin Receptor Knockout (LIRKO) mouse, a unique model that exhibits dramatic islet hyperplasia. Using complementary in vivo parabiosis and transplantation assays, and in vitro islet culture approaches, we demonstrate that humoral, non-neural, non-cell autonomous factor(s) induce β-cell proliferation in LIRKO mice. Furthermore, we report that a hepatocyte-derived factor(s) stimulates mouse and human β-cell proliferation in ex vivo assays, independent of ambient glucose and insulin levels. These data implicate the liver as a critical source of β-cell growth factors in insulin resistant states.

Analysis of Endothelial Adherence of Bartonella henselae and Acinetobacter baumannii Using a Dynamic Human Ex Vivo Infection Model

OpenAIRE

Weidensdorfer, Marko; Chae, Ju Ik; Makobe, Celestine; Stahl, Julia; Averhoff, Beate; Müller, Volker; Schürmann, Christoph; Brandes, Ralf P.; Wilharm, Gottfried; Ballhorn, Wibke; Christ, Sara; Linke, Dirk; Fischer, Doris; Göttig, Stephan; Kempf, Volkhard A. J.

2016-01-01

Bacterial adherence determines the virulence of many human-pathogenic bacteria. Experimental approaches elucidating this early infection event in greater detail have been performed using mainly methods of cellular microbiology. However, in vitro infections of cell monolayers reflect the in vivo situation only partially, and animal infection models are not available for many human-pathogenic bacteria. Therefore, ex vivo infection of human organs might represent an attractive method to overcome...

Ex-Vivo percutaneous absorption of enrofloxacin: Comparison of LMOG organogel vs. pentravan cream.

Science.gov (United States)

Kirilov, Plamen; Tran, Van Hung; Ducrotté-Tassel, Alban; Salvi, Jean-Paul; Perrot, Sébastien; Haftek, Marek; Boulieu, Roselyne; Pirot, Fabrice

2016-02-10

The objective of this study was to investigate the percutaneous absorption of enrofloxacin from two base formulations, Pentravan cream and LMOG organogel. Ex-vivo experiments were carried out on pig ear skin. The percutaneous permeation through pig skin of two formulations containing 5 wt% of enrofloxacin was measured and compared using Franz diffusion cells. At appropriate intervals up to 120 h, diffusion samples were taken and analyzed using HPLC assays. Permeation profiles were established and the parameters Tlag and flux values were calculated. In this ex-vivo study, the flux values were 0.35 μgcm(-2)h(-1) for Pentravan and 1.22 μgcm(-2)h(-1) for LMOG organogel, corresponding respectively to 7.9 % and 29.3 % of enrofloxacin absorbed after 120 h by these formulations. The lag time (T lag) of Pentravan and organogel were 6.32 and 0.015 h respectively. The absorption time to reach the antibiotic concentration of enrofloxacin (2 μgmL(-1)) in the receptor was 60 h with Pentravan and 30 h with the organogel, suggesting more effective treatment by the latter. Enrofloxacin contained in organogel could be absorbed through pig ear skin 3.7 times greater than that in Pentravan (commercial formulation). This study demonstrates the perspective of organogel formulations as potential drug delivery systems. Copyright © 2015 Elsevier B.V. All rights reserved.

Macro- and micronutrient disposition in an ex vivo model of extracorporeal membrane oxygenation.

Science.gov (United States)

Estensen, Kristine; Shekar, Kiran; Robins, Elissa; McDonald, Charles; Barnett, Adrian G; Fraser, John F

2014-12-01

Extracorporeal membrane oxygenation (ECMO) circuits have been shown to sequester circulating blood compounds such as drugs based on their physicochemical properties. This study aimed to describe the disposition of macro- and micronutrients in simulated ECMO circuits. Following baseline sampling, known quantities of macro- and micronutrients were injected post oxygenator into ex vivo ECMO circuits primed with the fresh human whole blood and maintained under standard physiologic conditions. Serial blood samples were then obtained at 1, 30 and 60 min and at 6, 12 and 24 h after the addition of nutrients, to measure the concentrations of study compounds using validated assays. Twenty-one samples were tested for thirty-one nutrient compounds. There were significant reductions (p single-dose ex vivo circuit study. Most significantly, there is potential for circuit loss of essential amino acid isoleucine and lipid soluble vitamins (A and E) in the ECMO circuit, and the mechanisms for this need further exploration. While the reductions in glucose concentrations and an increase in other macro- and micronutrient concentrations probably reflect cellular metabolism and breakdown, the decrement in arginine and glutamine concentrations may be attributed to their enzymatic conversion to ornithine and glutamate, respectively. While the results are generally reassuring from a macronutrient perspective, prospective studies in clinical subjects are indicated to further evaluate the influence of ECMO circuit on micronutrient concentrations and clinical outcomes.

Ex vivo fracture resistance of direct resin composite complete crowns with and without posts on maxillary premolars.

NARCIS (Netherlands)

Fokkinga, W.A.; Bell, A.M. Le; Kreulen, C.M.; Lassila, L.V.; Vallittu, P.K.; Creugers, N.H.J.

2005-01-01

AIM: To investigate ex vivo the fracture resistance and failure mode of direct resin composite complete crowns with and without various root canal posts made on maxillary premolars. METHODOLOGY: The clinical crowns of 40 human extracted single-rooted maxillary premolars were sectioned at the

Vasomotor function in rat arteries after ex vivo and intragastric exposure to food-grade titanium dioxide and vegetable carbon particles

DEFF Research Database (Denmark)

Jensen, Ditte Marie; Christophersen, Daniel Vest; Sheykhzade, Majid

2018-01-01

-grade particle exposure on vasomotor function and systemic oxidative stress in an ex vivo study and intragastrically exposed rats.Methods: In an ex vivo study, aorta rings from naive Sprague-Dawley rats were exposed for 30 min to food-grade TiO2 (E171), benchmark TiO2 (Aeroxide P25), food-grade vegetable carbon...... (E153) or benchmark carbon black (Printex 90). Subsequently, the vasomotor function was assessed in wire myographs. In an in vivo study, lean Zucker rats were exposed intragastrically once a week for 10 weeks to vehicle, E171 or E153. Doses were comparable to human daily intake. Vasomotor function...... no differences between groups.Conclusion: Gastrointestinal tract exposure to E171 and E153 was associated with modest albeit statistically significant alterations in the vasocontraction and vasorelaxation responses. Direct particle exposure to aorta rings elicited a similar type of response. The vasomotor...

Comparative kinetic characterization of catalases from Candida boidinii yeast and bovine liver.

Science.gov (United States)

Metelitza, D I; Eryomin, A N; Artzukevich, I M; Chernikevich, I P

1997-04-01

Catalase with molecular weight 230 +/- kD was isolated and purified from methylotrophic yeasts Candida boidinii by ion-exchange chromatography. The kinetic characteristics of yeast and bovine liver catalases were compared in the reaction of H2O2 decomposition using a wide range of H2O2 concentrations (up to 0.12 M) and PH (2-10). First order rates constants (k, sec-1) were determined for both enzymes from semi-logarithmic anamorphoses of kinetic curves of H2O2 utilization. Anamorphoses of complete kinetic curves as a function of 1/ln([H2O2]0/[H2O2]t) versus 1/t were used for calculation of the effective rate constants of catalase inactivation during the reaction (k(in), sec-1) and the rate constants of interaction of catalase complex I with the second molecule of H2O2 (k2, M-1.sec-1). The effects of initial catalase concentrations, H2O2, and pH on k, k2, and k(in) were similar for both enzymes. Catalytic constant, k2, and the efficacy expressed as a ratio kcat/Km were 1.87-, 1.45-, and 1.3-fold, respectively, higher for bovine catalase than that of yeast catalase. Operational stability of yeast catalase is 3.5-fold higher than the stability of bovine catalase and much higher during cyclic decomposition of 50 mM H2O2. Enhanced operational stability and inexpensive source of its preparation open prospects for practical applications of yeast catalase for co-immobilization with superoxide dismutase on non-toxic carriers.

Ex Vivo and In Vivo Evaluation of the Norepinephrine Transporter Ligand [11C]MRB for Brown Adipose Tissue Imaging

International Nuclear Information System (INIS)

Lin Shufei; Fan Xiaoning; Yeckel, Catherine Weikart; Weinzimmer, David; Mulnix, Tim; Gallezot, Jean-Dominique; Carson, Richard E.; Sherwin, Robert S.; Ding Yushin

2012-01-01

Introduction: It has been suggested that brown adipose tissue (BAT) in humans may play a role in energy balance and obesity. We conducted ex vivo and in vivo evaluation using [ 11 C]MRB, a highly selective NET (norepinephrine transporter) ligand for BAT imaging at room temperature, which is not achievable with [ 18 F]FDG. Methods: PET images of male Sprague–Dawley rats with [ 18 F]FDG and [ 11 C]MRB were compared. Relative [ 18 F]FDG or [ 11 C]MRB retention at 20, 40 and 60 min post-injection was quantified on awake rats after exposing to cold (4 °C for 4 h) or remaining at room temperature. Rats pretreated with unlabeled MRB or nisoxetine 30 min before [ 11 C]MRB injection were also assessed. The [ 11 C]MRB metabolite profile in BAT was evaluated. Results: PET imaging demonstrated intense [ 11 C]MRB uptake (SUV of 2.9 to 3.3) in the interscapular BAT of both room temperature and cold-exposed rats and this uptake was significantly diminished by pretreatment with unlabeled MRB; in contrast, [ 18 F]FDG in BAT was only detected in rats treated with cold. Ex vivo results were concordant with the imaging findings; i.e. the uptake of [ 11 C]MRB in BAT was 3 times higher than that of [ 18 F]FDG at room temperature (P = 0.009), and the significant cold-stimulated uptake in BAT with [ 18 F]FDG (10-fold, P = 0.001) was not observed with [ 11 C]MRB (P = 0.082). HPLC analysis revealed 94%–99% of total radioactivity in BAT represented unchanged [ 11 C]MRB. Conclusions: Our study demonstrates that BAT could be specifically labeled with [ 11 C]MRB at room temperature and under cold conditions, supporting a NET-PET strategy for imaging BAT in humans under basal conditions.

Curcumin loaded nano globules for solubility enhancement: preparation, characterization and ex vivo release study.

Science.gov (United States)

Kumar, Anil; Ahuja, Alka; Ali, Javed; Baboota, Sanjula

2012-11-01

Curcumin in spite of being an effective chemotherapeutic agent against different type of cancer, suffer from the problem of low systemic bioavailability due to low aqueous solubility, extensive intestinal metabolism and first-pass metabolism when administered via the oral route. The aim of present investigation was to evaluate the potential of nano globules based nanoemulsion formulation for the solubility enhancement of curcumin. The nano globules based formulation was developed using Labrafac Lipophile WL 1349, Unitop FFT 40, PEG 400 and distilled water as an oil, surfactant, co-surfactant and aqueous phase respectively using aqueous titration method. Furthermore, different formulations were subjected to physical stability and consequently evaluated for ex vivo permeation using small intestine. The optimized formulation had small average globule diameter of 58 nm with zeta potential of -32 mv which indicated long-term dispersion stability. The globules were spherical in shape as observed by Transmission electron microscopy. During ex vivo study, the release of curcumin from nanoemulsion was 96.21% and 98.1% in 6 h and 12 h respectively whereas CU suspension was release up to 28.2% at the end of 12 h. This indicated the enhancement of solubility of curcumin in aqueous solution which is the rate limiting step in the absorption of curcumin in the intestine.

Survival of cord blood haematopoietic stem cells in a hyaluronan hydrogel for ex vivo biomimicry.

Science.gov (United States)

Demange, Elise; Kassim, Yusra; Petit, Cyrille; Buquet, Catherine; Dulong, Virginie; Cerf, Didier Le; Buchonnet, Gérard; Vannier, Jean-Pierre

2013-11-01

Haematopoietic stem cells (HSCs) and haematopoietic progenitor cells (HPCs) grow in a specified niche in close association with the microenvironment, the so-called 'haematopoietic niche'. Scaffolds have been introduced to overcome the liquid culture limitations, mimicking the presence of the extracellular matrix (ECM). In the present study the hyaluronic acid scaffold, already developed in the laboratory, has been used for the first time to maintain long-term cultures of CD34⁺ haematopoietic cells obtained from human cord blood. One parameter investigated was the impact on ex vivo survival of CD34⁺ cord blood cells (CBCs) on the hyaluronic acid surface, immobilized with peptides containing the RGD motif. This peptide was conjugated by coating the hyaluronan hydrogel and cultured in serum-free liquid phase complemented with stem cell factor (SCF), a commonly indispensable cytokine for haematopoiesis. Our work demonstrated that these hyaluronan hydrogels were superior to traditional liquid cultures by maintaining and expanding the HPCs without the need for additional cytokines, and a colonization of 280-fold increment in the hydrogel compared with liquid culture after 28 days of ex vivo expansion. Copyright © 2012 John Wiley & Sons, Ltd.

Selective ex-vivo photothermal ablation of human pancreatic cancer with albumin functionalized multiwalled carbon nanotubes.

Science.gov (United States)

Mocan, Lucian; Tabaran, Flaviu A; Mocan, Teodora; Bele, Constantin; Orza, Anamaria Ioana; Lucan, Ciprian; Stiufiuc, Rares; Manaila, Ioana; Iulia, Ferencz; Dana, Iancu; Zaharie, Florin; Osian, Gelu; Vlad, Liviu; Iancu, Cornel

2011-01-01

The process of laser-mediated ablation of cancer cells marked with biofunctionalized carbon nanotubes is frequently called "nanophotothermolysis". We herein present a method of selective nanophotothermolisys of pancreatic cancer (PC) using multiwalled carbon nanotubes (MWCNTs) functionalized with human serum albumin (HSA). With the purpose of testing the therapeutic value of these nanobioconjugates, we have developed an ex-vivo experimental platform. Surgically resected specimens from patients with PC were preserved in a cold medium and kept alive via intra-arterial perfusion. Additionally, the HSA-MWCNTs have been intra-arterially administered in the greater pancreatic artery under ultrasound guidance. Confocal and transmission electron microscopy combined with immunohistochemical staining have confirmed the selective accumulation of HSA-MWCNTs inside the human PC tissue. The external laser irradiation of the specimen has significantly produced extensive necrosis of the malign tissue after the intra-arterial administration of HSA-MWCNTs, without any harmful effects on the surrounding healthy parenchyma. We have obtained a selective photothermal ablation of the malign tissue based on the selective internalization of MWCNTs with HSA cargo inside the pancreatic adenocarcinoma after the ex-vivo intra-arterial perfusion.

Effect of anticonvulsant drugs on (35S)t-butylbicyclophosphorothionate binding in vitro and ex vivo

International Nuclear Information System (INIS)

Pitkaenen, A.; Riekkinen, P.J.; Saano, V.; Tuomisto, L.

1987-01-01

Using several concentrations of eight anticonvulsant drugs in clinical use (carbamazepine, clonazepam, phenytoin, phenobarbital, ethosuximide, primidone, sodium valproate, and D,L-γ-vinyl GABA), we studied their abilities in vitro to displace ( 35 S)t-butylbicyclophosphorothionate ( 35 S-TBPS) from its binding site in a homogenate of rat brain. Thereafter ethosuximide (150 mg/kg), phenobarbital (30 mg/kg), clonazepam (0.3 mg/kg), or phenytoin (100 mg/kg) was injected intraperitoneally into rats for 16-20 days; and the effect of drug administration on 35 S-TBPS binding was studied in the cortex and hippocampus ex vivo. Phenobarbital (100 μM, P 35 S-TBPS binding in vitro by 10-16%. After drug administration of phenobarbital (concentration in plasma 168 μM), the number of binding sites decreased and the binding affinity (p 35 S-TBPS binding in vitro at the concentration analogous to therapeutic plasma levels or ex vivo at the dose used. These results suggest that the use of phenobarbital may modulate the TBPS binding site, but the role of the present findings in the anticonvulsant action of phenobarbital needs to be further studied. (author)

Phosphatidylcholine contributes to in vivo {sup 31}P MRS signal from the human liver

Energy Technology Data Exchange (ETDEWEB)

Chmelik, Marek; Bogner, Wolfgang; Gajdosik, Martin; Gruber, Stephan; Trattnig, Siegfried [Medical University of Vienna, MR Centre of Excellence, Department of Biomedical Imaging and Image-guided Therapy, Vienna (Austria); Valkovic, Ladislav [Medical University of Vienna, MR Centre of Excellence, Department of Biomedical Imaging and Image-guided Therapy, Vienna (Austria); Institute of Measurement Science, Slovak Academy of Sciences, Department of Imaging Methods, Bratislava (Slovakia); Wolf, Peter; Krebs, Michael [Medical University of Vienna, Division of Endocrinology and Metabolism, Department of Internal Medicine III, Vienna (Austria); Halilbasic, Emina; Trauner, Michael [Medical University of Vienna, Division of Gastroenterology and Hepatology, Department of Internal Medicine III, Vienna (Austria); Krssak, Martin [Medical University of Vienna, MR Centre of Excellence, Department of Biomedical Imaging and Image-guided Therapy, Vienna (Austria); Medical University of Vienna, Division of Endocrinology and Metabolism, Department of Internal Medicine III, Vienna (Austria)

2015-07-15

To demonstrate the overlap of the hepatic and bile phosphorus ({sup 31}P) magnetic resonance (MR) spectra and provide evidence of phosphatidylcholine (PtdC) contribution to the in vivo hepatic {sup 31}P MRS phosphodiester (PDE) signal, suggested in previous reports to be phosphoenolpyruvate (PEP). Phantom measurements to assess the chemical shifts of PEP and PtdC signals were performed at 7 T. A retrospective analysis of hepatic 3D {sup 31}P MR spectroscopic imaging (MRSI) data from 18 and five volunteers at 3 T and 7 T, respectively, was performed. Axial images were inspected for the presence of gallbladder, and PDE signals in representative spectra were quantified. Phantom experiments demonstrated the strong pH-dependence of the PEP chemical shift and proved the overlap of PtdC and PEP (∝2 ppm relative to phosphocreatine) at hepatic pH. Gallbladder was covered in seven of 23 in vivo 3D-MRSI datasets. The PDE{sub gall}/γ-ATP{sub liver} ratio was 4.8-fold higher (p = 0.001) in the gallbladder (PDE{sub gall}/γ-ATP{sub liver} = 3.61 ± 0.79) than in the liver (PDE{sub liver}/γ-ATP{sub liver} = 0.75 ± 0.15). In vivo 7 T {sup 31}P MRSI allowed good separation of PDE components. The gallbladder is a strong source of contamination in adjacent {sup 31}P MR hepatic spectra due to biliary phosphatidylcholine. In vivo {sup 31}P MR hepatic signal at 2.06 ppm may represent both phosphatidylcholine and phosphoenolpyruvate, with a higher phosphatidylcholine contribution due to its higher concentration. (orig.)

Ex vivo activity quantification in micrometastases at the cellular scale using the α-camera technique

DEFF Research Database (Denmark)

Chouin, Nicolas; Lindegren, Sture; Frost, Sofia H L

2013-01-01

Targeted α-therapy (TAT) appears to be an ideal therapeutic technique for eliminating malignant circulating, minimal residual, or micrometastatic cells. These types of malignancies are typically infraclinical, complicating the evaluation of potential treatments. This study presents a method of ex...... vivo activity quantification with an α-camera device, allowing measurement of the activity taken up by tumor cells in biologic structures a few tens of microns....

Umbilical cord bloods hematopoietic stem cells ex vivo expansion (the literature review

Directory of Open Access Journals (Sweden)

T. V. Shamanskaya

2012-01-01

Full Text Available Umbilical cord blood (CB is now an attractive source of hematopoietic stem cells (HSCs for transplantation in pediatric and adult patients with various malignant and non-malignant diseases. However, its clinical application is limited by low cells numbers in graft, which correlates with delayed engraftment, an extension of time to platelets and neutrophils recovery and increasing risk of infectious complications. Several strategies have been suggested to overcome this limitation, one of which is obtaining a sufficient number of hematopoietic progenitor cells by ex vivo expansion. Literature review about CB HSCs expansion in given article is presented.

In vitro and ex vivo evaluations on transdermal delivery of the HIV inhibitor IQP-0410.

Directory of Open Access Journals (Sweden)

Anthony S Ham

Full Text Available The aim of this study was to investigate the physicochemical and in vitro/ex vivo characteristics of the pyrmidinedione IQP-0410 formulated into transdermal films. IQP-0410 is a potent therapeutic anti-HIV nonnucleoside reverse transcriptase inhibitor that would be subjected to extensive first pass metabolism, through conventional oral administration. Therefore, IQP-0410 was formulated into ethyl cellulose/HPMC-based transdermal films via solvent casting. In mano evaluations were performed to evaluate gross physical characteristics. In vitro release studies were performed in both Franz cells and USP-4 dissolution vessels. Ex vivo release and permeability assays were performed on human epidermal tissue models, and the permeated IQP-0410 was collected for in vitro HIV-1 efficacy assays in CEM-SS cells and PBMCs. Film formulation D3 resulted in pliable, strong transdermal films that were loaded with 2% (w/w IQP-0410. Composed of 60% (w/w ethyl cellulose and 20% (w/w HPMC, the films contained < 1.2% (w/w of water and were hygroscopic resulting in significant swelling under humid conditions. The water permeable nature of the film resulted in complete in vitro dissolution and drug release in 26 hours. When applied to ex vivo epidermal tissues, the films were non-toxic to the tissue and also were non-toxic to HIV target cells used in the in vitro efficacy assays. Over a 3 day application, the films delivered IQP-0410 through the skin tissue at a zero-order rate of 0.94 ± 0.06 µg/cm(2/hr with 134 ± 14.7 µM collected in the basal media. The delivered IQP-0410 resulted in in vitro EC50 values against HIV-1 of 2.56 ± 0.40 nM (CEM-SS and 0.58 ± 0.03 nM (PBMC. The film formulation demonstrated no significant deviation from target values when packaged in foil pouches under standard and accelerated environmental conditions. It was concluded that the transdermal film formulation was a potentially viable method of administering IQP-0410 that warrants

Ex-vivo holographic microscopy and spectroscopic analysis of head and neck cancer

Science.gov (United States)

Holler, Stephen; Wurtz, Robert; Auyeung, Kelsey; Auyeung, Kris; Paspaley-Grbavac, Milan; Mulroe, Brigid; Sobrero, Maximiliano; Miles, Brett

2015-03-01

Optical probes to identify tumor margins in vivo would greatly reduce the time, effort and complexity in the surgical removal of malignant tissue in head and neck cancers. Current approaches involve visual microscopy of stained tissue samples to determine cancer margins, which results in the excision of excess of tissue to assure complete removal of the cancer. Such surgical procedures and follow-on chemotherapy can adversely affect the patient's recovery and subsequent quality of life. In order to reduce the complexity of the process and minimize adverse effects on the patient, we investigate ex vivo tissue samples (stained and unstained) using digital holographic microscopy in conjunction with spectroscopic analyses (reflectance and transmission spectroscopy) in order to determine label-free, optically identifiable characteristic features that may ultimately be used for in vivo processing of cancerous tissues. The tissue samples studied were squamous cell carcinomas and associated controls from patients of varying age, gender and race. Holographic microscopic imaging scans across both cancerous and non-cancerous tissue samples yielded amplitude and phase reconstructions that were correlated with spectral signatures. Though the holographic reconstructions and measured spectra indicate variations even among the same class of tissue, preliminary results indicate the existence of some discriminating features. Further analyses are presently underway to further this work and extract additional information from the imaging and spectral data that may prove useful for in vivo surgical identification.

Combination of high-intensity focused ultrasound irradiation and hydroxyapatite nanoparticle injection to injure normal goat liver tissue in vivo without costal bone incision.

Science.gov (United States)

Liu, L; Xiao, Z; Xiao, Y; Wang, Z; Li, F; Li, M; Peng, X

2014-10-20

The aims of this study were to evaluate the in vivo safety of intravenous nano-hydroxyapatite (nano-HA), to explore how nano-HA might influence the effects of high-intensity focused ultrasound (HIFU) on normal liver tissue, and to investigate whether intravenous nano-HA could enhance HIFU for hepatocellular carcinoma ablation in a goat model. The present study, for the first time, indicated that the delivery of abundant nano-HA into the body over short periods of time could be assembled by the hepatic reticuloendothelial system, subsequently leading to a rapid rise of ultrasound-induced overheating, and ultimately resulting in enlargement of the coagulation necrotic area for ablated hepatocellular carcinoma in goats both in vivo and ex vivo. On the other hand, therapeutic doses of nano-HA were much lower than the lethal dose, and consequently presented transient and mild abnormalities of hepatic enzymes and renal function during the first 24 h after nano-HA injection. These results suggested that the combined application of nano-HA and HIFU is potentially a more effective alternative option compared to surgery for hepatocellular carcinoma local ablation in a safe and feasible manner.

Guided access cavity preparation using cone-beam computed tomography and optical surface scans - an ex vivo study

DEFF Research Database (Denmark)

Buchgreitz, J; Buchgreitz, M; Mortensen, D

2016-01-01

AIM: To evaluate ex vivo, the accuracy of a preparation procedure planned for teeth with pulp canal obliteration (PCO) using a guide rail concept based on a cone-beam computed tomography (CBCT) scan merged with an optical surface scan. METHODOLOGY: A total of 48 teeth were mounted in acrylic bloc...

Aging of monolithic zirconia dental prostheses: Protocol for a 5-year prospective clinical study using ex vivo analyses.

Science.gov (United States)

Koenig, Vinciane; Wulfman, Claudine P; Derbanne, Mathieu A; Dupont, Nathalie M; Le Goff, Stéphane O; Tang, Mie-Leng; Seidel, Laurence; Dewael, Thibaut Y; Vanheusden, Alain J; Mainjot, Amélie K

2016-12-15

Recent introduction of computer-aided design/computer-aided manufacturing (CAD/CAM) monolithic zirconia dental prostheses raises the issue of material low thermal degradation (LTD), a well-known problem with zirconia hip prostheses. This phenomenon could be accentuated by masticatory mechanical stress. Until now zirconia LTD process has only been studied in vitro . This work introduces an original protocol to evaluate LTD process of monolithic zirconia prostheses in the oral environment and to study their general clinical behavior, notably in terms of wear. 101 posterior monolithic zirconia tooth elements (molars and premolars) are included in a 5-year prospective clinical trial. On each element, several areas between 1 and 2 mm 2 (6 on molars, 4 on premolars) are determined on restoration surface: areas submitted or non-submitted to mastication mechanical stress, glazed or non-glazed. Before prosthesis placement, ex vivo analyses regarding LTD and wear are performed using Raman spectroscopy, SEM imagery and 3D laser profilometry. After placement, restorations are clinically evaluated following criteria of the World Dental Federation (FDI), complemented by the analysis of fracture clinical risk factors. Two independent examiners perform the evaluations. Clinical evaluation and ex vivo analyses are carried out after 6 months and then each year for up to 5 years. For clinicians and patients, the results of this trial will justify the use of monolithic zirconia restorations in dental practice. For researchers, the originality of a clinical study including ex vivo analyses of material aging will provide important data regarding zirconia properties.Trial registration: ClinicalTrials.gov Identifier: NCT02150226.

Hyperreactivity of Blood Leukocytes in Patients with NAFLD to Ex Vivo Lipopolysaccharide Treatment Is Modulated by Metformin and Phosphatidylcholine but Not by Alpha Ketoglutarate.

Directory of Open Access Journals (Sweden)

Agnieszka Zwolak

Full Text Available Toll-like receptor 4 and proinflammatory cytokines play a central role in the progression of nonalcoholic fatty liver disease. We investigated IL-1, IL-6 and TNFα production and toll-like receptor 4 in both--obese and lean patients with non-alcoholic fatty liver disease who met different sets of metabolic syndrome criteria and linked the results with the disease burden.95 subjects were divided into four groups depending on the following criteria: presence or absence of metabolic syndrome and/or non-alcoholic fatty liver disease, glucose tolerance (prediabetes or normoglycemia and BMI value (obese or lean. We determined the levels of IL-1β, IL-6, TNFα, and monocyte toll-like receptor 4 expression in fresh blood as well as in blood cultures treated with lipopolysaccharide with or without metformin, alphaketoglutarate or phosphatidylcholine supplementation.The blood leukocytes of patients with non-alcoholic fatty liver disease are hypersensitive to lipopolysaccharide treatment and produce elevated levels of pro-inflammatory cytokines in response to ex vivo treatment with lipopolysaccharide. Moreover, they overexpress toll-like receptor-4. Hyperreactivity was typical mainly for obese patients with non-alcoholic fatty liver disease together with metabolic syndrome and decreased with the severity of disease. Metformin was the most effective in attenuation of hyperreactivity in all groups of patients with non-alcoholic fatty liver disease, but in obese patients the effectiveness of metformin was weaker than in lean. The reduction of cytokine level by metformin was accompanied by the decrease in toll-like receptor-4 expression. phosphatidylcholine also attenuated hyperreactivity to lipopolysaccharide but mainly in obese patients. Alpha ketoglutarate did not modulate cytokines' level and toll-like receptor 4 expression in non-alcoholic fatty liver disease patients.Metformin and phosphatidylcholine attenuated lipopolysaccharide induced toll

Precision cut intestinal slices are an appropriate ex vivo model to study NSAID-induced intestinal toxicity in rats

NARCIS (Netherlands)

Niu, Xiaoyu; de Graaf, Inge A. M.; van der Bij, Hendrik A.; Groothuis, Geny M. M.

2014-01-01

Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used therapeutic agents, however, they are associated with a high prevalence of intestinal side effects. In this investigation, rat precision cut intestinal slices (PCIS) were evaluated as an ex vivo model to study NSAID-induced intestinal

Primary Bovine Extra-Embryonic Cultured Cells: A New Resource for the Study of In Vivo Peri-Implanting Phenotypes and Mesoderm Formation.

Directory of Open Access Journals (Sweden)

Isabelle Hue

Full Text Available In addition to nourishing the embryo, extra-embryonic tissues (EETs contribute to early embryonic patterning, primitive hematopoiesis, and fetal health. These tissues are of major importance for human medicine, as well as for efforts to improve livestock efficiency, but they remain incompletely understood. In bovines, EETs are accessible easily, in large amounts, and prior to implantation. We took advantage of this system to describe, in vitro and in vivo, the cell types present in bovine EETs at Day 18 of development. Specifically, we characterized the gene expression patterns and phenotypes of bovine extra-embryonic ectoderm (or trophoblast; bTC, endoderm (bXEC, and mesoderm (bXMC cells in culture and compared them to their respective in vivo micro-dissected cells. After a week of culture, certain characteristics (e.g., gene expression of the in vitro cells were altered with respect to the in vivo cells, but we were able to identify "cores" of cell-type-specific (and substrate-independent genes that were shared between in vitro and in vivo samples. In addition, many cellular phenotypes were cell-type-specific with regard to extracellular adhesion. We evaluated the ability of individual bXMCs to migrate and spread on micro-patterns, and observed that they easily adapted to diverse environments, similar to in vivo EE mesoderm cells, which encounter different EE epithelia to form chorion, yolk sac, and allantois. With these tissue interactions, different functions arose that were detected in silico and corroborated in vivo at D21-D25. Moreover, analysis of bXMCs allowed us to identify the EE cell ring surrounding the embryonic disc (ED at D14-15 as mesoderm cells, which had been hypothesized but not shown prior to this study. We envision these data will serve as a major resource for the future in the analysis of peri-implanting phenotypes in response to the maternal metabolism and contribute to subsequent studies of placental/fetal development in

Lipid drug conjugate nanoparticle as a potential nanocarrier for the oral delivery of pemetrexed diacid: Formulation design, characterization, ex vivo, and in vivo assessment.

Science.gov (United States)

Soni, Kriti; Mujtaba, Ali; Kohli, Kanchan

2017-10-01

The present work was to develop lipid drug conjugated (LDC) nanoparticles for the potential oral delivery of pemetrexed diacid (PTX) and evaluation of its in vitro, ex vivo and in vivo potentials. The LDC was prepared by salt formation of PTX with stearic acid and followed by cold homogenization technique to produce the LDC nanoparticles. FTIR analysis of LDC proved the presence of amide bond in LDC powder indicating the conjugation between drug and lipid. LDC nanoparticles was found to have particle size 121.9±1.85nm and zeta potential -51.6mV±1.23 and entrapment efficiency 81.0±0.89%. TEM images revealed spherical morphology and were in corroboration with particle size measurements. Ex vivo gut permeation studies revealed a very good enhancement in permeation of drug present in the LDC as compared to plain drug solution and were confirmed by CLSM. MTT assay conformed significant% toxicity at the end of 24h and 48h. Furthermore, the AUC 0-24 of PTX from the optimized LDC nanoparticels was found to be 4.22 folds higher than that from PTX suspension on oral administration. Thus, LDC has high potential for the oral delivery of PTX in cancer therapy and future prospects for the industrial purpose. Copyright © 2017 Elsevier B.V. All rights reserved.

Transplante de células-tronco epiteliais límbicas alógenas expandidas ex vivo sobre membrana amniótica: relato de caso Transplantation of allogenic limbal epithelial stem cells cultivated ex vivo on amniotic membrane: case report

Directory of Open Access Journals (Sweden)

José Álvaro Pereira Gomes

2009-04-01

Full Text Available Paciente apresentou falência de transplante de limbo e conjuntiva de doador vivo alógeno no olho direito após ceratoconjuntivite epidêmica. Após alguns meses, foi submetida a transplante de células-tronco epiteliais límbicas alógenas cultivadas ex vivo sobre membrana amniótica (primeiro caso no Brasil, tendo evoluído com epitelização total da córnea e melhora da acuidade visual. Após o 3º mês da cirurgia, iniciou-se neovascularização superficial periférica com piora da transparência corneana. A visão manteve-se 0,1 após um ano de cirurgia.Case report of a patient who developed failure of an allogenic living related conjunctival limbal transplantation in the right eye after an episode of epidemic keratoconjunctivitis. After a few months, she underwent transplantation of allogenic limbal epithelial stem cells cultivated ex vivo on amniotic membrane (first case in Brazil. The patient evolved with total corneal epithelialization and improvement of the visual acuity. Three months after the surgery, peripheral superficial neovascularization with worsening of the corneal transparency was observed. The vision remained 0.1 after one year of the transplantation.

Inserting ex vivo fluorescence confocal microscopy perioperatively in Mohs micrographic surgery expedites bedside assessment of excision margins in recurrent basal cell carcinoma.

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Longo, Caterina; Ragazzi, Moira; Castagnetti, Fabio; Gardini, Stefano; Palmieri, Tamara; Lallas, Aimilios; Moscarella, Elvira; Piana, Simonetta; Pellacani, Giovanni; Zalaudek, Iris; Argenziano, Giuseppe

2013-01-01

Mohs micrographic surgery can be employed in recurrent basal cell carcinoma, although it is a time-consuming technique. Recently, ex vivo fluorescence confocal microscopy (FCM) has been employed to obtain a fast assessment of tumor margins at the bedside. In our case we successfully employed ex vivo FCM to assess the tumor margins and we treated the persistent tumor with intensity-modulated radiation therapy. Our case demonstrates that a multidisciplinary approach is very efficient in managing complex and recurrent tumors and highlights the benefits of FCM as a new technique that can be used in the surgical theater to speed up the entire procedure.

Immunomodulatory effect of Moringa peregrina leaves, ex vivo and in vivo study

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Al-Oran, Sawsan Atallah; Hassuneh, Mona Rushdie; Al-Qaralleh, Haitham Naief; Rayyan, Walid Abu; Al-Thunibat, Osama Yosef; Mallah, Eyad; Abu-Rayyan, Ahmed; Salem, Shadi

2017-01-01

This study was conducted to assess the in vivo and ex vivo immunomodulatory effect of the ethanol leaves extract of Moringa peregrina in Balb/c mice. For this study, five groups of 5 Balb/c mice were given a single acute subtoxic oral dose of the ethanolic extract at 1.13, 11.30, 23.40 and 113.4 mg/kg and the immunomodulatory effect was assessed on the 6th day following the ingestion. In the (non-functional) assessment, the effect of the extract on the body weight, relative lymphoid organ weight, splenic cellularity and peripheral blood hematologic parameters were evaluated. While in the immunomodulation assessment (functional), we investigated the effect of the extract on the proliferative capacity of splenic lymphocytes and peripheral T and B lymphocytes using mitogen blastogenesis, mixed allogeneic MLR and IgM-Plaque forming cells assays. The ingestion of M. peregrina extract caused a significant increase in the body weight, weight and number of cells of spleen and lymph nodes of the treated mice. Furthermore, the count of RBCs, WBCs, platelets, hemoglobin concentration and PCV % were increased by the extract treatment in a dose-dependent manner. M. peregrina enhanced the proliferative responses of splenic lymphocytes for both T cell and B-cell mitogens. Likewise, the mixed lymphocyte reaction MLR assay has revealed a T-cell dependent proliferation enhancement in the extract treated mice. Moreover, the oral administration of M. peregrina leaves extracts significantly increased PFCs/106 splenocytes in a dose-dependent manner. In conclusion, subtoxic acute doses of M. peregrina extract demonstrated significant potential as an immunomodulatory agent even at the lowest dose of 1.13 mg/kg. PMID:29204086

Ex Vivo and In Vivo Mice Models to Study Blastocystis spp. Adhesion, Colonization and Pathology: Closer to Proving Koch's Postulates.

Directory of Open Access Journals (Sweden)

Sitara S R Ajjampur

Full Text Available Blastocystis spp. are widely prevalent extra cellular, non-motile anerobic protists that inhabit the gastrointestinal tract. Although Blastocystis spp. have been associated with gastrointestinal symptoms, irritable bowel syndrome and urticaria, their clinical significance has remained controversial. We established an ex vivo mouse explant model to characterize adhesion in the context of tissue architecture and presence of the mucin layer. Using confocal microscopy with tissue whole mounts and two axenic isolates of Blastocystis spp., subtype 7 with notable differences in adhesion to intestinal epithelial cells (IEC, isolate B (ST7-B and isolate H (more adhesive, ST7-H, we showed that adhesion is both isolate dependent and tissue trophic. The more adhesive isolate, ST7-H was found to bind preferentially to the colon tissue than caecum and terminal ileum. Both isolates were also found to have mucinolytic effects. We then adapted a DSS colitis mouse model as a susceptible model to study colonization and acute infection by intra-caecal inoculation of trophic Blastocystis spp.cells. We found that the more adhesive isolate ST7-H was also a better colonizer with more mice shedding parasites and for a longer duration than ST7-B. Adhesion and colonization was also associated with increased virulence as ST7-H infected mice showed greater tissue damage than ST7-B. Both the ex vivo and in vivo models used in this study showed that Blastocystis spp. remain luminal and predominantly associated with mucin. This was further confirmed using colonic loop experiments. We were also successfully able to re-infect a second batch of mice with ST7-H isolates obtained from fecal cultures and demonstrated similar histopathological findings and tissue damage thereby coming closer to proving Koch's postulates for this parasite.

Ex-vivo response to blood products and haemostatic agents after paediatric cardiac surgery

DEFF Research Database (Denmark)

Hvas, Anne-Mette; Andreasen, Jo B; Christiansen, Kirsten

2013-01-01

cardiac surgery. The haemostatic potential of various factor concentrates (fibrinogen concentrate, recombinant factor VIIa and factor XIII), fresh frozen plasma (FFP), pooled platelets and tranexamic acid was investigated. After surgery, the coagulation profiles revealed significantly prolonged clotting...... of fibrinogen concentrate, FFP or tranexamic acid improved clot stability significantly. Whole blood coagulation was significantly impaired after cardiac surgery in children. Ex-vivo studies showed a total reversal of the coagulopathy after addition of pooled platelets and significantly improved clot stability...... after addition of fibrinogen concentrate, FFP and tranexamic acid, respectively....

Ultrasound imaging of Nd:YAG laser-induced tissue coagulation in porcine livers.

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Ritzel, U; Wietzke-Braun, P; Brinck, U; Leonhardt, U; Ramadori, G

2001-12-01

Absorption of laser light energy induces denaturation of proteins and thermocoagulation of irradiated tissue. Recently, MRI-guided laser coagulation in combination with MR thermometry was reported as a treatment of liver tumours. In the present study ultrasonographic imaging was evaluated for its suitability in laser induced tissue thermocoagulation. Fresh porcine livers were used for ex vivo examinations. Placement of the laser catheter and tissue coagulation during laser light emission were online monitored by ultrasonography. Nd:YAG laser-induced tissue damage was evaluated by macroscopical and microscopical examinations of histological sections. During laser light emission a marked hyperdense signal enhancement was observed by ultrasonography which strongly correlated with the extent of macroscopic tissue damage. The size of laser-induced coagulation zone depended on both the power setting and total energy delivered. Carbonization of the tissue surrounding the laser tip is a limiting factor because of laser light absorption. However our data indicate that using appropriate laser energy and exposure time prevent carbonization although carbonization can not be visualized by ultrasonography. It is concluded from the present ex vivo studies that laser coagulation can be effectively performed under ultrasonographic guidance.

Ex vivo nonlinear microscopy imaging of Ehlers-Danlos syndrome-affected skin.

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Kiss, Norbert; Haluszka, Dóra; Lőrincz, Kende; Kuroli, Enikő; Hársing, Judit; Mayer, Balázs; Kárpáti, Sarolta; Fekete, György; Szipőcs, Róbert; Wikonkál, Norbert; Medvecz, Márta

2018-07-01

Ehlers-Danlos syndrome (EDS) is the name for a heterogenous group of rare genetic connective tissue disorders with an overall incidence of 1 in 5000. The histological characteristics of EDS have been previously described in detail in the late 1970s and early 1980s. Since that time, the classification of EDS has undergone significant changes, yet the description of the histological features of collagen morphology in different EDS subtypes has endured the test of time. Nonlinear microscopy techniques can be utilized for non-invasive in vivo label-free imaging of the skin. Among these techniques, two-photon absorption fluorescence (TPF) microscopy can visualize endogenous fluorophores, such as elastin, while the morphology of collagen fibers can be assessed by second-harmonic generation (SHG) microscopy. In our present work, we performed TPF and SHG microscopy imaging on ex vivo skin samples of one patient with classical EDS and two patients with vascular EDS and two healthy controls. We detected irregular, loosely dispersed collagen fibers in a non-parallel arrangement in the dermis of the EDS patients, while as expected, there was no noticeable impairment in the elastin content. Based on further studies on a larger number of patients, in vivo nonlinear microscopic imaging could be utilized for the assessment of the skin status of EDS patients in the future.

Dermal uptake and percutaneous penetration of ten flame retardants in a human skin ex vivo model

DEFF Research Database (Denmark)

Frederiksen, Marie; Vorkamp, Katrin; Jensen, Niels Martin

2016-01-01

The dermal uptake and percutaneous penetration of ten organic flame retardants was measured using an ex vivo human skin model. The studied compounds were DBDPE, BTBPE, TBP-DBPE, EH-TBB, BEH-TEBP, α, β and γ-HBCDD as well as syn- and anti-DDC-CO. Little or none of the applied flame retardants...

2.5D Representations Combining in vivo 3D MRI and ex vivo 2D MSI Approaches to Study the Lipid Distribution in the Whole Sheep Brain

OpenAIRE

Labas , Valérie; Teixeira-Gomes , Ana Paula; Andersson , Frédéric; Ménigot , Sébastien; Batailler , Martine; Adriaensen , Hans; Migaud , Martine; Chaillou , Elodie

2015-01-01

National audience; Mass Spectrometry Imaging (MSI) provides easily high spatially resolved masses allowing characterization of endogenous lipids. These latter constitute about 70% of the composition of the white matter of the brain which can be implicated in developmental and/or cognitive troubles. In order to examine the molecular distribution of lipids in whole sheep brain, and especially in white/grey matter, we combined in vivo and ex vivo images, obtained in the same animals, using Magne...

In vivo versus in vitro produced bovine ova: similarities and differences relevant for practical application

DEFF Research Database (Denmark)

Holm, Peter; Callesen, Henrik

1998-01-01

- Abstract This present review describes some differences and similarities between bovine embryos produced in vivo and in vitro. The first part outlines the respective environments during maturation, fertilisation and early embryonic development of the two types of embryos and compares their mor...... differences between in vitro and in vivo produced embryos are also well documented. How- ever, improved culture conditions have been reported to minimise the differences. The second part focuses on the practical consequences of the differences in relation to embryo selection, cryo- preservation, sanitary...... risks and pregnancy following transfer as well as normality of calves. Lower viability following transfer and increased susceptibility to cryopreservation of in vitro produced embryos is discussed. Finally and most importantly, reported evidence of increased sanitary risks and abnormal foetal...

In vitro and ex vivo activity of Melaleuca alternifolia against protoscoleces of Echinococcus ortleppi.

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Monteiro, Danieli Urach; Azevedo, Maria Isabel; Weiblen, Carla; DE Avila Botton, Sônia; Funk, Nadine Lysyk; DE Bona DA Silva, Cristiane; Zanette, Régis Adriel; Schwanz, Thiago Guilherme; DE LA Rue, Mário Luiz

2017-02-01

Cystic echinococcosis is a zoonotic disease of difficult diagnosis and treatment. The use of protoscolicidal agents in procedures is of utmost importance for treatment success. This study was aimed at analysing the in vitro and ex vivo activity of Melaleuca alternifolia oil (tea tree oil - TTO), its nanoemulsion formulation (NE-TTO) and its major component (terpinen-4-ol) against Echinococcus ortleppi protoscoleces obtained from cattle. Concentrations of 2·5, 5 and 10 mg mL-1 of TTO, 10 mg mL-1 of NE-TTO and 1, 1·5 and 2 mg mL-1 of terpinen-4-ol were evaluated in vitro against protoscoleces at 5, 10, 15 and 30 min. TTO was also injected directly into hydatid cysts (ex vivo analysis, n = 20) and the viability of protoscoleces was evaluated at 5, 15 and 30 min. The results indicated protoscolicidal effect at all tested formulations and concentrations. Terpinen-4-ol (2 mg mL-1) activity was superior when compared with the highest concentration of TTO. NE-TTO reached a gradual protoscolicidal effect. TTO at 20 mg mL-1 showed 90% protoscolicidal action in hydatid cysts at 5 min. The results showed that TTO affects the viability of E. ortleppi protoscoleces, suggesting a new protoscolicidal option to the treatment of cystic equinococcosis.

A novel ex-vivo porcine renal xenotransplantation model using a pulsatile machine preservation system.

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Guarrera, James V; Stone, Jonathan; Tulipan, Jacob; Jhang, Jeffrey; Arrington, Ben; Boykin, Jason; Markowitz, Glen; Ratner, Lloyd E

2011-01-01

Animal models to investigate pathophysiology and xenotransplantation require complex techniques and significant animal utilization. The aim of the study was to develop a reliable ex-vivo technique to test xenotransplant interventions. Miniature Swine being utilized for a nonsurvival study acted as donor animals. Kidneys were flushed and rapidly explanted and chilled to 4°C. Kidneys were assigned to be the control (CK) (n=3) and the mate were used as a Xenograft Kidneys (XK) (n=3). Kidneys were perfused on separate Waters RM 3 perfusion devices. Perfusion temperature was 35-37°C and pressure was 100-110/60-70 mmHg at 60 pulses per minute. CKs were reperfused with autologous blood collected at the time of organ procurement. XKs were reperfused using freshly donated whole human blood. Physical characteristics, urine output were recorded. Core needle biopsies were obtained and examined by a blinded pathologist for evidence of antibody mediated rejection. XK kidneys demonstrated homogenous reperfusion which rapidly became patchy at 5-7 minutes. XK kidneys had become complete black and thrombosed by 60-70 minutes. XK biopsies demonstrated peritubular capillaritis. CK kidneys demonstrated homogenous reperfusion and urine production. H&E stain of CKs only demonstrated nonspecific inflammation. Our ex-vivo porcine xenotransplant model shows early promise as a tool for studying Xeno- associated hyperacute rejection. This technique saves resources and animal utilization.

An ex vivo porcine skin model to evaluate pressure-reducing devices of different mechanical properties used for pressure ulcer prevention.

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Yeung, Ching-Yan C; Holmes, David F; Thomason, Helen A; Stephenson, Christian; Derby, Brian; Hardman, Matthew J

2016-11-01

Pressure ulcers are complex wounds caused by pressure- and shear-induced trauma to skin and underlying tissues. Pressure-reducing devices, such as dressings, have been shown to successfully reduce pressure ulcer incidence, when used in adjunct to pressure ulcer preventative care. While pressure-reducing devices are available in a range of materials, with differing mechanical properties, understanding of how a material's mechanical properties will influence clinical efficacy remains limited. The aim of this study was to establish a standardized ex vivo model to allow comparison of the cell protection potential of two gel-like pressure-reducing devices with differing mechanical properties (elastic moduli of 77 vs. 35 kPa). The devices also displayed differing energy dissipation under compressive loading, and resisted strain differently under constant load in compressive creep tests. To evaluate biological efficacy we employed a new ex vivo porcine skin model, with a confirmed elastic moduli closely matching that of human skin (113 vs. 119 kPa, respectively). Static loads up to 20 kPa were applied to porcine skin ex vivo with subsequent evaluation of pressure-induced cell death and cytokine release. Pressure application alone increased the percentage of epidermal apoptotic cells from less than 2% to over 40%, and increased cellular secretion of the pro-inflammatory cytokine TNF-alpha. Co-application of a pressure-reducing device significantly reduced both cellular apoptosis and cytokine production, protecting against cellular damage. These data reveal new insight into the relationship between mechanical properties of pressure-reducing devices and their biological effects. After appropriate validation of these results in clinical pressure ulcer prevention with all tissue layers present between the bony prominence and external surface, this ex vivo porcine skin model could be widely employed to optimize design and evaluation of devices aimed at reducing pressure

Non-alcoholic fatty liver disease (NAFLD) models in drug discovery.

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Cole, Banumathi K; Feaver, Ryan E; Wamhoff, Brian R; Dash, Ajit

2018-02-01

The progressive disease spectrum of non-alcoholic fatty liver disease (NAFLD), which includes non-alcoholic steatohepatitis (NASH), is a rapidly emerging public health crisis with no approved therapy. The diversity of various therapies under development highlights the lack of consensus around the most effective target, underscoring the need for better translatable preclinical models to study the complex progressive disease and effective therapies. Areas covered: This article reviews published literature of various mouse models of NASH used in preclinical studies, as well as complex organotypic in vitro and ex vivo liver models being developed. It discusses translational challenges associated with both kinds of models, and describes some of the studies that validate their application in NAFLD. Expert opinion: Animal models offer advantages of understanding drug distribution and effects in a whole body context, but are limited by important species differences. Human organotypic in vitro and ex vivo models with physiological relevance and translatability need to be used in a tiered manner with simpler screens. Leveraging newer technologies, like metabolomics, proteomics, and transcriptomics, and the future development of validated disease biomarkers will allow us to fully utilize the value of these models to understand disease and evaluate novel drugs in isolation or combination.

Photoacoustic-based sO2 estimation through excised bovine prostate tissue with interstitial light delivery.

Science.gov (United States)

Mitcham, Trevor; Taghavi, Houra; Long, James; Wood, Cayla; Fuentes, David; Stefan, Wolfgang; Ward, John; Bouchard, Richard

2017-09-01

Photoacoustic (PA) imaging is capable of probing blood oxygen saturation (sO 2 ), which has been shown to correlate with tissue hypoxia, a promising cancer biomarker. However, wavelength-dependent local fluence changes can compromise sO 2 estimation accuracy in tissue. This work investigates using PA imaging with interstitial irradiation and local fluence correction to assess precision and accuracy of sO 2 estimation of blood samples through ex vivo bovine prostate tissue ranging from 14% to 100% sO 2 . Study results for bovine blood samples at distances up to 20 mm from the irradiation source show that local fluence correction improved average sO 2 estimation error from 16.8% to 3.2% and maintained an average precision of 2.3% when compared to matched CO-oximeter sO 2 measurements. This work demonstrates the potential for future clinical translation of using fluence-corrected and interstitially driven PA imaging to accurately and precisely assess sO 2 at depth in tissue with high resolution.

Host transcriptional responses following ex vivo re-challenge with Mycobacterium tuberculosis vary with disease status.

Directory of Open Access Journals (Sweden)

Elaine A Yu

Full Text Available The identification of immune correlates that are predictive of disease outcome for tuberculosis remains an ongoing challenge. To address this issue, we evaluated gene expression profiles from peripheral blood mononuclear cells following ex vivo challenge with Mycobacterium tuberculosis, among participants with active TB disease (ATBD, n = 10, latent TB infection (LTBI, n = 10, and previous active TB disease (after successful treatment; PTBD, n = 10, relative to controls (n = 10. Differential gene expression profiles were assessed by suppression-subtractive hybridization, dot blot, real-time polymerase chain reaction, and the comparative cycle threshold methods. Comparing ATBD to control samples, greater fold-increases of gene expression were observed for a number of chemotactic factors (CXCL1, CXCL3, IL8, MCP1, MIP1α. ATBD was also associated with higher IL1B gene expression, relative to controls. Among LTBI samples, gene expression of several chemotactic factors (CXCL2, CXCL3, IL8 was similarly elevated, compared to individuals with PTBD. Our results demonstrated that samples from participants with ATBD and LTBI have distinct gene expression profiles in response to ex vivo M. tuberculosis infection. These findings indicate the value in further characterizing the peripheral responses to M. tuberculosis challenge as a route to defining immune correlates of disease status or outcome.

Host transcriptional responses following ex vivo re-challenge with Mycobacterium tuberculosis vary with disease status.

Science.gov (United States)

Yu, Elaine A; John, Serene H; Tablante, Elizabeth C; King, Christine A; Kenneth, John; Russell, David G; Mehta, Saurabh

2017-01-01

The identification of immune correlates that are predictive of disease outcome for tuberculosis remains an ongoing challenge. To address this issue, we evaluated gene expression profiles from peripheral blood mononuclear cells following ex vivo challenge with Mycobacterium tuberculosis, among participants with active TB disease (ATBD, n = 10), latent TB infection (LTBI, n = 10), and previous active TB disease (after successful treatment; PTBD, n = 10), relative to controls (n = 10). Differential gene expression profiles were assessed by suppression-subtractive hybridization, dot blot, real-time polymerase chain reaction, and the comparative cycle threshold methods. Comparing ATBD to control samples, greater fold-increases of gene expression were observed for a number of chemotactic factors (CXCL1, CXCL3, IL8, MCP1, MIP1α). ATBD was also associated with higher IL1B gene expression, relative to controls. Among LTBI samples, gene expression of several chemotactic factors (CXCL2, CXCL3, IL8) was similarly elevated, compared to individuals with PTBD. Our results demonstrated that samples from participants with ATBD and LTBI have distinct gene expression profiles in response to ex vivo M. tuberculosis infection. These findings indicate the value in further characterizing the peripheral responses to M. tuberculosis challenge as a route to defining immune correlates of disease status or outcome.

Polarimetry based partial least square classification of ex vivo healthy and basal cell carcinoma human skin tissues.

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Ahmad, Iftikhar; Ahmad, Manzoor; Khan, Karim; Ikram, Masroor

2016-06-01

Optical polarimetry was employed for assessment of ex vivo healthy and basal cell carcinoma (BCC) tissue samples from human skin. Polarimetric analyses revealed that depolarization and retardance for healthy tissue group were significantly higher (ppolarimetry together with PLS statistics hold promise for automated pathology classification. Copyright © 2016 Elsevier B.V. All rights reserved.

Pathological findings of condemned bovine liver specimens and associated economic loss at Nyabugogo abattoir, Kigali, Rwanda.

Science.gov (United States)

Habarugira, Gervais; Mbasinga, Gloria; Mushonga, Borden; Chitura, Teedzai; Kandiwa, Erick; Ojok, Lonzy

2016-12-01

There are no published abattoir bovine hepatic lesion prevalence studies in cattle in Rwanda. This study estimated that 12.3% of the livers (n=4751) examined at Nyabugogo slaughterhouse in Kigali were condemned. Condemnation prejudiced the nation of 3492.00kg of meat with attendant economic losses of US$8932.40 during the study period. Risk factors for these lesions were also assessed. Male and female animals from 11 districts were used in this study. Hepatic lesions were higher in females (14.6%; n=1494) than in males (11.1%; n=3257). About 78.7% of the condemnations were due to fascioliasis, followed by abscesses (5.7%), hepatitis (5.3%), cirrhosis (4%) and other lesions (6.3%). Female animal livers showed more fascioliasis and abscesses (82.2% and 9.5%) than male animal livers (73.3% and 3.3%). The highest rate of condemnation was observed from Kayonza (40.2%; n=413) and the least was from Gakenke district (0.9%; n=1031). Cattle from the Eastern Province showed significantly (PRwanda. Copyright © 2016 Elsevier B.V. All rights reserved.

Ex vivo study of the home-use TriPollar RF device using an experimental human skin model.

Science.gov (United States)

Boisnic, Sylvie; Branchet, Marie Christine

2010-09-01

A wide variety of professional radio frequency (RF) aesthetic treatments for anti-aging are available aiming at skin tightening. A new home-use RF device for facial treatments has recently been developed based on TriPollar technology. To evaluate the mechanism of the new home-use device, in the process of collagen remodeling, using an ex vivo skin model. Human skin samples were collected in order to evaluate the anti-aging effect of a home-use device for facial treatments on an ex vivo human skin model. Skin tightening was evaluated by dermal histology, quantitative analysis of collagen fibers and dosage of collagen synthesis. Significant collagen remodeling following RF treatment with the device was found in the superficial and mid-deep dermis. Biochemical measurement of newly synthesized collagen showed an increase of 41% in the treated samples as compared to UV-aged control samples. The new home-use device has been demonstrated to affect significant collagen remodeling, in terms of the structural and biochemical improvement of dermal collagen on treated skin samples.

Radiosynthesis and ex vivo evaluation of [11C-carbonyl]carbamate- and urea-based monoacylglycerol lipase inhibitors

International Nuclear Information System (INIS)

Hicks, Justin W.; Parkes, Jun; Tong, Junchao; Houle, Sylvain; Vasdev, Neil; Wilson, Alan A.

2014-01-01

Introduction: Monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) are the two primary enzymes that regulate the tone of endocannabinoid signaling. Although new PET radiotracers have been discovered for imaging FAAH in vivo, no such radiotracer exists for imaging MAGL. Here we report the radiosynthesis of five candidate MAGL radiotracers and their ex vivo evaluations in mice and rats. Methods: Candidate carbamate and urea MAGL inhibitors were radiolabeled at the carbonyl position by [ 11 C]CO 2 fixation. Radiotracers were administered (tail-vein injection) to rodents and brain uptake of radioactivity measured at early and late time points ex vivo. Specificity of uptake was explored by pretreatment with unlabeled inhibitors (2 mg/kg, ip) 30 min prior to radiotracer administration. Results: All five candidate MAGL radiotracers were prepared in high specific activity (> 65 GBq/μmol) and radiochemical purity (> 98%). Moderate brain uptake (0.2–0.8 SUV) was observed for each candidate while pretreatment did not reduce uptake for four of the five tested. For two candidates ([ 11 C]12 and [ 11 C]14), high retention of radioactivity was observed in the blood (ca. 10 and 4 SUV at 40 min) which was blocked by pretreatment with unlabeled inhibitors. The most promising candidate, [ 11 C]18, demonstrated moderate brain uptake (ca. 0.8 SUV) which showed circa 50% blockade by pretreatment with unlabeled 18. Conclusion: One putative and four reported potent and selective MAGL inhibitors have been radiolabeled via [ 11 C]CO 2 fixation as radiotracers for this enzyme. Despite the promising in vitro pharmacological profile, none of the five candidate radiotracers exhibited in vivo behavior suitable for PET neuroimaging

Ultrasound elastography of the lower uterine segment in women with a previous cesarean section: Comparison of in-/ex-vivo elastography versus tensile-stress-strain-rupture analysis.

Science.gov (United States)

Seliger, Gregor; Chaoui, Katharina; Lautenschläger, Christine; Jenderka, Klaus-Vitold; Kunze, Christian; Hiller, Grit Gesine Ruth; Tchirikov, Michael

2018-06-01

The purpose of this study was to assess, if the biomechanical properties of the lower uterine segment (LUS) in women with a previous cesarean section (CS) can be determined by ultrasound (US) elastography. The first aim was to establish an ex-vivo LUS tensile-stress-strain-rupture(break point) analysis with the possibility of simultaneously using US elastography. The second aim was to investigate the relationship between measurement results of LUS stiffness using US elastography in-/ex-vivo with results of tensile-stress-strain-rupture analysis, and to compare different US elastography LUS-stiffness-measurement methods ex-vivo. An explorative experimental, in-/ex-vivo US study of women with previous CS was conducted. LUS elasticity was measured by point Shear Wave Elastography (pSWE) and bidimensional Shear-Wave-Elastography (2D-SWE) first in-vivo during preoperative examination within 24 h before repeat CS (including resection of the thinnest part of the LUS = uterine scar area during CS), second within 1 h after operation during the ex-vivo experiment, followed by tensile-stress-strain-rupture analysis. Pearson's correlation coefficient and scatter plots, Bland-Altman plots and paired T-tests, were used. Thirty three women were included in the study; elastography measurements n = 1412. The feasibility of ex-vivo assessment of LUS by quantitative US elastography using pSWE and 2D-SWE to detect stiffness of LUS was demonstrated. The strongest correlation with tensile-stress-strain analysis was found in the US elastography examination carried out with 2D-SWE (0.78, p break point - as a surrogate marker for the risk of rupture of the LUS after CS - is linearly dependent on the thickness of the LUS in the scar area (Coefficient of correlation: 0.79, p even at less stroke/strain than would be expected by their thickness. This study confirms that US elastography can help in determining viscoelastic properties of the LUS in women with a previous CS. The

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

OpenAIRE

MUDGIL, Meetali; PAWAR, Pravin

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 d...

Evaluating childhood obesity. Magnetic resonance-based quantification of abdominal adipose tissue and liver fat in children

International Nuclear Information System (INIS)

Raschpichler, M.C.; Leipzig Univ. Medical Center; Sorge, I.; Hirsch, W.; Mende, M.; Sergeyev, E.; Koerner, A.; Kruber, D.; Schick, F.

2012-01-01

The purpose of this study is to establish and validate a magnetic resonance (MR)-based fat quantification package that provides an accurate assessment of abdominal adipose tissue and liver fat in children. Ex vivo trials with a torso model and water-oil mixtures are conducted. Abdominal adipose tissue (AAT) is covered by magnetic resonance imaging (MRI) using a fat-selective sequence and is analyzed by a plug-in based on the open source software Image. Liver fat (LF) is measured with localized 1 H Magnetic Resonance Spectroscopy ( 1 H MRS) and the jMRUI (java-based Magnetic Resonance User Interface) software package. Evaluation of the clinical methodology involved a study of 10 children in this feasibility study (mean age and body mass index: 13.3 yr; 33.3 kg/m 2 ). To evaluate the method's validity, reference measurements were performed. Ex vivo trials with the torso model showed that adipose tissue was measured appropriately with a systematic underestimation by 9.3 ± 0.2 % (0.32 ± 0.064 kg). Coefficients of variation for both intra- and inter-observer measurements ranged between 0 - 2.7 % and repeated analyses showed significant equivalent results (p 1 H MRS ex vivo revealed significant equivalence with the predefined fat content in water-oil mixtures (p < 0.01). In vivo, the homemade plug-in significantly overestimated the AAT, with the visceral adipose tissue being most affected (+ 15.7 ± 8.4 %). Although an overestimation of the AAT by the presented plug-in should be taken into consideration, this children-friendly package enables the quantification of both LF and AAT within 30 min on a freeware-based platform. (orig.)

Some properties of purified hepatoredoxin from bovine liver mitochondria

International Nuclear Information System (INIS)

Gilevich, S.N.; Gurev, O.L.; Shkumatov, V.M.; Chashchin, V.L.; Akhrem, A.A.

1986-01-01

Some of the most important physicochemical properties of hepatoredoxin from bovine liver, purified to a homogeneous state, were determined for the first time. The protein contains a [2Fe-2S] cluster in its active site and in an oxidized state has absorption maxima at 280, 320, 415, and 455 nm. The spectrophotometric index of purity (A 415 /A 280 ) of the homogeneous native preparation is 0.84; the extinction coefficient (epsilon 415 ) is equal to 9800 M -1 cm -1 . According to the data of gel electrophoresis in the presence of SDS, hepatoredoxin has an M/sub r/ of 12,500; its isoelectric point (pI) is equal to 4.2. Hepatoredoxin is necessary for the reconstitution of the C 27 -steroid hydroxylase activity and can be replaced by the related protein, adrenodoxin. All the parameters listed above, as well as the CD spectra, the immunochemical properties, and sequence of the first five N-terminal amino acids of hepatoredoxin and adrenodoxin are very similar of identical. At the same time, the amino acid composition of the two ferredoxins, along with common properties, has some differences

Association of Parathyroid Gland Biopsy Excision Technique With Ex Vivo Radiation Counts During Radioguided Parathyroid Surgery.

Science.gov (United States)

Hinson, Andrew M; Lawson, Bradley R; Franco, Aime T; Stack, Brendan C

2017-06-01

Parathyroid biopsy represents a means for normal and hyperfunctional glands to be distinguished intraoperatively. However, no data exist to guide surgeons regarding how much of a parathyroid gland must be biopsied to satisfy the 20% rule. To quantify the relative proportion of a hyperfunctional parathyroid gland that must be evaluated with the gamma probe to satisfy the 20% rule. A retrospective review of surgical data for 24 consecutive patients (16 women, 18 men; mean [SD] age, 66.6 [10] years; range, 51-83 years) who underwent surgery for primary hyperparathyroidism between May and October, 2015, in a tertieary academic medical center. Extirpated parathyroid glands were sectioned into parallel or pie-shaped biopsies and evaluated ex vivo with a gamma probe to determine what percentage of a hyperfunctional gland must be sampled to meet the Norman 20% rule. The hypothesis was formulated during data collection. In total, 253 ex vivo biopsy specimens were obtained from 33 surgically removed parathyroid glands. Parathyroid biopsies satisfied the 20% rule with an accuracy that depended on the relative proportion of the parent gland represented: half or more (96.6%; 95% CI, 91.7%-100.0%), a quarter to one-half (87.0%; 95% CI, 79.3%-94.7%), less than a quarter (63.6%; 95% CI, 54.5%-72.8%). When less than a quarter of the gland was removed, pie-shaped biopsies were more likely to satisfy the 20% rule compared with parallel biopsies of the same weight (78.4% vs 56.2%; absolute difference, 22.2%; 95% CI, 4.7%-39.7%). Unless half of a parathyroid gland is biopsied during radioguided parathyroidectomy, the 20% rule cannot reliably rule out the presence of a hyperfunctional parathyroid lesion. Pie-shaped biopsies originating from the center of the gland are associated with a lower rate of false-negative results compared with peripheral biopsies of similar size. Pie-shaped biopsies and biopsy of half or more of each nonexcised parathyroid gland for ex vivo counts may increase

Ex Vivo Confocal Spectroscopy of Autofluorescence in Age-Related Macular Degeneration.

Directory of Open Access Journals (Sweden)

Joel Kaluzny

Full Text Available We investigated the autofluorescence (AF signature of the microscopic features of retina with age-related macular degeneration (AMD using 488 nm excitation.The globes of four donors with AMD and four age-matched controls were embedded in paraffin and sectioned through the macula. Sections were excited using a 488 nm argon laser, and the AF emission was captured using a laser scanning confocal microscope (496-610 nm, 6 nm resolution. The data cubes were then analyzed to compare peak emission spectra between the AMD and the controls. Microscopic features, including individual lipofuscin and melanolipofuscin granules, Bruch's Membrane, as well macroscopic features, were considered.Overall, the AMD eyes showed a trend of blue-shifted emission peaks compared with the controls. These differences were statistically significant when considering the emission of the combined RPE/Bruch's Membrane across all the tissue cross-sections (p = 0.02.The AF signatures of ex vivo AMD RPE/BrM show blue-shifted emission spectra (488 nm excitation compared with the control tissue. The magnitude of these differences is small (~4 nm and highlights the potential challenges of detecting these subtle spectral differences in vivo.

Recombinant TAT-BMI-1 fusion protein induces ex vivo expansion of human umbilical cord blood-derived hematopoietic stem cells.

Science.gov (United States)

Codispoti, Bruna; Rinaldo, Nicola; Chiarella, Emanuela; Lupia, Michela; Spoleti, Cristina Barbara; Marafioti, Maria Grazia; Aloisio, Annamaria; Scicchitano, Stefania; Giordano, Marco; Nappo, Giovanna; Lucchino, Valeria; Moore, Malcolm A S; Zhou, Pengbo; Mesuraca, Maria; Bond, Heather Mandy; Morrone, Giovanni

2017-07-04

Transplantation of hematopoietic stem cells (HSCs) is a well-established therapeutic approach for numerous disorders. HSCs are typically derived from bone marrow or peripheral blood after cytokine-induced mobilization. Umbilical cord blood (CB) represents an appealing alternative HSC source, but the small amounts of the individual CB units have limited its applications. The availability of strategies for safe ex vivo expansion of CB-derived HSCs (CB-HSCs) may allow to extend the use of these cells in adult patients and to avoid the risk of insufficient engraftment or delayed hematopoietic recovery.Here we describe a system for the ex vivo expansion of CB-HSCs based on their transient exposure to a recombinant TAT-BMI-1 chimeric protein. BMI-1 belongs to the Polycomb family of epigenetic modifiers and is recognized as a central regulator of HSC self-renewal. Recombinant TAT-BMI-1 produced in bacteria was able to enter the target cells via the HIV TAT-derived protein transduction peptide covalently attached to BMI-1, and conserved its biological activity. Treatment of CB-CD34+ cells for 3 days with repeated addition of 10 nM purified TAT-BMI-1 significantly enhanced total cell expansion as well as that of primitive hematopoietic progenitors in culture. Importantly, TAT-BMI-1-treated CB-CD34+ cells displayed a consistently higher rate of multi-lineage long-term repopulating activity in primary and secondary xenotransplants in immunocompromised mice. Thus, recombinant TAT-BMI-1 may represent a novel, effective reagent for ex vivo expansion of CB-HSC for therapeutic purposes.

Expression of Plasmodium vivax crt-o Is Related to Parasite Stage but Not Ex Vivo Chloroquine Susceptibility.

Science.gov (United States)

Pava, Zuleima; Handayuni, Irene; Wirjanata, Grennady; To, Sheren; Trianty, Leily; Noviyanti, Rintis; Poespoprodjo, Jeanne Rini; Auburn, Sarah; Price, Ric N; Marfurt, Jutta

2016-01-01

Chloroquine (CQ)-resistant Plasmodium vivax is present in most countries where P. vivax infection is endemic, but the underlying molecular mechanisms responsible remain unknown. Increased expression of P. vivax crt-o (pvcrt-o) has been correlated with in vivo CQ resistance in an area with low-grade resistance. We assessed pvcrt-o expression in isolates from Papua (Indonesia), where P. vivax is highly CQ resistant. Ex vivo drug susceptibilities to CQ, amodiaquine, piperaquine, mefloquine, and artesunate were determined using a modified schizont maturation assay. Expression levels of pvcrt-o were measured using a novel real-time quantitative reverse transcription-PCR method. Large variations in pvcrt-o expression were observed across the 51 isolates evaluated, with the fold change in expression level ranging from 0.01 to 59 relative to that seen with the P. vivax β-tubulin gene and from 0.01 to 24 relative to that seen with the P. vivax aldolase gene. Expression was significantly higher in isolates with the majority of parasites at the ring stage of development (median fold change, 1.7) compared to those at the trophozoite stage (median fold change, 0.5; P determinant of ex vivo drug susceptibility. A comprehensive transcriptomic approach is warranted for an in-depth investigation of the role of gene expression levels and P. vivax drug resistance. Copyright © 2015 Pava et al.

A Novel Ex Vivo Training Model for Acquiring Supermicrosurgical Skills Using a Chicken Leg.

Science.gov (United States)

Cifuentes, Ignacio J; Rodriguez, José R; Yañez, Ricardo A; Salisbury, María C; Cuadra, Álvaro J; Varas, Julian E; Dagnino, Bruno L

2016-11-01

Background  Supermicrosurgery is a technique used for dissection and anastomosis of submillimeter diameter vessels. This technique requires precise hand movements and superb eye-hand coordination, making continuous training necessary. Biological in vivo and ex vivo models have been described for this purpose, the latter being more accessible and cost-effective. The aim of this study is to present a new ex vivo training model using a chicken leg. Methods  In 28 chicken legs, an anatomical study was performed. An intramuscular perforator vessel was identified and dissected. Arterial diameters of 0.7, 0.5, and 0.3 mm were identified and consistency of the perforator was assessed. In additional 10 chicken legs, 25 submillimeter arteries were anastomosed using this perforator vessel. Five arteries of 0.3 and 10 of 0.5 mm were anastomosed with nylon 11-0 and 12-0 sutures. Intravascular stent (IVaS) technique and open guide (OG) technique were used in 0.5-mm arteries. A total of 10 arteries of 0.7 mm were anastomosed using 10-0 sutures in a conventional fashion. Dissection and anastomosis time were recorded and patency was tested. Results  We were able to identify 0.7 to 0.3 mm diameter arteries in all the specimens and confirm the consistency of the perforator. The median time for dissection was 13.4 minutes. The median time for anastomosis was 32.3 minutes for 0.3-mm arteries, 24.3 minutes for 0.5-mm arteries using IVaS, 29.5 minutes for the OG technique, and 20.9 minutes for the 0.7 mm diameter arteries. All the anastomoses were permeable. Conclusion  Due to its consistent and adequate diameter vessels, this model is adequate for training supermicrosurgical skills. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.

Ablation of clinically relevant kidney tissue volumes by high-intensity focused ultrasound: Preliminary results of standardized ex-vivo investigations.

Science.gov (United States)

Häcker, Axel; Peters, Kristina; Knoll, Thomas; Marlinghaus, Ernst; Alken, Peter; Jenne, Jürgen W; Michel, Maurice Stephan

2006-11-01

To investigate strategies to achieve confluent kidney-tissue ablation by high-intensity focused ultrasound (HIFU). Our model of the perfused ex-vivo porcine kidney was used. Tissue ablation was performed with an experimental HIFU device (Storz Medical, Kreuzlingen, Switzerland). Lesion-to-lesion interaction was investigated by varying the lesion distance (5 to 2.5 mm), generator power (300, 280, and 260 W), cooling time (10, 20, and 30 seconds), and exposure time (4, 3, and 2 seconds). The lesion rows were analyzed grossly and by histologic examination (hematoxylin-eosin and nicotinamide adenine dinucleotide staining). It was possible to achieve complete homogeneous ablation of a clinically relevant tissue volume but only by meticulous adjustment of the exposure parameters. Minimal changes in these parameters caused changes in lesion formation with holes within the lesions and lesion-to-lesion interaction. Our preliminary results show that when using this new device, HIFU can ablate a large tissue volume homogeneously in perfused ex-vivo porcine tissue under standardized conditions with meticulous adjustment of exposure parameters. Further investigations in vivo are necessary to test whether large tissue volumes can be ablated completely and reliably despite the influence of physiologic tissue and organ movement.

Plaque characterization in ex vivo MRI evaluated by dense 3D correspondence with histology

DEFF Research Database (Denmark)

van Engelen, Arna; de Bruijne, Marleen; Klein, Stefan

2011-01-01

registration of histology data with ex vivo MRI data, using non-rigid registration, both for training and evaluation. This is more objective than previously presented methods, as it eliminates selection bias that is introduced when 2D MRI slices are manually matched to histological slices before evaluation....... Histological slices of human atherosclerotic plaques were manually segmented into necrotic core, fibrous tissue and calcification. Classification of these three components was voxelwise evaluated. As features the intensity, gradient magnitude and Laplacian in four MRI sequences after different degrees...

Ex vivo imaging of early dental caries within the interproximal space

Science.gov (United States)

Choo-Smith, Lin-P'ing; Hewko, Mark D.; Dufour, Marc L.; Fulton, Crystal; Qiu, Pingli; Gauthier, Bruno; Padioleau, Christian; Bisaillon, Charles-Etienne; Dong, Cecilia; Cleghorn, Blaine M.; Lamouche, Guy; Sowa, Michael G.

2009-02-01

Optical coherence tomography (OCT) is emerging as a technology that can potentially be used for the detection and monitoring of early dental enamel caries since it can provide high-resolution depth imaging of early lesions. To date, most caries detection optical technologies are well suited for examining caries at facial, lingual, incisal and occlusal surfaces. The approximal surfaces between adjacent teeth are difficult to examine due to lack of visual access and limited space for these new caries detection tools. Using a catheter-style probe developed at the NRC-Industrial Materials Institute, the probe was inserted into the interproximal space to examine the approximal surfaces with OCT imaging at 1310 nm. The probe was rotated continuously and translated axially to generate depth images in a spiral fashion. The probe was used in a mock tooth arch model consisting of extracted human teeth mounted with dental rope wax in their anatomically correct positions. With this ex vivo model, the probe provided images of the approximal surfaces revealing morphological structural details, regions of calculus, and especially regions of early dental caries (white spot lesions). Results were compared with those obtained from OCT imaging of individual samples where the approximal surfaces of extracted teeth are accessible on a lab-bench. Issues regarding access, regions of interest, and factors to be considered in an in vivo setting will be discussed. Future studies are aimed at using the probe in vivo with patient volunteers.

A novel animal model for in vivo study of liver cancer metastasis

Institute of Scientific and Technical Information of China (English)

Shinsuke Fujiwara; Katsutoshi Yoshizato; Hikaru Fujioka; Chise Tateno; Ken Taniguchi; Masahiro Ito; Hiroshi Ohishi; Rie Utoh; Hiromi Ishibashi; Takashi Kanematsu

2012-01-01

AIM:To establish an animal model with human hepatocyte-repopulated liver for the study of liver cancer metastasis.METHODS:Cell transplantation into mouse livers was conducted using alpha-fetoprotein (AFP)-producing human gastric cancer cells (h-GCCs) and h-hepatocytes as donor cells in a transgenic mouse line expressing urokinase-type plasminogen activator (uPA) driven by the albumin enhancer/promoter crossed with a severe combined immunodeficient (SCID) mouse line (uPA/SCID mice).Host mice were divided into two groups (A and B).Group A mice were transplanted with h-GCCs alone,and group B mice were transplanted with h-GCCs and h-hepatocytes together.The replacement index (RI),which is the ratio of transplanted h-GCCs and h-hepatocytes that occupy the examined area of a histological section,was estimated by measuring h-AFP and h-albumin concentrations in sera,respectively,as well as by immunohistochemical analyses of h-AFP and human cytokeratin 18 in histological sections.RESULTS:The h-GCCs successfully engrafted,repopulated,and colonized the livers of mice in group A (RI =22.0％ ± 2.6％).These mice had moderately differentiated adenocarcinomatous lesions with disrupted glandular structures,which is a characteristics feature of gastric cancers.The serum h-AFP level reached 211.0 ± 142.2 g/mL (range,7.1-324.2 g/mL).In group B mice,the h-GCCs and h-hepatocytes independently engrafted,repopulated the host liver,and developed colonies (RI =12.0％ ± 6.8％ and 66.0％ ± 12.3％,respectively).h-GCC colonies also showed typical adenocarcinomatous glandular structures around the h-hepatocyte-colonies.These mice survived for the full 56day-study and did not exhibit any metastasis of h-GCCs in the extrahepatic regions during the observational period.The mice with an h-hepatocyte-repopulated liver possessed metastasized h-GCCs and therefore could be a useful humanized liver animal model for studying liver cancer metastasis in vivo.CONCLUSION:A novel animal model of

An Ex Vivo Imaging Pipeline for Producing High- Quality and High-Resolution Diffusion-Weighted Imaging Datasets

DEFF Research Database (Denmark)

Dyrby, Tim Bjørn; Baaré, William F.C.; Alexander, Daniel C.

2011-01-01

Diffusion tensor (DT) imaging and related multifiber reconstruction algorithms allow the study of in vivo microstructure and, by means of tractography, structural connectivity. Although reconstruction algorithms are promising imaging tools, high‐quality diffusion‐weighted imaging (DWI) datasets...... complexity, to establish an ex vivo imaging pipeline for generating high‐quality DWI datasets. Perfusion fixation ensured that tissue characteristics were comparable to in vivo conditions. There were three main results: (i) heat conduction and unstable tissue mechanics accounted for time‐varying artefacts...... in the DWI dataset, which were present for up to 15 h after positioning brain tissue in the scanner; (ii) using fitted DT, q‐ball, and persistent angular structure magnetic resonance imaging algorithms, any b‐value between ∼2,000 and ∼8,000 s/mm2, with an optimal value around 4,000 s/mm2, allowed...

Transplantation of human neonatal foreskin stromal cells in ex vivo organotypic cultures of embryonic chick femurs

DEFF Research Database (Denmark)

Aldahmash, Abdullah; Vishnubalaji, Radhakrishnan

2017-01-01

NSSCs in ex vivo organotypic cultures of embryonic chick femurs. Isolated embryonic chick femurs (E10 and E11) were cultured for 10 days together with micro-mass cell pellets of hNSSCs, human umbilical vein endothelial cells (HUVEC) or a combination of the two cell types. Changes in femurs gross morphology......We have previously reported that human neonatal foreskin stromal cells (hNSSCs) promote angiogenesis in vitro and in chick embryo chorioallantoic membrane (CAM) assay in vivo. To examine the in vivo relevance of this observation, we examined in the present study the differentiation potential of h......NSSC + HUVEC cultures. Our data suggest that organotypic cultures can be employed to test the differentiation potential of stem cells and demonstrate the importance of stem cell interaction with 3D-intact tissue microenvironment for their differentiation....

Effect of Er,Cr:YSGG laser irradiation on bovine enamel surface during in-office tooth bleaching ex vivo.

Science.gov (United States)

Dionysopoulos, Dimitrios; Strakas, Dimitrios; Koliniotou-Koumpia, Eugenia; Koumpia, Effimia

2017-07-01

The aim of this in vitro study was to evaluate the effect of using Er,Cr:YSGG laser during in-office tooth bleaching on bovine enamel surface to evaluate the safety of this therapy on tooth tissues. Thirty-six enamel specimens were prepared from bovine incisors and divided into three groups: Group 1 specimens (control) received no bleaching treatment; Group 2 received a conventional in-office bleaching treatment (40 % H 2 O 2 ); Group 3 received laser-assisted bleaching procedure (40 % H 2 O 2 ) utilizing an Er,Cr:YSGG laser. The specimens were stored for 10 days after the bleaching treatment in artificial saliva. Vickers hardness was determined using a microhardness tester, and measurements for surface roughness were done using a VSI microscope. Three specimens for each experimental group were examined under SEM and mineral composition of the specimens was evaluated using EDS. Data were statistically analyzed using one-way ANOVA, Tukey's post hoc, Wilcoxon signed rank and Kruskal-Wallis tests (a = 0.05). The Vickers hardness of the enamel was reduced after in-office bleaching procedures (p  0.05), and no changes in mineral composition of the enamel were detected after in-office bleaching procedures (p > 0.05). The laser-assisted bleaching treatment with Er,Cr:YSGG laser did not influence the enamel surface compared to the conventional bleaching technique. The safety of the use of Er,Cr:YSGG laser during in-office tooth bleaching regarding the surface properties of the enamel was confirmed under in vitro conditions.

Late Effects of Heavy Ion Irradiation on Ex Vivo Osteoblastogenesis and Cancellous Bone Microarchitecture

Science.gov (United States)

Tran, Luan Hoang; Alwood, Joshua; Kumar, Akhilesh; Limoli, C. L.; Globus, Ruth

2012-01-01

Prolonged spaceflight causes degeneration of skeletal tissue with incomplete recovery even after return to Earth. We hypothesize that heavy ion irradiation, a component of Galactic Cosmic Radiation, damages osteoblast progenitors and may contribute to bone loss during long duration space travel beyond the protection of the Earth's magnetosphere. Male, 16 week old C57BL6/J mice were exposed to high LET (56 Fe, 600MeV) radiation using either low (5 or 10cGy) or high (50 or 200cGy) doses at the NASA Space Radiation Lab and were euthanized 3 - 4, 7, or 35 days later. Bone structure was quantified by microcomputed tomography (6.8 micron pixel size) and marrow cell redox assessed using membrane permeable, free radical sensitive fluorogenic dyes. To assess osteoblastogenesis, adherent marrow cells were cultured ex vivo, then mineralized nodule formation quantified by imaging and gene expression analyzed by RT PCR. Interestingly, 3 - 4 days post exposure, fluorogenic dyes that reflect cytoplasmic generation of reactive nitrogen/oxygen species (DAF FM Diacetate or CM H2DCFDA) revealed irradiation (50cGy) reduced free radical generation (20-45%) compared to sham irradiated controls. Alternatively, use of a dye showing relative specificity for mitochondrial superoxide generation (MitoSOX) revealed an 88% increase compared to controls. One week after exposure, reactive oxygen/nitrogen levels remained lower(24%) relative to sham irradiated controls. After one month, high dose irradiation (200 cGy) caused an 86% decrement in ex vivo nodule formation and a 16-31% decrement in bone volume to total volume and trabecular number (50, 200cGy) compared to controls. High dose irradiation (200cGy) up regulated expression of a late osteoblast marker (BGLAP) and select genes related to oxidative metabolism (Catalase) and DNA damage repair (Gadd45). In contrast, lower doses (5, 10cGy) did not affect bone structure or ex vivo nodule formation, but did down regulate iNOS by 0.54 - 0.58 fold

Utilization of the organ care system as ex-vivo lung perfusion after cold storage transportation.

Science.gov (United States)

Mohite, P N; Maunz, O; Popov, A-F; Zych, B; Patil, N P; Simon, A R

2015-11-01

The Organ Care System (OCS) allows perfusion and ventilation of the donor lungs under physiological conditions. Ongoing trials to compare preservation with OCS Lung with standard cold storage do not include donor lungs with suboptimal gas exchange and donor lungs treated with OCS following cold storage transportation. We present a case of a 48-yr-old man who received such lungs after cold storage transportation treated with ex-vivo lung perfusion utilizing OCS. © The Author(s) 2015.

Maintenance of drug metabolism and transport functions in human precision-cut liver slices during prolonged incubation for 5 days

NARCIS (Netherlands)

Starokozhko, Viktoriia; Vatakuti, Suresh; Schievink, Bauke; Merema, Marjolijn T.; Asplund, Annika; Synnergren, Jane; Aspegren, Anders; Groothuis, Geny M. M.

Human precision-cut liver slices (hPCLS) are a valuable ex vivo model that can be used in acute toxicity studies. However, a rapid decline in metabolic enzyme activity limits their use in studies that require a prolonged xenobiotic exposure. The aim of the study was to extend the viability and

Machine Perfusion of Porcine Livers with Oxygen-Carrying Solution Results in Reprogramming of Dynamic Inflammation Networks

Directory of Open Access Journals (Sweden)

David Sadowsky

2016-11-01

Full Text Available Background: Ex vivo machine perfusion (MP can better preserve organs for transplantation. We have recently reported on the first application of a MP protocol in which liver allografts were fully oxygenated, under dual pressures and subnormothermic conditions, with a new hemoglobin-based oxygen carrier solution specifically developed for ex vivo utilization. In those studies, MP improved organ function post-operatively and reduced inflammation in porcine livers. Herein, we sought to refine our knowledge regarding the impact of MP by defining dynamic networks of inflammation in both tissue and perfusate. Methods: Porcine liver allografts were preserved either with MP (n = 6 or with cold static preservation (CSP; n = 6, then transplanted orthotopically after 9 h of preservation. Fourteen inflammatory mediators were measured in both tissue and perfusate during liver preservation at multiple time points, and analyzed using Dynamic Bayesian Network (DyBN inference to define feedback interactions, as well as Dynamic Network Analysis (DyNA to define the time-dependent development of inflammation networks.Results: Network analyses of tissue and perfusate suggested an NLRP3 inflammasome-regulated response in both treatment groups, driven by the pro-inflammatory cytokine interleukin (IL-18 and the anti-inflammatory mediator IL-1 receptor antagonist (IL-1RA. Both DyBN and DyNA suggested a reduced role of IL-18 and increased role of IL-1RA with MP, along with increased liver damage with CSP. DyNA also suggested divergent progression of responses over the 9 h preservation time, with CSP leading to a stable pattern of IL-18-induced liver damage and MP leading to a resolution of the pro-inflammatory response. These results were consistent with prior clinical, biochemical, and histological findings after liver transplantation. Conclusion: Our results suggest that analysis of dynamic inflammation networks in the setting of liver preservation may identify novel

Metabolism of 5-fluorouracil in human liver: an in vivo 19F NMR study

International Nuclear Information System (INIS)

Mohankrishnan, P.; Sprigg, J.; Cardwell, D.; Komoroski, R.A.; Hutchins, L.; Nauke, S.; Williamson, M.R.; Jagannathan, N.R.

1999-01-01

In vivo fluorine-19 nuclear magnetic resonance ( 19 F NMR) spectroscopy was used to study the metabolism and pharmacokinetics of 5-fluorouracil (5-FU) in human liver. Nine patients received 5-FU, and additional chemotherapeutic agents (methotrexate, leucovorin, or levamisole) either prophylactically after breast cancer surgery or for colorectal cancer. The time constant for the disappearance of 5-FU from the liver in vivo varied from 5 to 17 min, while the time constant for the appearance of α-fluoro-β-alanine (the major catabolite of 5 FU) varied from 7 to 86 min. The modulators of 5-FU metabolism did not appear to affect the time constant for the disappearance of 5-FU from the liver or for the appearance of α-fluoro-β-alanine. Results obtained indicate that the pharmacokinetics of 5-FU and α-fluoro-β-alanine may vary substantially at different times in a given individual. (author)

Extended normothermic extracorporeal perfusion of isolated human liver after warm ischaemia: a preliminary report.

Science.gov (United States)

Bellomo, Rinaldo; Marino, Bruno; Starkey, Graeme; Fink, Michael; Wang, Bao Zhong; Eastwood, Glenn M; Peck, Leah; Young, Helen; Houston, Shane; Skene, Alison; Opdam, Helen; Jones, Robert

2014-09-01

Donation after circulatory death (DCD) livers are at markedly increased risk of primary graft dysfunction and biliary tract ischaemia. Normothermic extracorporeal liver perfusion (NELP) may increase the ability to transplant DCD livers and may allow their use for artificial extracorporeal liver support of patients with fulminant liver failure. We conducted two proof-of-concept experiments using human livers after DCD to assess the feasibility and functional efficacy of NELP over an extended period. We applied extracorporeal membrane oxygenation, parenteral nutrition, separate hepatic artery and portal vein perfusion and physiological perfusion pressures to two livers obtained after DCD. We achieved NELP and evidence of liver function (bile production, paracetamol removal and maintenance of normal lactate levels) in both livers; one for 24 hours and the other for 43 hours. Histological examination showed areas of patchy ischaemia but preserved biliary ducts and canaliculi. Our experiments justify further investigations of the feasibility and efficacy of extended DCD liver preservation by ex-vivo perfusion.

The role of Hibiscus sabdariffa L. (Roselle) in maintenance of ex vivo murine bone marrow-derived hematopoietic stem cells.

Science.gov (United States)

Abdul Hamid, Zariyantey; Lin Lin, Winnie Hii; Abdalla, Basma Jibril; Bee Yuen, Ong; Latif, Elda Surhaida; Mohamed, Jamaludin; Rajab, Nor Fadilah; Paik Wah, Chow; Wak Harto, Muhd Khairul Akmal; Budin, Siti Balkis

2014-01-01

Hematopoietic stem cells- (HSCs-) based therapy requires ex vivo expansion of HSCs prior to therapeutic use. However, ex vivo culture was reported to promote excessive production of reactive oxygen species (ROS), exposing HSCs to oxidative damage. Efforts to overcome this limitation include the use of antioxidants. In this study, the role of Hibiscus sabdariffa L. (Roselle) in maintenance of cultured murine bone marrow-derived HSCs was investigated. Aqueous extract of Roselle was added at varying concentrations (0-1000 ng/mL) for 24 hours to the freshly isolated murine bone marrow cells (BMCs) cultures. Effects of Roselle on cell viability, reactive oxygen species (ROS) production, glutathione (GSH) level, superoxide dismutase (SOD) activity, and DNA damage were investigated. Roselle enhanced the survival (P Roselle increased (P Roselle showed significant cellular genoprotective potency against H2O2-induced DNA damage. Conclusively, Roselle shows novel property as potential supplement and genoprotectant against oxidative damage to cultured HSCs.

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

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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

Maltitol inhibits small intestinal glucose absorption and increases insulin mediated muscle glucose uptake ex vivo but not in normal and type 2 diabetic rats.

Science.gov (United States)

Chukwuma, Chika Ifeanyi; Ibrahim, Mohammed Auwal; Islam, Md Shahidul

2017-02-01

This study investigated the effects of maltitol on intestinal glucose absorption and muscle glucose uptake using ex vivo and in vivo experimental models. The ex vivo experiment was conducted in isolated jejunum and psoas muscle from normal rats. The in vivo study investigated the effects of a single bolus dose of maltitol on gastric emptying, intestinal glucose absorption and digesta transit in normal and type 2 diabetic rats. Maltitol inhibited glucose absorption in isolated rat jejunum and increased glucose uptake in isolated rat psoas muscle in the presence of insulin but not in the absence of insulin. In contrast, maltitol did not significantly (p > 0.05) alter small intestinal glucose absorption or blood glucose levels as well as gastric emptying and digesta transit in normal or type 2 diabetic rats. The results suggest that maltitol may not be a suitable dietary supplement for anti-diabetic food and food products to improve glycemic control.

CT Fluoroscopy-Guided Lung Biopsy with Novel Steerable Biopsy Canula: Ex-Vivo Evaluation in Ventilated Porcine Lung Explants

International Nuclear Information System (INIS)

Schaefer, Philipp J.; Fabel, Michael; Bolte, Hendrik; Schaefer, Fritz K. W.; Jahnke, Thomas; Heller, Martin; Lammer, Johannes; Biederer, Juergen

2010-01-01

The purpose was to evaluate ex-vivo a prototype of a novel biopsy canula under CT fluoroscopy-guidance in ventilated porcine lung explants in respiratory motion simulations. Using an established chest phantom for porcine lung explants, n = 24 artificial lesions consisting of a fat-wax-Lipiodol mixture (approx. 70HU) were placed adjacent to sensible structures such as aorta, pericardium, diaphragm, bronchus and pulmonary artery. A piston pump connected to a reservoir beneath a flexible silicone reconstruction of a diaphragm simulated respiratory motion by rhythmic inflation and deflation of 1.5 L water. As biopsy device an 18-gauge prototype biopsy canula with a lancet-like, helically bended cutting edge was used. The artificial lesions were punctured under CT fluoroscopy-guidance (SOMATOM Sensation 64, Siemens, Erlangen, Germany; 30mAs/120 kV/5 mm slice thickness) implementing a dedicated protocol for CT fluoroscopy-guided lung biopsy. The mean-diameter of the artificial lesions was 8.3 ± 2.6 mm, and the mean-distance of the phantom wall to the lesions was 54.1 ± 13.5 mm. The mean-displacement of the lesions by respiratory motion was 14.1 ± 4.0 mm. The mean-duration of CT fluoroscopy was 9.6 ± 5.1 s. On a 4-point scale (1 = central; 2 = peripheral; 3 = marginal; 4 = off target), the mean-targeted precision was 1.9 ± 0.9. No misplacement of the biopsy canula affecting adjacent structures could be detected. The novel steerable biopsy canula proved to be efficient in the ex-vivo set-up. The chest phantom enabling respiratory motion and the steerable biopsy canula offer a feasible ex-vivo system for evaluating and training CT fluoroscopy-guided lung biopsy adapted to respiratory motion.

First phase 1 double-blind, placebo-controlled, randomized rectal microbicide trial using UC781 gel with a novel index of ex vivo efficacy.

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Peter A Anton

Full Text Available Successful control of the HIV/AIDS pandemic requires reduction of HIV-1 transmission at sexually-exposed mucosae. No prevention studies of the higher-risk rectal compartment exist. We report the first-in-field Phase 1 trial of a rectally-applied, vaginally-formulated microbicide gel with the RT-inhibitor UC781 measuring clinical and mucosal safety, acceptability and plasma drug levels. A first-in-Phase 1 assessment of preliminary pharmacodynamics was included by measuring changes in ex vivo HIV-1 suppression in rectal biopsy tissue after exposure to product in vivo.HIV-1 seronegative, sexually-abstinent men and women (N = 36 were randomized in a double-blind, placebo-controlled trial comparing UC781 gel at two concentrations (0.1%, 0.25% with placebo gel (1∶1∶1. Baseline, single-dose exposure and a separate, 7-day at-home dosing were assessed. Safety and acceptability were primary endpoints. Changes in colorectal mucosal markers and UC781 plasma drug levels were secondary endpoints; ex vivo biopsy infectibility was an ancillary endpoint.All 36 subjects enrolled completed the 7-14 week trial (100% retention including 3 flexible sigmoidoscopies, each with 28 biopsies (14 at 10 cm; 14 at 30 cm. There were 81 Grade 1 adverse events (AEs and 8 Grade 2; no Grade 3, 4 or procedure-related AEs were reported. Acceptability was high, including likelihood of future use. No changes in mucosal immunoinflammatory markers were identified. Plasma levels of UC781 were not detected. Ex vivo infection of biopsies using two titers of HIV-1(BaL showed marked suppression of p24 in tissues exposed in vivo to 0.25% UC781; strong trends of suppression were seen with the lower 0.1% UC781 concentration.Single and 7-day topical rectal exposure to both concentrations of UC781 were safe with no significant AEs, high acceptability, no detected plasma drug levels and no significant mucosal changes. Ex vivo biopsy infections demonstrated marked suppression of HIV

Recognition algorithm for assisting ovarian cancer diagnosis from coregistered ultrasound and photoacoustic images: ex vivo study

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Alqasemi, Umar; Kumavor, Patrick; Aguirre, Andres; Zhu, Quing

2012-12-01

Unique features and the underlining hypotheses of how these features may relate to the tumor physiology in coregistered ultrasound and photoacoustic images of ex vivo ovarian tissue are introduced. The images were first compressed with wavelet transform. The mean Radon transform of photoacoustic images was then computed and fitted with a Gaussian function to find the centroid of a suspicious area for shift-invariant recognition process. Twenty-four features were extracted from a training set by several methods, including Fourier transform, image statistics, and different composite filters. The features were chosen from more than 400 training images obtained from 33 ex vivo ovaries of 24 patients, and used to train three classifiers, including generalized linear model, neural network, and support vector machine (SVM). The SVM achieved the best training performance and was able to exclusively separate cancerous from non-cancerous cases with 100% sensitivity and specificity. At the end, the classifiers were used to test 95 new images obtained from 37 ovaries of 20 additional patients. The SVM classifier achieved 76.92% sensitivity and 95.12% specificity. Furthermore, if we assume that recognizing one image as a cancer is sufficient to consider an ovary as malignant, the SVM classifier achieves 100% sensitivity and 87.88% specificity.

Proteomic Profiling of Ex Vivo Expanded CD34-Positive Haematopoetic Cells Derived from Umbilical Cord Blood

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Heiner Falkenberg

2013-01-01

Full Text Available Ex vivo expansion of haematopoetic cells by application of specific cytokines is one approach to overcome boundaries in cord blood transplantation due to limited numbers of haematopoetic stem cells. While many protocols describe an effective increase of total cell numbers and the amount of CD34-positive cells, it still remains unclear if and how the procedure actually affects the cells’ properties. In the presented publications, CD34-positive cells were isolated from cord blood and expanded for up to 7 days in media supplemented with stem cell factor (SCF, thrombopoietin (THPO, interleukin 6 (IL-6, and fms-related tyrosine kinase 3 ligand (FLT3lg. At days 3 and 7, expanded cells were harvested and analyzed by flow cytometry and quantitative proteomics. 2970 proteins were identified, whereof proteomic analysis showed 440 proteins significantly changed in abundance during ex vivo expansion. Despite the fact that haematopoetic cells still expressed CD34 on the surface after 3 days, major changes in regard to the protein profile were observed, while further expansion showed less effect on the proteome level. Enrichment analysis of biological processes clearly showed a proteomic change toward a protein biosynthesis phenotype already within the first three days of expression.

Comparative ex vivo study on humidifying function of three speaking valves with integrated heat and moisture exchanger for tracheotomised patients

NARCIS (Netherlands)

van den Boer, C.; Lansaat, L.; Muller, S.H.; van den Brekel, M.W.M.; Hilgers, F.J.M.

2015-01-01

Objective Assessment of humidifying function of tracheotomy speaking valves with integrated heat and moisture exchanger. Design Ex vivo measurement of water exchange and storage capacity of three tracheotomy speaking valves: Humidiphon Plus, Spiro and ProTrach DualCare (with two different heat and

Comparative ex vivo study on humidifying function of three speaking valves with integrated heat and moisture exchanger for tracheotomised patients

NARCIS (Netherlands)

van den Boer, C.; Lansaat, L.; Muller, S. H.; van den Brekel, M. W. M.; Hilgers, F. J. M.

2015-01-01

Assessment of humidifying function of tracheotomy speaking valves with integrated heat and moisture exchanger. Ex vivo measurement of water exchange and storage capacity of three tracheotomy speaking valves: Humidiphon Plus, Spiro and ProTrach DualCare (with two different heat and moisture

Angiopoietin-like protein 3 promotes preservation of stemness during ex vivo expansion of murine hematopoietic stem cells.

Science.gov (United States)

Farahbakhshian, Elnaz; Verstegen, Monique M; Visser, Trudi P; Kheradmandkia, Sima; Geerts, Dirk; Arshad, Shazia; Riaz, Noveen; Grosveld, Frank; van Til, Niek P; Meijerink, Jules P P

2014-01-01

Allogeneic hematopoietic stem cell (HSC) transplantations from umbilical cord blood or autologous HSCs for gene therapy purposes are hampered by limited number of stem cells. To test the ability to expand HSCs in vitro prior to transplantation, two growth factor cocktails containing stem cell factor, thrombopoietin, fms-related tyrosine kinase-3 ligand (STF) or stem cell factor, thrombopoietin, insulin-like growth factor-2, fibroblast growth factor-1 (STIF) either with or without the addition of angiopoietin-like protein-3 (Angptl3) were used. Culturing HSCs in STF and STIF media for 7 days expanded long-term repopulating stem cells content in vivo by ∼6-fold and ∼10-fold compared to freshly isolated stem cells. Addition of Angptl3 resulted in increased expansion of these populations by ∼17-fold and ∼32-fold, respectively, and was further supported by enforced expression of Angptl3 in HSCs through lentiviral transduction that also promoted HSC expansion. As expansion of highly purified lineage-negative, Sca-1+, c-Kit+ HSCs was less efficient than less pure lineage-negative HSCs, Angptl3 may have a direct effect on HCS but also an indirect effect on accessory cells that support HSC expansion. No evidence for leukemia or toxicity was found during long-term follow up of mice transplanted with ex vivo expanded HSCs or manipulated HSC populations that expressed Angptl3. We conclude that the cytokine combinations used in this study to expand HSCs ex vivo enhances the engraftment in vivo. This has important implications for allogeneic umbilical cord-blood derived HSC transplantations and autologous HSC applications including gene therapy.

Effect of irradiation on gene expression of rat liver adhesion molecules. In vivo and in vitro studies

International Nuclear Information System (INIS)

Moriconi, Federico; Malik, Ihtzaz; Ahmad, Ghayyor; Dudas, Joszef; Ramadori, Giuliano; Rave-Fraenk, Margret; Vorwerk, Hilke; Hille, Andrea; Hess, Clemens Friedrich; Christiansen, Hans

2009-01-01

Background and purpose: Migration of leukocytes into tissue is a key element of innate and adaptive immunity. An animal study showed that liver irradiation, in spite of induction of chemokine gene expression, does not lead to recruitment of leukocytes into the parenchyma. The aim of this study was to analyze gene expression of adhesion molecules, which mediate leukocyte recruitment into organs, in irradiated rat liver in vivo and rat hepatocytes in vitro. Material and methods: Rat livers in vivo were irradiated selectively at 25 Gy. Isolated hepatocytes in vitro were irradiated at 8 Gy. RNA extracted within 48 h after irradiation in vivo and in vitro was analyzed by real-time PCR (polymerase chain reaction) and Northern blot. Adhesion molecule concentration in serum was measured by ELISA (enzyme-linked immunosorbent assay). Cryostat sections of livers were used for immunohistology. Results: Significant radiation-induced increase of ICAM-1 (intercellular adhesion molecule-1), VCAM-1 (vascular cell adhesion molecule-1), JAM-1 (junctional adhesion molecule-1), β 1 -integrin, β 2 -integrin, E-cadherin, and P-selectin gene expression could be detected in vivo, while PECAM-1 (platelet-endothelial cell adhesion molecule-1) gene expression remained unchanged. In vitro, β 1 -integrin, JAM-1, and ICAM-2 showed a radiation-induced increased expression, whereas the levels of P-selectin, ICAM-1, PECAM-1, VCAM-1, Madcam-1 (mucosal addressin cell adhesion molecule-1), β 2 -integrin, and E-cadherin were downregulated. However, incubation of irradiated hepatocytes with either tumor necrosis factor-(TNF-)α, interleukin-(IL-)1β, or IL-6 plus TNF-α led to an upregulation of P-selectin, ICAM-1 and VCAM-1. Conclusion: The findings suggest that liver irradiation modulates gene expression of the main adhesion molecules in vivo and in cytokine-activated hepatocytes, with the exception of PECAM-1. This may be one reason for the lack of inflammation in the irradiated rat liver. (orig.)

Profiling of Cytokines Secreted by Conventional Aqueous Outflow Pathway Endothelial Cells Activated In Vitro and Ex Vivo With Laser Irradiation.

Science.gov (United States)

Alvarado, Jorge A; Chau, Phuonglan; Wu, Jianfeng; Juster, Richard; Shifera, Amde Selassie; Geske, Michael

2015-11-01

To profile which cytokine genes are differentially expressed (DE) as up- or downregulated by cultured human trabecular meshwork (TMEs) and Schlemm's canal endothelial cells (SCEs) after three experimental treatments consisting of selective laser trabeculoplasty (SLT) irradiation, exposure to media conditioned either by SLT-irradiated TMEs (TME-cm) or by SCEs (SCE-cm). Also, to profile which cytokines are upregulated ex vivo in SLT-irradiated human conventional aqueous outflow pathway (CAOP) tissues. After each treatment, Affymetrix microarray assays were used to detect upregulated and downregulated genes for cytokines and their receptors in TMEs and SCEs. ELISA and protein antibody arrays were used to detect upregulated cytokines secreted in SLT-irradiated CAOP tissues ex vivo. The SLT irradiation upregulated numerous cytokine genes in TMEs, but only a few in SCEs. Exposure to TME- and SCE-cm induced SCEs to upregulate many more cytokine genes than TMEs. Selective laser trabeculoplasty irradiation and exposure to TME-cm downregulated several cytokine genes in TMEs but none in SCEs. Selective laser trabeculoplasty irradiation induced one upregulated and three downregulated cytokine-receptor genes in TMEs but none in SCEs. Exposure to TME-cm induced upregulation of one and downregulation of another receptor gene in TMEs, whereas two unique cytokine-receptor genes were upregulated in SCEs. Cytokine protein expression analysis showed that at least eight cytokines were upregulated in SLT-irradiated human CAOP tissues in situ/ex vivo. This study has helped us identify a cytokine signaling pathway and to consider newly identified mechanisms regulating aqueous outflow that may lay the foundation for the future development of cytokine-based glaucoma therapies.

Ex Vivo Normothermic Perfusion Induces Donor-Derived Leukocyte Mobilization and Removal Prior to Renal Transplantation

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John P. Stone

2016-11-01

Discussion: We demonstrate that ex vivo normothermic perfusion initiates an inflammatory cytokine storm and release of mitochondrial and genomic DNA. This is likely to be responsible for immune cell activation and mobilization into the circuit prior to transplantation. Interestingly this did not have an impact on renal function. These data therefore suggest that normothermic perfusion can be used to immunodeplete and to saturate the pro-inflammatory capacity of donor kidneys prior to transplantation.

[68Ga]pentixafor for CXCR4 imaging in a PC-3 prostate cancer xenograft model - comparison with [18F]FDG PET/CT, MRI and ex vivo receptor expression.

Science.gov (United States)

Schwarzenböck, Sarah M; Stenzel, Jan; Otto, Thomas; Helldorff, Heike V; Bergner, Carina; Kurth, Jens; Polei, Stefan; Lindner, Tobias; Rauer, Romina; Hohn, Alexander; Hakenberg, Oliver W; Wester, Hans J; Vollmar, Brigitte; Krause, Bernd J

2017-11-10

The aim was to characterize the properties of [ 68 Ga]Pentixafor as tracer for prostate cancer imaging in a PC-3 prostate cancer xenograft mouse model and to investigate its correlation with [ 18 F]FDG PET/CT, magnetic resonance imaging (MRI) and ex vivo analyses. Static [ 68 Ga]Pentixafor and [ 18 F]FDG PET as well as morphological/ diffusion weighted MRI and 1 H MR spectroscopy was performed. Imaging data were correlated with ex vivo biodistribution and CXCR4 expression in PC-3 tumors (immunohistochemistry (IHC), mRNA analysis). Flow cytometry was performed for evaluation of localization of CXCR4 receptors ( in vitro PC-3 cell experiments). Tumor uptake of [ 68 Ga]Pentixafor was significantly lower compared to [ 18 F]FDG. Ex vivo CXCR4 mRNA expression of tumors was shown by PCR. Only faint tumor CXCR4 expression was shown by IHC (immuno reactive score of 3). Accordingly, flow cytometry of PC-3 cells revealed only a faint signal, cell membrane permeabilisation showed a slight signal increase. There was no significant correlation of [ 68 Ga]Pentixafor tumor uptake and ex vivo receptor expression. Spectroscopy showed typical spectra of prostate cancer. PC-3 tumor uptake of [ 68 Ga]Pentixafor was existent but lower compared to [ 18 F]FDG. No significant correlation of ex vivo tumor CXCR4 receptor expression and [ 68 Ga]Pentixafor tumor uptake was shown. CXCR4 receptor expression on the surface of PC-3 cells was existent but rather low possibly explaining the limited [ 68 Ga]Pentixafor tumor uptake; receptor localization in the interior of PC-3 cells is presumable as shown by cell membrane permeabilisation. Further studies are necessary to define the role of [ 68 Ga]Pentixafor in prostate cancer imaging.

Measurement of shear wave speed dispersion in the placenta by transient elastography: A preliminary ex vivo study.

Science.gov (United States)

Simon, Emmanuel G; Callé, Samuel; Perrotin, Franck; Remenieras, Jean-Pierre

2018-01-01

Placental elasticity may be modified in women with placental insufficiency. Shear wave elastography (SWE) can measure this, using acoustic radiation force, but the safety of its use in pregnant women has not yet been demonstrated. Transient elastography (TE) is a safer alternative, but has not yet been applied to the placenta. Moreover, the dispersion of shear wave speed (SWS) as a function of frequency has received relatively little study for placental tissue, although it might improve the accuracy of biomechanical assessment. To explore the feasibility and reproducibility of TE for placental analysis, to compare the values of SWS and Young's modulus (YM) from TE and SWE, and to analyze SWS dispersion as a function of frequency ex vivo in normal placentas. Ten normal placentas were analyzed ex vivo by an Aixplorer ultrasound system as shear waves were generated by a vibrating plate and by using an Aixplorer system. The frequency analysis provided the value of the exponent n from a fractional rheological model applied to the TE method. We calculated intra- and interobserver agreement for SWS and YM with 95% prediction intervals, created Bland-Altman plots with 95% limits of agreement, and estimated the intraclass correlation coefficient (ICC). The mean SWS was 1.80 m/s +/- 0.28 (standard deviation) with the TE method at 50 Hz and 1.82 m/s +/-0.13 with SWE (P = 0.912). No differences were observed between the central and peripheral regions of placentas with either TE or SWE. With TE, the intraobserver ICC for SWS was 0.68 (0.50-0.82), and the interobserver ICC for SWS 0.65 (0.37-0.85). The mean parameter n obtained from the fractional rheological model was 1.21 +/- 0.12, with variable values of n for any given SWS. TE is feasible and reproducible on placentas ex vivo. The frequency analysis of SWS provides additional information about placental elasticity and appears to be able to distinguish differences between placental structures.

Evaluating childhood obesity. Magnetic resonance-based quantification of abdominal adipose tissue and liver fat in children

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Raschpichler, M.C. [Leipzig Univ. (Germany). Dept. of Paediatric Radiology; Leipzig Univ. Medical Center (Germany). IFB Adiposity Diseases; Sorge, I.; Hirsch, W. [Leipzig Univ. (Germany). Dept. of Paediatric Radiology; Mende, M. [Leipzig Univ. (Germany). Clinical Trial Centre Leipzig; Sergeyev, E.; Koerner, A. [Leipzig Univ. (Germany). University Hospital for Children and Adolescents; Kruber, D. [Leipzig Univ. (Germany). Dept. of Oral, Craniomaxillofacial and Facial Plastic Surgery; Schick, F. [Univ. Hospital Tuebingen (Germany). Section on Experimental Radiology

2012-04-15

The purpose of this study is to establish and validate a magnetic resonance (MR)-based fat quantification package that provides an accurate assessment of abdominal adipose tissue and liver fat in children. Ex vivo trials with a torso model and water-oil mixtures are conducted. Abdominal adipose tissue (AAT) is covered by magnetic resonance imaging (MRI) using a fat-selective sequence and is analyzed by a plug-in based on the open source software Image. Liver fat (LF) is measured with localized {sup 1}H Magnetic Resonance Spectroscopy ({sup 1}H MRS) and the jMRUI (java-based Magnetic Resonance User Interface) software package. Evaluation of the clinical methodology involved a study of 10 children in this feasibility study (mean age and body mass index: 13.3 yr; 33.3 kg/m{sup 2}). To evaluate the method's validity, reference measurements were performed. Ex vivo trials with the torso model showed that adipose tissue was measured appropriately with a systematic underestimation by 9.3 {+-} 0.2 % (0.32 {+-} 0.064 kg). Coefficients of variation for both intra- and inter-observer measurements ranged between 0 - 2.7 % and repeated analyses showed significant equivalent results (p < 0.01). The lipid content obtained by {sup 1}H MRS ex vivo revealed significant equivalence with the predefined fat content in water-oil mixtures (p < 0.01). In vivo, the homemade plug-in significantly overestimated the AAT, with the visceral adipose tissue being most affected (+ 15.7 {+-} 8.4 %). Although an overestimation of the AAT by the presented plug-in should be taken into consideration, this children-friendly package enables the quantification of both LF and AAT within 30 min on a freeware-based platform. (orig.)

Normothermic extracorporeal perfusion of isolated porcine liver after warm ischaemia: a preliminary report.

Science.gov (United States)

Bellomo, Rinaldo; Suzuki, Satoshi; Marino, Bruno; Starkey, Graeme K; Chambers, Brenton; Fink, Michael A; Wang, Bao Zhong; Houston, Shane; Eastwood, Glenn; Calzavacca, Paolo; Glassford, Neil; Skene, Alison; Jones, Daryl A; Jones, Robert

2012-09-01

Liver transplantation is a major life-saving procedure, and donation after cardiac death (DCD) has increased the pool of potential liver donors. However, DCD livers are at increased risk of primary graft dysfunction and biliary tract ischaemia. Normothermic extracorporeal liver perfusion (NELP) may increase the ability to protect, evaluate and, in future, transplant DCD livers. We conducted proof-of-concept experiments using a DCD model in the pig to assess the short-term (4 hours) feasibility and functional efficacy of NELP. Using extracorporeal membrane oxygenation, parenteral nutrition, separate hepatic artery and portal vein perfusion, and physiological perfusion pressures, we achieved NELP and evidence of function (bile production, paracetamol removal, maintenance of normal ammonia and lactate levels) for 4 hours in pig livers subjected to 15 and 30 minutes of cardiac arrest before explantation. Our experiments justify further investigations of the feasibility and efficacy of human DCD liver preservation by ex-vivo perfusion.

mRNA levels of imprinted genes in bovine in vivo oocytes, embryos and cross species comparisons in humans, mice and pigs

Science.gov (United States)

Twenty-six confirmed imprinted genes in the bovine were quantified in in vivo produced oocytes and embryos. Eighteen were detectable and their transcriptional abundance were categorized into five patterns: largely decreased (MEST and PLAGL1); first decreased and then increased (CDKN1C and IGF2R); p...

Measurements of fluorine in contemporary urban Canadians: a comparison of the levels found in human bone using in vivo and ex vivo neutron activation analysis

International Nuclear Information System (INIS)

Mostafaei, F; McNeill, F E; Chettle, D R; Prestwich, W V; Wainman, B C; Pidruczny, A E

2015-01-01

of the tea drinkers and the non-tea drinkers were found to be 0.127 (± 0.029) and 0.050 (± 0.009) mg F/g Ca per year respectively. Finally, we also obtained twelve bone samples from cadavers’ skulls. Neutron activation analysis was used to determine the fluorine levels in these ex vivo samples. The rate of increase of fluorine content versus age for in vivo and ex vivo measurements were found to be 0.078  ±  0.014 and 0.078  ±  0.050 mg F/g Ca per year respectively. Excellent agreement was found between the fluorine levels determined in vivo and ex vivo using the two separate systems, providing confidence in the fluorine concentration data being measured in vivo. (paper)

Clinical value of CT/MR-US fusion imaging for radiofrequency ablation of hepatic nodules

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Lee, Jae Young, E-mail: leejy4u@snu.ac.kr [Department of Radiology, Seoul National University Hospital, Seoul (Korea, Republic of); Institute of Radiation Medicine, Seoul National University Hospital, Seoul (Korea, Republic of); Choi, Byung Ihn [Department of Radiology, Seoul National University Hospital, Seoul (Korea, Republic of); Institute of Radiation Medicine, Seoul National University Hospital, Seoul (Korea, Republic of); Chung, Yong Eun [Department of Radiology, Research Institute of Radiological Science, Severance Hospital, Yonsei University College of Medicine, Seoul (Korea, Republic of); Kim, Min Wook; Kim, Se Hyung; Han, Joon Koo [Department of Radiology, Seoul National University Hospital, Seoul (Korea, Republic of); Institute of Radiation Medicine, Seoul National University Hospital, Seoul (Korea, Republic of)

2012-09-15

Objective: The aim of this study was to determine the registration error of an ultrasound (US) fusion imaging system during an ex vivo study and its clinical value for percutaneous radiofrequency ablation (pRFA) during an in vivo study. Materials and methods: An ex vivo study was performed using 4 bovine livers and 66 sonographically invisible lead pellets. Real-time CT-US fusion imaging was applied to assist the targeting of pellets with needles in each liver; the 4 sessions were performed by either an experienced radiologist (R1, 3 sessions) or an inexperienced resident (R2, 1 session). The distance between the pellet target and needle was measured. An in vivo study was retrospectively performed with 51 nodules (42 HCCs and 9 metastases; mean diameter, 16 mm) of 37 patients. Fusion imaging was used to create a sufficient safety margin (>5 mm) during pRFA in 24 nodules (group 1), accurately target 21 nodules obscured in the US images (group 2) and precisely identify 6 nodules surrounded by similar looking nodules (group 3). Image fusion was achieved using MR and CT images in 16 and 21 patients, respectively. The reablation rate, 1-year local recurrence rate and complications were assessed. Results: In the ex vivo study, the mean target–needle distances were 2.7 mm ± 1.9 mm (R1) and 3.1 ± 3.3 mm (R2) (p > 0.05). In the in vivo study, the reablation rates in groups 1–3 were 13%, 19% and 0%, respectively. At 1 year, the local recurrence rate was 11.8% (6/51). In our assessment of complications, one bile duct injury was observed. Conclusion: US fusion imaging system has an acceptable registration error and can be an efficacious tool for overcoming the major limitations of US-guided pRFA.

In vivo NMR spectroscopy of the liver. Spectroscopie RMN du foie in vivo

Energy Technology Data Exchange (ETDEWEB)

Jehenson, P.; Cuenod, C.A.; Syrota, A. (CEA, 91 - Orsay (FR). Service Hospitalier Frederic Joliot)

1989-01-01

The application of in vivo MR spectroscopy to the study of the liver is currently an expanding field of research. Owing to technical difficulties, the results obtained thus far were mainly those of animal observations. Several nuclei have been considered: hydrogen, phosphorus, carbon or fluorine. This non-traumatic method allows following and quantifying the various metabolic pathways, especially during hepatic diseases. The major metabolic pathways, i.e. neoglycogenesis, glycogenolysis, Krebs' cycle, etc., are studied, as well as their alterations during diseases such as ischemia, diabetes or alcoholism. The development of this promising technique requires the cooperation of various clinical and fundamental disciplines.

Gene expression of bovine embryos developing at the air-liquid interface on oviductal epithelial cells (ALI-BOEC).

Science.gov (United States)

van der Weijden, Vera A; Chen, Shuai; Bauersachs, Stefan; Ulbrich, Susanne E; Schoen, Jennifer

2017-11-25

We recently developed an air-liquid interface long-term culture of differentiated bovine oviductal epithelial cells (ALI-BOEC). This ex vivo oviduct epithelium is capable of supporting embryo development in co-culture up to the blastocyst stage without addition of embryo culture medium. However, blastocyst rates in co-culture were markedly lower than in conventional in vitro embryo production procedures. In the present study, we assessed target gene expression of ALI-BOEC derived embryos to test their similarity to embryos from conventional in vitro embryo culture. We screened previously published data from developing bovine embryos and selected 41 genes which are either differentially expressed during embryo development, or reflect differences between various in vitro culture conditions or in vitro and in vivo embryos. Target gene expression was measured in 8-cell embryos and blastocysts using a 48.48 Dynamic Array™ on a Biomark HD instrument. For comparison with the ALI-BOEC system, we generated embryos by two different standard IVP protocols. The culture conditions lead to differential gene expression in both 8-cell embryos and blastocysts. Across the expression of all target genes the embryos developing on ALI-BOEC did not depart from conventional IVP embryos. These first results prove that gene expression in ALI-BOEC embryos is not largely aberrant. However, there was no clear indication for a more in vivo-like target gene expression of these embryos. This calls for further optimization of the ALI-BOEC system to increase its efficiency both quantitatively and qualitatively.

Ex vivo evaluation of the serotonin 1A receptor partial agonist [³H]CUMI-101 in awake rats

DEFF Research Database (Denmark)

Palner, Mikael; Underwood, Mark D; Kumar, Dileep J S

2011-01-01

[³H]CUMI-101 is a 5-HT(1A) partial agonist, which has been evaluated for use as a positron emission tracer in baboon and humans. We sought to evaluate the properties of [³H]CUMI-101 ex vivo in awake rats and determine if [³H]CUMI-101 can measure changes in synaptic levels of serotonin after diffe...

Bioactive compounds fractionated from endophyte Streptomyces SUK 08 with promising ex-vivo antimalarial activity

Directory of Open Access Journals (Sweden)

Noraziah Mohamad Zin

2017-12-01

Full Text Available Objective: To determine ex vivo antimalarial activity and cytotoxicity of endophytic Streptomyces SUK 08 as well as the main core structure fractionated from its crude extract. Methods: The activities of SUK 08 crude extract were evaluated by using the Plasmodium lactate dehydrogenase assay and synchronization test against rodent malaria parasite Plasmodium berghei, instead of human malarial parasite Plasmodium falciparum. The cytotoxicity of the crude extract was determined by MTT assay. The crude extract was analyzed by thin-layer chromatography and gas chromatography–mass spectrophotometry. Results: The ethyl acetate crude extract showed very promising antimalarial activity with IC50 of 1.25 mg/mL. The synchronization tests showed that ethyl acetate extraction could inhibit all stages of the Plasmodium life cycle, but it was most effective at the Plasmodium ring stage. On the basis of a MTT assay on Chang Liver cells, ethyl acetate and ethanol demonstrated IC50 values of >1.0 mg/mL. The IC50 of parasitemia at 5% and 30% for this extract was lower than chloroquine. Thin-layer chromatography, with 1: 9 ratio of ethyl acetate: hexane, was used to isolate several distinct compounds. Based on gas chromatography–mass spectrophotometry analysis, three core structures were identified as cyclohexane, butyl propyl ester, and 2,3-heptanedione. Structurally, these compounds were similar to currently available antimalarial drugs. Conclusions: The results suggest that compounds isolated from Streptomyces SUK 08 are viable antimalarial drug candidates that require further investigations. Keywords: Butyl–propyl–ester, Cyclohexane, 2,3-Heptanedione, Endophyte, Streptomyces, Antimalarial

Diclofenac toxicity in human intestine ex vivo is not related to the formation of intestinal metabolites

NARCIS (Netherlands)

Niu, Xiaoyu; de Graaf, Inge A. M.; Langelaar-Makkinje, Miriam; Horvatovich, Peter; Groothuis, Geny M. M.

The use of diclofenac (DCF), a nonsteroidal anti-inflammatory drug, is associated with a high prevalence of gastrointestinal side effects. In vivo studies in rodents suggested that reactive metabolites of DCF produced by the liver or the intestine might be responsible for this toxicity. In the

A novel technique for impaction bone grafting in acetabular reconstruction of revision total hip arthroplasty using an ex vivo compaction device

International Nuclear Information System (INIS)

Akiyama, Haruhiko; Takemoto, Mitsuru; Morishima, Takkan

2011-01-01

Impaction bone grafting allows restoration of the acetabular bone stock in revision hip arthroplasty. The success of this technique depends largely on achieving adequate initial stability of the component. To obtain well-compacted, well-graded allograft aggregates, we developed an ex vivo compaction device to apply it in revision total hip arthroplasty on the acetabular side, and characterized mechanical properties and putative osteoconductivity of allograft aggregates. Morselized allograft bone chips were compacted ex vivo using the creep technique and subsequent impaction technique to form the bone aggregates. Impaction allograft reconstruction of the acetabulum using an ex vivo compaction device was performed on eight hips. The mechanical properties and three-dimensional micro-CT-based structural characteristics of the bone aggregates were investigated. In clinical practice, this technique offered good reproducibility in reconstructing the cavity and the segmental defects of the acetabulum, with no migration and no loosening of the component. In vitro analysis showed that the aggregates generated from 25 g fresh-frozen bone chips gained compression stiffness of 13.5-15.4 MPa under uniaxial consolidation strain. The recoil of the aggregates after compaction was 2.6-3.9%. The compression stiffness and the recoil did not differ significantly from those measured using a variety of proportions of large- and small-sized bone chips. Micro-CT-based structural analysis revealed average pore sizes of 268-299 μm and average throat diameter of pores in the bone aggregates of more than 100 μm. These sizes are desirable for osteoconduction, although large interconnected pores of more than 500 μm were detectable in association with the proportion of large-sized bone chips. Cement penetration into the aggregates was related to the proportion of large-sized bone chips. This study introduces the value of an ex vivo compaction device in bone graft compaction in clinical

Ex vivo expansion of bone marrow stromal cells by platelet-rich plasma: a promising strategy in maxillo-facial surgery.

Science.gov (United States)

Oliva, A; Passaro, I; Di Pasquale, R; Di Feo, A; Criscuolo, M; Zappia, V; Della Ragione, F; D'Amato, S; Annunziata, M; Guida, L

2005-01-01

The aim of our study is to evaluate in vitro the response of bone marrow stromal cells (BMSCs) to platelet-rich plasma (PRP), in order to clarify the potential role of their combined use in a preclinical phase preceding BMSCs transplantation for bone repair and regeneration procedures. The incubation of BMSCs with PRP promoted a remarkable, dose- and time- dependent, growth stimulation, that was paralleled to a strong increase in the quantity of type I collagen and to a significant decrease in the activity of the early osteoblastic differentiation marker, alkaline phosphatase (AP). Once PRP was removed and osteogenic inducers were added, AP returned to levels comparable to the control, while the late phenotypic markers, osteocalcin and matrix calcification, were enhanced to higher levels than in controls. Our data demonstrate that PRP induces a remarkable ex vivo enrichment of BMSCs maintaining their differentiative potential. Thus PRP represents a valid preclinical tool for obtaining an effective, rapid and safe ex vivo expansion of BMSCs prior to their clinical utilization in bone engineering.