Sirolimus modifies cholesterol homeostasis in hepatic cells: a potential molecular mechanism for sirolimus-associated dyslipidemia.
BACKGROUND
Sirolimus is a potent immunosuppressive agent, which is associated with dyslipidemia in clinical transplantation. The present study was undertaken to investigate the potential hepatocyte mechanisms by which sirolimus causes dyslipidemia.


METHODS
Using both a quantitative assay of intracellular cholesterol and an [3H]-labeled cholesterol efflux assay, we studied the effect of sirolimus on cholesterol accumulation and cholesterol efflux in HepG2 cells in the absence or presence of inflammatory stress induced by interleukin-1beta. The gene and protein expression of molecules involved in cholesterol homeostasis were examined by real-time reverse-transcription polymerase chain reaction and Western blotting.


RESULTS
Sirolimus inhibited low-density lipoprotein (LDL) receptor (LDLr)-mediated cholesterol ester accumulation induced by interleukin-1beta in HepG2 cells. This inhibitory effect was mediated by down-regulation of sterol regulatory element-binding proteins (SREBP) cleavage activating protein (SCAP) and SREBP-2 mRNA expression. Using confocal microscopy, we demonstrated that sirolimus reduced translocation of SCAP-SREBP2 complex from endoplasmic reticulum to Golgi for activation, thereby inhibiting LDLr gene transcription. Reduction of LDLr in the liver may result in a delay of LDL-cholesterol clearance from circulation causing an increase of plasma cholesterol concentration. Furthermore, sirolimus increased cholesterol efflux mediated by adenosine triphosphate-binding cassette transporter A1 gene expression by increasing peroxisome proliferator-activated receptor-alpha and liver X receptor-alpha gene and protein expression. Increased cholesterol efflux from HepG2 cells may increase high-density lipoprotein cholesterol level and also contribute to apolipoprotein B lipoprotein formation by enhancing transfer of high-density lipoprotein cholesterol to apolipoprotein B lipoproteins.


CONCLUSIONS
This study demonstrates that the increase of LDL cholesterol by sirolimus is partly due to the reduction of LDLr on hepatocytes.
Ma
Kun L
KL
Centre for Nephrology, Royal Free and University College Medical School, Royal Free Campus, London, United Kingdom.
Ruan
Xiong Z
XZ
Powis
Stephen H
SH
Chen
Yaxi
Y
Moorhead
John F
JF
Varghese
Zac
Z
D002460:Cell Line; D008078:Cholesterol, LDL / Q000378:metabolism*; D050171:Dyslipidemias / Q000139:chemically induced* / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000737:chemistry / Q000378:metabolism; D006056:Golgi Apparatus / Q000737:chemistry / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006706:Homeostasis / Q000187:drug effects; D006801:Humans; D007166:Immunosuppressive Agents / Q000009:adverse effects / Q000494:pharmacology*; D047908:Intracellular Signaling Peptides and Proteins / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism; D008565:Membrane Proteins / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism; D021381:Protein Transport; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism; D011973:Receptors, LDL / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D020123:Sirolimus / Q000009:adverse effects / Q000494:pharmacology*; D051782:Sterol Regulatory Element Binding Protein 2 / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism
D008078:Cholesterol, LDL; D007166:Immunosuppressive Agents; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D012333:RNA, Messenger; D011973:Receptors, LDL; C497390:SREBF2 protein, human; C102196:SREBP cleavage-activating protein; D051782:Sterol Regulatory Element Binding Protein 2; D020123:Sirolimus