Parthenolide inhibits proliferation of vascular smooth muscle cells through induction of G₀/G₁ phase cell cycle arrest

Parthenolide inhibits proliferation of vascular smooth muscle cells through induction of G 0 /G 1 phase cell cycle arrest 1 first-author †E-mail:wengshaox@sina.com †E-mail:wengshaox@sina.com wengshaox@sina.com 2 3 1 1 1 1 1 1 Department of Cardiology, the Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310009, China Department of Cardiology, the Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou 310009, China Department of Chemical and Biochemical Engineering, Zhejiang University, Hangzhou 310027, China Department of Chemical and Biochemical Engineering, Zhejiang University, Hangzhou 310027, China Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC 29425, USA Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC 29425, USA Objective: 2 This study is to determine the effect of the natural product parthenolide, a sesquiterpene lactone isolated from extracts of the herb Tanacetum parthenium , on the proliferation of vascular smooth muscle cells (VSMCs). Methods: 2 Rat aortic VSMCs were isolated and cultured in vitro, and treated with different concentrations of parthenolide (10, 20 and 30 μmol/L). [ 3 H]thymidine incorporation was used as an index of cell proliferation. Cell cycle progression and distribution were determined by flow cytometric analysis. Furthermore, the expression of several regulatory proteins relevant to VSMC proliferation including IκBα, cyclooxygenase-2 (Cox-2), p21, and p27 was examined to investigate the potential molecular mechanism. Results: 2 Treatment with parthenolide significantly decreased the [ 3 H]thymidine incorporation into DNA by 30%~56% relative to control values in a dose-dependent manner ( P < 0.05). Addition of parthenolide also increased cell population at G 0 /G 1 phase by 19.2%~65.7% ( P < 0.05) and decreased cell population at S phase by 50.7%~84.8% ( P < 0.05), which is consistent with its stimulatory effects on p21 and p27. In addition, parthenolide also increased IκBα expression and reduced Cox-2 expression in a time-dependent manner. Conclusion: 2 Our results show that parthenolide significantly inhibits the VSMC proliferation by inducing G 0 /G 1 cell cycle arrest. IκBα and Cox-2 are likely involved in such inhibitory effect of parthenolide on VSMC proliferation. These findings warrant further investigation on potential therapeutic implications of parthenolide on VSMC proliferation in vivo. Parthenolide Vascular smooth muscle cell (VSMC) Cell proliferation IκBα Cyclooxygenase-2 (Cox-2) p21 INTRODUCTION 1 The proliferation of vascular smooth muscle cells (VSMCs) is a central event during the pathogenesis of vascular lesions, including postangioplasty restenosis, transplant arteriosclerosis and vein graft occlusion (Rivard and Andres, 2000 2000 ; Hedin et al., 2004 2004 ). Identification of novel drugs that may inhibit the proliferation of VSMCs has become an area of great interest in both research and clinical settings (Spyridopoulos and Andres, 1998 1998 ; Weissberg et al., 1993 1993 ). The natural product parthenolide, a sesquiterpene lactone isolated from extracts of the Mexican-Indian medicinal herb Tanacetum parthenium , has been commonly used for the treatment of various infectious and inflammatory conditions such as arthritis, asthma and migraines for many years (Heinrich et al., 1998 1998 ). Furthermore, an increasing number of studies have indicated that parthenolide could inhibit tumor growth (Oka et al., 2007 2007 ; Ross et al., 1999 1999 ), exert beneficial effects in myocardial reperfusion injury (Zingarelli et al., 2002 2002 ), and interfere with a number of cellular processes including oxidative phosphorylation, platelet aggregation, and histamine and serotonin release, as well as neutrophil chemotaxis (Groenewegen and Heptinstall, 1990 1990 ; Sheehan et al., 2002 2002 ). Parthenolide has also been found to inhibit activation of transcription factor nuclear factor-κB (NF-κB) (Zingarelli et al., 2002 2002 ; Sheehan et al., 2002 2002 ; Hehner et al., 1998 1998 ; 1999 1999 ). In this study, we determined the effects of parthenolide on the proliferation and cell cycle dynamics of rat aortic VSMCs. Moreover, the regulatory effects of parthenolide on p21 and p27 were also examined, since p21 and p27 are two important members of Cip/Kip family that negatively regulates G 1 /S phase and inhibits cell proliferation (Maddika et al., 2007 2007 ; Hengst and Reed, 1998 1998 ). Previous studies have shown that alpha inhibitor of nuclear factor-κB (IκBα) may mediate the anti-inflammatory effect of parthenolide (Zingarelli et al., 2002 2002 ; Hehner et al., 1998 1998 ; 1999 1999 ) and that cyclooxygenase-2 (Cox-2) is regulated by NF-κB (Dubois et al., 1998 1998 ; Moon et al., 2004 2004 ). Both IκBα and Cox-2 may play roles in VSMC proliferation (Young et al., 2000 2000 ; Tzeng et al., 2007 2007 ); therefore, the effects of parthenolide on these two molecules were also investigated. MATERIALS AND METHODS 1 Cell culture 2 Rat aortic smooth muscle cells were isolated from the thoracic aorta of 2- to 3-month-old Harlan Sprague-Dawley rats by using the explant technique according to a modified method (Thyberg et al., 1983 1983 ). Cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS) and 1% (v/v) penicillin/streptomycin. The purity of VSMCs was evaluated by morphology and immunocytochemistry staining with monoclonal antibodies against α-smooth muscle actin (SM-α actin). Prior to the experimental treatments, cells were grown to ~80% confluency followed by incubation with serum-free medium for 24 h to make cells quiescent. VSMCs from passages 4~6 were used in this study. Reagents 2 DMEM, FBS and penicillin/streptomycin were obtained from Gibco Life Technologies Inc. (Rockville, MD, USA). Parthenolide was purchased from Jingmei Biotech. Co. Ltd. (Shenzhen, China). [ 3 H]thymidine was supplied by Chinese Academy of Sciences Shanghai Atomic Institute (Shanghai, China). Polyvinylidine difluoride transfer membranes were purchased from Boehringer Mannheim Corp. (Indianapolis, IN, USA). Anti-IκBα, Cox-2, p21, p27 and β-actin antibodies, as well as enhanced chemiluminescence (ECL) kit were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). SM-α actin monoclonal antibody and horseradish peroxidase conjugated goat anti-rabbit and rabbit anti-mouse secondary antibodies were purchased from Beijing Chemical Corp. (Beijing, China). All other chemicals used in this study were purchased from Sigma (St. Louis, MO, USA). Determination of DNA synthesis 2 [ 3 H]thymidine incorporation into DNA synthesis was used as an index of cell proliferation (Nilsson et al., 1983 1983 ). Cells were seeded in 24-well plates and grown to ~80% confluency. After serum starvation for 24 h, cells were treated with parthenolide at different concentrations (10, 20, and 30 μmol/L) in DMEM containing 10% FBS for 24 h. 1 μCi/ml [ 3 H]thymidine was coincubated with the cells for the last 8 h of treatment. Incubations were terminated by removing the culture medium and washing the cells three times with cold phosphate buffer saline (PBS), followed by the addition of ice-cold 10% (w/v) trichloroacetic acid for 20 min at 4 °C to remove the unincorporated [ 3 H]thymidine and protein precipitation. Next, the cells were washed three times with ice-cold water, solubilized with 1 ml of 0.5 mol/L NaOH, and incubated for 30 min at 37 °C. After solubilization, 0.5 ml/well aliquot samples were removed, transferred to scintillation vials with 10 ml of scintillation cocktail. The radioactivity of incorporated [ 3 H]thymidine was quantified by a liquid scintillation counter (Beckman Coulter Inc., Fullerton, CA, USA). Three independent experiments were performed with triplicate samples for each group in each experiment. Flow cytometric analysis 2 Cell cycle progression and distribution were determined by flow cytometric analysis. After serum starvation for 24 h, cells were stimulated with different concentrations of parthenolide (10, 20 and 30 μmol/L) in medium supplemented with 10% FBS for 24 h. At the end of time-point, all the cells were harvested by trypsinization, washed with ice-cold PBS, and fixed in 70% ice-cold ethanol diluted in PBS. Around 1×10 5 VSMCs were incubated in PBS containing 100 g/ml RNase and 40 g/ml propidium iodide at room temperature for 0.5~1.0 h before flow cytometry analysis. Cell cycle distribution and DNA content were determined by using a Berkman Coulter cytometer (Fullerton, CA, USA). Western blots 2 After VSMCs were stimulated with 20 μmol/L of parthenolide in medium containing 10% FBS for 2, 6, 12, and 24 h, cells were harvested and cellular protein was isolated with a protein extraction buffer. Protein concentrations were determined using Bio-Rad DC Protein Assay (Bio-Rad Laboratories, Milan, Italy). Equal amounts (30 g/lane) of proteins were fractionated on 10%~12% (w/v) sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) gels and transferred to polyvinylidene difluoride membranes. The membranes were incubated with anti-IκBα, Cox-2, p21 and p27 primary antibodies (all in 1:400 dilution), respectively, overnight or with anti-actin (1:1000) for 1 h. After washing with PBS containing 0.1% (v/v) Tween 20, the membranes were incubated with corresponding horseradish peroxidase-conjugated anti-rabbit or anti-mouse secondary antibodies (1:1000) for 1 h, followed by exposure using enhanced chemiluminescent reagent. The images of Western blots were scanned and quantitated using the software Quantity One (Bio-Rad Laboratories). Results were normalized by the respective level of β-actin. Statistical analysis 2 Data are presented as mean± SD of three independent experiments. Student’s t -test was used to determine the statistical difference between various experimental and control groups. Differences were considered statistically significant at a level of P < 0.05. RESULTS 1 Identification and characterization of VSMCs 2 VSMCs isolated by using the explant method in this study grew in the characteristic hill and valley pattern. As shown in Fig. 1b 1b , the purity of the VSMCs was determined by immunostaining for monoclonal anti-SM-α-actin. Primary cultures with less than 95% VSMCs were discarded. Cellular viability was determined by cell counting, morphology examinations, and trypan blue exclusion, and the concentrations of parthenolide (10, 20, and 30 μmol/L) used in this study were not toxic to VSMCs. Effects of parthenolide on DNA synthesis in VSMCs 2 First, we determined the effect of parthenolide on VSMC proliferation assessed by determination of [ 3 H]thymidine incorporation. As indicated in Fig. 2 2 , the incubation of 10, 20 and 30 μmol/L of parthenolide decreased the [ 3 H]thymidine incorporation into DNA by 30.3%, 35.93%, and 56.1%, respectively, relative to respective control levels ( P < 0.05 or P < 0.01). Thus, the treatment of parthenolide resulted in significant and dose-dependent inhibition on DNA synthesis in VSMCs cultured under 10% FBS. VSMCs were serum-starved for 24 h to arrest the cells at G 0 /G 1 phase. Then cells were stimulated with different concentrations of parthenolide (0, 10, 20 and 30 μmol/L) in DMEM containing 10% FBS for 16 h, followed by pulsing cells with 1 μCi/ml [ 3 H]thymidine for an additional 8 h. The radioactivity of incorporated [ 3 H]thymidine was quantified as counts per minute (cpm) by a liquid scintillation counter. Data are the representative of three independent experiments with similar results and are expressed as mean± SD ( n =3). * P < 0.05 and ** P < 0.01, compared with the corresponding control group Effects of parthenolide on cell cycle progression in VSMCs 2 Next, we determined the effect of parthenolide on cell cycle progression in VSMCs by flow cytometry. Treatment with 10, 20, and 30 μmol/L parthenolide significantly increased the cell population at G 0 /G 1 phase by 19.2%, 45.3%, and 65.7%, respectively, and decreased the cell population at S phase by 50.7%, 76.2%, and 84.8%, respectively ( P < 0.05 or P < 0.01 compared with the control group) (Fig. 3 3 and Table 1 1 ). These results indicate that parthenolide induces cell cycle arrest at G 0 /G 1 phase and decreases the cell population at S phase, which is consistent with the finding that treatment of parthenolide inhibits [ 3 H]thymidine from being incorporated into DNA in VSMCs. After serum starvation, VSMCs were stimulated with different concentrations of parthenolide ((a) 0, (b) 10, (c) 20, and (d) 30 μmol/L) in medium supplemented with 10% FBS for 24 h. Cells were harvested and DNA contents were stained with propidium iodide for flow cytometric analysis. The cell populations corresponding to G 0 /G 1 , G 2 /M and S phases of the cell cycle are summarized in Table 1 1 Data are the representative of three independent experiments with similar results and are expressed as mean± SD ( n =3). P < 0.05, compared with the corresponding control group P < 0.01, compared with the corresponding control group 2 Parthenolide (μmol/L) 3 Cell cycle (%) G 0 /G 1 S G 2 /M 0 49.5±0.8 48.7±3.3 1.9±3.3 10 59.0±2.4 * * 24.0±6.5 * * 16.1±8.4 20 71.9±5.4 * * 11.6±8.8 * * 16.5±9.8 30 82.0±1.7 ** ** 7.4±0.4 ** ** 10.4±1.2 Effects of parthenolide on expression of IκBα, Cox-2, p21 and p27 proteins 2 To investigate the potential molecular mechanisms that may be involved in the inhibitory effect of parthenolide on VSMCs, we examined several regulatory proteins that are associated with cell proliferation, differentiation, and cell cycle progression, including IκBα, Cox-2, p21 and p27. As shown in Fig. 4 4 , 10% FBS induced degradation of IκBα in the control group at 12 and 24 h of incubation. However, the treatment with 20 μmol/L parthenolide completely blocked the serum-induced IκBα degradation at 12 and 24 h incubation ( P < 0.01). In contrast, 10% FBS alone induced up-regulation of Cox-2, particularly, at 6 h of incubation, but the presence of parthenolide signifycantly reduced the protein levels of Cox-2 at 2, 6, and 12 h of incubation ( P < 0.01). Moreover, while there were no remarkable changes on protein levels of p21 and p27 in the control group, the treatment of parthenolide significantly up-regulated both proteins at 12 and 24 h of incubation ( P < 0.01). (a) Cells were cultured in 10% FBS and treated with 20μmol/L of parthenolide after serum starvation. Cellular proteins were extracted from cells after 2, 6, 12 and 24 h of treatment. Equal amounts (30 μg/lane) of cellular proteins were fractionated on 10%~12% SDS-PAGE and transferred to polyvinylidene difluoride membranes, followed by immunoblotting with anti-IκBα, Cox-2, p21 and p27 primary antibodies. β-actin protein was blotted as a control (Lane 1: control 2 h; 2: parthenolide 2 h; 3: control 6 h; 4: parthenolide 6 h; 5: control 12 h; 6: parthenolide 12 h; 7: control 24 h; 8: parthenolide 24 h); (b) The quantification of Western blots was done by densitometry and protein levels were expressed as product of OD and area (mm 2 ). Data are the representative of three independent experiments with similar results and are expressed as mean± SD ( n =3). ** P < 0.01, compared with the corresponding control group DISCUSSION 1 Parthenolide is a sesquiterpene lactone found as the major active component in feverfew ( Tanacetum parthenium ), a herbal medicine that has been used to treat migraine and rheumatoid arthritis for centuries (Heinrich et al., 1998 1998 ; Knight, 1995 1995 ). Recently, it has been reported that parthenolide possesses antitumor activity in a number of cultured cancer cell lines and in xenograft models (Oka et al., 2007 2007 ; Ross et al., 1999 1999 ). Moreover, parthenolide has been suggested to protect against myocardial ischemia and reperfusion injury in rats (Zingarelli et al., 2002 2002 ), inhibit the proliferation of vascular endothelial cells (Parada-Turska et al., 2007 2007 ; Shanmugam et al., 2006 2006 ), and inhibit inducible nitric oxide synthase gene expression in cultured rat aortic smooth muscle cells (Wong and Menendez, 1999 1999 ). Thus, interest has arisen in investigating the potential roles of parthenolide in preventing or treating cardiovascular diseases. In the present study, we evaluated the effects of parthenolide on the proliferation of rat aortic smooth muscle cells. We found that treatment of parthenolide, at non-toxic concentrations between 10~30 μmol/L, significantly suppressed the [ 3 H]thymidine incorporation into DNA by 30.3%~56.1% in a dose-dependent manner (Fig. 2 2 ), and increased the cell population at G 0 /G 1 phase while decreased the cell population at S phase (Fig. 3 3 ). These results suggest that parthenolide inhibits VSMC proliferation by inducing cell cycle arrest at G 0 /G 1 phase. We further found that parthenolide increased IκBα, p21, and p27, while decreased Cox-2 in VSMCs, providing some clues on the mechanisms of inhibitory effects of parthenolide on VSMC proliferation. Cellular proliferation is regulated primarily by regulation of the cell cycle, which consists of four distinct sequential phases (G 0 /G 1 , S, G 2 , and M). As two important members of Cip/Kip family, p21 and p27 negatively regulate G 1 /S phase and inhibit cell proliferation (Maddika et al., 2007 2007 ; Hengst and Reed, 1998 1998 ). Our results indicate that parthenolide significantly increased the expression levels of both p21 and p27 (Fig. 4 4 ), which is consistent with parthenolide-induced inhibition of DNA synthesis and G 0 /G 1 cell cycle arrest in VSMCs. Although the mechanisms mediating the various effects of parthenolide in different diseases are not entirely clear, several studies have shown that an important part of the anti-inflammatory action of this compound appears to be related to its activity in inhibiting the NF-κB signal pathway through preventing the degradation of IκBα. NF-κB is a dimeric transcription factor, and genes activated by NF-κB play central roles in cell differentiation, proliferation, inflammation, and apoptosis (Baeuerle and Henkel, 1994 1994 ; Chen et al., 1999 1999 ). This transcription factor is involved in the pathogenesis of many vascular diseases such as atherosclerosis (Collins and Cybulsky, 2001 2001 ). IκBα is the specific cytoplasmic inhibitory protein of NF-κB, and degradation of IκBα is generally believed to be the critical step for the activation of NF-κB (Brown et al., 1993 1993 ). A recent study indicated that in vascular smooth muscle cells and monocytes stimulated with lipopolysaccharide (LPS), parthenolide reduced IκBα degradation and NF-κB activation (López-Franco et al., 2006 2006 ). By Western blots, we observed that 10% FBS induced IκBα degradation, which may be related to the activation of NF-κB signal pathway and the proliferation of VSMCs stimulated by enriched serum. However, the presence of parthenolide completely antagonized this effect and resulted in upregulation of IκBα protein, which is in agreement with the previous reports showing parthenolide as an inhibitor of NF-κB (Zingarelli et al., 2002 2002 ; Sheehan et al., 2002 2002 ; Hehner et al., 1998 1998 ; 1999 1999 ; López-Franco et al., 2006 2006 ). As an inducible isoform of enzyme, Cox-2 is upregulated in response to inflammatory factors and some peptides (Bishop-Bailey et al., 1999 1999 ). Cox-2 metabolites can regulate a number of cellular processes involved in inflammation and remodeling, including the production of chemokines, matrix metalloproteinase, and angiogenic factors (Bishop-Bailey et al., 1999 1999 ; Marnett et al., 1999 1999 ; Pang et al., 1998 1998 ). Emerging evidence has also shown that Cox-2 enhances VSMC proliferation (Young et al., 2000 2000 ; Tzeng et al., 2007 2007 ). In the present study, we observed that the protein level of Cox-2 was significantly reduced in VSMCs treated with parthenolide for 2~12 h compared with cells cultured with 10% FBS alone (Fig. 4 4 ). Recent studies have reported inhibitory effect of parthenolide on Cox-2 expression in monocytic cells and lipopolysaccharide-stimulated macrophages and in cancers (Gomez-Hernandez et al., 2006 2006 ; Smolinski and Pestka, 2005 2005 ). Since the expression of Cox-2 has been shown to be regulated by NF-κB (Dubois et al., 1998 1998 ; Moon et al., 2004 2004 ), our results may suggest that parthenolide upregulates the expression of IκBα protein and subsequently inhibits NF-κB activity, thus reducing the expression of Cox-2 protein. Our results also indicate that NF-κB/IκBα signaling pathway and Cox-2 are likely involved in the inhibitory effect of parthenolide on VSMC proliferation. In summary, this study has evaluated the effects of parthenolide on proliferation and cell cycle progression in rat aortic VSMCs in vitro. Our results show that the treatment of parthenolide, in a dose-dependent manner, significantly inhibited the proliferation and induced cell cycle arrest at G 0 /G 1 phase in VSMCs, which was supported by the finding that parthenolide significantly increased the protein levels of p21 and p27. Furthermore, parthenolide also significantly increased the protein level of IκBα, while reduced the expression of Cox-2 in a time-dependent manner, suggesting that NF-κB/IκBα signaling pathway may play an important role in this process. Taken together, these findings merit further preclinical studies as well as clinical investigations on the potential therapeutic applications of parthenolide on vascular disorders with overgrowth of VSMCs, such as restenosis and atherosclerosis. References PA Baeuerle , T Henkel . Function and activation of NF-kappaB in the immune system . Annu Rev Immunol , 1994 . 12 ( 1 ): 141 - 179 . 8011280 DOI: 10.1146/annurev.immunol.12.1.141 http://doi.org/10.1146/annurev.immunol.12.1.141 . Function and activation of NF-kappaB in the immune system D Bishop-Bailey , T Hla , JA Mitchell . Cyclo-oxygenase-2 in vascular smooth muscle . Int J Mol Med , 1999 . 3 ( 1 ): 41 - 48 . 9864384 . Cyclo-oxygenase-2 in vascular smooth muscle K Brown , S Park , T Kanno , 等 . Mutual regulation of the transcriptional activator NF-kappa B and its inhibitor, I kappa B-alpha . Proc Natl Acad Sci USA , 1993 . 90 ( 6 ): 2532 - 2536 . DOI: 10.1073/pnas.90.6.2532 http://doi.org/10.1073/pnas.90.6.2532 . Mutual regulation of the transcriptional activator NF-kappa B and its inhibitor, I kappa B-alpha F Chen , V Castranova , X Shi , LM Demers . New insights into the role of nuclear factor-kappaB, a ubiquitous transcription factor in the initiation of diseases . Clin Chem , 1999 . 45 ( 1 ): 7 - 17 . 9895331 . New insights into the role of nuclear factor-kappaB, a ubiquitous transcription factor in the initiation of diseases T Collins , MI Cybulsky . NF-kappaB: pivotal mediator or innocent bystander in atherogenesis? . J Clin Invest , 2001 . 107 ( 3 ): 255 - 264 . 11160146 DOI: 10.1172/JCI10373 http://doi.org/10.1172/JCI10373 . NF-kappaB: pivotal mediator or innocent bystander in atherogenesis? RN Dubois , SB Abramson , L Crofford , RA Gupta , LS Simon , LB van de Putte , PE Lipsky . Cyclooxygenase in biology and disease . FASEB J , 1998 . 12 ( 12 ): 1063 - 1073 . 9737710 . Cyclooxygenase in biology and disease A Gomez-Hernandez , JL Martin-Ventura , E Sanchez-Galan , C Vidal , M Ortego , LM Blanco-Colio , L Ortegan , J Tunon , J Egido . Overexpression of COX-2, prostaglandin E synthase-1 and prostaglandin E receptors in blood mononuclear cells and plaque of patients with carotid atherosclerosis: regulation by nuclear factor-kappaB . Atherosclerosis , 2006 . 187 ( 1 ): 139 - 149 . 16212965 DOI: 10.1016/j.atherosclerosis.2005.08.035 http://doi.org/10.1016/j.atherosclerosis.2005.08.035 . Overexpression of COX-2, prostaglandin E synthase-1 and prostaglandin E receptors in blood mononuclear cells and plaque of patients with carotid atherosclerosis: regulation by nuclear factor-kappaB WA Groenewegen , S Heptinstall . A comparison of the effects of an extract of feverfew and parthenolide, a component of feverfew, on human platelet activity in-vitro . J Pharm Pharmacol , 1990 . 42 ( 8 ): 553 - 557 . 1981582 . A comparison of the effects of an extract of feverfew and parthenolide, a component of feverfew, on human platelet activity in-vitro U Hedin , J Roy , PK Tran . Control of smooth muscle cell proliferation in vascular disease . Curr Opin Lipidol , 2004 . 15 ( 5 ): 559 - 565 . 15361792 DOI: 10.1097/00041433-200410000-00010 http://doi.org/10.1097/00041433-200410000-00010 . Control of smooth muscle cell proliferation in vascular disease SP Hehner , M Heinrich , PM Bork , M Vogt , F Ratter , V Lehmann , K Schulze-Osthoff , W Droge , ML Schmitz . Sesquiterpene lactones specifically inhibit activation of NF-kappaB by preventing the degradation of I kappa B-alpha and I kappa B-beta . J Biol Chem , 1998 . 273 ( 3 ): 1288 - 1297 . 9430659 DOI: 10.1074/jbc.273.3.1288 http://doi.org/10.1074/jbc.273.3.1288 . Sesquiterpene lactones specifically inhibit activation of NF-kappaB by preventing the degradation of I kappa B-alpha and I kappa B-beta SP Hehner , TG Hofmann , W Droge , ML Schmitz . The antiinflammatory sesquiterpene lactone parthenolide inhibits NF-kappa B by targeting the I kappa B kinase complex . J Immunol , 1999 . 163 ( 10 ): 5617 - 5623 . 10553091 . The antiinflammatory sesquiterpene lactone parthenolide inhibits NF-kappa B by targeting the I kappa B kinase complex M Heinrich , M Robles , JE West , BR Ortiz de Montellano , E Rodriguez . Ethnopharmacology of Mexican asteraceae (Compositae) . Annu Rev Pharmacol Toxicol , 1998 . 38 ( 1 ): 539 - 565 . 9597165 DOI: 10.1146/annurev.pharmtox.38.1.539 http://doi.org/10.1146/annurev.pharmtox.38.1.539 . Ethnopharmacology of Mexican asteraceae (Compositae) L Hengst , SI Reed . Inhibitors of the Cip/Kip family . Curr Top Microbiol Immunol , 1998 . 227 25 - 41 . 9479824 . Inhibitors of the Cip/Kip family DW Knight . Feverfew: chemistry and biological activity . Nat Prod Rep , 1995 . 12 ( 3 ): 271 - 276 . 7792073 DOI: 10.1039/np9951200271 http://doi.org/10.1039/np9951200271 . Feverfew: chemistry and biological activity O López-Franco , P Hernández-Vargas , G Ortiz-Muñoz , G Sanjuán , Y Suzuki , L Ortega , J Blanco , J Egido , C Gómez-Guerrero . Parthenolide modulates the NF-kappaB-mediated inflammatory responses in experimental atherosclerosis . Arterioscler Thromb Vasc Biol , 2006 . 26 ( 8 ): 1864 - 1870 . 16741149 DOI: 10.1161/01.ATV.0000229659.94020.53 http://doi.org/10.1161/01.ATV.0000229659.94020.53 . Parthenolide modulates the NF-kappaB-mediated inflammatory responses in experimental atherosclerosis S Maddika , SR Ande , S Panigrahi , T Paranjothy , K Weglarczyk , A Zuse , M Eshraghi , KD Manda , E Wiechec , M Los . Cell survival, cell death and cell cycle pathways are interconnected: implications for cancer therapy . Drug Resist Update , 2007 . 10 ( 1 ): 13 - 29 . DOI: 10.1016/j.drup.2007.01.003 http://doi.org/10.1016/j.drup.2007.01.003 . Cell survival, cell death and cell cycle pathways are interconnected: implications for cancer therapy LJ Marnett , SW Rowlinson , DC Goodwin , AS Kalgutkar , CA Lanzo . Arachidonic acid oxygenation by COX-1 and COX-2. Mechanisms of catalysis and inhibition . J Biol Chem , 1999 . 274 ( 33 ): 22903 - 22906 . 10438452 DOI: 10.1074/jbc.274.33.22903 http://doi.org/10.1074/jbc.274.33.22903 . Arachidonic acid oxygenation by COX-1 and COX-2. Mechanisms of catalysis and inhibition SK Moon , BY Cha , CH Kim . ERK1/2 mediates TNF-alpha-induced matrix metalloproteinase-9 expression in human vascular smooth muscle cells via the regulation of NF-kappaB and AP-1: involvement of the ras dependent pathway . J Cell Physiol , 2004 . 198 ( 3 ): 417 - 427 . 14755547 DOI: 10.1002/jcp.10435 http://doi.org/10.1002/jcp.10435 . ERK1/2 mediates TNF-alpha-induced matrix metalloproteinase-9 expression in human vascular smooth muscle cells via the regulation of NF-kappaB and AP-1: involvement of the ras dependent pathway J Nilsson , T Ksiazek , J Thyberg . Effects of colchicine on DNA synthesis, endocytosis and fine structure of cultivated arterial smooth muscle cells . Exp Cell Res , 1983 . 143 ( 2 ): 367 - 375 . 6832223 DOI: 10.1016/0014-4827(83)90063-0 http://doi.org/10.1016/0014-4827(83)90063-0 . Effects of colchicine on DNA synthesis, endocytosis and fine structure of cultivated arterial smooth muscle cells D Oka , K Nishimura , M Shiba , Y Nakai , Y Arai , M Nakayama , H Takayama , H Inoue , A Okuyama , N Nonomura . Sesquiterpene lactone parthenolide suppresses tumor growth in a xenograft model of renal cell carcinoma by inhibiting the activation of NF-kappaB . Int J Cancer , 2007 . 120 ( 12 ): 2576 - 2581 . 17290398 DOI: 10.1002/ijc.22570 http://doi.org/10.1002/ijc.22570 . Sesquiterpene lactone parthenolide suppresses tumor growth in a xenograft model of renal cell carcinoma by inhibiting the activation of NF-kappaB L Pang , E Holland , AJ Knox . Role of cyclo-oxygenase-2 induction in interleukin-1beta induced attenuation of cultured human airway smooth muscle cell cyclic AMP generation in response to isoprenaline . Br J Pharmacol , 1998 . 125 ( 6 ): 1320 - 1328 . 9863663 DOI: 10.1038/sj.bjp.0702193 http://doi.org/10.1038/sj.bjp.0702193 . Role of cyclo-oxygenase-2 induction in interleukin-1beta induced attenuation of cultured human airway smooth muscle cell cyclic AMP generation in response to isoprenaline J Parada-Turska , R Paduch , M Majdan , M Kandefer-Szerszen , W Rzeski . Antiproliferative activity of parthenolide against three human cancer cell lines and human umbilical vein endothelial cells . Pharmacol Rep , 2007 . 59 ( 2 ): 233 - 237 . 17556802 . Antiproliferative activity of parthenolide against three human cancer cell lines and human umbilical vein endothelial cells A Rivard , V Andres . Vascular smooth muscle cell proliferation in the pathogenesis of atherosclerotic cardiovascular diseases . Histol Histopathol , 2000 . 15 ( 2 ): 557 - 571 . 10809377 . Vascular smooth muscle cell proliferation in the pathogenesis of atherosclerotic cardiovascular diseases JJ Ross , JT Arnason , HC Birnboim . Low concentrations of the feverfew component parthenolide inhibit in vitro growth of tumor lines in a cytostatic fashion . Planta Med , 1999 . 65 ( 2 ): 126 - 129 . 10193202 DOI: 10.1055/s-1999-13972 http://doi.org/10.1055/s-1999-13972 . Low concentrations of the feverfew component parthenolide inhibit in vitro growth of tumor lines in a cytostatic fashion R Shanmugam , V Jayaprakasan , Y Gokmen-Polar , S Kelich , KD Miller , M Yipschneider , L Cheng , P Bhat-Nakshatri , GWJr Sledge , H Nakshatri , 等 . Restoring chemotherapy and hormone therapy sensitivity by parthenolide in a xenograft hormone refractory prostate cancer model . Prostate , 2006 . 66 ( 14 ): 1498 - 1511 . 16921510 DOI: 10.1002/pros.20482 http://doi.org/10.1002/pros.20482 . Restoring chemotherapy and hormone therapy sensitivity by parthenolide in a xenograft hormone refractory prostate cancer model M Sheehan , HR Wong , PW Hake , V Malhotra , M O′Connor , B Zingarelli . Parthenolide, an inhibitor of the nuclear factor-kappaB pathway, ameliorates cardiovascular derangement and outcome in endotoxic shock in rodents . Mol Pharmacol , 2002 . 61 ( 5 ): 953 - 963 . 11961112 DOI: 10.1124/mol.61.5.953 http://doi.org/10.1124/mol.61.5.953 . Parthenolide, an inhibitor of the nuclear factor-kappaB pathway, ameliorates cardiovascular derangement and outcome in endotoxic shock in rodents AT Smolinski , JJ Pestka . Comparative effects of the herbal constituent parthenolide (Feverfew) on lipopolysaccharide-induced inflammatory gene expression in murine spleen and liver . J Inflamm (Lond) , 2005 . 2 ( 1 ): 6 15987517 DOI: 10.1186/1476-9255-2-6 http://doi.org/10.1186/1476-9255-2-6 . Comparative effects of the herbal constituent parthenolide (Feverfew) on lipopolysaccharide-induced inflammatory gene expression in murine spleen and liver I Spyridopoulos , V Andres . Control of vascular smooth muscle and endothelial cell proliferation and its implication in cardiovascular disease . Front Biosci , 1998 . 3 ( 1 ): d269 - d287 . 9473210 . Control of vascular smooth muscle and endothelial cell proliferation and its implication in cardiovascular disease J Thyberg , L Palmberg , J Nilsson , T Ksiazek , M Sjolund . Phenotype modulation in primary cultures of arterial smooth muscle cells. On the role of platelet-derived growth factor . Differentiation , 1983 . 25 ( 2 ): 156 - 167 . 6686563 . Phenotype modulation in primary cultures of arterial smooth muscle cells. On the role of platelet-derived growth factor HP Tzeng , RS Yang , TH Ueng , SH Liu . Upregulation of cyclooxygenase-2 by motorcycle exhaust particulate-induced reactive oxygen species enhances rat vascular smooth muscle cell proliferation . Chem Res Toxicol , 2007 . 20 ( 8 ): 1170 - 1176 . 17645304 DOI: 10.1021/tx700084z http://doi.org/10.1021/tx700084z . Upregulation of cyclooxygenase-2 by motorcycle exhaust particulate-induced reactive oxygen species enhances rat vascular smooth muscle cell proliferation PL Weissberg , DJ Grainger , CM Shanahan , JC Metcalfe . Approaches to the development of selective inhibitors of vascular smooth muscle cell proliferation . Cardiovasc Res , 1993 . 27 ( 7 ): 1191 - 1198 . 8252577 DOI: 10.1093/cvr/27.7.1191 http://doi.org/10.1093/cvr/27.7.1191 . Approaches to the development of selective inhibitors of vascular smooth muscle cell proliferation HR Wong , IY Menendez . Sesquiterpene lactones inhibit inducible nitric oxide synthase gene expression in cultured rat aortic smooth muscle cells . Biochem Biophys Res Commun , 1999 . 262 ( 2 ): 375 - 380 . 10462483 DOI: 10.1006/bbrc.1999.1207 http://doi.org/10.1006/bbrc.1999.1207 . Sesquiterpene lactones inhibit inducible nitric oxide synthase gene expression in cultured rat aortic smooth muscle cells W Young , K Mahboubi , A Haider , I Li , NR Ferreri . Cyclooxygenase-2 is required for tumor necrosis factor-alpha- and angiotensin II-mediated proliferation of vascular smooth muscle cells . Circ Res , 2000 . 86 ( 8 ): 906 - 914 . 10785514 . Cyclooxygenase-2 is required for tumor necrosis factor-alpha- and angiotensin II-mediated proliferation of vascular smooth muscle cells B Zingarelli , PW Hake , A Denenberg , HR Wong . Sesquiterpene lactone parthenolide, an inhibitor of IkappaB kinase complex and nuclear factor-kappaB, exerts beneficial effects in myocardial reperfusion injury . Shock , 2002 . 17 ( 2 ): 127 - 134 . 11837788 DOI: 10.1097/00024382-200202000-00008 http://doi.org/10.1097/00024382-200202000-00008 . Sesquiterpene lactone parthenolide, an inhibitor of IkappaB kinase complex and nuclear factor-kappaB, exerts beneficial effects in myocardial reperfusion injury 10.1631/jzus.B0820351 Project (No. 491020-W50315) supported by the Foundation of the Health Bureau of Zhejiang, China 2008-11-02 2009-06-08 2009-03-03 2022-01-05 Copyright © 2009, Journal of Zhejiang University Science 2009 Journal of Zhejiang University-SCIENCE B (Biomedicine and Biotechnology) 07 10