P glycoprotein-mediated cholesterol cycling determines proximal tubular cell viability

Abstract

Background. MDR P glycoproteins may help transport plasma membrane free cholesterol (FC) to the endoplasmic reticulum (ER), where it undergoes acylation, forming cholesterol esters (CE). This study assessed whether P glycoprotein inhibitors alter renal tubular FC/CE expression, thereby altering cell integrity. Methods. Mouse proximal tubule segments (PTS) were exposed to chemically dissimilar P glycoprotein inhibitors [progesterone (prog), trifluoperazine (TFP), or cyclosporine A (CsA)]. Their effects on FC/CE and adenosine 5′-triphosphate (ATP) levels, phospholipid expression, lipid peroxidation, and cell viability (lactate dehydrogenase release; LDH) were assessed. P glycoprotein inhibitor effects on cultured proximal tubular (HK-2) cell viability and susceptibility to Fe-induced oxidant stress were also addressed. Results. When applied to PTS, prog, TFP, and, to lesser extent, CsA induced dose-dependent ATP reductions (≤90%), CE decrements (∼40%), and LDH release (≤60%). No concomitant changes in lipid peroxidation or phospholipid profiles were observed. Ouabain did not preserve tubular ATP, suggesting that decreased ATP production, rather than increased consumption, was operative. Mechanisms leading to cell lysis were not identical, as glycine and arachidonic acid blocked prog- but not TFP-mediated cell death. When prog-driven CE reductions were attenuated in PTS with a procycling agent (cholesterol oxidase), decreased cell death resulted. P glycoprotein inhibitors also caused dose-dependent HK-2 cell death. Blocking Fe-mediated CE formation (∼ × 10) with sublethal CsA doses led to a marked increase in Fe-mediated cell death. Conclusions. P glycoproteins may be critical to tubule cholesterol transport. If blocked with pharmacologic agents, decreased ATP production, overt cell lysis, and/or a marked propensity to superimposed tubular cell injury can result.