Natriuretic peptide receptors regulate cytoprotective effects in a human ex vivo 3D/bioreactor model

© 2013 Peake et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited

Cx43 regulates mechanotransduction mechanisms in human preterm amniotic membrane defects.

OBJECTIVE: The effects of mechanical stimulation in preterm amniotic membrane (AM) defects were explored. METHODS: Preterm AM was collected from women undergoing planned preterm caesarean section (CS) due to fetal growth restriction or emergency CS after spontaneous preterm prelabour rupture of the membranes (sPPROM). AM explants near the cervix or placenta were subjected to trauma and/or mechanical stimulation with the Cx43 antisense. Markers for nuclear morphology (DAPI), myofibroblasts (αSMA), migration (Cx43), inflammation (PGE2 ) and repair (collagen, elastin and transforming growth factor β [TGFβ1 ]) were examined by confocal microscopy, second harmonic generation, qPCR and biochemical assays. RESULTS: In preterm AM defects, myofibroblast nuclei were highly deformed and contractile and expressed αSMA and Cx43. Mechanical stimulation increased collagen fibre polarisation and the effects on matrix markers were dependent on tissue region, disease state, gestational age and the number of fetuses. PGE2 levels were broadly similar but reduced after co-treatment with Cx43 antisense in late sPPROM AM defects. TGFβ1 and Cx43 gene expression were significantly increased after trauma and mechanical stimulation but this response dependent on gestational age. CONCLUSION: Mechanical stimulation affects Cx43 signalling and cell/collagen mechanics in preterm AM defects. Establishing how Cx43 regulates mechanosignalling could be an approach to repair tissue integrity after trauma

Multicomponent Hydrogels for the Formation of Vascularized Bone-like Constructs In Vitro.

The native extracellular matrix (ECM) is a complex gel-like system with a broad range of structural features and biomolecular signals. Hydrogel platforms that can recapitulate the complexity and signaling properties of this ECM would have enormous impact in fields ranging from tissue engineering to drug discovery. Here, we report on the design, synthesis, and proof-of-concept validation of a microporous and nanofibrous hydrogel exhibiting multiple bioactive epitopes designed to recreate key features of the bone ECM. The material platform integrates self-assembly with orthogonal enzymatic cross-linking to create a supramolecular environment comprising hyaluronic acid modified with tyramine (HA-Tyr) and peptides amphiphiles (PAs) designed to promote cell adhesion (RGDS-PA), osteogenesis (Osteo-PA), and angiogenesis (Angio-PA). Through individual and co-cultures of human adipose derived mesenchymal stem cells (hAMSCs) and human umbilical vascular endothelial cells (HUVECs), we confirmed the capacity of the HA-Tyr/RGDS-PA/Osteo-PA/Angio-PA hydrogel to promote cell adhesion as well as osteogenic and angiogenic differentiation in both 2D and 3D setups. Furthermore, using immunofluorescent staining and reverse transcription-quantitative polymerase chain reaction (RT-qPCR), we demonstrated co-differentiation and organization of hAMSCs and HUVECs into 3D aggregates resembling vascularized bone-like constructs

Magnetically responsive layer-by-layer microcapsules can be retained in cells and under flow conditions to promote local drug release without triggering ROS production.

Nanoengineered vehicles have the potential to deliver cargo drugs directly to disease sites, but can potentially be cleared by immune system cells or lymphatic drainage. In this study we explore the use of magnetism to hold responsive particles at a delivery site, by incorporation of superparamagnetic iron oxide nanoparticles (SPIONs) into layer-by-layer (LbL) microcapsules. Microcapsules with SPIONs were rapidly phagocytosed by cells but did not trigger cellular ROS synthesis within 24 hours of delivery nor affect cell viability. In a non-directional cell migration assay, SPION containing microcapsules significantly inhibited movement of phagocytosing cells when placed in a magnetic field. Similarly, under flow conditions, a magnetic field retained SPION containing microcapsules at a physiologic wall shear stress of 0.751 dyne cm-2. Even when the SPION content was reduced to 20%, the majority of microcapsules were still retained. Dexamethasone microcrystals were synthesised by solvent evaporation and underwent LbL encapsulation with inclusion of a SPION layer. Despite a lower iron to volume content of these structures compared to microcapsules, they were also retained under shear stress conditions and displayed prolonged release of active drug, beyond 30 hours, measured using a glucocorticoid sensitive reporter cell line generated in this study. Our observations suggest use of SPIONs for magnetic retention of LbL structures is both feasible and biocompatible and has potential application for improved local drug delivery

Connexin 43 is overexpressed in human fetal membrane defects after fetoscopic surgery

This project was funded by the RoseTrees Trust (M400, TTC), the QMUL Life Sciences Initiative, Institutional Strategic Support Fund from the Wellcome Trust (105626/Z/14/Z, TTC) and supported by researchers at the National Institute for Health Research, University College London Hospitals Biomedical Research Centre (ALD)

Potential sealing and repair of human FM defects after trauma with peptide amphiphiles and Cx43 antisense

OBJECTIVE: We examined whether peptide amphiphiles functionalised with adhesive, migratory or regenerative sequences could be combined with amniotic fluid (AF) to form plugs that repair fetal membrane (FM) defects after trauma and co-culture with connexin 43 (Cx43) antisense. METHODS: We assessed interactions between peptide amphiphiles and AF and examined the plugs in FM defects after trauma and co-culture with the Cx43antisense. RESULTS: Confocal microscopy confirmed directed self-assembly of peptide amphiphiles with AF to form a plug within minutes, with good mechanical properties. SEM of the plug revealed a multi-layered, nanofibrous network that sealed the FM defect after trauma. Co-culture of the FM defect with Cx43 antisense and plug increased collagen levels but reduced GAG. Culture of the FM defect with peptide amphiphiles incorporating regenerative sequences for 5 days, increased F-actin and nuclear cell contraction, migration and polarization of collagen fibers across the FM defect when compared to control specimens with minimal repair. CONCLUSIONS: Whilst the nanoarchitecture revealed promising conditions to seal iatrogenic FM defects, the peptide amphiphiles need to be designed to maximize repair mechanisms and promote structural compliance with high mechanical tolerance that maintains tissue remodeling with Cx43 antisense for future treatment

Targeting mechanotransduction mechanisms and tissue weakening signals in the human amniotic membrane

Mechanical and inflammatory signals in the fetal membrane play an important role in extracellular matrix (ECM) remodelling in order to dictate the timing of birth. We developed a mechanical model that mimics repetitive stretching of the amniotic membrane (AM) isolated from regions over the placenta (PAM) or cervix (CAM) and examined the effect of cyclic tensile strain (CTS) on mediators involved in mechanotransduction (Cx43, AKT), tissue remodelling (GAGs, elastin, collagen) and inflammation (PGE2, MMPs). In CAM and PAM specimens, the application of CTS increased GAG synthesis, PGE2 release and MMP activity, with concomitant reduction in collagen and elastin content. Co-stimulation with CTS and pharmacological agents that inhibit either Cx43 or AKT, differentially influenced collagen, GAG and elastin in a tissue-dependent manner. SHG confocal imaging of collagen fibres revealed a reduction in SHG intensity after CTS, with regions of disorganisation dependent on tissue location. CTS increased Cx43 and AKT protein and gene expression and the response could be reversed with either CTS, the Cx43 antisense or AKT inhibitor. We demonstrate that targeting Cx43 and AKT prevents strain-induced ECM damage and promotes tissue remodelling mechanisms in the AM. We speculate that a combination of inflammatory and mechanical factors could perturb typical mechanotransduction processes mediated by Cx43 signalling. Cx43 could therefore be a potential therapeutic target to prevent inflammation and preterm premature rupture of the fetal membranes

Route planning with transportation network maps: an eye-tracking study.

Planning routes using transportation network maps is a common task that has received little attention in the literature. Here, we present a novel eye-tracking paradigm to investigate psychological processes and mechanisms involved in such a route planning. In the experiment, participants were first presented with an origin and destination pair before we presented them with fictitious public transportation maps. Their task was to find the connecting route that required the minimum number of transfers. Based on participants' gaze behaviour, each trial was split into two phases: (1) the search for origin and destination phase, i.e., the initial phase of the trial until participants gazed at both origin and destination at least once and (2) the route planning and selection phase. Comparisons of other eye-tracking measures between these phases and the time to complete them, which depended on the complexity of the planning task, suggest that these two phases are indeed distinct and supported by different cognitive processes. For example, participants spent more time attending the centre of the map during the initial search phase, before directing their attention to connecting stations, where transitions between lines were possible. Our results provide novel insights into the psychological processes involved in route planning from maps. The findings are discussed in relation to the current theories of route planning