The dependence of residence time distribution on flow in co-axial cylinder devices

Numerical simulations are presented showing the effects of operating and geometrical parameters on the transition of laminar to Taylor vortex flow for induced rotational-axial flow in the gap of a pair of rotating cylinders. These simulations indicate that annular rotational flow becomes more stable in the presence of a small degree of axial flow and as gap width increases. The effect of rotational speed on the breakdown of laminar flow is more complex and for given radius ratio and axial flow rate depends on both the speed ratio and the direction of the rotation of the cylinders, counter-rotating flow generally producing a more stable flow than co-rotating. Limited experimental data are provided on the residence time distribution for flow of Newtonian liquids through the gap of two rotating cylinders. The data include results from experiments in which flow transition occurred from laminar to Taylor vortex flow. The findings from these experiments are successfully analyzed and assessed using the simulations studies.     

Endothelial intercellular cell adhesion molecule 1 contributes to cell aggregate formation in CHO cells cultured in serum-free media

Chinese hamster ovary (CHO) cells have been adapted to grow in serum-free media and in suspension culture to facilitate manufacturing needs. Some CHO cell lines, however, tend to form cell aggregates while being cultured in suspension. This can result in reduced viability and capacity for single cell cloning (SCC) via limiting dilution, and process steps to mitigate cell aggregate formation, for example, addition of anti-cell-aggregation agents. In this study, we have identified endothelial intercellular cell adhesion molecule 1 (ICAM-1) as a key protein promoting cell aggregate formation in a production competent CHO cell line, which is prone to cell aggregate formation. Knocking out (KO) the ICAM-1 gene significantly decreased cell aggregate formation in the culture media without anti-cell-aggregation reagent. This trait can simplify the process of transfection, selection, automated clone isolation, and so on. Evaluation in standard cell line development of ICAM-1 KO and wild-type CHO hosts did not reveal any noticeable impacts on titer or product quality. Furthermore, analysis of a derived nonaggregating cell line showed significant reductions in expression of cell adhesion proteins. Overall, our data suggest that deletion of ICAM-1 and perhaps other cell adhesion proteins can reduce cell aggregate formation and improve clonality assurance during SCC.     

Cover Image,Volume 116, Number 5, May 2019

Human pluripotent stem cell-derived endothelial cells (hPSC-ECs) present an attractive alternative to primary EC sources for vascular grafting. However, there is a need to mature them towards either an arterial or venous subtype. A vital environmental factor involved in the arteriovenous specification of ECs during early embryonic development is fluid shear stress; therefore, there have been attempts to employ adult arterial shear stress conditions to mature hPSC-ECs. However, hPSC-ECs are naïve to fluid shear stress, and their shear responses are still not well understood. Here, we used a multiplex microfluidic platform to systematically investigate the dose-time shear responses on hPSC-EC morphology and arterial-venous phenotypes over a range of magnitudes coincidental with physiological levels of embryonic and adult vasculatures. The device comprised of six parallel cell culture chambers that were individually linked to flow-setting resistance channels, allowing us to simultaneously apply shear stress ranging from 0.4 to 15 dyne/cm ². We found that hPSC-ECs required up to 40 hr of shear exposure to elicit a stable phenotypic change. Cell alignment was visible at shear stress <1 dyne/cm ², which was independent of shear stress magnitude and duration of exposure. We discovered that the arterial markers NOTCH1 and EphrinB2 exhibited a dose-dependent increase in a similar manner beyond a threshold level of 3.8 dyne/cm ², whereas the venous markers COUP-TFII and EphB4 expression remained relatively constant across different magnitudes. These findings indicated that hPSC-ECs were sensitive to relatively low magnitudes of shear stress, and a critical level of ~4 dyne/cm ² was sufficient to preferentially enhance their maturation into an arterial phenotype for future vascular tissue engineering applications.     

GTP hydrolysis by transitional endoplasmic reticulum from rat liver inhibited by all-trans-retinol

GTP hydrolysis by an endoplasmic reticulum fraction from rat liver enriched in part-rough, part-smooth transition elements was inhibited by all-trans-retinol half maximally at a concentration of about 10 micrograms/ml. Similar results were obtained with GTPase activity partially purified by ion-exchange (DE-52) chromatography. The inhibition was non-competitive and given by both retinol and retinaldehyde but not by retinoic acid or alpha-tocopheryl acetate. The hydrolysis of other nucleoside di- and triphosphates was much less affected by retinol. The activity was inhibited by detergents but at much higher concentrations than by retinol. The results suggest that enhancement of cell-free transfer from endoplasmic reticulum to Golgi apparatus by retinol observed previously at low concentrations of cytosol may be mediated through an interaction with GTP.