A new implantable tool for repeated assessment of supraventricular electrophysiology and atrial fibrillation susceptibility in freely moving rats

2020 ◽  
Author(s):  
Michael Murninkas ◽  
Roni Gillis ◽  
Danielle I Lee ◽  
Sigal Elyagon ◽  
Nikhil Suresh Bhandarkar ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Ex Vivo ◽  
Adult Rats ◽  
Electrophysiological Properties ◽  
Multiple Risk Factors ◽  
Electrophysiological Studies ◽  
Cycle Lengths ◽  
Major Factors

The complex pathophysiology of atrial fibrillation (AF) is governed by multiple risk factors in ways that are still elusive. Basic electrophysiological properties including atrial effective refractory period (AERP) and conduction velocity are major factors determining the susceptibility of the atrial myocardium to AF. Although there is a great need for affordable animal models in this field of research, in-vivo rodent studies are limited by technical challenges. Recently, we introduced an implantable system for long-term assessment of AF susceptibility in ambulatory rats. However, technical considerations did not allow us to perform concomitant supraventricular electrophysiology measurements. Here, we designed a novel quadripolar-electrode specifically adapted for comprehensive atrial studies in ambulatory rats. Electrodes were fabricated from medical-grade silicone, four platinum-iridium poles and stainless steel fixating pins. Initial quality validation was performed ex-vivo, followed by implantation in adult rats and repeated electrophysiological studies 1, 4 and 8 weeks post implantation. Capture threshold was stable. Baseline AERP values (38.1±2.3 and 39.5±2.0 using 70ms and 120ms S1-S1 cycle lengths, respectively) confirmed the expected absence of rate-adaptation in the unanesthetized state and validated our prediction that markedly higher values reported under anesthesia are non-physiological. Evaluation of AF substrate in parallel with electrophysiological parameters validated our recent finding of a gradual increase in AF susceptibility over-time and demonstrated that this phenomenon is associated with an electrical remodeling process characterized by AERP shortening. Our findings indicate that the miniature quadripolar-electrode is a potent new tool, which opens a window of opportunities for better utilization of rats in AF research.

Strategies to Protect Hematopoietic Stem Cells from Culture-Induced Stress Conditions

2020 ◽  
Vol 15 ◽  
Author(s):  
Fatima Aerts-Kaya
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Ex Vivo ◽  
Stress Conditions ◽  
Stress Factors ◽  
Hematopoietic Stem ◽  
Induced Stress

: In contrast to their almost unlimited potential for expansion in vivo and despite years of dedicated research and optimization of expansion protocols, the expansion of Hematopoietic Stem Cells (HSCs) in vitro remains remarkably limited. Increased understanding of the mechanisms that are involved in maintenance, expansion and differentiation of HSCs will enable the development of better protocols for expansion of HSCs. This will allow procurement of HSCs with long-term engraftment potential and a better understanding of the effects of the external influences in and on the hematopoietic niche that may affect HSC function. During collection and culture of HSCs, the cells are exposed to suboptimal conditions that may induce different levels of stress and ultimately affect their self-renewal, differentiation and long-term engraftment potential. Some of these stress factors include normoxia, oxidative stress, extra-physiologic oxygen shock/stress (EPHOSS), endoplasmic reticulum (ER) stress, replicative stress, and stress related to DNA damage. Coping with these stress factors may help reduce the negative effects of cell culture on HSC potential, provide a better understanding of the true impact of certain treatments in the absence of confounding stress factors. This may facilitate the development of better ex vivo expansion protocols of HSCs with long-term engraftment potential without induction of stem cell exhaustion by cellular senescence or loss of cell viability. This review summarizes some of available strategies that may be used to protect HSCs from culture-induced stress conditions.

scholarly journals Aη-α and Aη-β peptides impair LTP ex vivo within the low nanomolar range and impact neuronal activity in vivo

2021 ◽  
Vol 13 (1) ◽  
Author(s):  
Maria Mensch ◽  
Jade Dunot ◽  
Sandy M. Yishan ◽  
Samuel S. Harris ◽  
Aline Blistein ◽  
...  
Keyword(s):  
Neuronal Activity ◽  
Ex Vivo ◽  
Long Term Potentiation ◽  
Network Activity ◽  
Strongly Correlated ◽  
App Processing ◽  
Term Potentiation ◽  
Hippocampal Network

Abstract Background Amyloid precursor protein (APP) processing is central to Alzheimer’s disease (AD) etiology. As early cognitive alterations in AD are strongly correlated to abnormal information processing due to increasing synaptic impairment, it is crucial to characterize how peptides generated through APP cleavage modulate synapse function. We previously described a novel APP processing pathway producing η-secretase-derived peptides (Aη) and revealed that Aη–α, the longest form of Aη produced by η-secretase and α-secretase cleavage, impaired hippocampal long-term potentiation (LTP) ex vivo and neuronal activity in vivo. Methods With the intention of going beyond this initial observation, we performed a comprehensive analysis to further characterize the effects of both Aη-α and the shorter Aη-β peptide on hippocampus function using ex vivo field electrophysiology, in vivo multiphoton calcium imaging, and in vivo electrophysiology. Results We demonstrate that both synthetic peptides acutely impair LTP at low nanomolar concentrations ex vivo and reveal the N-terminus to be a primary site of activity. We further show that Aη-β, like Aη–α, inhibits neuronal activity in vivo and provide confirmation of LTP impairment by Aη–α in vivo. Conclusions These results provide novel insights into the functional role of the recently discovered η-secretase-derived products and suggest that Aη peptides represent important, pathophysiologically relevant, modulators of hippocampal network activity, with profound implications for APP-targeting therapeutic strategies in AD.

scholarly journals Assessment of Electrospun Pellethane-Based Scaffolds for Vascular Tissue Engineering

Materials ◽  
2021 ◽  
Vol 14 (13) ◽  
pp. 3678
Author(s):  
Vera Chernonosova ◽  
Alexandr Gostev ◽  
Ivan Murashov ◽  
Boris Chelobanov ◽  
Andrey Karpenko ◽  
...  
Keyword(s):  
Vascular Tissue ◽  
Ex Vivo ◽  
Vascular Grafts ◽  
Wistar Rat ◽  
Graft Occlusion ◽  
Suitable Candidate ◽  
Large Animals ◽  
Cell Adhesion And Proliferation

We examined the physicochemical properties and the biocompatibility and hemocompatibility of electrospun 3D matrices produced using polyurethane Pellethane 2363-80A (Pel-80A) blends Pel-80A with gelatin or/and bivalirudin. Two layers of vascular grafts of 1.8 mm in diameter were manufactured and studied for hemocompatibility ex vivo and functioning in the infrarenal position of Wistar rat abdominal aorta in vivo (n = 18). Expanded polytetrafluoroethylene (ePTFE) vascular grafts of similar diameter were implanted as a control (n = 18). Scaffolds produced from Pel-80A with Gel showed high stiffness with a long proportional limit and limited influence of wetting on mechanical characteristics. The electrospun matrices with gelatin have moderate capacity to support cell adhesion and proliferation (~30–47%), whereas vascular grafts with bivalirudin in the inner layer have good hemocompatibility ex vivo. The introduction of bivalirudin into grafts inhibited platelet adhesion and does not lead to a change hemolysis and D-dimers concentration. Study in vivo indicates the advantages of Pel-80A grafts over ePTFE in terms of graft occlusion, calcification level, and blood velocity after 6 months of implantation. The thickness of neointima in Pel-80A–based grafts stabilizes after three months (41.84 ± 20.21 µm) and does not increase until six months, demonstrating potential for long-term functioning without stenosis and as a suitable candidate for subsequent preclinical studies in large animals.

Abstract 13383: The Vascular Endothelial Barrier: A Novel Therapeutic Target for Preventing Atrial Fibrillation

Circulation ◽  
2020 ◽  
Vol 142 (Suppl_3) ◽  
Author(s):  
Louisa Mezache ◽  
Heather Struckman ◽  
Anna Phillips ◽  
Stephen Baine ◽  
Amara Greer-short ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Ex Vivo ◽  
Vascular Dysfunction ◽  
Super Resolution ◽  
Vascular Endothelial ◽  
Vascular Leak ◽  
Barrier Breakdown ◽  
Endothelial Growth Factor ◽  
Intercalated Disk

Atrial fibrillation (AF), the most common arrhythmia, is associated with inflammation and vascular dysfunction. AF patients have elevated levels of vascular endothelial growth factor (VEGF; 90-580 pg/ml), which promotes vascular leak and edema. We have previously identified edema-induced disruption of sodium channel (Na V 1.5) -rich intercalated disk (ID) nanodomains as a novel arrhythmia mechanism. We hypothesized that (i) elevated VEGF levels promote AF by disrupting ID nanodomains, and slowing atrial conduction, and (ii) protection of the vascular barrier can prevent these arrhythmias. Clinically-relevant VEGF levels (500 pg/ml, 60 minutes) increased FITC-dextran extravasation (99.3% vs. 24.3% in vehicle controls) in WT mouse hearts, consistent with increased vascular leak. Electron microscopy revealed ID nanodomain swelling, near both gap junctions (perinexi; 64±9nm vs 17±1nm) and mechanical junctions (63±4nm vs 27±2nm) in VEGF-treated hearts relative to controls. Super-resolution STORM microscopy revealed Na V 1.5 enrichment at perinexi (9±2 fold) and N-cadherin-rich sites (7±1 fold) relative to non-junctional ID sites in control hearts. VEGF reduced Na V 1.5 enrichment at both sites (6±1 and 4±1 fold, respectively), consistent with Na V 1.5 translocation from ID nanodomains. Atrial conduction, assessed by optical mapping, was slowed by VEGF (10±0.4 cm/s vs 21.3±1.3 cm/s at baseline). VEGF increased atrial arrhythmia burden both ex vivo (80% vs 0% in vehicle controls) and in vivo (70% vs 20% in vehicle controls). Next, we tested two strategies shown to prevent vascular barrier breakdown. Blocking connexin43 hemichannels (αCT11 peptide) decreased both incidence (40%) and duration (1.45±3.42s) of VEGF-induced arrhythmias. Likewise, blocking pannexin1 channels (Panx1-IL2 peptide) shortened VEGF-induced arrhythmias (2.48±0.83s). Mefloquine and spironolactone, which are small molecules that respectively inhibit Cx43 hemichannels and pannexin channels, were also found to effectively prevent VEGF-induced atrial arrhythmias. These results highlight VEGF-induced vascular leak as a novel mechanism for AF, and suggest vascular barrier protection as an anti-arrhythmic strategy.

scholarly journals Isolated superfused rats atrial model for the investigation of atrial fibrillation mechanisms and treatment

2021 ◽  
Vol 42 (Supplement_1) ◽  
Author(s):  
S Glatstein ◽  
M Ghiringhelli ◽  
L Maizels ◽  
E Heller ◽  
E Maor ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
High Resolution ◽  
Sinus Rhythm ◽  
Optical Mapping ◽  
Ex Vivo ◽  
Anatomical Landmarks ◽  
Ex Vivo Model ◽  
Electrophysiological Properties ◽  
Custom Made ◽  
Isolated Atria

Abstract Background One of the major barriers to an improved mechanistic understanding of atrial fibrillation (AF), and thus in the pipeline of drug development, has been a lack of appropriate tissue models, especially in small animals. Aim We propose an advanced anatomical ex-vivo model based on rat atria for acute assessment of AF susceptibility. This novel model could yield a better understanding of arrhythmia mechanisms as well as the development of potential therapeutic strategies for the prevention or termination of atrial arrhythmias. Methods Wistar rats atria (N=25) were isolated, flattened and pinned to a custom-made silicon plate. Atria were superfused with an oxygenized Tyrode's solution. Tissues were then loaded with a voltage-sensitive dye and mapped using a high-resolution optical mapping system. AF was induced with 1uM carbamylcholine (N=23) coupled with pacing maneuvers and treated with 30uM Vernakalant (N=10) or 10uM Flecainide (N=10). Finally, the feasibility of a new ablation technique (electroporation) was evaluated. Results Optical mapping results suggested that the superfusion procedure led to a fast atrial recovery. Sinus activity was conserved for all atria for a long period. All the anatomical landmarks were clearly visualized. The acquired optical signals were analyzed during sinus rhythm and pacing, which allowed the creation of detailed activation maps and measurements of action potential duration (APD) and conduction velocity (CV) at different pacing rates. The resulting APD restitution curves revealed electrical excitation at high pacing rates (cycle length between 50ms and 300ms) with a relatively flattened curve. AF was successfully induced and optically mapping confirmed the presence of reentrant activity. AF was successfully treated using Vernacalant and Flecainide. Finally, we demonstrated the feasibility of a new ablation approach (electroporation) for creation of a continuous linear lesion serving as a functional block. Conclusion The isolated superfused atria model, coupled with voltage-sensitive dyes, can be utilized for long-term high-resolution functional imaging of the atria during sinus rhythm, pacing and arrhythmogenic activity. This allows the study of the atrial electrophysiological properties, the mechanisms involved in AF initiation, perpetuation, and termination as well as the study of drug and new ablation modalities. FUNDunding Acknowledgement Type of funding sources: Public grant(s) – EU funding. Main funding source(s): European Research Council (ERC) Spontaneous activation of isolated atria

BioMEMS Technologies for Regenerative Medicine

2008 ◽  
Vol 1139 ◽  
Author(s):  
Jeffrey T. Borenstein
Keyword(s):  
Drug Delivery ◽  
Regenerative Medicine ◽  
Drug Delivery Systems ◽  
Ex Vivo ◽  
Delivery Systems ◽  
Organ Shortage ◽  
Engineered Tissues ◽  
Drug Concentrations

AbstractThe emergence of BioMEMS fabrication technologies such as soft lithography, micromolding and assembly of 3D structures, and biodegradable microfluidics, are already making significant contributions to the field of regenerative medicine. Over the past decade, BioMEMS have evolved from early silicon laboratory devices to polymer-based structures and even biodegradable constructs suitable for a range of ex vivo and in vivo applications. These systems are still in the early stages of development, but the long-term potential of the technology promises to enable breakthroughs in health care challenges ranging from the systemic toxicity of drugs to the organ shortage. Ex vivo systems for organ assist applications are emerging for the liver, kidney and lung, and the precision and scalability of BioMEMS fabrication techniques offer the promise of dramatic improvements in device performance and patient outcomes.Ultimately, the greatest benefit from BioMEMS technologies will be realized in applications for implantable devices and systems. Principal advantages include the extreme levels of achievable miniaturization, integration of multiple functions such as delivery, sensing and closed loop control, and the ability of precision microscale and nanoscale features to reproduce the cellular microenvironment to sustain long-term functionality of engineered tissues. Drug delivery systems based on BioMEMS technologies are enabling local, programmable control over drug concentrations and pharmacokinetics for a broad spectrum of conditions and target organs. BioMEMS fabrication methods are also being applied to the development of engineered tissues for applications such as wound healing, microvascular networks and bioartificial organs. Here we review recent progress in BioMEMS-based drug delivery systems, engineered tissue constructs and organ assist devices for a range of ex vivo and in vivo applications in regenerative medicine.

scholarly journals Acute (24 h) activation of peroxisome proliferator-activated receptor-alpha (PPARalpha) reverses high-fat feeding-induced insulin hypersecretion in vivo and in perifused pancreatic islets

2003 ◽  
Vol 177 (2) ◽  
pp. 197-205 ◽  
Author(s):  
MJ Holness ◽  
ND Smith ◽  
GK Greenwood ◽  
MC Sugden
Keyword(s):  
Insulin Secretion ◽  
Ex Vivo ◽  
Peroxisome Proliferator ◽  
High Fat ◽  
Intravenous Glucose ◽  
Perifused Islets ◽  
Glucose Stimulated Insulin Secretion ◽  
Fat Feeding

Abnormal depletion or accumulation of islet lipid may be important for the development of pancreatic beta cell failure. Long-term lipid sensing by beta cells may be co-ordinated via peroxisome proliferator-activated receptors (PPARs). We investigated whether PPARalpha activation in vivo for 24 h affects basal and glucose-stimulated insulin secretion in vivo after intravenous glucose administration and ex vivo in isolated perifused islets. Insulin secretion after intravenous glucose challenge was greatly increased by high-fat feeding (4 weeks) but glucose tolerance was minimally perturbed, demonstrating insulin hypersecretion compensated for insulin resistance. The effect of high-fat feeding to enhance glucose-stimulated insulin secretion was retained in perifused islets demonstrating a stable, long-term effect of high-fat feeding to potentiate islet glucose stimulus-secretion coupling. Treatment of high-fat-fed rats with WY14,643 for 24 h reversed insulin hypersecretion in vivo without impairing glucose tolerance, suggesting improved insulin action, and ex vivo in perfused islets. PPARalpha activation only affected hypersecretion of insulin since glucose-stimulated insulin secretion was unaffected by WY14,643 treatment in vivo in control rats or in perifused islets from control rats. Our data demonstrate that activation of PPARalpha for 24 h can oppose insulin hypersecretion elicited by high-fat feeding via stable long-term effects exerted on islet function. PPARalpha could, therefore, participate in ameliorating abnormal glucose homeostasis and hyperinsulinaemia in dietary insulin resistance via modulation of islet function, extending the established requirement for PPARalpha for normal islet lipid homeostasis.

Designed multifunctional polymeric nanomedicines: long-term biodistribution and tumour accumulation of aptamer-targeted nanomaterials

2018 ◽  
Vol 54 (82) ◽  
pp. 11538-11541 ◽  
Author(s):  
N. L. Fletcher ◽  
Z. H. Houston ◽  
J. D. Simpson ◽  
R. N. Veedu ◽  
K. J. Thurecht
Keyword(s):  
Polyethylene Glycol ◽  
Pet Imaging ◽  
Hyperbranched Polymer ◽  
Ex Vivo

We report a novel multifunctional hyperbranched polymer based on polyethylene glycol (PEG) as a nanomedicine platform that facilitates longitudinal and quantitative 89Zr-PET imaging, enhancing knowledge of nanomaterial biodistribution and pharmacokinetics/pharmacodynamics both in vivo and ex vivo.

Anti-nociceptive and anti-inflammatory activity of synthesized novel benzoxazole derivatives

2021 ◽  
Vol 20 ◽  
Author(s):  
Mansi L. Patil ◽  
Swati S. Gaikwad ◽  
Naresh J. Gaikwad
Keyword(s):  
Cytotoxic Activity ◽  
Research Study ◽  
Ex Vivo ◽  
Immunological Response ◽  
Inflammatory Activity ◽  
Inflammatory Drugs ◽  
Anti Inflammatory ◽  
Inflammatory Effect

Introduction: Pain is an immunological response to any infection or inflammation and long term use of pain management therapy includes use of Nonsteroidal anti-inflammatory drugs which is associated with occurrence of toxicity as well as gastrointestinal bleeding. Therefore, the investigation of new analgesic and anti-inflammatory agents remains a major challenge. Aims: The objective of this research study is to undergo the pharmacological evaluation of newly synthesized benzoxazole derivatives. These novel derivatives were evaluated for anti-nociceptive, anti-inflammatory and cytotoxic activity using various in-vivo and ex-vivo methods. Methods: The study was carried out using swiss mice (adult male) weighing between 20gm to 30gm and were divided into groups containing (n=6) six animals in each group for treatment. The anti-nociceptive activity was performed by using 0.1ml of 0.6% v/v acetic acid as nociception inducer and evaluated by the diminished number of abdominal writhes. The anti-inflammatory activity was done using 0.1 ml of 2% w/v Carrageenan induced paw edema method was observed which was evaluated by calculating the percent maximum possible effect. Histopathological evaluation and cytotoxic activity of the compounds was carried out. Results: The results of this research study revealed that synthesized derivatives (a, b, c, d and e) showed promising anti-nociceptive and anti-inflammatory effect along significantly higher cytotoxic activity in MCF-7 cell lines. Conclusion: It can be concluded that synthesized derivatives (a, b, c, d and e) have potential anti-nociceptive and anti-inflammatory effect along with cytotoxic activity and certain modification in structure may result in potent activity.

Ex Vivo Expansion of Genetically Marked Rhesus Peripheral Blood Progenitor Cells Results in Diminished Long-Term Repopulating Ability

Blood ◽  
1998 ◽  
Vol 92 (4) ◽  
pp. 1131-1141 ◽  
Author(s):  
J.F. Tisdale ◽  
Y. Hanazono ◽  
S.E. Sellers ◽  
B.A. Agricola ◽  
M.E. Metzger ◽  
...  
Keyword(s):  
Peripheral Blood ◽  
Ex Vivo ◽  
Retroviral Vectors ◽  
Ex Vivo Expansion ◽  
Us Government ◽  
Government Work ◽  
Stromal Support ◽  
Short And Long Term

Abstract The possibility of primitive hematopoietic cell ex vivo expansion is of interest for both gene therapy and transplantation applications. The engraftment of autologous rhesus peripheral blood (PB) progenitors expanded 10 to 14 days were tracked in vivo using genetic marking. Stem cell factor (SCF)/granulocyte colony-stimulating factor (G-CSF)–mobilized and CD34-enriched PB cells were divided into two equal aliquots and transduced with one of two retroviral vectors carrying the neomycin-resistance gene (neo) for 4 days in the presence of interleukin-3 (IL-3), IL-6, and SCF in the first 5 animals, IL-3/IL-6/SCF/Flt-3 ligand (FLT) in 2 subsequent animals, or IL-3/IL-6/SCF/FLT plus an autologous stromal monolayer (STR) in the final 2. At the end of transduction period, one aliquot (nonexpanded) from each animal was frozen, whereas the other was expanded under the same conditions but without vector for a total of 14 days before freezing. After total body irradiation, both the nonexpanded and expanded transduced cells were reinfused. Despite 5- to 13-fold higher cell and colony-forming unit (CFU) doses from the expanded fraction of marked cells, there was greater short- and long-term marking from the nonexpanded cells in all animals. In animals receiving cells transduced and expanded in the presence of IL-3/IL-6/SCF/FLT, engraftment by the marked expanded cells was further diminished. This discrepancy was even more pronounced in the animals who received cells transduced and expanded in the presence of FLT and autologous stroma, with no marking detectable from the expanded cells. Despite lack of evidence for expansion of engrafting cells, we found that the addition of FLT and especially STR during the initial brief transduction period increased engraftment with marked cells into a clinically relevant range. Levels of marked progeny cells originating from the nonexpanded aliqouts were significantly higher than that seen in previous 4 animals receiving cells transduced in the presence of IL-3/IL-6/SCF, with levels of 10% to 20% confirmed by Southern blotting from the nonexpanded IL-3/IL-6/SCF/FLT/STR graft compared with 0.01% in the original IL-3/IL-6/SCF cohort. These results suggest that, although expansion of PB progenitors is feasible ex vivo, their contribution towards both short- and long-term engraftment is markedly impaired. However, a brief transduction in the presence of specific cytokines and stromal support allows engraftment with an encouraging number of retrovirally modified cells. This is a US government work. There are no restrictions on its use.

Sign in / Sign up

Export Citation Format

Share Document