14 research outputs found
Intimal and medial contributions to the hydraulic resistance of the arterial wall at different pressures: a combined computational and experimental study
The hydraulic resistances of the intima and media determine water flux and the advection of macromolecules into and across the arterial wall. Despite several experimental and computational studies, these transport processes and their dependence on transmural pressure remain incompletely understood. Here, we use a combination of experimental and computational methods to ascertain how the hydraulic permeability of the rat abdominal aorta depends on these two layers and how it is affected by structural rearrangement of the media under pressure. Ex vivo experiments determined the conductance of the whole wall, the thickness of the media and the geometry of medial smooth muscle cells (SMCs) and extracellular matrix (ECM). Numerical methods were used to compute water flux through the media. Intimal values were obtained by subtraction. A mechanism was identified that modulates pressure-induced changes in medial transport properties: compaction of the ECM leading to spatial reorganization of SMCs. This is summarized in an empirical constitutive law for permeability and volumetric strain. It led to the physiologically interesting observation that, as a consequence of the changes in medial microstructure, the relative contributions of the intima and media to the hydraulic resistance of the wall depend on the applied pressure; medial resistance dominated at pressures above approximately 93 mmHg in this vessel
Role of endothelial permeability hotspots and endothelial mitosis in determining age-related patterns of macromolecule uptake by the rabbit aortic wall near branch points
AbstractBackground and aimsTransport of macromolecules between plasma and the arterial wall plays a key role in atherogenesis. Scattered hotspots of elevated endothelial permeability to macromolecules occur in the aorta; a fraction of them are associated with dividing cells. Hotspots occur particularly frequently downstream of branch points, where lesions develop in young rabbits and children. However, the pattern of lesions varies with age, and can be explained by similar variation in the pattern of macromolecule uptake. We investigated whether patterns of hotspots and mitosis also change with age.MethodsEvans’ Blue dye-labeled albumin was injected intravenously into immature or mature rabbits and its subsequent distribution in the aortic wall around intercostal branch ostia examined by confocal microscopy and automated image analysis. Mitosis was detected by immunofluorescence after adding 5-bromo-2-deoxiuridine to drinking water.ResultsHotspots were most frequent downstream of branches in immature rabbits, but a novel distribution was observed in mature rabbits. Neither pattern was explained by mitosis. Hotspot uptake correlated spatially with the much greater non-hotspot uptake (p < 0.05), and the same pattern was seen when only the largest hotspots were considered.ConclusionsThe pattern of hotspots changes with age. The data are consistent with there being a continuum of local permeabilities rather than two distinct mechanisms. The distribution of the dye, which binds to elastin and collagen, was similar to that of non-binding tracers and to lesions apart from a paucity at the lateral margins of branches that can be explained by lower levels of fibrous proteins in those regions
Noradrenaline has opposing effects on the hydraulic conductance of arterial intima and media.
The uptake of circulating macromolecules by the arterial intima is thought to be a key step in atherogenesis. Such transport is dominantly advective, so elucidating the mechanisms of water transport is important. The relation between vasoactive agents and water transport in the arterial wall is incompletely understood. Here we applied our recently-developed combination of computational and experimental methods to investigate the effects of noradrenaline (NA) on hydraulic conductance of the wall (Lp), medial extracellular matrix volume fraction (Ď•(ECM)) and medial permeability (K1(1)) in the rat abdominal aorta. Experimentally, we found that physiological NA concentrations were sufficient to induce SMC contraction and produced significant decreases in Lp and increases in Ď•(ECM). Simulation results based on 3D confocal images of the extracellular volume showed a corresponding increase in K1(1), attributed to the opening of the ECM. Conversion of permeabilities to layer-specific resistances revealed that although the total wall resistance increased, medial resistance decreased, suggesting an increase in intimal resistance upon application of NA
Targeted Molecular Iron Oxide Contrast Agents for Imaging Atherosclerotic Plaque.
Overview: Cardiovascular disease remains a leading cause of death worldwide, with vulnerable plaque rupture the underlying cause of many heart attacks and strokes. Much research is focused on identifying an imaging biomarker to differentiate stable and vulnerable plaque. Magnetic Resonance Imaging (MRI) is a non-ionising and non-invasive imaging modality with excellent soft tissue contrast. However, MRI has relatively low sensitivity (micromolar) for contrast agent detection compared to nuclear imaging techniques. There is also an increasing emphasis on developing MRI probes that are not based on gadolinium chelates because of increasing concerns over associated systemic toxicity and deposits1. To address the sensitivity and safety concerns of gadolinium this project focused on the development of a high relaxivity probe based on superparamagnetic iron oxide nanoparticles for the imaging of atherosclerotic plaque with MRI. With development, this may facilitate differentiating stable and vulnerable plaque in vivo. Aim: To develop a range of MRI contrast agents based on superparamagnetic iron oxide nanoparticles (SPIONs), and test them in a murine model of advanced atherosclerosis. Methods: Nanoparticles of four core sizes were synthesised by thermal decomposition and coated with poly(maleicanhydride-alt-1-octadecene) (PMAO), poly(ethyleneimine) (PEI) or alendronate, then characterised for core size, hydrodynamic size, surface potential and relaxivity. On the basis of these results, one candidate was selected for further studies. In vivo studies using 10 nm PMAO-coated SPIONs were performed in ApoE -/- mice fed a western diet and instrumented with a perivascular cuff on the left carotid artery. Control ApoE -/- mice were fed a normal chow diet and were not instrumented. Mice were scanned on a 3T MR scanner (Philips Achieva) with the novel SPION contrast agent, and an elastin-targeted gadolinium agent that was shown previously to enable visualisation of plaque burden. Histological analysis was undertaken to confirm imaging findings through staining for macrophages, CX3CL1, elastin, tropoelastin, and iron. Results: The lead SPION agent consisted of a 10 nm iron oxide core with poly(maleicanhydride-alt-1-octadecene), (-36.21 mV, r2 18.806 mmol-1/s-1). The irregular faceting of the iron oxide core resulted in high relaxivity and the PMAO provided a foundation for further functionalisation on surface -COOH groups. The properties of the contrast agent, including the negative surface charge and hydrodynamic size, were designed to maximise circulation time and evade rapid clearance through the renal system or phagocytosis. In vitro testing showed that the SPION agent was non-toxic. In vivo results show that the novel contrast agent accumulates in similar vascular regions to a gadolinium-based contrast agent (Gd-ESMA) targeted to elastin, which accumulates in plaque. There was a significant difference in SPION signal between the instrumented and the contralateral non-instrumented vessels in diseased mice (p = 0.0411, student's t-test), and between the instrumented diseased vessel and control vessels (p = 0.0043, 0.0022, student's t-test). There was no significant difference between the uptake of either contrast agent between stable and vulnerable plaques (p = 0.3225, student's t-test). Histological verification was used to identify plaques, and Berlin Blue staining confirmed the presence of nanoparticle deposits within vulnerable plaques and co-localisation with macrophages. Conclusion: This work presents a new MRI contrast agent for atherosclerosis which uses an under-explored surface ligand, demonstrating promising properties for in vivo behaviour, is still in circulation 24 hours post-injection with limited liver uptake, and shows good accumulation in a murine plaque model
New developments in mechanotransduction: Cross talk of the Wnt, TGF-β and Notch signalling pathways in reaction to shear stress
Mechanotransduction, the ability of cells to detect and react to mechanical forces, is increasingly playing a critical role in a variety of physiological and pathophysiological processes. While the focus has previously been on the MAPK, NF-8B and ROS generating pathways, ancient embryological pathways have reached little attention. Recently, a surge of new studies have been published on these pathways and their role in mechanotransduction and this review paper aims to provide a concise overview on the latest studies and brings them in to a larger perspective. Special emphasis is on the non-canonical aspects of the Wnt, TGF-b and Notch pathways and their role in flow
Fabrication of multi-component spinal motion segment-like construct using mesenchymal stem cells and collagen
Concurrent Session 3Oral Presentatio
Engineering a Multicomponent Spinal Motion Segment-Like Construct from Mesenchymal Stem Cells
Conference theme: The Intervertebral Disc - from Degeneration to Therapeutic Motion PreservationThe abstract can be viewed at http://www.spineresearchforum.org/WFSR_2014_Thieme_AbstractBook_with_Cover.pdfOral PresentationIntroduction
The task of engineering the intervertebral disc is challenging
as the complex tissue needs to integrate with the host tissue
and performits function after the implantation. The vertebrae
connected to the endplates are essential to integrate with the
host vertebrae tissue which had been shown by Luk et al in
whole disc transplantation.1 Hence, engineering the complex
tissue needs to integrate the different components of the
vertebrae (VB), cartilaginous endplate (CEP), nucleus pulposus
(NP), and annulus fibrosus (AF); both biologically and mechanically.
In this study, the multiple component spinal
motion segments were fabricated by integrating these components.
The construct was then loaded in a bioreactor and
supplied with mechanical and biological stimulation. The
functional aspect of the fabricated endplate-like construct
was evaluated by a permeability test.
Materials and Methods
Rabbit mesenchymal stem cells (rMSCs) were encapsulated in
collagen and induced to differentiate toward osteogenic and
chondrogenic lineages before fabricating trilayered osteochondral
(OC) constructs as previously mentioned. To test
the nutritional function of the OC construct which acts as the
endplate, rabbit nucleus pulposus cells (rNPCs)-encapsulated
collagen microspheres were trapped in a sealed chamber
formed with the OC construct such that the nutrients have
to diffuse through the OC construct to reach the inside of the
chamber. Cell viability of the rNPCs was then evaluated. To
fabricate the multiple component construct, a rMSCs encapsulated
collagen-GAG precipitate was added in between two
OC construct and placed in between the shaft of the bioreactor.
Then a layer of rMSC encapsulated collagen was formed
around the construct to form the AF-like lamella. Torsional
loading was applied onto the construct to study its effect on cell alignment in the AF-like lamella. Finally, one to three
layers of AF-like lamellae were added to the spinal motion
segment construct and cultured in the bioreactor with complex
loading for 14 days. Histological, ultrastructural, and
mechanical evaluation was done on the construct.
Results
In the custom developed functionality test for nutrient transport,
the rNPCs in the chamber were viable at the end of the
culture showed that nutrientswere able to diffuse through the
OC construct. For the effect of torsional loading on cell
alignment in the AF-like lamella, alignment analysis showed
that the cells were aligned along a preferred axis under
torsional loading compared with control group without loading.
However, no collagen fibers alignment was found in this
study. The multiple component construct was fabricated with
each component similar to the spinal motion segment. The
different components of the construct were well integrated
throughout the culture and were shown by histology. Mean
torsional stiffness of the constructs significantly increased as
the number of rMSC encapsulated AF-like layer increased.
Conclusion
This study demonstrated the feasibility to engineer a spinal
motion segment-like tissue with collagen and MSC. The OC
constructs demonstrated its nutritional function and can be
used as a vertebra-endplate construct in this model. rMSC
encapsulated in collagen gel can be induced to re-orientate
and align in a certain direction by applying cyclic torsional
force on the tubular structure. This can be a tissue engineered
model to study the effects of various strategies in functional
remodeling and maturation of the intervertebral disc.
Disclosure of Interest
None declared
Reference
1. Luk KD, Ruan DK. Intervertebral disc transplantation: a
biological approach to motion preservation. Eur Spine J
2008;17(Suppl 4):504–51
Mechanosensitive pathways are regulated by mechanosensitive miRNA clusters in endothelial cells.
Shear stress is known to affect many processes in (patho-) physiology through a complex, multi-molecular mechanism, termed mechanotransduction. The sheer complexity of the process has raised questions how mechanotransduction is regulated. Here, we comprehensively evaluate the literature about the role of small non-coding miRNA in the regulation of mechanotransduction. Regulation of mRNA by miRNA is rather complex, depending not only on the concentration of mRNA to miRNA, but also on the amount of mRNA competing for a single mRNA. The only mechanism to counteract the latter factor is through overarching structures of miRNA. Indeed, two overarching structures are present miRNA families and miRNA clusters, and both will be discussed in details, regarding the latest literature and a previous conducted study focussed on mechanotransduction. Both the literature and our own data support a new hypothesis that miRNA-clusters predominantly regulate mechanotransduction, affecting 65% of signalling pathways. In conclusion, a new and important mode of regulation of mechanotransduction is proposed, based on miRNA clusters. This finding implicates new avenues for treatment of mechanotransduction and atherosclerosis
Bioengineering a multicomponent spinal motion segment construct – A 3D model for complex tissue engineering
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