The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Liang, S.-L.
Right arrow Articles by Zhou, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Liang, S.-L.
Right arrow Articles by Zhou, A.
Right arrowPubmed/NCBI databases
*Compound via MeSH
*Substance via MeSH
Hazardous Substances DB
*LOVASTATIN
The Journal of Immunology, 2006, 177: 651-656.
Copyright © 2006 by The American Association of Immunologists

Lovastatin-Induced Apoptosis in Macrophages through the Rac1/Cdc42/JNK Pathway

Shu-Ling Liang, Hongli Liu and Aimin Zhou1

Clinical Chemistry Program, Department of Chemistry, Cleveland State University, Cleveland, OH 44115


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Statins, inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase, have been used successfully in the treatment of hypercholesterolemia for more than a decade. Statins also exhibit overall clinical benefits on cardiovascular diseases independent of their effects on lowering serum cholesterol levels. These beneficial effects of statin therapy are believed to be due, at least in part, to the anti-inflammatory and immunomodulatory roles of statins. Statin treatment reduces the levels of inflammatory markers, decreases the activation and recruitment of immune cells, and delays the progression of atherosclerosis, a chronic inflammatory disease. However, little is known about the direct impact of statins on immune cells, particularly on macrophages. We report that lovastatin, a member of the statin family, effectively induces apoptosis in macrophages. Further investigation of the molecular mechanism has revealed that Rac1 and Cdc42, the small GTPase family members, may play an important role in lovastatin-induced macrophage apoptosis. Moreover, the activation of the JNK pathway may contribute to this event. Our findings provide a better understanding of the molecular basis underlying the anti-inflammatory clinical benefits of statin therapy in cardiovascular diseases.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
The statin family of drugs has been used successfully in the treatment of hypercholesterolemia for more than a decade. To date, they are still the most powerful drugs for lowering cholesterol levels in blood (1, 2, 3). In addition to their potential role in lowering serum cholesterol levels, clinical trials and in vitro studies have shown that statins have pleiotropic effects on a wide range of cell functions and exhibit overall clinical benefits on cardiovascular diseases. Statin therapy significantly reduces ischemic stroke for patients with established coronary artery diseases (4, 5, 6, 7, 8), and is associated with improved patency of autogenous infrainguinal bypass grafts (9).

Atherosclerosis as a chronic inflammatory disease leads to a variety of cardiovascular disorders, such as myocardial infarction, stroke, peripheral vascular disease, and aortic aneurysm in aged people. Clinically, statins slow plaque progression and promote regression of atherosclerotic lesions (10, 11, 12, 13, 14, 15). These beneficial effects of statin therapy are believed to be due, at least in part, to the anti-inflammatory and immunomodulatory properties of statins. Treatment with statins in patients with acute coronary syndromes significantly reduces the levels of inflammatory markers, such as C-reactive protein and serum amyloid A (16, 17, 18, 19, 20). Statins have been found to decrease the activation and recruitment of immune cells, an immune response that contributes to the progression of atherosclerotic development (21, 22). However, the direct impact of statins on immune cells, particularly on macrophages, remains largely unexplored.

Statins are inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase, the rate-limiting enzyme of the mevalonate pathway. The mevalonate pathway produces isoprenoids that are critical for diverse cellular functions, ranging from cholesterol synthesis to growth control. Farnesylpyrophosphate (FPP)2 and geranylgeranylpyrophosphate (GGPP), the isoprenoid intermediates of the mevalonate pathway, are also important for the posttranslational modification of a variety of proteins, including Ras and small GTPases, such as RhoA, Rac, and Cdc42. Prenylation of these proteins with the isoprenoids is required for their translocation to the membrane and for cellular functions (23).

Rho, Rac, and Cdc42 are among the best characterized small GTPases. Although they display similar biological activities in most cases, each of the GTPases can also mediate distinct cellular functions through interaction with its own downstream effector proteins in different cell types. For example, the serum response element of the c-fos promoter is activated by Rho A, but not Rac1 and Cdc42 (24). Rac1 regulates the activation of NADPH oxidase for the purposes of innate immunity in phagocytes. However, Cdc42 acts as an antagonist in the formation of reactive oxygen species in these cells (25, 26). Evidence for extensive cross talks and cooperation between GTPases and other signal transduction pathways is well documented. Rac1 and Cdc42 can synergize with Raf to activate ERK (27, 28, 29). Furthermore, Rac1 cross talks with PI3K in controlling cell migration and polarity (30). Interestingly, these small GTPases are found to mediate apoptosis in a wide range of cell types. Overexpression of the active form of Cdc42 in Jurkat T lymphocytes induces an increase of ceramide levels, resulting in cell death (31, 32). Both Rac1 and Cdc42 mediate apoptosis induced by diverse stimuli (33). It seems that the contribution of Rac1 and Cdc42 to apoptosis is thought to mainly regulate the activation of the JNK pathways (34, 35, 36).

In this study, we investigated the direct impact of lovastatin on macrophages. Our data have shown that lovastatin induces apoptosis in macrophages through enhancing the expression of Rac1 and Cdc42, resulting in the activation of the JNK pathway.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Reagents and Abs

Lovastatin (Mevinolin), FPP, GGPP, cycloheximide, and SP600125 were from Sigma-Aldrich. Ab specific to the apoptotically cleaved form (86 kDa) of poly(ADP-ribose) polymerase (PARP) was purchased from Cell Signaling Technology. Abs to total PARP, Ras, RhoA, Rac1, Cdc42, phospho-c-Jun, and beta-actin were from Santa Cruz Biotechnology. Activation of lovastatin was performed, as described previously (37). Briefly, lovastatin (25 mg) was dissolved in 0.625 ml of absolute ethanol. Freshly made 0.1 N NaOH (0.938 ml) was added and mixed, and the solution was then heated at 50°C for 2 h. The pH was adjusted to 7.2 by using 0.1 N HCl, and dd H2O was added to a final volume of 6.25 ml. The solution of 10 mM lovastatin was aliquoted and stored at –20°C.

Cell culture

Raw 264.7 (Raw) (American Type Culture Collection) cells were grown in DMEM (Cleveland Clinic Foundation Core Facility) supplemented with 10% cosmic calf serum (HyClone) and antibiotics in a humidified atmosphere of 5% CO2 at 37°C. Mouse bone marrow cells were isolated from C57BJ/6B mice (The Jackson Laboratory) and grown in RPMI 1640 supplemented with 15% FBS (BioSource International) and 15% L cell medium plus antibiotics for 1 wk to be differentiated into macrophages.

Assay for cell viability

The viability of Raw cells was determined using the colorimetric CellTiter 96 aqueous cell proliferation assay (MTT), according to the instructions provided by the manufacturer (Promega). Briefly, cells (1 x 104 cells/well) were seeded in 96-well plates. One day after seeding, they were treated with or without 10 µM lovastatin in the presence or absence of 5 µM GGPP or 5 µM FPP for 22 h. At the end of incubation, 50 µl of CellTiter 96 aqueous reagent (40% v/v dilution in 1x PBS) was added to each well. Plates were incubated at 37°C for 2 h, and absorbance was measured at 490 nm with a 96-well plate reader (model Spectra Max 340; Molecular Devices).

Caspase assays

The activities of caspase 3 and 9 in the cells treated with lovastatin were examined by using the Caspase-GloTM 3/7 and 9 assay kits (Promega). In these assays, caspase cleaves the proluminescent substrate linked with the tetrapeptide DEVD (for caspase 3/7) or LEHD (for caspase 9), resulting in the generation of a glow-type luminescent signal, produced by luciferase. In brief, cells were treated as indicated, and cytosolic extracts were prepared by suspension of cell pellets in Nonidet P-40 lysis buffer (10 mM Tris-HCl (pH 8.0), 5 mM Mg(OAc)2, 90 mM KCl, 0.2 mM PMSF, 100 U/ml aprotinin, 10 µg/ml leupeptin, and 2% Nonidet P-40). After centrifugation at 10,000 x g for 5 min, cell extracts containing 40 µg of proteins were transferred into a 96-well plate, and 50 µl of Caspase-Glo3/7 or Caspase-Glo9 Reagents was added to the wells. After incubation for 1 h at 37°C, caspase activities were determined by a fluorescent plate reader (Microtiter Plate Luminometer; Dynex Technologies).

Determination of DNA fragmentation

DNA in the cells, after the indicated treatment, was isolated using an apoptotic DNA ladder kit (Roche). Briefly, 200 µl of cell suspensions in PBS was mixed with 200 µl of binding buffer (Roche). After incubation for 10 min at room temperature, 100 µl of isopropanol was added to the sample and mixed by vortexing. DNA was then purified by using glass fibers. DNA samples were separated on a 2% agarose gel and visualized with ethidine bromide staining under UV light.

TUNEL assay

TUNEL for DNA degradation was performed using an apo-BrdU kit (BD Biosciences). Briefly, cells were scraped using a scraper with medium containing floating cells after the indicated treatments. The cells were fixed in 1% paraformaldehyde for 30 min and stored in 70% ethanol at –20°C until staining and analysis. DNA fragmentation was examined by incorporating BrdU and staining with a labeled anti-bromodeoxyuridine mAb. The total DNA content was determined with propidium iodine, and the labeled cells were sorted by flow cytometry in a FACS.

Western blot analysis

After treatments, cells were washed twice with ice-cold PBS and collected with a scraper. Cytoplasmic extracts were prepared by suspension of cell pellets in Nonidet P-40 lysis buffer, as described above. After centrifugation at 10,000 x g in a microcentrifuge at 4°C for 10 min, cell extracts (100 µg/sample) were fractionated on SDS-10% polyacrylamide gels and transferred to polyvinylidene difluoride membranes (Millipore). The membranes were blocked with 5% nonfat milk in PBS containing 0.02% sodium azide and 0.2% (v/v) Tween 20, and incubated with different primary Abs for 1 h at room temperature. The membranes were then washed with PBS containing 0.2% (v/v) Tween 20 and incubated with specific secondary Abs conjugated with HRP (Cell Signaling Technology) for 1 h at room temperature. After washing, these proteins were detected by a chemiluminescence method, according to the manufacturer’s specification (Amersham).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
Lovastatin-induced decrease of macrophage viability was prevented by GGPP and FPP

To determine the direct effect of statins on immune cells, we treated Raw cells, a mouse macrophage cell line, with varied concentrations of lovastatin, a member of the statin family. Cell viability was analyzed using the MTT assay. As shown in Fig. 1A, lovastatin at 10 µM reduced cell viability to 25.6% of that of untreated cells in 24 h. To determine whether lovastatin-induced macrophage apoptosis is cell type specific, we treated Ba/F3 cells, a mouse pro-B cell line, and Jurkat cells, a human T cell line, with lovastatin. Although lovastatin was able to induce apoptosis in these cells, a higher concentration (30–50 µM) and a prolonged incubation time (48–72 h) were required to achieve the same level of apoptosis (data not shown). These results are consistent with the previous reports that lovastatin at a higher concentration is needed to induce apoptosis in smooth muscle cells, endothelial cells, and plasma cells (38, 39, 40), suggesting that macrophages are particularly vulnerable to lovastatin-induced apoptosis.


Figure 1
View larger version (63K):
[in this window]
[in a new window]
  		FIGURE 1. Effect of lovastatin on macrophage viability. Raw cells were treated with different amounts of lovastatin (A) or 10 µM lovastatin for 24 h in the presence or absence of 5 µM GGPP or FPP (B). The percentage of viable cells was measured by using MTT assay (Promega). Experiments were performed three times in triplicates. Data are presented as mean ± SD. C, Photomicrographs of Raw cells and BMM after lovastatin treatment. Photographs of the unstained cells were taken at 30 h for Raw cells (C-I) and 48 h for BMM (C-II) after lovastatin was added under Olympus model CKX31 at x100 magnification: a, untreated; b, treated with 10 µM lovastatin; c, treated with 10 µM lovastatin plus 5 µM GGPP; d, treated with 10 µM lovastatin plus 5 µM FPP. D, Effect of lovastatin on the PARP cleavage in Raw cells. Raw cells were treated with 10 µM lovastatin for the times indicated, and the PARP cleavage was determined by Western blot analysis using a specific mAb to the 86-kDa fragment (Cell Signaling Technology).

 
Inhibition of 3-hydroxy-3-methylglutaryl coenzyme A reductase by statins results in the depletion of several intermediate products in the mevalonate pathway. These intermediates, including FPP and GGPP, are isoprenoids that serve as the substrates for the prenylation of Ras and small GTPases, such as RhoA, Rac, and Cdc42. To determine whether supplementation of GGPP or FPP is able to prevent cells from death, Raw cells were coincubated with 10 µM lovastatin and 5 µM GGPP or FPP. Treatment of Raw cells with GGPP or FPP almost completely prevented cell death induced by lovastatin, suggesting that inhibition of protein prenylation accounts for lovastatin-induced macrophage death (Fig. 1B). The lovastatin-induced Raw cell death was observed by phase contrast microscopy (Fig. 1, C-I). To rule out the possibility that this result is cell line specific, we treated J774 cells (data not shown), another murine macrophage cell line, and primary mouse bone marrow-derived macrophages (BMM) (Fig. 1, C-II) with lovastatin. Similar results were obtained. Therefore, our data with all macrophage cells tested indicated that lovastatin is able to induce macrophage death in both cell lines and primary cells.

Lovastatin induces macrophage apoptosis

To determine whether the decreased viability of Raw cells is due to lovastatin-induced apoptosis, the PARP cleavage was measured after the cells were treated with lovastatin. PARP is an enzyme responsible for DNA damage surveillance, and PARP cleavage is considered to be one of the classical characteristics of apoptosis. During apoptosis, PARP (116 kDa) is cleaved by the activated caspase family members, such as caspase 3, into 25- and 86-kDa fragments. In this experiment, Raw cells were treated with 10 µM Lovastatin for different times, and PARP cleavage in the cells was determined by Western blot analysis using a mAb recognizing 86-kDa PARP. As shown in Fig. 1D, PARP cleavage was detected in a time-dependent fashion. To further characterize lovastatin-induced macrophage apoptosis, we determined the activity of caspases in Raw cells after being treated with lovastatin. The activity of caspase 3 and 9 in the cells treated with lovastatin was examined by using the Caspase-GloTM 3/7 and 9 assay kits (Promega). In these assays, caspase cleaves the proluminescent substrate linked with the tetrapeptide DEVD (for caspase 3/7) or LEHD (for caspase 9), resulting in the generation of a glow-type luminescent signal, produced by luciferase. Luminescence is proportional to the amount of caspase activity present. Clearly, lovastatin could greatly induce the activity of caspase 3 in Raw cells. However, the supplementation of GGPP and FPP was able to suppress the caspase 3 activity induced by lovastatin, implicating that the blockage of the mevalonate pathway triggers the apoptotic cascade (Fig. 2A). Notably, caspase 9 is not activated in lovastatin-induced macrophage apoptosis (Fig. 2B).


Figure 2
View larger version (28K):
[in this window]
[in a new window]
  		FIGURE 2. Induction of caspase 3 activity by lovastatin. Raw cells were treated with 10 µM lovastatin in the presence or absence of 5 µM GGPP, 5 µM FPP, 10 µM caspase 3 inhibitor (Ac-DEVD-CHO), and 10 µM caspase 9 inhibitor (LEHD-CHO) for 22 h. The activity of caspase 3 (A) and caspase 9 (B) was measured using Caspase-GloTM 3/7 and 9 assay kits (Promega). Experiments were performed two times in triplicates. Data are presented as mean ± SD.

 
DNA fragmentation is another typical feature of cell apoptosis. The genomic DNA extracted from Raw cells treated with lovastatin was analyzed by agarose electrophoresis. The degree of ladder formation was related to the concentration of lovastatin (Fig. 3A), and this event can be blocked by adding GGPP or FPP (Fig. 3B). We also performed the TUNEL to determine apoptotic cells. The TUNEL assay detects DNA strand breaks induced during the process of apoptosis. Treatment of Raw cells with 10 µM lovastatin for 24 h produced 68% TUNEL-positive cells, indicating that Raw cells are very sensitive to lovastatin. To determine whether inhibition of protein synthesis impacts lovastatin-induced apoptosis, we treated Raw cells with lovastatin in the presence or absence of cycloheximide, a common inhibitor of protein synthesis. Cycloheximide at a concentration of 0.5 µM remarkably inhibited lovastatin-induced apoptosis in Raw cells, suggesting that a novel protein synthesis is required for the apoptotic process (Fig. 4).


Figure 3
View larger version (25K):
[in this window]
[in a new window]
  		FIGURE 3. Induction of DNA fragmentation in Raw cells by lovastatin. Raw cells were treated for 24 h with different concentration of lovastatin (A) or with 10 µM lovastatin plus 5 µM GGPP or FPP (B). The DNA fragments were purified using an apoptotic DNA ladder kit (Roche), and separated by 2% agarose gel electrophoresis and stained by ethidium bromide.

 

Figure 4
View larger version (7K):
[in this window]
[in a new window]
  		FIGURE 4. Inhibition of protein synthesis suppresses lovastatin-induced macrophage apoptosis. Raw cells were incubated with 10 µM lovastatin in the presence or absence of 0.5 µM cycloheximide for 24 h, and apoptotic cells were analyzed by TUNEL assay (Roche) and the results were displayed as a histogram.

 
Lovastatin induces the expression of Rac1 and Cdc42 in macrophages

Blockage of the mevalonate pathway by statins results in the depletion of isoprenoids, which are required for the posttranslational modification of Ras and Ras-like GTPase family, such as RhoA, Rac, and Cdc42. These small GTPases act as molecular switches to regulate diverse biochemical functions in all eukaryotic cells through mediating gene expressions. Therefore, we investigated the effect of lovastatin on the expression of certain proteins, including the members of small GTPase family. Intriguingly, we found that lovastatin significantly induced the expression of Rac1 and Cdc42, but not RhoA, in a dose- and time-dependent manner (Fig. 5, A and B). Lovasatin treatment did not affect the level of Ras in the cells (data not shown). The role of Rac1 and Cdc42 in activation of JNK has been well established (34, 35, 36). Thus, we examined the activation of JNK pathway. Interestingly, we found that the high levels of Rac1 and Cdc42 proteins in the cells appeared along with the phosphorylation of c-Jun (Fig. 5B) and the cleavage of PARP (Fig. 5, A and B). These data suggest that an increase of the expression of both proteins may be the cause to trigger the apoptotic cascade through the activation of JNK. The supplementation of GGPP and FPP prevented apoptosis in Raw cells induced by lovastatin while suppressing the expression of Rac1 and Cdc42 (Fig. 5C). To determine whether there was a causal relationship between JNK activation and macrophage apoptosis induced by lovastatin, we treated Raw cells with 10 µM lovastatin in the presence or absence of 10 µM SP600125, an inhibitor of JNK. Cell viability was measured by MTT assays. The presence of SP600125 prevented 60% cell death compared with lovastatin alone. This result suggests that JNK activity is responsible for lovastatin-induced apoptosis, although SP600125 was not as effective as GGPP and FPP in preventing cell death. Western blot analysis indicated that SP600125, GGPP, and FPP prevented apoptosis in Raw cells through suppressing c-jun phosphorylation (Fig. 5D).


Figure 5
View larger version (39K):
[in this window]
[in a new window]
  		FIGURE 5. Lovastatin induced the expression of Rac1/Cdc42 and activation of JNK. Raw cells were treated with different doses of lovastatin for 20 h (A); treated with 10 µM lovastatin at different times (B); treated with 10 µM lovastatin in the presence or absence of 5 µM GGPP or FPP for 20 h (C); and treated with 10 µM lovastatin in the presence or absence of 10 µm SP600125, 5 µM GGPP, and 5 µM FPP for 24 h (D). Proteins in the extracts were separated by 10% SDS-PAGE and analyzed by Western blot using Abs to PARP, Rac1, Cdc42, RhoA, p-c-Jun, and beta-actin (Santa Cruz Biotechnology).

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
Our findings demonstrate that lovastatin induces apoptosis in macrophages, correlating with enhanced expression of Rac1 and Cdc42 and activation of the JNK pathway. Although it has been reported that statin therapy decreases inflammatory markers in blood and modulates the functions of immune cells, our results provide the first evidence that statins may reduce inflammation via elimination of immune cells. Apparently, the mevalonate pathway plays a pivotal role in macrophage survival because supplementation of GGPP or FPP can effectively rescue cells from lovastatin-induced apoptosis. Lovastatin also induces apoptosis in smooth muscle cells, endothelial cells, and plasma cells (38, 39, 40). In contrast to the observation by others that FPP is ineffective in preventing cells from apoptosis (38, 41), both GGPP and FPP were able to prevent lovastatin-induced macrophage apoptosis in our experiments. GGPP and FPP are the intermediates in the mevalonate pathway that leads to the synthesis of sterols and ubiquinone (42). It has been well established that GGPP and FPP are very important for the posttranslational modification of a variety of proteins, such as Ras and other GTPase family members. Small GTPases display their biological function through binding GTP, which leads to the interaction with specific molecular targets in the membrane, and subsequently initiates the signaling cascade that results in gene expression. Prenylation of the GTPases is a critical step for them to translocate into cellular membrane. As other groups noted, GGPP only partially restores the translocation of RhoA, but it can completely prevent lovastatin-induced apoptosis, suggesting that this event is unrelated to inhibition of membrane translocation of these GTPases (43, 44). Furthermore, inhibition of protein synthesis by cycloheximide, which does not have any effect on the prenylation of GTPases, efficiently blocks lovastatin-induced macrophage apoptosis. This finding suggests that the induction of cell apoptosis by lovastatin is not due to the depletion of isoprenoids, but that other pathways are involved.

The role of Rac1 and Cdc42 in cell apoptosis has been demonstrated, although the molecular mechanism remains elusive. Both Rac1 and Cdc42 can activate the JNK pathway, resulting in cell apoptosis (34, 35). Our data are consistent with these observations. In our experiments, lovastatin induced the expression of Rac1 and Cdc42 along with the phosphorylation of c-Jun and PARP cleavage (Fig. 5, A and B). GGPP and FPP prevent cells from death through suppressing the expression of Rac1 and Cdc42 (Fig. 5C). The basal level of Rac1 and Cdc42 in Raw cells is very low in comparison with Ras and RhoA. Interestingly, lovastatin selectively induces the expression of Rac1 and Cdc42, but not Ras (data not shown) and RhoA, in the cells and subsequently triggers the JNK pathway, resulting in cell apoptosis. It is conceivable that lovastatin may activate a specific pathway that is responsible for the expression of Rac1 and Cdc42 in macrophages. The specific pathway involved has not been determined, but is under investigation in our laboratory.

A major function of JNKs is to regulate the AP-1 transcription factor through phosphorylation of c-Jun and related proteins. JNKs play a dual role in apoptotic signaling. In some cell types, activation of JNK/AP-1 prevents cells death (45). In other cells, JNKs regulate the activities of pre-existing Bcl-2 family proteins that mediate mitochondrial release of cytochrome c, resulting in caspase activation (46). For example, UV-stimulated proteolytic activation of Bid, a proapoptotic BH3-only Bcl-2-related protein, is suppressed in similarly treated JNK1/2–/– mouse embryonic fibroblasts. Furthermore, JNK activation failed to induce apoptosis in cells deficient in the proapoptotic proteins, Bax and Bak (47). The effect of lovastatin on the activity of Bcl-2 family members in macrophages is the subject of future investigations in our laboratory.

Statin therapy exerts beneficial pleiotropic effects on a wide range of cardiovascular disorders, including delaying the progression of atherosclerosis independent of serum cholesterol levels in patients. Our present findings provide a better understanding of the molecular basis underlying the clinical benefits of statin therapy in cardiovascular diseases and its role in anti-inflammation. Furthermore, these results could have implication in the design of novel therapeutic strategies for other inflammatory diseases, such as cancer, diabetes, and Alzheimer’s disease.


   Acknowledgments
 
We thank Drs. Robert H. Silverman, Bret A. Hassel, and David Quirk for critical reading of the manuscript, and Sudha Gundugola for technical support.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 Address correspondence and reprint requests to Dr. Aimin Zhou, Clinical Chemistry Program, Department of Chemistry, SI 424, Cleveland State University, Cleveland, OH 44115. E-mail address: a.zhou{at}csuohio.edu Back

2 Abbreviations used in this paper: FPP, farnesylpyrophosphate; BMM, bone marrow-derived macrophage; GGPP, geranylgeranylpyrophosphate; PARP, poly(ADP-ribose) polymerase. Back

Received for publication March 28, 2005. Accepted for publication April 7, 2006.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Caslake, M. J., C. J. Packard. 2004. Phenotypes, genotypes and response to statin therapy. Curr. Opin. Lipidol. 15: 387-392. [Medline]
  2. Stein, E. A.. 2003. The power of statins: aggressive lipid lowering. Clin. Cardiol. 26: (Suppl. 3):III25-III31. [Medline]
  3. Schmitz, G., W. Drobnik. 2003. Pharmacogenomics and pharmacogenetics of cholesterol-lowering therapy. Clin. Chem. Lab. Med. 41: 581-589. [Medline]
  4. Amarenco, P., P. Lavallee, P. J. Touboul. 2004. Stroke prevention, blood cholesterol, and statins. Lancet Neurol. 3: 271-278. [Medline]
  5. Nixon, J. V.. 2004. Cholesterol management and the reduction of cardiovascular risk. Prev. Cardiol. 7: 34-39. [Medline]
  6. Amarenco, P., P. Lavallee, P. J. Touboul. 2004. Statins and stroke prevention. Cerebrovasc. Dis. 1: 81-88.
  7. Tsiara, S., M. Elisaf, D. P. Mikhailidis. 2003. Early vascular benefits of statin therapy. Curr. Med. Res. Opin. 19: 540-556. [Medline]
  8. Vaughan, C. J.. 2003. Prevention of stroke and dementia with statins: effects beyond lipid lowering. Am. J. Cardiol. 91: 23B-29B. [Medline]
  9. Abbruzzese, T. A., J. Havens, J. M. Belkin, M. C. Donaldson, A. D. Whittemore, J. K. Liao, M. S. Conte. 2004. Statin therapy is associated with improved patency of autogenous infrainguinal bypass grafts. J. Vasc. Surg. 39: 1178-1185. [Medline]
  10. Napoli, C., V. Sica. 2004. Statin treatment and the natural history of atherosclerotic-related diseases: pathogenic mechanisms and the risk-benefit profile. Curr. Pharm. Des. 10: 425-432. [Medline]
  11. Ganesh, S. K., C. M. Nass, R. S. Blumenthal. 2003. Anti-atherosclerotic effects of statins: lessons from prevention trials. J. Cardiovasc. Risk 10: 155-159. [Medline]
  12. Kwak, B. R., F. Mulhaupt, F. Mach. 2003. Atherosclerosis: anti-inflammatory and immunomodulatory activities of statins. Autoimmun. Rev. 2: 332-338. [Medline]
  13. Halcox, J. P., J. E. Deanfield. 2004. Beyond the laboratory: clinical implications for statin pleiotropy. Circulation 109: (Suppl. 1):II42-48. [Medline]
  14. Marz, W., W Koenig. 2003. HMG-CoA reductase inhibition: anti-inflammatory effects beyond lipid lowering?. J. Cardiovasc. Risk 10: 169-179. [Medline]
  15. Langer, A., C. Constance, J. G. Fodor, J. J. Frohlich, J. Gregoire, D. C. Lau, L. A. Leiter, G. B. Mancini, D. Marr, R. McPherson, et al 2003. Statin therapy and the management of acute coronary syndromes. Can. J. Cardiol. 19: 921-927. [Medline]
  16. Mach, F.. 2003. Statins as novel immunomodulators: from cell to potential clinical benefit. Thromb. Haemostasis 90: 607-610. [Medline]
  17. Blake, G. J.. 2003. Inflammatory biomarkers of the patient with myocardial insufficiency. Curr. Opin. Crit. Care 9: 369-374. [Medline]
  18. Stuve, O., S. Youssef, L. Steinman, S. S. Zamvil. 2003. Statins as potential therapeutic agents in neuroinflammatory disorders. Curr. Opin. Neurol. 16: 393-401. [Medline]
  19. Crisby, M.. 2003. Modulation of the inflammatory process by statins. Drugs Today 39: 137-143. [Medline]
  20. Callahan, A. S., III. 2003. Vascular pleiotropy of statins: clinical evidence and biochemical mechanisms. Curr. Atheroscler. Rep. 5: 33-37. [Medline]
  21. Schonbeck, U., P. Libby. 2004. Inflammation, immunity, and HMG-CoA reductase inhibitors: statins as antiinflammatory agents?. Circulation 109: (Suppl. 1):II18-II26. [Medline]
  22. Fenton, J. W., II, W. P. Jeske, J. L. Catalfamo, D. V. Brezniak, D. G. Moon, G. X. Shen. 2002. Statin drugs and dietary isoprenoids down-regulate protein prenylation in signal transduction and are antithrombotic and prothrombolytic agents. Biochemistry 67: 85-91.
  23. Zhang, F. L., P. J. Casey. 1996. Protein prenylation: molecular mechanisms and functional consequences. Annu. Rev. Biochem. 65: 241-269. [Medline]
  24. Hill, C. S., J. Wynne, R. Treisman. 1995. The Rho family GTPases RhoA, Rac1, and CDC42Hs regulate transcriptional activation by SRF. Cell 81: 1159-1170. [Medline]
  25. Diebold, B. A., B. Fowler, J. Lu, M. C. Dinauer, G. M. Bokoch. 2004. Antagonistic cross-talk between Rac and Cdc42 GTPases regulates generation of reactive oxygen species. J. Biol. Chem. 279: 28136-28142. [Abstract/Free Full Text]
  26. Bokoch, G. M., B. A. Diebold. 2002. Current molecular models for NADPH oxidase regulation by Rac GTPase. Blood 100: 2692-2696. [Abstract/Free Full Text]
  27. Rul, W., O. Zugasti, P. Roux, C. Peyssonnaux, A. Eychene, T. F. Franke, P. Lenormand, P. Fort, U. Hibner. 2002. Activation of ERK, controlled by Rac1 and Cdc42 via Akt, is required for anoikis. Ann. NY Acad. Sci. 973: 145-148. [Medline]
  28. Frost, J. A., H. Steen, P. Shapiro, T. Lewis, N. Ahn, P. E. Shaw, M. H. Cobb. 1997. Cross-cascade activation of ERKs and ternary complex factors by Rho family proteins. EMBO J. 16: 6426-6438. [Medline]
  29. Clerk, A., F. H. Pham, S. J. Fuller, E. Sahai, K. Aktories, R. Marais, C. Marshall, P. H. Sugden. 2001. Regulation of mitogen-activated protein kinases in cardiac myocytes through the small G protein Rac1. Mol. Cell. Biol. 21: 1173-1184. [Abstract/Free Full Text]
  30. Keely, P. J., J. K. Westwick, I. P. Whitehead, C. J. Der, L. V. Parise. 1997. Cdc42 and Rac1 induce integrin-mediated cell motility and invasiveness through PI(3)K. Nature 390: 632-636. [Medline]
  31. Aznar, S., J. C. Lacal. 2001. Rho signals to cell growth and apoptosis. Cancer Lett. 165: 1-10. [Medline]
  32. Chuang, T. H., K. M. Hahn, J. D. Lee, D. E. Danley, G. M. Bokoch. 1997. The small GTPase Cdc42 initiates an apoptotic signaling pathway in Jurkat T lymphocytes. Mol. Biol. Cell 8: 1687-1698. [Abstract]
  33. Subauste, M. C., M. Von Herrath, V. Benard, C. E. Chamberlain, T. H. Chuang, K. Chu, G. M. Bokoch, K. M. Hahn. 2000. Rho family proteins modulate rapid apoptosis induced by cytotoxic T lymphocytes and Fas. J. Biol. Chem. 275: 9725-9733. [Abstract/Free Full Text]
  34. Minden, A., A. Lin, F. X. Claret, A. Abo, M. Karin. 1995. Selective activation of the JNK signaling cascade and c-Jun transcriptional activity by the small GTPases Rac and Cdc42Hs. Cell 81: 1147-1157. [Medline]
  35. Coso, O. A., M. Chiariello, J. C. Yu, H. Teramoto, P. Crespo, N. Xu, T. Miki, J. S. Gutkind. 1995. The small GTP-binding proteins Rac1 and Cdc42 regulate the activity of the JNK/SAPK signaling pathway. Cell 81: 1137-1146. [Medline]
  36. Teramoto, H., O. A. Coso, H. Miyata, T. Igishi, T. Miki, J. S. Gutkind. 1996. Signaling from the small GTP-binding proteins Rac1 and Cdc42 to the c-Jun N-terminal kinase/stress-activated protein kinase pathway: a role for mixed lineage kinase 3/protein-tyrosine kinase 1, a novel member of the mixed lineage kinase family. J. Biol. Chem. 271: 27225-27228. [Abstract/Free Full Text]
  37. Jakobisiak, M., S. Bruno, J. S. Skierski, Z. Darzynkiewicz. 1991. Cell cycle-specific effects of lovastatin. Proc. Natl. Acad. Sci. USA 88: 3628-3632. [Abstract/Free Full Text]
  38. Van de Donk, N. W., M. M. Kamphuis, B. van Kessel, H. M. Lokhorst, A. C. Bloem. 2003. Inhibition of protein geranylgeranylation induces apoptosis in myeloma plasma cells by reducing Mcl-1 protein levels. Blood 102: 3354-3362. [Abstract/Free Full Text]
  39. Erl, W., M. Hristov, M. Neureuter, Z. Q. Yan, G. K. Hansson, P. C. Weber. 2003. HMG-CoA reductase inhibitors induce apoptosis in neointima-derived vascular smooth muscle cells. Atherosclerosis 169: 251-258. [Medline]
  40. Kaneta, S., K. Satoh, S. Kano, M. Kanda, K. Ichihara. 2003. All hydrophobic HMG-CoA reductase inhibitors induce apoptotic death in rat pulmonary vein endothelial cells. Atherosclerosis 170: 237-243. [Medline]
  41. Jiang, Z., X. Zheng, R. A. Lytle, R. Higashikubo, K. M. Rich. 2004. Lovastatin-induced up-regulation of the BH3-only protein, Bim, and cell death in glioblastoma cells. J. Neurochem. 89: 168-178. [Medline]
  42. Goldstein, J. L., M. S. Brown. 1990. Regulation of the mevalonate pathway. Nature 343: 425-430. [Medline]
  43. Zhong, W. B., C. Y. Wang, T. C. Chang, W. S. Lee. 2003. Lovastatin induces apoptosis of anaplastic thyroid cancer cells via inhibition of protein geranylgeranylation and de novo protein synthesis. Endocrinology 144: 3852-3859. [Abstract/Free Full Text]
  44. Agarwal, B., B. Halmos, A. S. Feoktistov, P. Protiva, W. G. Ramey, M. Chen, C. Pothoulakis, J. T. Lamont, P. R. Holt. 2002. Mechanism of lovastatin-induced apoptosis in intestinal epithelial cells. Carcinogenesis 23: 521-528. [Abstract/Free Full Text]
  45. Huang, Y., J. C. Keen, E. Hager, R. Smith, A. Hacker, B. Frydman, A. L. Valasinas, V. K. Reddy, L. J. Marton, R. A. Casero, Jr, N. E. Davidson. 2004. Regulation of polyamine analogue cytotoxicity by c-Jun in human MDA-MB-435 cancer cells. Mol. Cancer Res. 2: 81-88. [Abstract/Free Full Text]
  46. Tournier, C., P. Hess, D. D. Yang, J. Xu, T. K. Turner, A. Nimnual, D. Bar-Sagi, S. N. Jones, R. A. Flavell, R. J. Davis. 2000. Requirement of JNK for stress-induced activation of the cytochrome c-mediated death pathway. Science 288: 870-874. [Abstract/Free Full Text]
  47. Lei, K., A. Nimnual, W. X. Zong, N. J. Kennedy, R. A. Flavell, C. B. Thompson, D. Bar-Sagi, R. J. Davis. 2002. The Bax subfamily of Bcl2-related proteins is essential for apoptotic signal transduction by c-Jun NH2-terminal kinase. Mol. Cell. Biol. 22: 4929-4942. [Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Mol. Pharmacol.Home page
A. Kromer and B. Moosmann
Statin-Induced Liver Injury Involves Cross-Talk between Cholesterol and Selenoprotein Biosynthetic Pathways
Mol. Pharmacol., June 1, 2009; 75(6): 1421 - 1429.
[Abstract] [Full Text] [PDF]

Home page
 Nephrol Dial TransplantHome page
F. C. Cormack-Aboud, P. T. Brinkkoetter, J. W. Pippin, S. J. Shankland, and R. V. Durvasula
Rosuvastatin protects against podocyte apoptosis in vitro
Nephrol. Dial. Transplant., February 1, 2009; 24(2): 404 - 412.
[Abstract] [Full Text] [PDF]

Home page
 Cancer Res.Home page
A. Sassano, E. Katsoulidis, G. Antico, J. K. Altman, A. J. Redig, S. Minucci, M. S. Tallman, and L. C. Platanias
Suppressive Effects of Statins on Acute Promyelocytic Leukemia Cells
Cancer Res., May 1, 2007; 67(9): 4524 - 4532.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Liang, S.-L.
Right arrow Articles by Zhou, A.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Liang, S.-L.
Right arrow Articles by Zhou, A.
Right arrowPubmed/NCBI databases
*Compound via MeSH
*Substance via MeSH
Hazardous Substances DB
*LOVASTATIN

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS