Restoration of MiR-34a Expression by 5-Azacytidine Augments Alimta -Induced Cell Death in Non-Small Lung Cancer Cells by Downregulation of HMG B1, A2 and Bcl-2 Pathway.

OBJECTIVE: Alimta (Pemetrexed) as an antifolate drug has been approved for the treatment of lung cancer. The aim of the present study was to investigate the combination effect of 5-Azacytidine (5-aza) and Alimta on the miR-34a and its target genes expression and induction of apoptotic cell death in non-small lung cancer A549 cells. MATERIALS AND METHODS: In this experimental study, lung cancer A549 cells were treated with various concentrations of Alimta alone and combined with 5-Aza. Then, viability was assessed by trypan blue and MTT assays. mRNA expressions were performed by real time-polymerase chain reaction (PCR) and western blot. Flow cytometry used to detect apoptotic/ necrotic cells and cell cycle arrest. RESULTS: Alimta alone reduced viability of the cells in a dose dependent manner with the half-maximal inhibitory concentration (IC50) value of 12 µM. Pretreatment of the cells with 5-aza (5 µM) induced a synergistic cytotoxic effect with IC50 of 3 µM. Sequential exposure of the cells to 5-aza and Alimta enhanced miR-34a expression and significantly downregulated HMGB1, HMGA2 and BCL-2 expressions. Also, it was associated with reduction of nuclear HMGB1 and HMGA2 content. Caspase-3 activation, HMGB1 release into extracellular space and staining of the cells with annexine V/PI suggested that 5-aza reduced late apoptotic/necrotic cell death induced by Alimta. In addition, combination of 5-aza and Alimta arrested the cells at S and sub-G1 phases and inhibited colony formation. CONCLUSION: 5-aza synergistically enhances Alimta induced apoptotic cell death through HMG proteins regulation, MIR34A gene expression and intrinsic apoptosis mechanism, providing a promising combination therapy in clinical lung cancer therapy.

Protective effect of bleomycin on 5-azacitidine induced cytotoxicity and apoptosis in mice hematopoietic stem cells via Bcl-2/Bax and HMGB1 signaling pathway.

Antineoplastic drugs cause severe cytotoxicity for normal cells, especially hematopoietic stem cells (HSCs). However, bleomycin (BLM) is glycopeptide antibiotic that is effective on various cancers and has either low or no myelosuppression effects. The aim of the present study was to investigate the effect of BLM on 5-Azacitidine (5-AZA) induced cytotoxicity in bone marrow HSCs. 5-AZA reduced HSC cell viability in a time and dose-dependent manner with an IC50 value of 16 µM. However, pretreatment of the cells with BLM for 4 h induced an antagonistic cytotoxicity with an increased IC50 of 64 µM. 5-AZA decreased the colony formation ability of HSC cells in semi-solid agar culture and this effect was attenuated by BLM. 5-AZA significantly downregulated high mobility group Box1 (HMGB1) and Bcl-2 gene expression but upregulated Bax gene expression, while BLM impeded the action of 5-AZA. Pretreatment with BLM remarkably decreased HMGB1 release into culture media that was induced by 5-AZA. The cells were distribution at the sub/G1 phase. Annexin/PI staining of the cells, poly (ADP-ribose) polymerase (PARP) cleavage, and anion superoxide production indicated that BLM limited 5-AZA induced apoptotic cell death. In conclusion, BLM in combination with 5-AZA effectively reduces the adverse cytotoxic effects of 5-AZA on bone marrow hematopoietic stem cells, providing a new chemotherapeutic strategy.

Synergy of theophylline reduces necrotic effect of berberine, induces cell cycle arrest and PARP, HMGB1, Bcl-2 family mediated apoptosis in MDA-MB-231 breast cancer cells.

Berberine, is a plant alkaloid, proved to have anticancer effect on various cancers. Theophylline (TH), a natural product, is widely used in the treatment of respiratory difficulties. The present study designed to elucidate the effects of theophylline and berberine combination on breast cancer cells cytotoxicity, gene expression and cell cycle. MTT assay revealed that berberine inhibited MDA-MB-231 breast cancer cells viability in a time and dose dependent manner (IC50 of 100 µM) but theophylline had no considerably effect on the cells. Combined treatment of berberine and theophylline showed a synergistic anti-proliferation effect, IC50 of berberine reduced to 50 µM and the cells were arrested at G2/M phase. Combined treatment of Berberine and theophylline reduced extracellular level of HMGB1 and down regulated HMGB1 and MMP-9 mRNA expression. The results of flow cytometry using annexin/PI staining of the cells, HMGB1 release, and poly ADP ribose polymerase cleavage demonstrated that theophylline attenuated necrotic effect of berberine and increased the level of apoptotic cell death. Enhancement of Bax content detected by ELISA and upregulation of Bax mRNA expression, down-regulation of Bcl-2 expression and increase of anion superoxide production confirmed induction of apoptosis via intrinsic apoptotic pathway. Replacement of theophylline with exogenous cyclic AMP in combination treatment represented similar effect on berberine cytotoxicity. From the results it is concluded that synergistic anticancer effect of theophylline and berberine suggests that combination of these two drugs may be an effective therapeutic agent against breast cancer cell.

Lithium chloride attenuates mitomycin C induced necrotic cell death in MDA-MB-231 breast cancer cells via HMGB1 and Bax signaling.

The clinical use of potent anticancer drug mitomycin C (MMC) has limited due to side effects and resistance of cancer cells. The aim of this study was to investigate whether lithium chloride (LiCl), as a mood stabilizer, can affect the sensitivity of MDA-MB-231 breast cancer cells to mitomycin C. The cells were exposed to various concentrations of mitomycin C alone and combined with LiCl and the viability determined by trypan blue and MTT assays. Proteins were analyzed by western blot and mRNA expression of HMGB1 MMP9 and Bcl-2 were analyzed by RT-PCR. Flow cytometry was used to determine the cell cycle arrest and percent of apoptotic and necrotic cells. Concentration of Bax assessed by ELISA. Exposure of the cells to mitomycin C revealed IC50 value of 20 µM, whereas pretreatment of the cells with LiCl induced synergistic cytotoxicity and IC50 value declined to 5 µM. LiCl combined with mitomycin C significantly down-regulated HMGB1, MMP9 and Bcl-2 gene expression but significantly increased the level of Bax protein. In addition, the content of HMGB1 in the nuclei decreased and pretreatment with LiCl reduced the content of HMGB1 release induced by MMC. LiCl increased mitomycin C-induced cell shrinkage and PARP fragmentation suggesting induction of apoptosis in these cells. LiCl prevented mitomycin C-induced necrosis and changed the cell death arrest at G2/M-phase. Taking all together, it is suggested that LiCl efficiently enhances mitomycin C-induced apoptosis and HMGB1, Bax and Bcl-2 expression may play a major role in this process, the findings that provide a new therapeutic strategy for LiCl in combination with mitomycin C.

Evaluation of Apoptosis in Multipotent Hematopoietic Cells of Bone Marrow by Anthracycline Antibiotics.

Anthracycline antibiotics are potent anticancer drugs widely used in the treatment of solid tumors and hematological malignancies. Because of their extensive clinical use and their toxic effect on normal cells, in the present study the effect of these drugs on multipotent hematopoietic bone marrow cells was investigated employing, viability tests, PARP cleavage, Hoechst 33258 staining, DNA fragmentation and superoxide anion production techniques. The results revealed that daunorubicin and doxorubicin exhibited time and dose dependent cytotoxicity against the cells and upon increasing the drugs concentrations, apoptosis was occurred after 4 h of incubation and at low concentration of the drugs. The cleavage of poly ADP-ribose polymerase (PARP) demonstrated by daunorubicin and doxorubicin treatment of the cells, suggest that the apoptotic process is PARP dependent. The drugs induced DNA fragmentation and also anion superoxide production was increased upon rising drugs concentrations. From the results it is concluded that anthracycline antibiotics represent cytotoxic effect on hematopoietic progenitor/stem cells of bone marrow, inducing apoptosis and in this process toxicity of daunorubicin is more pronounced compared to doxorubicin.

Effect of irinotecan on HMGB1, MMP9 expression, cell cycle, and cell growth in breast cancer (MCF-7) cells.

Irinotecan is a natural alkaloid agent widely used in cancer therapy. High-mobility group protein B1 as a non-histone chromosomal protein plays a fundamental role in gene expression and inflammation. In this study, the effect of irinotecan on high-mobility group protein B1 and MMP9 content, gene expression, cell cycle, and cell growth in human breast cancer cells (MCF-7) was investigated. The cells were exposed to various concentrations of irinotecan and the viability determined by trypan blue exclusion and 3-(4,5-dimethylthiazal-2-yl)-2,5-diphenyltetrazolium bromide assays. High-mobility group B proteins were extracted from the control and drug-treated cells and analyzed by immunoblot. High-mobility group protein B1 and MMP9 messenger RNA expression was studied by reverse transcription polymerase chain reaction. The results demonstrated reduction of cell viability upon increasing irinotecan concentration, up-regulated high-mobility group protein B1 gene expression, and down-regulated MMP9 mRNA. Although the content of high-mobility group protein B1 was decreased in chromatin extract upon drug action, no high-mobility group protein B1 release to extracellular space was detected by immunoblot analysis. Irinotecan decreased H3K9 acetylation and increased poly ADP-ribose polymerase fragmentation to 89 kDa and anion superoxide production suggesting induction of apoptosis in these cells. Propidium iodide staining of the cells 24 h after the drug treatment revealed arrest of the cells in S-phase. From the results, it is concluded that overexpression of high-mobility group protein B1 in the presence of irinotecan precedes breast cancer cells into apoptosis and in this response the binding of irinotecan to chromatin or high-mobility group protein B1 may condense/aggregate chromatin, preventing high-mobility group protein B1 release from chromatin.

Spectroscopic analysis of the interaction of valproic acid with histone H1 in solution and in chromatin structure.

Histone H1 is a basic chromosomal protein which links adjacent nucleosomes in chromatin structure. Valproic acid (VPA), a histone deacetylase inhibitor, is widely used as an antiepileptic drug for the treatment of various cancers. In this study the interaction between VPA and histone H1, chromatin and DNA in solution was investigated employing spectroscopic techniques. The results showed that VPA binds cooperatively to histone H1 and chromatin but exhibited very weak interaction with DNA. The association constants demonstrated higher affinity of VPA to H1 compared to chromatin. Fluorescence emission intensity was reduced by quenching value (Ksv) of 2.3 and 0.83 for H1 and chromatin respectively. VPA also altered ellipticity of chromatin and H1 at 220nm indicating increase in α-helix content of H1/chromatin proteins suggesting that the protein moiety of chromatin is the site of VPA action. Moreover, thermal denaturation revealed hypochromicity in chromatin Tm profiles with small shift in Tm values without any significant change in DNA pattern. It is concluded that VPA, apart from histone deacetylase inhibition activity, binds strongly to histone H1 in chromatin structure, demonstrating that VPA may also exert its anticancer activity by influencing chromatin proteins which opens new insight into the mechanism of VPA action.

Investigation of the interaction between berberine and nucleosomes in solution: Spectroscopic and equilibrium dialysis approach.

Berberine is a natural plant alkaloid with high pharmacological potential. Although its interaction with free DNA has been the subject of several reports, to date there is no work concerning the effect of berberine on nucleoprotein structure of DNA, the nucleosomes. The present study focuses on the binding affinity of berberine to nucleosomes and histone H1 employing various spectroscopic techniques, fluorescence, circular dichroism, thermal denaturation as well as equilibrium dialysis. The results showed that the binding of berberine to nucleosomes is positive cooperative with Ka=5.57×103M-1. Berberine quenched with the chromophores of protein moiety of nucleosomes and reduced fluorescence emission intensity at 335nm with Ksv value of 0.135. Binding of berberine to nucleosomes decreased the absorbance at 210 and 260nm, produced hypochromicity in thermal denaturation profiles and its affinity to nucleoprotein structure of nucleosomes was much higher than to free DNA. Berberine also exhibited high affinity to histone H1 in solution and the binding was positive cooperative with. Ka=3.61×103M-1. Moreover berberine decreased fluorescence emission intensity of H1 by quenching with tyrosine residue in its globular core domain. The circular dichroism profiles demonstrated that the binding of drug induced secondary structural changes in both DNA stacking and histone H1. It is concluded that berberine is genotoxic drug, interacts with nucleosomes and in this process histone H1 is involved to exert its anticancer activity.

Lithium attenuates lead induced toxicity on mouse non-adherent bone marrow cells.

Lead is a poisonous heavy metal that occurs in all parts of environment and causes serious health problems in humans. The aim of the present study was to investigate the possible protective effect of lithium against lead nitrate induced toxicity in non-adherent bone marrow stem cells. Trypan blue and MTT assays represented that exposure of the cells to different concentrations of lead nitrate decreased viability in a dose dependent manner, whereas, pretreatment of the cells with lithium protected the cells against lead toxicity. Lead reduced the number and differentiation status of bone marrow-derived precursors when cultured in the presence of colony stimulating factor (CSF), while the effect was attenuated by lithium. The cells treated with lead nitrate exhibited cell shrinkage, DNA fragmentation, anion superoxide production, but lithium prevented lead action. Moreover, apoptotic indexes such as PARP cleavage and release of HMGB1 induced by lead, were protected by lithium, suggesting anti-apoptotic effect of lithium. Immunoblot analysis of histone H3K9 acetylation indicated that lithium overcame lead effect on acetylation. In conclusion, lithium efficiently reduces lead toxicity suggesting new insight into lithium action which may contribute to increased cell survival. It also provides a potentially new therapeutic strategy for lithium and a cost-effective approach to minimize destructive effects of lead on bone marrow stem cells.

Binding of topotecan to chromatin: Insights into cooperative binding and comparison with DNA.

Topotecan (TPT) is an anticancer drug widely used in cancer therapy. Although the interaction of TPT with DNA is a subject of few reports, no work has been reported on the binding affinity of TPT to DNA-histone complex in chromatin structure. In the present study we have focused on the effect of TPT on chromatin employing various types of spectroscopy and equilibrium dialysis techniques. The results showed that TPT quenched with chromatin chromophores and decreased fluorescence emission intensity corresponding to aromatic residues of histone proteins. The UV absorbance at 260 and 210 nm in decreased in a dose dependent manner. Upon binding of the drug, ellipticity at 222 nm in the circular dichroism profile became more positive implying reduction of α-helix content of histones. The binding is positive cooperative with association constant (Ka) of 2.65×10(2) M(-1) and 1.11×10(2) M(-1) for chromatin and DNA respectively indicating higher affinity of TPT to chromatin compared to DNA. From the results it is concluded that in the cell nucleus, TPT, as a potent anticancer drug, exerts its biological action through binding to chromatin and in this process not only DNA but also histone proteins play a fundamental role.

Exploring binding affinity of oxaliplatin and carboplatin, to nucleoprotein structure of chromatin: spectroscopic study and histone proteins as a target.

Platinum drugs are potent chemotherapeutic agents widely used in cancer therapy. They exert their biological activity by binding to DNA, producing DNA adducts; however, in the cell nucleus, DNA is complexed with histone proteins into a nucleoprotein structure known as chromatin. The aim of this study was to explore the binding affinity of oxaliplatin and carboplatin to chromatin using spectroscopic as well as thermal denaturation and equilibrium dialysis techniques. The results showed that the drugs quenched with chromophores of chromatin and the quenching effect for oxaliplatin (Ksv = 3.156) was higher than carboplatin (Ksv = 0.28). The binding of the drugs exhibited hypochromicity both in thermal denaturation profiles and UV absorbance at 210 nm. The binding was positive cooperation with spontaneous reaction and oxaliplatin (Ka = 5.3 × 10(3) M(-1), n = 1.7) exhibited higher binding constant and number of binding sites than carboplatin (Ka = 0.33 × 10(3) M(-1), n = 1.0) upon binding to chromatin. Also secondary structure of chromatin proteins was altered upon drugs binding. It is concluded that oxaliplatin represents higher binding affinity to chromatin compared to carboplatin. In chromatin where DNA is compacted into nucleosomes structure with histones, the affinity of the platinated drugs is reduced and histone proteins may play a fundamental role in this binding process.

An in vitro study on the effect of vinca alkaloid, vinorelbine, on chromatin histone, HMGB proteins and induction of apoptosis in mice non-adherent bone marrow cells.

CONTEXT: Vinoreline is a vinca alkaloid anticancer drug widely used in cancer therapy. Drugs are not target specific, therefore might affect normal tissues/cells, in which bone marrow is the important one. OBJECTIVE: To elucidate the cytotoxic and genotoxic effect of vinca alkaloid anti cancer drug, vinorelbine, on mice non-adherent bone marrow cells in vitro. MATERIALS AND METHODS: Non-adherent bone marrow cells were isolated and exposed to various concentrations (0-160 µg/ml) for 4 h at 23 °C. The chromatin proteins were analyzed by SDS PAGE and western blot. Fluorescent dye staining of the cells, anion superoxide and DNA fragmentations assays were also employed. RESULT: The results from MTT and trypan blue exclusion assays represented reduction of the cells viability. Extractability of histones and HMG proteins contrasted with difficulty as their content was decreased on SDS-gel upon increasing drug concentration as western blots confirmed it. The amount of degradation form of PARP (89 KD) increased significantly in a dose dependent manner. Increase in anion superoxide production and DNA fragmentation together with cytological detection of chromatin condensation and cellular damage upon exposure of the cells to vinorelbine were indicative of apoptosis induction in these normal cells. CONCLUSION: Vinorelbine is genotoxic in non-adherent bone marrow cells as affects chromatin components, DNA, histone and HMGB1 proteins and induces apoptosis.

Spectroscopic detection of etoposide binding to chromatin components: the role of histone proteins.

Chromatin has been introduced as a main target for most anticancer drugs. Etoposide is known as a topoisomerase II inhibitor, but its effect on chromatin components is unknown. This report, for the first time, describes the effect of etoposide on DNA, histones and DNA-histones complex in the structure of nucleosomes employing thermal denaturation, fluorescence, UV absorbance and circular dichroism spectroscopy techniques. The results showed that the binding of etoposide decreased UV absorbance and fluorescence emission intensity, altered secondary structure of chromatin and hypochromicity was occurred in thermal denaturation profiles. The drug exhibited higher affinity to chromatin compared to DNA. Quenching of drug chromophores with tyrosine residues of histones indicated that globular domain of histones is the site of etoposide binding. Moreover, the binding of etoposide to histones altered their secondary structure accompanied with hypochromicity revealing compaction of histones in the presence of the drug. From the results it is concludes that apart from topoisomerase II, chromatin components especially its protein moiety can be introduced as a new site of etoposide binding and histone proteins especially H1 play a fundamental role in this process and anticancer activity of etoposide.

Investigation on the chromium oxide interaction with soluble chromatin and histone H1: a spectroscopic study.

Chromatin has been introduced as a tool for studying heavy metals action in nuclei. Chromium oxide is highly soluble and toxic with chronic exposure leading to mutagenesis and carcinogenesis. In the present study, for the first time, the binding affinity of chromium oxide to rat liver chromatin and histone proteins was investigated. Reduction of chromatin absorbencies at 210 and 260 nm (hypochromicity) and fluorescence emission intensity upon metal binding represented quenching of the metal with chromatin chromophores. Binding isotherms demonstrated a positive cooperative binding pattern revealing higher affinity of the metal to chromatin compared to DNA as confirmed by the binding constants. Melting temperature of chromatin was altered in a dose dependent manner and suggests partial removal of histones from the chromatin at metal concentrations higher than 15 µg/ml. Chromium oxide decreased the absorbance of histone H1 at 210 nm (hypochromicity) and fluorescence emission intensity revealed quenching of the metal with tyrosine residue located in the core domain of H1. Also the interaction of chromium oxide with histone H1 increased its secondary structures. The results suggest toxic effect of very low concentrations of chromium oxide on chromatin and in this reaction both DNA and histones are involved.

Insights into the structural stability of nuclear matrix ribonucleoprotein, LMG160: thermodynamic and spectroscopic analysis.

Low-mobility group nonhistone chromatin protein, LMG160, is a nuclear matrix ribonucleoprotein particle (RNP) which has a RNA molecule with approximately 300 bases. In this study, structural stability of the intact LMG160 (I-LMG160) was investigated at different ionic strength and in the absence of its RNA moiety (T-LMG160) employing spectroscopic and thermodynamic techniques. The UV absorption spectra showed hypochromicity and red shift under increasing ionic strength for both forms of LMG160 but in different extents. The fluorescence emission intensity was decreased as ionic strength was increased and the Stern-Volmer quenching constant (Ksv) for T-LMG160 was 3.7 times less than for I-LMG160. In the absence of sodium chloride, I-LMG160 exhibited a very stable structure against the temperature change compared to T-LMG160. The thermodynamic parameters showed that the positive values of ΔHm and ΔSm increased by increasing ionic strength in both forms of LMG160. Removal of the RNA moiety altered secondary structure: as T-LMG160 showed more helical content than I-LMG160. From the results, it is concluded that I-LMG160 is more sensitive to alteration of environment and the RNA has an important role in this RNP conformation. Also, interaction of both I- and T-LMG160 with sodium chloride is entropy driven and is usually accompanied by surface hydrophobicity.

Mechanism of the interaction of plant alkaloid vincristine with DNA and chromatin: spectroscopic study.

Chromatin has been successfully used as a tool for the study of genome function in cancers. Vincristine as a vinca alkaloid anticancer drug exerts its action by binding to tubulins. In this study the effect of vincristine on DNA and chromatin was investigated employing various spectroscopy techniques as well as thermal denaturation, equilibrium dialysis and DNA-cellulose affinity. The results showed that the binding of vincristine to DNA and chromatin reduced absorbance at both 260 and 210 nm with different extent. Chromopheres of chromatin quenched with the drug and fluorescence emission intensity decreased in a dose-dependent manner. Chromatin exhibited higher emission intensity changes compared to DNA. Upon addition of vincristine, Tm of DNA and chromatin exhibited hypochromicity without any shift in Tm. The binding of the drug induced structural changes in both positive and negative extremes of circular dichroism spectra and exhibited a cooperative binding pattern as illustrated by a positive slope observed in low r values of the binding isotherm. Vincristine showed higher binding affinity to double stranded DNA compared to single stranded one. The results suggest that vincristine binds with higher affinity to chromatin compared to DNA. The interaction is through intercalation along with binding to phosphate sugar backbone and histone proteins play fundamental role in this process. The binding of the drug to chromatin opens a new insight into vincristine action in the cell nucleus.

Studies on the genotoxic effect of chromium oxide (Cr VI): interaction with deoxyribonucleic acid in solution.

Chromium is a toxic and carcinogenic compound widely distributed in environment. In the present study we have investigated the interaction of chromium oxide with DNA employing UV/vis and fluorescence spectroscopy as well as Circular dichroism, thermal denaturation, retardation polyacrylamide gel electrophoresis and DNA-cellulose affinity techniques. The results showed that the binding of chromium oxide to DNA is concentration dependent; at low concentration shows a little effect but ant higher concentrations (>100µg/ml) reduced the absorbance at 260 and 210nm producing hypochromicity. Also λ(max) of the metal at 210, 260 and 350nm was reduced. DNA chromophores quenched with the chromium oxide and decreased fluorescence emission intensity. Upon binding of the metal to DNA the elliplicity at positive extreme was decreased (275nm) and increased the ellipticity of the DNA at negative extreme 245nm. Thermal denaturation profile of DNA shifted to higher degrees upon chromium oxide binding which accompanied by hypochromicity. Also, affinity of chromium oxide to double stranded DNA was higher than single stranded DNA. From the result it is concluded that chromium oxide interacts with DNA via two modes of interaction inducing structural changes and DNA compaction evidence providing chromium oxide genotoxicity.

Evidence for the binding affinity of daunomycin to HMGB1 protein in chromatin and in solution.

In this study the interaction of daunomycin with HMGB1 nonhistone chromatin protein in the chromatin context using hydroxyapatite (HAP) column chromatography and free in solution was investigated employing fluorescence, circular dichroism spectroscopy and thermal denaturation techniques. The results demonstrate that HMGB1 fraction eluted from HAP column contained the most amount of daunomycin. Upon addition of daunomycin to HMGB1 solution, fluorescence emission intensity was dependent on the drug concentration used whereas the ellipticity in CD spectra was decreased at both 205 and 220 nm extremes implying that quenching of the drug with the HMGB1 chromospheres alters secondary structure of the protein. Although daunomycin slightly increased the melting point of HMGB1, but exhibited a significant hyperchromicity at low concentrations and hypochromicity at higher concentrations of daunomycin. The results suggest that daunomycin binds to HMGB1 protein which may influence its interaction with DNA in nucleosomes and other cellular processes.

Prostaglandin E2 alteration in contraceptive consumers: as a risk factor for inflammatory diseases.

PURPOSE: The aim of this study was to evaluate the effect of oral combined contraceptive pills on Prostaglandin E2 levels and lipid profiles. METHODS: The enzyme-linked immune absorbent assay method and spectrophotometric assay were used for the evaluation of PGE2 levels and lipid profiles, respectively, in 50 healthy women with normal menstrual cycles who served as the control group and 50 women taking contraceptive pill. RESULTS: The data obtained for serum Prostaglandin E2, LDL-C, and cholesterol concentrations in contraceptive pill consumers were significantly upper (P = 0.04, 0.002, and 0.05, respectively) than control group. The age of contraceptive pill consumption and the duration of pill intake beyond 36 months had no significant effect on the prostaglandin E2 concentration. CONCLUSION: It is suggested that the increase of Prostaglandin E2 and atherogenic lipid levels may be related to their probable effects in response to various pathological and physiological properties of COCs.