26 research outputs found
A Multiwell Electrochemical Biosensor for Real-Time Monitoring of the Behavioural Changes of Cells in Vitro
We report the development of a multiwell biosensor for detecting changes in the electrochemical open circuit potential (OCP) generated by viable human cells in vitro. The instrument features eight culture wells; each containing three gold sensors around a common silver/silver chloride reference electrode, prepared using screen-printed conductive inks. The potential applications of the device were demonstrated by monitoring rheumatoid synovial fibroblasts (RSF) and HepG2 hepatocarcinoma cells in response to chemical and biological treatments. This technology could provide an alternative to conventional end-point assays used in the fields of chemotherapy, toxicology and drug discovery
Vitamin D attenuates sphingosine-1-phosphate (S1P)-mediated inhibition of extravillous trophoblast migration.
Failure of trophoblast invasion and remodelling of maternal blood vessels leads to the pregnancy complication pre-eclampsia (PE). In other systems, the sphingolipid, sphingosine-1-phosphate (S1P), controls cell migration therefore this study determined its effect on extravillous trophoblast (EVT) function.A transwell migration system was used to assess the behaviour of three trophoblast cell lines, Swan-71, SGHPL-4, and JEG3, and primary human trophoblasts in the presence or absence of S1P, S1P pathway inhibitors and 1,25(OH)2D3. QPCR and immunolocalisation were used to demonstrate EVT S1P receptor expression.EVTs express S1P receptors 1, 2 and 3. S1P inhibited EVT migration. This effect was abolished in the presence of the specific S1PR2 inhibitor, JTE-013 (p < 0.05 versus S1P alone) whereas treatment with the S1R1/3 inhibitor, FTY720, had no effect. In other cell types S1PR2 is regulated by vitamin D; here we found that treatment with 1,25(OH)2D3 for 48 or 72 h reduces S1PR2 (4-fold; <0.05), but not R1 and R3, expression. Moreover, S1P did not inhibit the migration of cells exposed to 1,25(OH)2D3 (p < 0.05).This study demonstrates that although EVT express three S1P receptor isoforms, S1P predominantly signals through S1PR2/Gα12/13 to activate Rho and thereby acts as potent inhibitor of EVT migration. Importantly, expression of S1PR2, and therefore S1P function, can be down-regulated by vitamin D. Our data suggest that vitamin D deficiency, which is known to be associated with PE, may contribute to the impaired trophoblast migration that underlies this condition
Core–shell–shell cytocompatible polymer dot-based particles with near-infrared emission and enhanced dispersion stability
Transcriptional response of endometrial cells to insulin, cultured using microfluidics
Obesity is a rapidly growing public health issue among women of reproductive age associated with decreased reproductive function including implantation failure. This can result from a myriad of factors including impaired gametes and endometrial dysfunction. The mechanisms of how obesity-related hyperinsulinaemia disrupts endometrial function are poorly understood. We investigated potential mechanisms by which insulin alters endometrial transcript expression. Ishikawa cells were seeded into a microfluidics device attached to a syringe pump to deliver a constant flow rate of 1 ÎĽL/min of the following: (i) control (ii) vehicle control (acidified PBS), or (iii) insulin (10 ng/mL) for 24 h (n = 3 biological replicates). Insulin-induced transcriptomic response of endometrial epithelial cells was determined via RNA sequencing, and DAVID and Webgestalt to identify Gene Ontology (GO) terms and signalling pathways. A total of 29 transcripts showed differential expression levels across two comparison groups (control vs vehicle control; vehicle control vs insulin). Nine transcripts were differentially expressed in vehicle control vs insulin comparison (P < 0.05). Functional annotation analysis of transcripts altered by insulin (n = 9) identified three significantly enriched GO terms: SRP-dependent co-translational protein targeting to membrane, poly(A) binding, and RNA binding (P < 0.05). The overrepresentation analysis found three significantly enriched signalling pathways relating to insulin-induced transcriptomic response: protein export, glutathione metabolism, and ribosome pathways (P < 0.05). Transfection of siRNA for RAPSN successfully knocked down expression (P < 0.05), but this did not have any effect on cellular morphology. Insulin-induced dysregulation of biological functions and pathways highlights potential mechanisms by which high insulin concentrations within maternal circulation may perturb endometrial receptivity
Responsive Nanogel Probe for Ratiometric Fluorescent Sensing of pH and Strain in Hydrogels
In this study a new pH-responsive nanogel probe containing a complementary nonradiative resonance energy transfer (NRET) fluorophore pair is investigated and its ability to act as a versatile probe of network-related changes in three hydrogels demonstrated. Fluorescent sensing using NRET is a powerful method for studying relationships between Angstrom length-scale structure and macroscopic properties of soft matter. Unfortunately, inclusion of NRET fluorophores into such materials requires material-specific chemistry. Here, low concentrations of preformed nanogel probes were included into hydrogel hosts. Ratiometric photoluminescence (PL) data for the gels labeled with the nanogel probes enabled pH-triggered swelling and deswelling to be studied as well as Ca2+-triggered collapse and solute release. PL measurements during compression of a nanogel probe-labeled nanocomposite gel demonstrated mechanochromic behavior and strain sensing. The new nanogel probes have excellent potential for investigating the internal structures of gels and provide a versatile ratiometric fluorescent platform for studying pH and strain
Anisotropic pH-Responsive Hydrogels Containing Soft or Hard Rod-Like Particles Assembled Using Low Shear
A simple and versatile low-shear approach for assembling hydrogels containing aligned rod-like particles (RLPs) that are birefringent and exhibit pH-triggered anisotropic swelling is developed. Anisotropic composite hydrogels are prepared by applying low shear (0.1 s–1) to mixtures of pH-responsive nanogels (NGs) and RLPs. The NGs, which contained high methacrylic acid contents, acted as both shear transfer vehicles and macro-cross-linkers for anisotropic gel formation. Three model RLP systems are investigated: (i) soft triblock copolymer worms, (ii) stiff self-assembled β-sheet peptide fibers, and (iii) ultrahigh modulus nanocrystalline cellulose fibers. RLP alignment was confirmed using polarized light imaging, atomic force microscopy, and small-angle X-ray scattering as well as modulus and anisotropic swelling experiments. Unexpectedly, the composite gel containing the soft copolymer worms showed the most pronounced anisotropy swelling. The copolymer worms enabled higher RLP loadings than was possible for the stiffer RLPs. For fixed RLP loading, the extent of anisotropic swelling increased with intra-RLP bonding strength. The facile and versatile approach to anisotropic gel construction demonstrated herein is expected to enable new applications for strain sensing or biomaterials for soft tissue repair
Electrochemical monitoring of rat mammary adenocarcinoma cells: An in vitro assay for anticancer drug selection
Hydrogel Composites Containing Sacrificial Collapsed Hollow Particles as Dual Action pH-Responsive Biomaterials
In this study hydrogel
composites are investigated that contain
sacrificial pH-responsive collapsed hollow particles (CHPs) entrapped
within a polyÂ(acrylamide) (PAAm) network. The CHPs were prepared using
a scalable (mainly) water-based method and had a bowl-like morphology
that was comparable to that of red blood cells. The CHPs were constructed
from polyÂ(methyl methacrylate-<i>co</i>-methacrylic acid),
which is a pH-responsive copolymer. The PAAm/CHP composite morphology
was probed with optical microscopy, CLSM and SEM. These data showed
the CHPs were dispersed throughout the PAAm network. Inclusion of
the CHPs within the gel composites increased the modulus in a tunable
manner. The CHPs fragmented at pH values greater than the p<i>K</i><sub><i>a</i></sub> of the particles, and this
process decreased the gel modulus to values similar to that of the
parent PAAm hydrogel. CHPs containing a model drug were used to demonstrate
pH-triggered release from PAAm/CHP and the release kinetics obeyed
Fickian diffusion. The composite gels had low cytotoxicity as evidenced
by Live/Dead and MTT assays. The hydrogel composites showed dual action
pH-triggered softening with simultaneous drug release which occurred
without a volume increase. The hydrogel composites may have potential
application as enteric gels or for intra-articular drug delivery