scholarly journals Kinetic Cytokine Secretion Profile of LPS-Induced Inflammation in the Human Skin Organ Culture

Pharmaceutics ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 299 ◽  
Author(s):  
Raanan Gvirtz ◽  
Navit Ogen-Shtern ◽  
Guy Cohen
Keyword(s):  
Organ Culture ◽  
Human Skin ◽  
Comprehensive Evaluation ◽  
Ex Vivo ◽  
Cytokine Secretion ◽  
Peak Time ◽  
Local Skin ◽  
The Impact ◽  
Skin Organ Culture

Several in vitro models that mimic different aspects of local skin inflammation exist. The use of ex vivo human skin organ culture (HSOC) has been reported previously. However, comprehensive evaluation of the cytokine secretory capacity of the system and its kinetics has not been performed. Objective: the aim of the current study was to investigate the levels and secretion pattern of key cytokine from human skin tissue upon lipopolysaccharide (LPS) stimulation. HSOC maintained in an air–liquid interface was used. Epidermal and tissue viability was monitored by MTT and Lactate Dehydrogenase (LDH) activity assay, respectively. Cytokine levels were examined by ELISA and multiplex array. HSOCs were treated without or with three different LPS subtypes and the impact on IL-6 and IL-8 secretion was evaluated. The compounds enhanced the secreted levels of both cytokines. However, differences were observed in their efficacy and potency. Next, a kinetic multiplex analysis was performed on LPS-stimulated explants taken from three different donors to evaluate the cytokine secretion pattern during 0–72 h post-induction. The results revealed that the pro-inflammatory cytokines IL-6, IL-8, TNFα and IL-1β were up-regulated by LPS stimuli. IL-10, an anti-inflammatory cytokine, was also induced by LPS, but exhibited a different secretion pattern, peak time and maximal stimulation values. IL-1α and IL-15 showed donor-specific changes. Lastly, dexamethasone attenuated cytokine secretion in five independent repetitions, supporting the ability of the system to be used for drug screening. The collective results demonstrate that several cytokines can be used as valid inflammatory markers, regardless of changes in the secretion levels due to donor’s specific alterations.

scholarly journals 086 Human skin organ culture as a clinically relevant model for studying VEGF-dependent angiogenesis ex vivo

2019 ◽  
Vol 139 (9) ◽  
pp. S229
Author(s):  
A. Luengas-Martinez ◽  
J. Hardman ◽  
D. Rutkowski ◽  
R. Paus ◽  
H. Young
Keyword(s):  
Organ Culture ◽  
Human Skin ◽  
Ex Vivo ◽  
Relevant Model ◽  
Skin Organ Culture

scholarly journals Cell–cell coupling and DNA methylation abnormal phenotypes in the after-hours mice

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Federico Tinarelli ◽  
Elena Ivanova ◽  
Ilaria Colombi ◽  
Erica Barini ◽  
Edoardo Balzani ◽  
...  
Keyword(s):  
Gene Expression ◽  
Dna Methylation ◽  
Neuronal Networks ◽  
Molecular Mechanisms ◽  
Ex Vivo ◽  
Neuronal Cultures ◽  
Functional Impairments ◽  
After Hours ◽  
The Impact

Abstract Background DNA methylation has emerged as an important epigenetic regulator of brain processes, including circadian rhythms. However, how DNA methylation intervenes between environmental signals, such as light entrainment, and the transcriptional and translational molecular mechanisms of the cellular clock is currently unknown. Here, we studied the after-hours mice, which have a point mutation in the Fbxl3 gene and a lengthened circadian period. Methods In this study, we used a combination of in vivo, ex vivo and in vitro approaches. We measured retinal responses in Afh animals and we have run reduced representation bisulphite sequencing (RRBS), pyrosequencing and gene expression analysis in a variety of brain tissues ex vivo. In vitro, we used primary neuronal cultures combined to micro electrode array (MEA) technology and gene expression. Results We observed functional impairments in mutant neuronal networks, and a reduction in the retinal responses to light-dependent stimuli. We detected abnormalities in the expression of photoreceptive melanopsin (OPN4). Furthermore, we identified alterations in the DNA methylation pathways throughout the retinohypothalamic tract terminals and links between the transcription factor Rev-Erbα and Fbxl3. Conclusions The results of this study, primarily represent a contribution towards an understanding of electrophysiological and molecular phenotypic responses to external stimuli in the Afh model. Moreover, as DNA methylation has recently emerged as a new regulator of neuronal networks with important consequences for circadian behaviour, we discuss the impact of the Afh mutation on the epigenetic landscape of circadian biology.

Corticosteroids alter alveolar macrophage control of Lichtheimia corymbifera spores in an ex vivo mouse model

2020 ◽  
Author(s):  
Kévin Brunet ◽  
François Arrivé ◽  
Jean-Philippe Martellosio ◽  
Isabelle Lamarche ◽  
Sandrine Marchand ◽  
...  
Keyword(s):  
Mouse Model ◽  
Alveolar Macrophage ◽  
Oxidative Burst ◽  
Ex Vivo ◽  
Fungal Growth ◽  
Invasive Infection ◽  
Ex Vivo Model ◽  
Lichtheimia Corymbifera ◽  
The Impact

Abstract Alveolar macrophages (AM) are the first-line lung defense against Mucorales in pulmonary mucormycosis. Since corticosteroid use is a known risk factor for mucormycosis, the aim of this study was to describe the role of corticosteroids on AM capacities to control Lichtheimia corymbifera spore growth using a new ex vivo model. An in vivo mouse model was developed to determine the acetate cortisone dose able to trigger pulmonary invasive infection. Then, in the ex vivo model, male BALB/c mice were pretreated with the corticosteroid regimen triggering invasive infection, before AM collection through bronchoalveolar lavage. AMs from corticosteroid-treated mice and untreated control AMs were then exposed to L. corymbifera spores in vitro (ratio 1:5). AM control of fungal growth, adherence/phagocytosis, and oxidative burst were assessed using optical densities by spectrophotometer, flow cytometry, and 2', 7'-dichlorofluoresceine diacetate fluorescence, respectively. Cortisone acetate at 500 mg/kg, at D-3 and at D0, led to pulmonary invasive infection at D3. Co-incubated spores and AMs from corticosteroid-treated mice had significantly higher absorbance (fungal growth) than co-incubated spores and control AMs, at 24 h (P = .025), 36 h (P = .004), and 48 h (P = .001). Colocalization of spores with AMs from corticosteroid-treated mice was significantly lower than for control AMs (7.6 ± 1.9% vs 22.3 ± 5.8%; P = .003), reflecting spore adherence and phagocytosis inhibition. Finally, oxidative burst was significantly increased when control AMs were incubated with spores (P = 0.029), while corticosteroids hampered oxidative burst from treated AMs (P = 0.321). Corticosteroids enhanced fungal growth of L. corymbifera through AM phagocytosis inhibition and burst oxidative decrease in our ex vivo model. Lay Summary The aim of this study was to describe the impact of corticosteroids on alveolar macrophage (AM) capacities to control Mucorales growth in a new murine ex vivo model. Corticosteroids enhanced fungal growth of L. corymbifera through AM phagocytosis inhibition and burst oxidative decrease.

Bench-Top Validation Tests for Tissue-Engineered Arteries

2000 ◽  
Author(s):  
Shawn Chin Quee ◽  
Hai-Chao Han ◽  
David N. Ku
Keyword(s):  
Organ Culture ◽  
Culture System ◽  
Comprehensive Evaluation ◽  
Ex Vivo ◽  
Vascular Grafts ◽  
Living System ◽  
Organ Culture System ◽  
Flow Pressure ◽  
Validation Tests

Abstract Standard tests are needed for evaluating and comparing the mechanical and biological functions of tissue engineered arteries and other vascular grafts. We propose an ex vivo organ culture system as a living system for testing tissue-engineered vascular grafts. This bench-top organ culture system mimics the physiological environment of arteries including the flow, pressure, and the axial stretch. Arterial mechanical properties and physiologic functions including compliance, burst pressure, and contractile functions can be assessed before an expensive long-term animal test is initiated. Test results of natural arteries indicate that organ culture is a valid model for comprehensive evaluation of tissue-engineered vascular grafts.

scholarly journals Enhanced Transdermal Delivery of Pranoprofen Using a Thermo-Reversible Hydrogel Loaded with Lipid Nanocarriers for the Treatment of Local Inflammation

2021 ◽  
Vol 15 (1) ◽  
pp. 22
Author(s):  
María Rincón ◽  
Marcelle Silva-Abreu ◽  
Lupe Carolina Espinoza ◽  
Lilian Sosa ◽  
Ana Cristina Calpena ◽  
...  
Keyword(s):  
Human Skin ◽  
Topical Application ◽  
Topical Treatment ◽  
Transdermal Delivery ◽  
Ex Vivo ◽  
Skin Permeation ◽  
In Vitro Release ◽  
Heterogeneous Structure ◽  
Clinical Environment ◽  
Local Skin

A biocompatible topical thermo-reversible hydrogel containing Pranoprofen (PF)-loaded nanostructured lipid carriers (NLCs) was studied as an innovative strategy for the topical treatment of skin inflammatory diseases. The PF-NLCs-F127 hydrogel was characterized physiochemically and short-time stability tests were carried out over 60 days. In vitro release and ex vivo human skin permeation studies were carried out in Franz diffusion cells. In addition, a cytotoxicity assay was studied using the HaCat cell line and in vivo tolerance study was performed in humans by evaluating the biomechanical properties. The anti-inflammatory effect of the PF-NLCs-F127 was evaluated in adult male Sprague Daw-ley® rats using a model of inflammation induced by the topical application of xylol for 1 h. The developed PF-NLCs-F127 exhibited a heterogeneous structure with spherical PF-NLCs in the hydrogel. Furthermore, a thermo-reversible behaviour was determined with a gelling temperature of 32.5 °C, being close to human cutaneous temperature and thus favouring the retention of PF. Furthermore, in the ex vivo study, the amount of PF retained and detected in human skin was high and no systemic effects were observed. The hydrogel was found to be non-cytotoxic, showing cell viability of around 95%. The PF-NLCs-F127 is shown to be well tolerated and no signs of irritancy or alterations of the skin’s biophysical properties were detected. The topical application of PF-NLCs-F127 hydrogel was shown to be efficient in an inflammatory animal model, preventing the loss of stratum corneum and reducing the presence of leukocyte infiltration. The results from this study confirm that the developed hydrogel is a suitable drug delivery carrier for the transdermal delivery of PF, improving its dermal retention, opening the possibility of using it as a promising candidate and safer alternative to topical treatment for local skin inflammation and indicating that it could be useful in the clinical environment.

scholarly journals Emulsion of Chloramphenicol: an Overwhelming Approach for Ocular Delivery

Folia Medica ◽  
2017 ◽  
Vol 59 (1) ◽  
pp. 23-30 ◽  
Author(s):  
Kalpesh C. Ashara ◽  
Ketan V. Shah
Keyword(s):  
Ternary Phase Diagram ◽  
Ex Vivo ◽  
Sterility Testing ◽  
Ocular Irritation ◽  
Peg 400 ◽  
Significant Difference ◽  
Oil Mixtures ◽  
Student’S T ◽  
The Impact

Abstract Background: Ophthalmic formulations of chloramphenicol have poor bioavailability of chloramphenicol in the ocular cavity. Aim: The present study aimed at exploring the impact of different oil mixtures in the form of emulsion on the permeability of chloramphenicol after ocular application. Materials and methods: Selection of oil mixture and ratio of the components was made by an equilibrium solubility method. An emulsifier was chosen according to its emulsification properties. A constrained simplex centroid design was used for the assessment of the emulsion development. Emulsions were evaluated for physicochemical properties; zone of inhibition, in-vitro diffusion and ex-vivo local accumulation of chloramphenicol. Validation of the design using check-point batch and reduced polynomial equations were also developed. Optimization of the emulsion was developed by software Design® expert 6.0.8. Assessment of the osmolarity, ocular irritation, sterility testing and isotonicity of optimized batch were also made. Results: Parker Neem®, olive and peppermint oils were selected as an oil phase in the ratio 63.64:20.2:16.16. PEG-400 was selected as an emulsifier according to a pseudo-ternary phase diagram. Constrained simplex-centroid design was applied in the range of 25-39% water, 55-69% PEG-400, 5-19% optimized oil mixture, and 1% chloramphenicol. Unpaired Student’s t-test showed for in-vitro and ex-vivo studies that there was a significant difference between the optimized batch of emulsion and Chloramphenicol eye caps (a commercial product) according to both were equally safe. Conclusion: The optimized batch of an emulsion of chloramphenicol was found to be as safe as and more effective than Chloramphenicol eye caps.

scholarly journals Neuroinflammation in Aged Brain: Impact of the Oral Administration of Ellagic Acid Microdispersion

2020 ◽  
Vol 21 (10) ◽  
pp. 3631 ◽  
Author(s):  
Raffaella Boggia ◽  
Federica Turrini ◽  
Alessandra Roggeri ◽  
Guendalina Olivero ◽  
Francesca Cisani ◽  
...  
Keyword(s):  
Oral Administration ◽  
Ellagic Acid ◽  
Ex Vivo ◽  
Behavioral Skills ◽  
Aged Mice ◽  
Age Related ◽  
The Central Nervous System ◽  
The Impact ◽  
Old Mice

The immune system and the central nervous system message each other to preserving central homeostasis. Both systems undergo changes during aging that determine central age-related defects. Ellagic acid (EA) is a natural product which is beneficial in both peripheral and central diseases, including aging. We analyzed the impact of the oral administration of a new oral ellagic acid micro-dispersion (EAm), that largely increased the EA solubility, in young and old mice. Oral EAm did not modify animal weight and behavioral skills in young and old mice, but significantly recovered changes in “ex-vivo, in vitro” parameters in old animals. Cortical noradrenaline exocytosis decreased in aged mice. EAm administration did not modify noradrenaline overflow in young animals, but recovered it in old mice. Furthermore, GFAP staining was increased in the cortex of aged mice, while IBA-1 and CD45 immunopositivities were unchanged when compared to young ones. EAm treatment significantly reduced CD45 signal in both young and old cortical lysates; it diminished GFAP immunopositivity in young mice, but failed to affect IBA-1 expression in both young and old animals. Finally, EAm treatment significantly reduced IL1beta expression in old mice. These results suggest that EAm is beneficial to aging and represents a nutraceutical ingredient for elders.

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