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Lovastatin Induces Apoptosis through Deprivation of Intracellular Geranylgeranyl Pyrophosphate and Induction of Puma Expression in C6 Glioma Cells
Author(s) -
Choi Jae Won
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.03286
Subject(s) - geranylgeranyl pyrophosphate , farnesyl pyrophosphate , lovastatin , puma , mevalonic acid , viability assay , apoptosis , cycloheximide , chemistry , biology , biochemistry , microbiology and biotechnology , reductase , cholesterol , enzyme , protein biosynthesis , atp synthase
Lovastatin is widely used to decrease serum cholesterol levels and reduce the risk of cardiovascular diseases. Lovastatin is an inhibitor of 3‐hydroxy‐3‐methylglutaryl‐coenzyme A (HMG‐CoA) reductase, an enzyme that converts HMG‐CoA to mevalonate. Mevalonate is a precursor of various metabolic intermediates such as isopentenyl pyrophosphate, farnesyl pyrophosphate, geranylgeranyl pyrophosphate, and cholesterol. It has been reported that statins can induce apoptosis in various cell lines. In this study, we examined the mechanism of lovastatin‐induced apoptosis in C6 and HCT116 cell lines. C6 is a rat glioma cell line and HCT116 is a human colon cancer cell line. Cells were maintained in Dulbecco's minimal essential medium containing 10% (v/v) fetal bovine serum. After treatment, the number of viable cells was estimated using a colorimetric assay based on tetrazolium salt. The percentage of dead cells was estimated using trypan blue cell viability assay or lactate dehydrogenase assay. The levels of Puma mRNA were estimated by reverse transcription quantitative polymerase chain reaction assay. The levels of PUMA were estimated by Western blot assay. Our data showed that lovastatin increased both mRNA and protein levels of Puma in C6 cells, which could be prevented in the presence of 100 μM geranylgeranyl pyrophosphate. Treatment with geranylgeranyl pyrophosphate, cycloheximide, and siPuma (siRNA against Puma ) prevented the death of lovastatin‐treated cells. Pifithrin‐a, an inhibitor of tumor protein p53, neither prevented cell death nor decreased PUMA levels in lovastatin‐treated cells. Pifithrin‐a attenuated cell death and reduced PUMA levels in etoposide‐treated C6 cells. Etoposide, an anticancer drug, has been known to induce p53‐dependent apoptosis. To evaluate the role of p53 on the induction of Puma expression in lovastatin‐treated cells, cytotoxic effects of lovastatin were examined in HCTm and HCTw cells. HCTm is a HCT116 cell line expressing inactive mutant p53 and HCTw is the wild‐type. When cells were cultured in the presence of 25 μM lovastatin for 60 hours, the values of the percentage of cell death were 46% and 43% in HCTm and HCTw groups respectively: there was little difference in lovastatin‐induced cell death between two groups. Etoposide‐induced cell death was exacerbated in HCTw cells as compared to HCTm cells. When cells were cultured with 100 μM etoposide for 72 hours, the values of the percentage of cell death were 35% and 45% in HCTm and HCTw groups respectively. Pifithrin‐a reduced etoposide‐induced cell death in HCTw cells from 45% to 25%. Pifithrin‐a did not attenuate etoposide‐induced cell death in HCTm cells. Our data show that lovastatin depletes intracellular geranylgeranyl pyrophosphate to increase Puma expression in a p53‐independent manner, resulting in apoptosis.