Upregulation of selective cholesteryl ester uptake pathway in mice with deletion of low-density lipoprotein receptor function

Abstract

This study examines the effect of mutation of the low-density lipoprotein receptor (LDLR) on cholesterol metabolism, and especially lipoprotein-derived cholesteryl ester uptake, in murine ovarian granulosa cells. Although the tests were conducted on cells prepared by two different procedures, the results are similar. Deletion of LDLR function did not noticeably affect key enzymes of the steroidogenic pathway or affect progestin production and secretion in granulosa cells. No change was found in expression of LDL-related protein (LRP). These data suggested that cholesterol turnover in cells from the knockout animals is within normal limits and that the cells are not stressed to acquire more cholesterol. Both biochemical and morphological data indicate that unstimulated granulosa cells from LDLR-/- mice are nonetheless programmed to take in double the amount of lipoprotein-derived cholesteryl ester (via the selective cholesteryl ester uptake pathway) and to process (hydrolyze, re-esterify, or utilize) more than twofold the cholesteryl ester processed by cells from wildtype (LDLR+/+) animals. Bt2cAMP stimulation of the murine granulosa cells increases the mass of cholesteryl ester taken up by the selective pathway by an additional 38%. To determine to what extent this increase is related to high-density lipoprotein (HDL) scavenger receptor protein (SR-BI) or caveolin function, Western blots and immunohistochemical studies were performed under a variety of conditions. SR-BI levels are found to be low in unstimulated cells of both LDLR+/+ and LDLR-/- animals, but highly expressed (approximately 20-fold increase over basal levels) in stimulated (Bt2cAMP) cells of both animal models. Thus, the functional relationship between selective cholesteryl ester uptake and SR-BI receptor protein is not as tight as in previously reported studies, suggesting a requirement for other tissue factors. Caveolin expression did not change under any of the conditions tested and appears not to be functionally involved in this process.

Caveolin-1 does not affect SR-BI-mediated cholesterol efflux or selective uptake of cholesteryl ester in two cell lines

“…Surprisingly, treatment of rat ovarian granulosa cells with dibutyryl cAMP or folliclestimulating hormone down-regulated caveolin-1, though both SR-BI and selective uptake of CE were strongly induced (25). Caveolin-1 expression was not stimulated by similar hormone treatment of murine ovarian granulosa cells, although SR-BI expression and CE selective uptake were both induced (26). Similarly, in preliminary experiments, we found that though adrenocorticotropin hormone treatment of Y1 mouse adrenocortical cells stimulated SR-BI expression and CE selective uptake, caveolin-1 expression was not induced and may even have been reduced (M. A. Connelly and D. L. Williams, unpublished observations).…”

Experimental Glomerulopathy Alters Renal Cortical Cholesterol, SR-B1, ABCA1, and HMG CoA Reductase Expression

“…SR-B1 (or the high-density lipoprotein receptor) is thought to be a dominant pathway for cellular CE uptake. [12][13][14][15][16][17] It can also impact FC levels, potentially via a direct action 40,41 and/or by inhibiting the ABCA1 efflux pathway. 12 Given that CE overload was the dominant cholesterol change observed in these experiments, we hypothesized that this would induce a compensatory decrease in SR-B1 expression, because cells might attempt to limit the CE overload by specifically decreasing CE uptake.…”

“…8 -10 Secondly, increased filtration of FC-and CE-bearing lipoproteins might increase tubular cell cholesterol uptake, eg, via the high-density lipoprotein scavenger receptor-B1 (SR-B1) receptor. [11][12][13][14][15][16][17] The latter is particularly relevant for rats, given that high-density lipoprotein, rather than low-density lipoprotein, is the primary cholesterol carrier. 11 An alternative scenario might be that once cholesterol accumulation within glomerular and/or tubular cells is initiated, secondary compensatory mechanisms come into play that act to hold further cholesterol accumulation in check.…”

Expression of scavenger receptor class B type 1 (SR-BI) promotes microvillar channel formation and selective cholesteryl ester transport in a heterologous reconstituted system

“…Antibody-treated cells were subjected to standard immunochemical-labeling techniques (6,14,15,17,18) and viewed by confocal microscopy (Stanford University Cell Science Imaging Facility), and the resulting images were subsequently colorized by using PHOTOSHOP technology (Adobe Systems, Mountain View, CA).…”

“…For uptake and internalization studies, hHDL 3 preparations were conjugated with residualizing labels, i.e., (6,7,21). For fluorescence microscopy, reconstituted BODIPY-CE-HDL particles were prepared as described (9,14,15,17,18). ]COE-hHDL 3 (33 g͞ml medium) in the presence or absence of excess unlabeled hHDL 3 500 g͞ml medium, to determine nonspecific binding (6,11,21).…”