Search

The polymer polydimethylsiloxane (PDMS) is widely used to build microfluidic devices compatible with cell culture. Whilst convenient in manufacture, PDMS has the disadvantage that it can absorb small molecules such as drugs. In microfluidic devices like “Organs-on-Chip”, designed to examine cell behavior and test the effects of drugs, this might impact drug bioavailability. Here we developed an assay to compare the absorption of a test set of four cardiac drugs by PDMS based on measuring the residual non-absorbed compound by High Pressure Liquid Chromatography (HPLC). We showed that absorption was variable and time dependent and not determined exclusively by hydrophobicity as claimed previously. We demonstrated that two commercially available lipophilic coatings and the presence of cells affected absorption. The use of lipophilic coatings may be useful in preventing small molecule absorption by PDMS., Graphical abstract Image 1, Highlights • Binding of different compounds to PDMS varies greatly. • Previous reported correlations of absorption and LogP values could not be repeated. • Topological polar surface area possibly related to compound absorption. • A lipid based coating partially obviates compound absorption. • Presence of cultured cells affects free drug concentration, but less than substrate.

Insights into the complex stem cell niche have identified the cell–material interface to be a potent regulator of stem cell fate via material properties such as chemistry, topography and stiffness. In light of this, materials scientists have the opportunity to develop bioactive materials for stem cell culture that elicit specific cellular responses. To accelerate materials discovery, high throughput screening platforms have been designed which can rapidly evaluate combinatorial material libraries in two and three-dimensional environments. In this review, we present screening platforms for the discovery of material properties that influence stem cell behavior., Engineering and Physical Sciences Research Council, National Institutes of Health (U.S.) (Ruth L. Kirschstein National Research Service Award F32HL122009), National Institutes of Health (U.S.) (Grant R01 DE016516)

Background Online label-free monitoring of in-vitro differentiation of stem cells remains a major challenge in stem cell research. In this paper we report the use of Raman micro-spectroscopy (RMS) to measure time- and spatially-resolved molecular changes in intact embryoid bodies (EBs) during in-vitro cardiogenic differentiation. Methods EBs formed by aggregation of human embryonic stem cells (hESCs) were cultured in defined medium to induce differentiation towards cardiac phenotype and maintained in purpose-built micro-bioreactors on the Raman microscope for 5 days (between days 5 and 9 of differentiation) and spatially-resolved spectra were recorded at 24 h intervals. Results The Raman spectra showed that the onset of spontaneous beating of EBs at day 7 coincided with an increase in the intensity of the Raman bands at 1340 cm− 1, 1083 cm− 1, 937 cm− 1, 858 cm− 1, 577 cm− 1 and 482 cm− 1. The spectral maps corresponding to these bands had a high positive correlation with the expression of the cardiac-specific α-actinin obtained by immuno-fluorescence imaging of the same EBs. The spectral markers obtained here are also in agreement with previous studies performed on individual live hESC-derived CMs. Conclusions The intensity profile of these Raman bands can be used for label-free in-situ monitoring of EBs to estimate the efficacy of cardiogenic differentiation. General significance As the acquisition of the time-course Raman spectra did not affect the viability or the differentiation potential of the hESCs, this study demonstrates the feasibility of using RMS for on-line non-invasive continuous monitoring of such processes inside bioreactor culture systems.

Cardiotoxicity arising from cell death or altered electrophysiology is a major cause of late stage drug failure, posing a significant burden on the pharmaceutical industry. Refinement and development of new early stage in vitro screens that can reliably detect cardiotoxicological events will be necessary to eliminate compounds that would otherwise fail at the preclinical and clinical stages. Because these developments will probably require a reproducible source of functional human cardiomyocytes, we consider the suitability of human embryonic stem cell (hESC)-derived cardiomyocytes by reviewing their characteristics and maturation. We highlight expression of >40 cardiac-related genes and responsiveness to >30 agents, concluding that these cells warrant further investigation in assessing cardiotoxicity.

To realize the full scientific and clinical potential of human embryonic stem cell (hESC)-cardiomyocytes, strategies to overcome the high degree of heterogeneity of differentiated populations are required. Here we demonstrate the utility of two transgenic approaches in enrichment of cardiomyocytes derived from HUES-7 cells: (i) negative selection of proliferating cells with the herpes simplex virus thymidine kinase/ganciclovir (HSVtk/GCV) suicide gene system; and (ii) positive selection of cardiomyocytes expressing a bicistronic reporter [green fluorescent protein (GFP)-internal ribosome entry site (IRES)-puromycin-N-acetyltransferase (PAC)] from the human alphamyosin heavy chain promoter. Parental and transgenic HUES-7 cells were similar with regard to morphology, pluripotency marker expression, differentiation, and cardiomyocyte electrophysiology. Whereas immunostaining of dissociated cardiomyocyte preparations expressing HSVtk or PAC contained7% cardiomyocytes, parallel cultures treated with GCV or puromycin, respectively, contained 33.4 +/- 2.1% or 91.5 +/- 4.3% cardiomyocytes corresponding to an enrichment factor of 6.7- or 14.5-fold. Drug-selected cardiomyocytes responded to chronotropic stimulation and displayed cardiac-specific action potentials, demonstrating that functionality was retained. Both transgenic strategies will be generically applicable and should readily translate to the enrichment of many other differentiated lineages derived from hESCs.

Our laboratory is evaluating whether an epigenetic mechanism involving alterations in DNA methylation can alter the trajectory of embryonic/fetal development in response to maternal nutrients. A similar mechanism may operate in embryo culture environments commonly used in human assisted conception. Since developmental studies on early human embryos in utero are obviously not possible, we have begun to investigate the utility of human embryonic stem cells (hESC) to uncover potential programming mechanisms. This review highlights some of the advantages and problems associated with such a model and suggests that these issues are also broadly applicable to utility of hESC for more general toxicology and drug screening applications.

The classic question in corporate governance is how to manage the conflicts-of-interest that arise from fundamental principal-agent problems. One of the most popular methods of solving the problem is by separating ownership and management. Since 2002, the Sarbanes-Oxley Act has set new standards for board composition by using independent directors, especially for audit committees. This article analyzes the impact of board composition, related to a majority of independent directors, on returns. For this purpose we construct a panel dataset of 2919 stocks over a 10 year period. We find that a majority of independent directors on the board has an overall negative effect on stock returns. We also use our panel data set to show the consequences of the Sarbanes-Oxley Act on the defense or offense mechanisms that companies might pursue over their lifespan.