Differential multidrug resistance-associated protein 1 through 6 isoform expression and function in human intestinal epithelial Caco-2 cells

Multidrug resistance-associated protein (MRP) isoforms 1 through 6 mRNA are expressed in the human intestine and Caco-2 cells. In Caco-2 cells, the rank order for mRNA expression was MRP2 ≥ MRP6 > MRP4 ≥ MRP3 > MRP1 = MRP5. The functional expression of MRP-like activity was quantified as the efflux of the fluorescent probe calcein from confluent, polarized monolayers of Caco-2 cells. Calcein efflux was sensitive to temperature, energy depletion, and the MRP antagonist MK571 [3-[[3-[2-(7-chloroquinolin-2-yl)vinyl]phenyl]-(2- dimethylcarbamoylethylsulfanyl)methylsulfanyl] propionic acid]. Calcein efflux across the apical membrane of Caco-2 cells exceeded that across the basolateral by approximately 2-fold, correlating with the apical localization of MRP2 visualized by immunocytochemical staining. T84 cells do not express MRP2 and show a predominance of basolateral calcein efflux over apical efflux. MRP3 was localized by immunocytochemical staining to the basolateral membrane. MRP1 staining was not localized to either membrane domain and MRP5 staining was not detected. Thus, basolateral calcein efflux may reflect a function of MRP3 or MRP4 and 6 inferred by their basolateral localization in other tissues. Basolateral, but not apical, calcein efflux was sensitive to glutathione depletion with buthionine-sulfoximine, indicating that whereas MRP2-mediated apical efflux is independent of glutathione, basolateral efflux is glutathione-dependent. Benzbromarone, probenecid, pravastatin, and diclofenac were able to inhibit both apical and basolateral calcein efflux. The apical calcein efflux in Caco-2 cells was selectively sensitive to indomethacin and propranolol, but not verapamil or erythromycin, whereas the converse was observed for basal efflux. The differential pharmacological sensitivity of apical (MRP2) and basolateral calcein efflux provides tools for dissecting MRP isoform functional roles

The UK coalition and the civil service: A half-term report

Drawing upon a range of policy documents, government papers and parliamentary reports, as well as published academic and think tank analyses, this article sets out the civil service inheritance bequeathed to the 2010 UK coalition government by its predecessors, examines the pre-election civil service policies of the Conservatives and the Liberal Democrats, illustrates the policy challenges facing the civil service in serving two masters, and analyses the reasons for and early impact of organisational and leadership changes in Whitehall since 2010. The background to, and potential significance of, the 2012 Civil Service Reform Plan are set out. The article provides a succinct overview of the impact of the coalition government on Whitehall during the first two and a half years of its term

The Misalignment of Policy and Practice in Sustainable Urban Design

The urban renaissance that took place in major UK cities during the late 1990’s was seen as a response to counter-urbanisation and inner city decline. This chapter will argue that there has been an implementation gap between the intent of urban design policy and its impact on the ground. Drawing upon the experiences of the lead author as an urban designer in the North-East of England over this period, the text will present examples of the unforeseen consequences that have accompanied the policy trajectory of sustainability and quality within the built environment, specifically looking at a move towards community-led development away from centralisation, by passing local government in the process. The chapter will examine how this urban renaissance has emerged through policies focused on the renewal of the housing market and more sustainable communities (e.g. HMR Pathfinder) to the creation of the Big Society with devolved services and funding, initiatives aimed at individual households (e.g. the Green Deal) and stakeholder engagement. In analysing this policy journey the chapter will consider why demolition and new build invariably occurred instead of refurbishment and community development; why the accepted need for evidence largely ignored qualitative and anecdotal insight from local communities and why end state planning was pursued in preference to the need for a more adaptive and dynamic process