Search results for: induced apoptosis

Authors: Meng-Jou Chien, Jheng-Cheng Chen, Chang-Hsiung Tsai

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Let maxζG(m) denote the maximum number of edges in a subgraph of graph G induced by m nodes. The n-dimensional augmented cube, denoted as AQn, a variation of the hypercube, possesses some properties superior to those of the hypercube. We study the cases when G is the augmented cube AQn.

Keywords: interconnection network, augmented cube, induced subgraph, bisection width

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Authors: Nahid Eskandari, Marjan Ghagozolo, Ehsan Almasi

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Introduction: Silymarin, as a polyphenolic flavonoid derived from milk thistle (Silybum marianum), is known to have antioxidant, immunomodulatory, antiproliferative, antifibrotic, and antiviral effects. The goal of this study was to determine immunosuppressive effect of Silymarin on proliferation and apoptosis of human T cells in comparison with Rapamycin and FK506. Methods: Peripheral Blood Mononuclear Cells (PBMCs) from healthy individuals were activated with Con A (5µg/ml) and then treated with Silymarin, Rapamycin and FK506 in various concentrations (0.001, 0.01, 0.1, 1, 10,100 and 200M) for 5 days. PBMCs were examined for proliferation using CFSE assay and the concentration that inhibited 50% of the cell proliferation (IC50) was determined for each treatment. For apoptosis assay using flow cytometry, PBMCs were activated with Con A and treated with IC50 dose of Silymarin, Rapamycin and FK506 for 5 days, then cell apoptosis was analysed by FITC-annexin V/PI staining and flow cytometry. The effects of Silymarin, Rapamycin and FK506 on the activation of PARP (poly ADP ribose polymerase) pathway in PBMCs stimulated with Con A and treated with IC50 dose of drugs for 5 days evaluated using the PathScan cleaved PARP sandwich ELISA kit. Results: This study showed that Silymarin had the ability to inhibit T cell proliferation in vitro. Moreover, our results indicated that 100 μM (P < 0.001) and 200 μM (P < 0.001) of Silymarin has more inhibitory effect on T cells proliferation than FK506 and Rapamycin. Our data showed that the effective doses (IC50) of Silymarin, FK506 and Rapamycin were 3×10-5 µM, 10-8 µM and 10-6 µM respectively. Data showed that the inhibitory effect of Silymarin, FK506 and Rapamycin on T cell proliferation was not due to cytotoxicity and none of these drugs at IC50 concentration had not affected the level of cleaved PARP. Conclusion: Silymarin could be a good candidate for immunosuppressive therapy for certain medical conditions with superior efficacy and lesser toxicity in comparison with other immunosuppressive drugs.

Keywords: silymarin, immunosuppressive effect, rapamycin, immunology

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Authors: Lalita Subedi, Jae Kyoung Chae, Yong Un Park, Cho Kyo Hee, Lee Jae Hyuk, Kang Min Cheol, Sun Yeou Kim

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Neuroinflammation may mediate the relationship between low levels of estrogens and neurodegenerative disease. Estrogens are neuroprotective and anti-inflammatory in neurodegenerative disease models. Due to the long term side effects of estrogens, researches have been focused on finding an effective phytoestrogens for biological activities. Daidzein present in soybeans and its active metabolite equol (7-hydroxy-3-(4'-hydroxyphenyl)-chroman) bears strong antioxidant and anticancer showed more potent anti-inflammatory and neuroprotective role in neuroinflammatory model confirmed its in vitro activity with molecular mechanism through NF-κB pathway. Three major CNS cells Microglia (BV-2), Astrocyte (C6), Neuron (N2a) were used to find the effect of equol in inducible nitric oxide synthase (iNOS), cyclooxygenase (COX-2), MAPKs signaling proteins, apoptosis related proteins by western blot analysis. Nitric oxide (NO) and prostaglandin E2 (PGE2) was measured by the Gries method and ELISA, respectively. Cytokines like tumor necrosis factor-α (TNF-α) and IL-6 were also measured in the conditioned medium of LPS activated cells with or without equol. Equol inhibited the NO production, PGE-2 production and expression of COX-2 and iNOS in LPS-stimulated microglial cells at a dose dependent without any cellular toxicity. At the same time Equol also showed promising effect in modulation of MAPK’s and nuclear factor kappa B (NF-κB) expression with significant inhibition of the production of proinflammatory cytokine like interleukin -6 (IL-6), and tumor necrosis factor -α (TNF-α). Additionally, it inhibited the LPS activated microglia-induced neuronal cell death by downregulating the apoptotic phenomenon in neuronal cells. Furthermore, equol increases the production of neurotrophins like NGF and increase the neurite outgrowth as well. In conclusion the natural daidzein metabolite equol are more active than daidzein, which showed a promising effectiveness as an anti-neuroinflammatory and neuroprotective agent via downregulating the LPS stimulated microglial activation and neuronal apoptosis. This work was supported by Brain Korea 21 Plus project and High Value-added Food Technology Development Program 114006-4, Ministry of Agriculture, Food and Rural Affairs.

Keywords: apoptosis, equol, neuroinflammation, phytoestrogen

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Authors: Manjunatha Bangeppagari, Lee Sang Joon

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Recently, graphene-related nanomaterials are receiving fast-increasing attention with augmented applications in various fields. Especially, graphene-related materials have been widely applied to the biomedical field in the past years. In the present study, we evaluated the adverse effects of pristine graphene (pG) in zebrafish (Danio rerio) embryos in various aspects, such as mortality rate, heart rate, hatching rate, cardiotoxicity, cardiovascular defect, cardiac looping, apoptosis, and globin expression. For various trace concentrations of pG (1, 5, 10, 15, 20, 25, 30, 35, 40, 45, and 50 μg/L), early life-stage parameters were observed at 24, 48, 72, and 96 hpf. As a result, pG induces significant developmental defects including yolk sac edema, pericardial edema, embryonic mortality, delayed hatching, heartbeat, several morphological defects, pericardial toxicity, and bradycardia. Moreover, the exposure to pG was found to be a potential risk factor to the cardiovascular system of zebrafish embryos. However, further study on their properties which vary according to production methods and surface functionalization is essentially required. In addition, the possible risks of pG flakes to aquatic animals, and public health should be evaluated before releasing them to the surrounding environment.

Keywords: apoptosis, cardiovascular toxicity, globin expression, pristine graphene, zebrafish embryos

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Authors: Nima Samie, Batoul Sadat Haerian, Sekaran Muniandy, M. S. Kanthimathi

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Colon and breast cancers are categorized as the most prevalent types of cancer worldwide. Recently, the broad clinical application of metal complex compounds has led to the discovery of potential therapeutic drugs. The aim of this study was to evaluate the cytotoxic action of a selected nickel complex compound (NCC) against human colon and breast cancer cells. In this context, we determined the potency of the compound in the induction of apoptosis, cell cycle arrest, and cytoskeleton rearrangement. HT-29, WiDr, CCD-18Co, MCF-7 and Hs 190.T cell lines were used to determine the IC50 of the compound using the MTT assay. Analysis of apoptosis was carried out using immunofluorescence, acridine orange/ propidium iodide double staining, Annexin-V-FITC assay, evaluation of the translocation of NF-kB, oxygen radical antioxidant capacity, quenching of reactive oxygen species content , measurement of LDH release, caspase-3/-7, -8 and -9 assays and western blotting. The cell cycle arrest was examined using flowcytometry and gene expression was assessed using qPCR array. Results showed that our nickel complex compound displayed a potent suppressive effect on HT-29, WiDr, MCF-7 and Hs 190.T after 24 h of treatment with IC50 value of 2.02±0.54, 2.13±0.65, 3.76±015 and 3.14±0.45 µM respectively. This cytotoxic effect on normal cells was insignificant. Dipping in the mitochondrial membrane potential and increased release of cytochrome c from the mitochondria indicated induction of the intrinsic apoptosis pathway by the nickel complex compound. Activation of this pathway was further evidenced by significant activation of caspase 9 and 3/7.The nickel complex compound (NCC) was also shown activate the extrinsic pathways of apoptosis by activation of caspase-8 which is linked to the suppression of NF-kB translocation to the nucleus. Cell cycle arrest in the G1 phase and up-regulation of glutathione reductase, based on excessive ROS production were also observed. The results of this study suggest that the nickel complex compound is a potent anti-cancer agent inducing both intrinsic and extrinsic pathways as well as cell cycle arrest in colon and breast cancer cells.

Keywords: nickel complex, apoptosis, cytoskeletal rearrangement, colon cancer, breast cancer

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Authors: Sakhon Woothipatanapan, Surasit Prakobkit

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This paper presents a model for analysis the induced voltage of transmission lines (energized) acting on neighboring distribution lines (de-energized). From environmental restrictions, 22 kV distribution lines need to be installed under 115 kV transmission lines. With the installation of the two parallel circuits like this, they make the induced voltage which can cause harm to operators. This work was performed with the ATP-EMTP modeling to analyze such phenomenon before field testing. Simulation results are used to find solutions to prevent danger to operators who are on the pole.

Keywords: transmission system, distribution system, induced voltage, off-line operation

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Authors: Monika Rezacova

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Radiation-induced breast changes are a consequence of radiotherapy toxicity over the breast tissues either related to targeted breast cancer treatment or other thoracic malignancies (eg. lung cancer). This study has created an overview of different changes and their pathophysiology. The main conditions included were skin thickening, interstitial oedema, fat necrosis, dystrophic calcifications, skin retractions, glandular atrophy, breast fibrosis and radiation induced breast cancer. This study has performed focused literature search through multiple databases including pubmed, medline and embase. The study has reviewed English as well as non English publications. As a result of the literature the study provides comprehensive overview of radiation-induced breast changes and their pathophysiology with small focus on new development and prevention.

Keywords: radiotherapy toxicity, breast tissue changes, breast cancer treatment, radiation-induced breast changes

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Authors: Angel Champion

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Chemotherapy and radiation use an acidified approach to induce apoptosis, which only kills mature cancer cells while resulting in gene and cell damage with significant levels of toxicity in tumor-affected tissues and organs. The acid approach, where the cells exterminated are not differentiated, induces the disappearance of white blood cells from the blood. This increases susceptibility to infection in severe forms of cancer spread. However, chemotherapy and radiation cannot kill cancer stem cells that metastasize, being the leading cause of 98% of cancer fatalities. With over 12 million new cancer cases symptomatic each year, including common malignancies such as Hepatocellular Carcinoma (HCC), this study aims to assess the bioactive constituents and phytochemical composition of Taraxacum Officinale (Dandelion). This analysis enables pharmaceutical quality and potency to be applied to studies on cancer cell proliferation and apoptosis. A phytochemical screening is carried out to identify the antioxidant components of Dandelion root, stem, and flower extract. The constituents tested for are phlorotannins, carbohydrates, glycosides, saponins, flavonoids, alkaloids, sterols, triterpenes, and anthraquinone glycosides. To conserve the existing phenolic compounds, a portion of the constituent tests will be examined with an acid, alcohol, or aqueous solvent. As a result, the qualitative and quantitative variations within the Dandelion extract that measure uniform effective potency are vital to the conformity for producing medicinal products. These medicines will be constructed with a consistent, uniform composition that physicians can use to control and effectively eradicate malignant diseases safely. Taraxacum Officinale's phytochemical composition comprises a highly-graded potency due to present bioactive contents that will essentially drive out malignant disease within the human body. Its high potency rate is powerful enough to eliminate both mature cancer cells and cancer stem cells without the cell and gene damage induced by chemotherapy and radiation. Correspondingly, the high margins of cancer mortality on a global scale are mitigated. This remarkable contribution to modern therapeutics will essentially optimize the margins of natural products and their derivatives, which account for 50% of pharmaceuticals in modern therapeutics, while preventing the adverse effects of radiation and chemotherapy drugs.

Keywords: antioxidant, apoptosis, metastasize, phytochemical, proliferation, potency

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Authors: Mohamed Abdel-Hamid, Olfat Gamil Shaker, Doha El-Sayed Ellakwa, Eman Fathy Abdel-Maksoud

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Introduction: Despite advances in the knowledge of the molecular virology of hepatitis C virus (HCV), the mechanisms of hepatocellular injury in HCV infection are not completely understood. Hepatitis C viral infection (HCV) influences the susceptibility to apoptosis. This could lead to insufficient antiviral immune response and persistent viral infection. Aim of this study: was to examine whether BCL-2 gene polymorphism at codon 43 (+127G/A or Ala43Thr) has an impact on development of hepatocellular carcinoma caused by chronic hepatitis C Egyptian patients. Subjects and Methods: The study included three groups; group 1: composing of 30 patients with hepatocellular carcinoma (HCC), group 2 composing of 30 patients with HCV, group 3 composing of 30 healthy subjects matching the same age and socioeconomic status were taken as a control group. Gene polymorphism of BCL2 (Ala43Thr) were evaluated by PCR-RFLP technique and measured for all patients and controls. Results: The summed 43Thr genotype was more frequent and statistically significant in HCC patients as compared to control group. This genotype of BCL2 gene may inhibit the programmed cell death which leads to disturbance in tissue and cells homeostasis and reduction in immune regulation. This result leads to viral replication and HCV persistence. Moreover, virus produces variety of mechanisms to block genes participated in apoptosis. This mechanism proves that HCV patients who have 43Thr genotype are more susceptible to HCC. Conclusion: The data suggest for the first time that the BCL2 polymorphism is associated with the susceptibility to HCC in Egyptian populations and might be used as molecular markers for evaluating HCC risk. This study clearly demonstrated that Chronic HCV exhibit a deregulation of apoptosis with the disease progression. This provides an insight into the pathogenesis of chronic HCV infection, and may contribute to the therapy.

Keywords: BCL2 gene, Hepatitis C Virus, Hepatocellular carcinoma, sensitivity, specificity, apoptosis

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Authors: Isha Ranadive, Sonam Patel, Suresh Balakrishnan

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It has been observed that in lizards wound can be healed by either a scar free mechanism or by scarring. The animal model used to study both these healing processes was Northern House Gecko. In lizard, the tail when amputated heals by scar free mechanism which allows it to regenerate, the same is not seen when the limb is amputated. Proliferation, apoptosis, and angiogenesis are the main events which succeed an injury. We observed that proliferation of the cells beneath the wound epidermis was much higher in case of wound healing in tail. This could be because after the wound gets covered by the epithelium, it enters in to a cross-talk with the underlying mesenchyme to recruit a pool of blastemal cells which proliferate and later differentiate to form the lost part through epimorphic regeneration. This was substantiated by mRNA expression levels of various FGFs which facilitate the cross-talk and also by PCNA which is a marker for proliferation. Western blot result reaffirms the same notion. However, in case of the limb, the rate of apoptosis was more than proliferation as there are a lot of debris that needs to be removed. We came to this conclusion as we observed that p53 the apoptotic gene was highly upregulated in case of the scarred tissue. Further, we confirmed this result by checking the anti-apoptotic gene bcl2 and found it to be significantly down-regulated. As we noticed heightened proliferation in the case of scar-free wound healing in tail, angiogenesis was targeted for the study. This is because, when the cells are proliferating they require constant supply of blood and hence neo-vascularization is inevitable. It was observed that the marker of angiogenesis, VEGF, was expressed more during wound healing as compared to the resting stage of tail. Moreover, a high up-regulation was seen in KDR, a receptor of VEGF. Thus, this study reveals how proliferation, apoptosis, and angiogenesis play a key role in the scar-free as well as scarred wound healing.

Keywords: epimorphic regeneration, injury, northern house gecko, wound healing

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Authors: Amr A. Fouad, Waleed H. Albuali, Iyad Jresat

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The protective effect of hesperidin was investigated in rats exposed to liver injury induced by a single intraperitoneal injection of cyclophosphamide (CYP) at a dose of 150 mg kg-1. Hesperidin treatment (100 mg kg-1/day, orally) was applied for seven days, starting five days before CYP administration. Hesperidin significantly decreased the CYP-induced elevations of serum alanine aminotransferase, and hepatic malondialdehyde and myeloperoxidase activity, significantly prevented the depletion of hepatic glutathione peroxidase activity resulted from CYP administration. Also, hesperidin ameliorated the CYP-induced liver tissue injury observed by histopathological examination. In addition, hesperidin decreased the CYP-induced expression of inducible nitric oxide synthase, tumor necrosis factor-α, cyclooxygenase-2, Fas ligand, and caspase-9 in liver tissue. It was concluded that hesperidin may represent a potential candidate to protect against CYP-induced hepatotoxicity.

Keywords: hesperidin, cyclophosphamide, liver, rats

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Authors: Hamed Karimian, Noraziah Nordin, Mohamad Ibrahim Noordin, Syam Mohan, Mahboubeh Razavi, Najihah Mohd Hashim, Happipah Mohd Ali

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Satureja bachtiarica Bunge is a perennial medicinal plant belonging to the Lamiaceae family and endemic species in Iran. Satureja bachtiarica Bunge with the local name of Marzeh koohi is edible vegetable use as flavoring agent, anti-bacterial and to relieve cough and indigestion. In this study, the anti-cancer effect of Satureja bachtiarica Bunge on the MDA-MB-231 cell line as an Breast cancer cell model has been analyzed for the first time. Satureja bachtiarica Bunge was extracted using different solvents in the order of increasing polarity. Cytotoxicity activity of hexane extract of Satureja bachtiarica Bunge (SBHE) was observed using MTT assay. Acridine orange/Propidium iodide staining was used to detect early apoptosis; Annexin-V-FITC assay was carried out to observe the detection of cell-surface Phosphatidylserine (PS), with Annexin-Vserving as a marker for apoptotic cells. Caspase 3/7, 8 and-9 assays showed significantly activation of caspase-9 where lead intrinsic mitochondrial pathway. Bcl-2/Bax expressions and cell cycle arrest were also investigated. SBHE had exhibited significantly higher cytotoxicity against MDA-MB-231 Cell line compare to other cell lines. A significant increase in chromatin condensation in the cell nucleus was observed by fluorescence analysis. Treatment of MDA-MB-231 cells with SBHE encouraged apoptosis, by down-regulating Bcl-2 and up-regulating Bax, which lead the activation of caspase 9. Moreover, SBHE treatment significantly arrested MDA-MB-231 cells in the G1 phase. Together, the results presented in this study demonstrated that SBHE inhibited the proliferation of MDA-MB-231 cells, leading cell cycle arrest and programmed cell death, which was confirmed to be through the mitochondrial pathway.

Keywords: Satureja bachtiarica Bunge, MDA-MB-231, apoptosis, annexin-V, cell cycle

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Authors: Shao-Ju Chien, Yi-Chien Wu, Ting-Ying Huang, Li-Tsen Li, You-Jin Chen, Cheng-Nan Chen

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Homocysteine and pro-inflammatory mediators such as cyclooxygenase-2 (COX-2) have been linked to vascular dysfunction and risks of cardiovascular diseases. Fulvic acid (FA) is class of compounds of humic substances and possesses various pharmacological properties. However, the effect of FA on inflammatory responses of the monocytes remains unclear. We investigated the regulatory effect of FA on homocysteine-induced COX-2 expression in human monocytes. Peripheral blood monocytes and U937 cells were kept as controls or pre-treated with FA, and then stimulated with homocysteine. The results show that pretreating monocytes with FA inhibited the homocysteine-induced COX-2 expression in a dose-dependent manner. The inhibitor for nuclear factor-kB (NF-kB) attenuated homocysteine-induced COX-2 expression. Our findings provide a molecular mechanism by which FA inhibit homocysteine-induced COX-2 expression in monocytes, and a basis for using FA in pharmaceutical therapy against inflammation.

Keywords: homocysteine, monocytes, cyclooxygenase-2, fulvic acid, anti-inflammation

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Authors: Kakali Bhadra

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Harmalol administration caused remarkable reduction in proliferation of HepG₂ cells with GI₅₀ of 14.2 mM, without showing much cytotoxicity in embryonic liver cell line, WRL-68. Data from circular dichroism and differential scanning calorimetric analysis of harmalol-CT DNA complex shows conformational changes with prominent CD perturbation and stabilization of CT DNA by 8 ^oC. Binding constant and stoichiometry was also calculated using the above biophysical techniques. Further, dose dependent apoptotic induction ability of harmalol was studied in HepG₂ cells using different biochemical assays. Generation of ROS, DNA damage, changes in cellular external and ultramorphology, alteration of membrane, formation of comet tail, decreased mitochondrial membrane potential and a significant increase in Sub G_o/G₁ population made the cancer cell, HepG₂, prone to apoptosis. Up regulation of p53 and caspase 3 further indicated the apoptotic role of harmalol.

Keywords: apoptosis, beta carboline alkaloid, comet assay, cytotoxicity, ROS

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Authors: Maryam Zahedifard, Fadhil Lafta Faraj, Maryam Hajrezaie, Nazia Abdul Majid, Mahmood Ameen Abdulla, Hapipah Mohd Ali

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In the current study, we prepared two new quinazoline schiff bases through condensation reaction of 2-aminobenzhydrazide with 5-bromosalicylaldehyde and 3-methoxy-5-bromosalicylaldehyde. The chemical structures of both newly synthesized compounds (1 and 2) were confirmed by FT-IR and X-ray crystallography studies. The cytotoxic effect of compounds was investigated against MCF-7 human breast cancer cells. MTT results showed that (1) and (2) decreased the viability of MCF-7 cells in a time-dependent manner, exhibiting an IC50 value of 3.23 ± 0.28 µg/mL and 3.41 ± 0.34 µg/mL, respectively, after a 72-hours treatment period. In contrast, they did not show significant anti-proliferative effect towards MCF-10A normal breast cells and WRL-68 normal liver cells. We found a perturbation in mitochondrial membrane potential and increased cytochrome c release from the mitochondria to the cytosol, suggesting an activation of apoptosis by compounds, which was confirmed by activation of the initiator caspase-9 and the executioner caspases-3/7. (1) was also able to trigger extrinsic pathway via activation of caspase-8 and inhibition of NF-κB translocation. The acute toxicity test showed no toxicity effect of the compounds in rats. Our results showed that the selected synthesized compounds are highly potent to induce apoptosis in MCF-7 cells via either intrinsic or extrinsic mitochondrial pathway.

Keywords: Quinazoline Schiff base, apoptosis, MCF-7 human breast cancer cell line, caspase, NF-κB translocation

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Authors: Qi LI, Xu Lin, Nengming Lin

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Objective: The aim of this study was to investigate the role of desmocollin 2 (DSC2) protein in the proliferation, migration and drug resistance of lung cancer cells. Method: CCK-8 assays and colony formation assays were used to evaluate the effect of dsc2 regulation on cancer cell viability and colony formation. Transwell assays and wound healing assays were also performed. Cell flow double staining was used to detect the apoptosis rate of cells with DSC2, which was added cisplatin. Western blot assay was used to detect cell cycle, PI3k/Akt and apoptosis-related proteins. Results: Our data showed that dsc2 is upregulated in clinical lung cancer tissues compared with pericarcinomatous tissues, and it is differentially expressed in lung cancer cell lines. The down-regulation of dsc2 in A549 and H358 lung cancer cells significantly suppressed the cell proliferation, metastasis, and motility. In contrast, the opposite effects were observed in overexpression of dsc2 both in H23 and PC9 cell lines. In addition to lung adenocarcinoma cell lines, we also examined its expression in lung squamous cell lines, such as H226. Western blotting showed that dsc2 could reduce the level of phosphorylated Akt (Ser 473) and p-mTOR. Thus, it is speculated that dsc2 up-regulation promotes proliferation and invasiveness through activation of the PI3K/AKT pathway. Also, knockdown of dsc2 in A549 and H226 could significantly decreased in the levels of cyclinB and wee1 protein. Additionally, flow cytometry showed that dsc2 knockdown combined with cisplatin could significantly enhance cell apoptosis rate. Conclusion: These data suggest that dsc2 promotes the proliferation and migration of lung cancer cells in vitro. Also, the results suggested that dsc2 could affect the cell cycle and apoptosis of lung cells. Furthermore, knockdown of dsc2 could sensitize cisplatin in both lung adenocarcinoma and lung squamous cell lines. Thus we suggested that dsc2 can be used as a therapeutic target for lung cancer.

Keywords: desmocollin 2, cisplatin, lung cancer, PI3K/AKT, lung squamous cell

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Authors: Vinaya Kambappa, Chinnadurai Mani, Komaraiah Palle

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Non-small cell-lung cancer (NSCLC) cells have increased expression of EGFR, which makes them a potential target for cancer therapy. Based on molecular docking and previous reports, we designed and synthesized quinazoline derivatives as potent EGFR inhibitors. Among the derivatives, three compounds showed good antiproliferative activity against A-549 and H-1299 cells. Furthermore, these compounds inhibited EGFR signaling exhibiting diminishing p-EGFR and its downstream proteins like p-Akt, p-Erk1/2, and p-mTOR; however, it did not alter the levels of EGFR, Akt, Erk1/2 and mTOR proteins. Flow cytometric analysis indicated the accumulation of cells at G1 phase suggesting induction of apoptosis, which was further confirmed by annexin V/propidium iodide staining. Our study suggested that quinazoline scaffold can be developed as novel EGFR kinase inhibitors for cancer therapy.

Keywords: apoptosis, non-small cell-lung cancer cells, EGFR, quinazoline

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Authors: Shuo Huang, Huomiao Guo, Wenrui Huang

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In navigation channels located in coasts and estuaries as the waterways connecting coastal water to ports or harbors, density-induced flow often exist due to the density-gradient or gravity gradient as the results of mixing between fresh water from coastal rivers and saline water in the coasts. The density-induced flow often carries sediment transport into navigation channels and causes sediment depositions in the channels. As a result, expensive dredging may need to maintain the water depth required for navigation. In our study, we conduct a series of experiments to investigate the characteristics of density-induced flow in the estuarine navigation channels under different density gradients. Empirical equations between density flow and salinity gradient were derived. Effects of coastal structures for regulating navigation channel on density-induced flow have also been investigated. Results will be very helpful for improving the understanding of the characteristics of density-induced flow in estuarine navigation channels. The results will also provide technical support for cost-effective waterway regulation and management to maintain coastal and estuarine navigation channels.

Keywords: density flow, estuarine, navigation channel, structure

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Authors: Mohammadjavad Sotoudeheian

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Heart failure (HF) prevalence, as a global cardiovascular problem, is increasing gradually. A variety of molecular mechanisms contribute to HF. Proteins involved in cardiac contractility regulation, such as ion channels and calcium handling proteins, are altered. Additionally, epigenetic modifications and gene expression can lead to altered cardiac function. Moreover, inflammation and oxidative stress contribute to HF. The progression of HF can be attributed to mitochondrial dysfunction that impairs energy production and increases apoptosis. Molecular mechanisms such as these contribute to the development of cardiomyocyte defects and HF and can be therapeutically targeted. The heart's contractile function is controlled by cardiomyocytes. Calpain, and its related molecules, including Bax, VEGF, and AMPK, are among the proteins involved in regulating cardiomyocyte function. Apoptosis is facilitated by Bax. Cardiomyocyte apoptosis is regulated by this protein. Furthermore, cardiomyocyte survival, contractility, wound healing, and proliferation are all regulated by VEGF, which is produced by cardiomyocytes during inflammation and cytokine stress. Cardiomyocyte proliferation and survival are also influenced by AMPK, an enzyme that plays an active role in energy metabolism. They all play key roles in apoptosis, angiogenesis, hypertrophy, and metabolism during myocardial inflammation. The role of calpains has been linked to several molecular pathways. The calpain pathway plays an important role in signal transduction and apoptosis, as well as autophagy, endocytosis, and exocytosis. Cell death and survival are regulated by these calcium-dependent cysteine proteases that cleave proteins. As a result, protein fragments can be used for various cellular functions. By cleaving adhesion and motility proteins, calcium proteins also contribute to cell migration. HF may be brought about by calpain-mediated pathways. Many physiological processes are mediated by the calpain molecular pathways. Signal transduction, cell death, and cell migration are all regulated by these molecular pathways. Calpain is activated by calcium binding to calmodulin. In the presence of calcium, calmodulin activates calpain. Calpains are stimulated by calcium, which increases matrix metalloproteinases (MMPs). In order to develop novel treatments for these diseases, we must understand how this pathway works. A variety of myocardial remodeling processes involve calpains, including remodeling of the extracellular matrix and hypertrophy of cardiomyocytes. Calpains also play a role in maintaining cardiac homeostasis through apoptosis and autophagy. The development of HF may be in part due to calpain-mediated pathways promoting cardiomyocyte death. Numerous studies have suggested the importance of the Ca2+ -dependent protease calpain in cardiac physiology and pathology. Therefore, it is important to consider this pathway to develop and test therapeutic options in humans that targets calpain in HF. Apoptosis, autophagy, endocytosis, exocytosis, signal transduction, and disease progression all involve calpain molecular pathways. Therefore, it is conceivable that calpain inhibitors might have therapeutic potential as they have been investigated in preclinical models of several conditions in which the enzyme has been implicated that might be treated with them. Ca 2+ - dependent proteases and calpains contribute to adverse ventricular remodeling and HF in multiple experimental models. In this manuscript, we will discuss the calpain molecular pathway's important roles in HF development.

Keywords: calpain, heart failure, autophagy, apoptosis, cardiomyocyte

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Authors: Latife Merve Oktay, Berrin Tugrul

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Hydroxytyrosol (HT) is a phenolic phytochemical molecule derived from the hydrolysis of oleuropein, which originates during the maturation of the olives. It has recently received particular attention because of its antioxidant, anti-proliferative, pro-apoptotic and anti-inflammatory activities. In this study, we investigated the cytotoxic and apoptotic effects of hydroxytyrosol and its effects on phosphatidylinositol 3-kinase/Akt (PI3K/Akt) and extracellular signal-regulated kinase 1/2 (ERK 1/2) signaling pathways in human ovarian cancer cell lines OVCAR-3 and MDAH-2774. XTT cell proliferation kit, Cell Death Detection Elisa Plus Kit (Roche) and Human Apoptosis Array (R&D Systems) were used to determine the cytotoxic and apoptotic effects of HT in OVCAR-3 and MDAH-2774 cell lines at 24, 48, 72, and 96 h. Effect of HT on PI3K/Akt and ERK 1/2 signaling pathways were investigated by using specific inhibitors of these pathways. IC50 values of HT were found to be 102.3 µM in MDAH-2774 cells at 72 h and 51.5 µM in OVCAR-3 cells at 96 h. Apoptotic effect of HT in MDAH-2774 cells was the highest at 50 µM at 72 h, and kept decreasing at 100 and 150 µM concentrations and was not seen at 200 µM and higher concentrations. Highest apoptotic effect was seen at 100 µM concentration in OVCAR-3 cells at 96 h, however apoptotic effect was decreased over 100 µM concentrations. According to antibody microarray results, HT increased the levels of pro-apoptotic molecules Bad, Bax, active caspase-3, Htra2/Omi by 2.0-, 1.4-, 1.2-, 4.2-fold, respectively and also increased the levels of pro-apoptotic death receptors TRAIL R1/DR4, TRAIL R2/DR5, FAS/TNFRSF6 by 2.1-, 1.7-, 1.6-fold, respectively, however, it decreased the level of Survivin by 1.6-fold which is one of the inhibitor of apoptosis protein (IAP) family in MDAH-2774 cells. In OVCAR-3 cells, HT decreased the levels of anti-apoptotic proteins Bcl-2, pro-caspase 3 by 3.1-, 8.2-fold, respectively and IAP family proteins CIAP-1, CIAP-2, XIAP, Livin, Survivin by 6.5-, 6.0-, 3.2-, 2.2-, 2.7-fold, respectively and increased the level of cytochrome-c by 1.2-fold. We have shown that HT shows its cytotoxic and apoptotic effect through inhibiting ERK 1/2 signaling pathway in both OVCAR-3 and MDAH-2774 cells. Further studies are needed to investigate molecular mechanisms and modulatory effects of hydroxytyrosol.

Keywords: apoptosis, cytotoxicity, hydroxytyrosol, ovarian cancer

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Authors: Oluwafemi Ojo, Omotade Oloyede

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This study seeks to investigate the antioxidative properties and the ability of aqueous, ethanolic and ethyl acetate extracts from Ocimum gratissimum (OG) leaves to inhibit some pro-oxidants (Fe2+ and sodium nitroprusside) induced lipid peroxidation in rat’s liver homogenates in vitro. The ability of the extracts to inhibit 25 µM FeSO4 and 7.0 µM sodium nitroprusside induced lipid peroxidation in isolated rat’s liver was determined. The results of the study revealed that both pro-oxidants caused a significantly decrease in (p < 0.05) accumulation of lipid peroxides. However, aqueous extract of OG shows a high ability to inhibit lipid production in the liver induced with SNP than Fe2+. Ethanolic and ethyl acetate extract of OG which shows a high ability to inhibit lipid production more when induced with Fe2+ than SNP. However, ethyl acetate fraction of OG shows a higher inhibitory effect on both Fe2+ and SNP induced lipid peroxidation in rat’s liver. This applies to its significantly higher extractable phytochemicals. Therefore, Fe II and sodium nitroprusside induced oxidative stress could be managed by dietary intake of Ocimum gratissimum leaves.

Keywords: antioxidative, pro-oxidants, lipid peroxidation, Ocimum gratissimum

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Authors: Hien Thi Thu Le, Nuno Rafael Candeias, Olli Yli-Harja, Meenakshisundaram Kandhavelu

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Purinergic receptor 1 (P2Y1R) is the potential therapeutic target for inducing prostate cancer (PCa) cell death. Recently, 1-indolinoalkyl 2-phenolic derivative, HIC, was identified as a P2Y1R agonist that increases apoptosis and inhibits cell proliferation of PCa. However, the biological effects of HIC have not been extensively studied at the molecular level. In the present study, we have investigated the anticancer effects of HIC and the molecular mechanisms underlying in PCa cells. Half maximal inhibitory concentration (IC₅₀) of HIC was measured as 15.98 μM and 15.64 μM for DU145 and PC3 cells, respectively. In addition, we found that HIC inhibited cell growth and metastasis of PC3 and DU145 cells colonies, spheroid areas, and migrated cells. RNA seq analysis revealed significant changes of over 3000 genes (p value < 0.05) upon HIC treatment in PC3 and DU145 cells. Genes involved in DNA damage, apoptosis, cell cycle arrest at G1/S phase were modulated by HIC treatment. MAPK and NF-κB protein array revealed the increased expression of ERK1/2, JNK1/2, p53 phosphorylation, and p53 protein. ERK1/2 and JNK1/2 activations are known to increase the stabilization of p53, a tumor suppressor protein, which is required to arrest the cell cycle at G1/S phase and cause cell death of PCa cells. Overall, our results suggest that HIC can serve as a multi-dimensional chemotherapeutic agent possessing strong cytotoxic, anti-cancer, and anti-metastasis against PCa growth.

Keywords: prostate cancer, P2Y1 receptor, apoptosis, metastasis

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Authors: Monika Verma, Renuka Sharma, B. R. Gulati, Namita Singh

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In combination therapy, two chemotherapeutic agents work together in a collaborative action. It has appeared as one of the promising approaches to improve anti-cancer treatment efficacy. In the present investigation, piperine (P-NPS), paclitaxel (PTX NPS), and a combination of both, piperine-paclitaxel nanoparticle (Pip-PTX NPS), were made by the nanoprecipitation method and later characterized by PSA, DSC, SEM, TEM, and FTIR. All nanoparticles exhibited a monodispersed size distribution with a size of below 200 nm, zeta potential ranges from (-30-40mV) and a narrow polydispersity index (>0.3) of the drugs. The average encapsulation efficiency was found to be between 80 and 90%. In vitro release of drugs for nanoparticles was done spectrophotometrically. FTIR and DSC results confirmed the presence of the drug. The Pip-PTX NPS significantly inhibit cell proliferation as compared to the native drugs nanoparticles in the breast cancer cell line MCF-7. In addition, Pip-PTX NPS suppresses cells in colony formation and soft gel agar assay. Scratch migration and Transwell chamber invasion assays revealed that combined nanoparticles reduce the migration and invasion of breast cancer cells. Morphological studies showed that Pip-PTX NPS penetrates the cells and induces apoptosis, which was further confirmed by DNA fragmentation, SEM, and western blot analysis. Taken together, Pip-PTX NPS inhibits cell proliferation, anchorage dependent and anchorage independent cell growth, reduces migration and invasion, and induces apoptosis in cells. These findings support that combination therapy using Pip-PTX NPS represents a potential approach and could be helpful in the future for breast cancer therapy.

Keywords: piperine, paclitaxel, breast cancer, apoptosis

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Authors: Fadia Gafri, Kathryn Mckintosh, Louise Young, Alan Harvey, Simon Mackay, Andrew Paul, Robin Plevin

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Nuclear factor-kappa B (NF-κB) is a ubiquitous transcription factor found originally to play a key role in regulating inflammation. However considerable evidence links this pathway to the suppression of apoptosis, cellular transformation, proliferation and invasion (Aggarwal et al., 2006). Moreover, recent studies have also linked inflammation to cancer progression making NF-κB overall a promising therapeutic target for drug discovery (Dobrovolskaia & Kozlov, 2005). In this study we examined the effect of the natural product canthin-6-one (SU182) as part of a CRUK small molecule drug discovery programme for effects upon the NF-κB pathway. Initial studies demonstrated that SU182 was found to have good potency against the inhibitory kappa B kinases (IKKs) at 30M in vitro. However, at concentrations up to 30M, SU182 had no effect upon TNFα stimulated loss in cellular IκBα or p65 phosphorylation in the keratinocyte cell line NCTC2544. Nevertheless, 30M SU182 reduced TNF-α / PMA-induced NF-κB-linked luciferase reporter activity to (22.9 ± 5%) and (34.6± 3 %, P<0.001) respectively, suggesting an action downstream of IKK signalling. Indeed, SU182 neither decreased NF-κB-DNA binding as assayed by EMSA nor prevented the translocation of p65 (NF-κB) to the nucleus assessed by immunofluorescence and subcellular fractionation. In addition to the inhibition of transcriptional activity of TNFα-induced NF-κB reporter activity SU182 significantly reduced PMA-induced AP-1-linked luciferase reporter activity to about (48± 9% at 30M, P<0.001) . This mode of inhibition was not sufficient to prevent the activation of NF-κB dependent induction of other proteins such as COX-2 and iNOS, or activated MAP kinases (p38, JNK and ERK1/2) in LPS stimulated RAW 264.7 macrophages. Taken together these data indicate the potential for SU182 to interfere with the transcription factors NF-κB and AP-1 at transcriptional level. However, no potential anti-inflammatory effect was indicated, further investigation for other NF-κB dependent proteins linked to survival are also required to identify the exact mechanism of action.

Keywords: Canthin-6-one, NF-κB, AP-1, phosphorylation, Nuclear translocation, DNA-binding activity, inflammatory proteins.

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Authors: Elsie M. Nolte, Annie M. Joubert, Roy Lakier, Maryke Etsebeth, Jolene M. Helena, Marcel Verwey, Laurence Lafanechere, Anne E. Theron

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Treatment regimens for breast- and lung cancers may include both radiation- and chemotherapy. Ideally, a pharmaceutical agent which selectively sensitizes cancer cells to gamma (γ)-radiation would allow administration of lower doses of each modality, yielding synergistic anti-cancer benefits and lower metastasis occurrence, in addition to decreasing the side-effect profiles. A range of 2-methoxyestradiol (2-ME) analogues, namely 2-ethyl-3-O-sulphamoyl-estra-1,3,5 (10) 15-tetraene-3-ol-17one (ESE-15-one), 2-ethyl-3-O-sulphamoyl-estra-1,3,5(10),15-tetraen-17-ol (ESE-15-ol) and 2-ethyl-3-O-sulphamoyl-estra-1,3,5(10)16-tetraene (ESE-16) were in silico-designed by our laboratory, with the aim of improving the parent compound’s bioavailability in vivo. The main effect of these compounds is the disruption of microtubule dynamics with a resultant mitotic accumulation and induction of programmed cell death in various cancer cell lines. This in vitro study aimed to determine the cellular responses involved in the radiation sensitization effects of these analogues at low doses in breast- and lung cancer cell lines. The oestrogen receptor positive MCF-7-, oestrogen receptor negative MDA-MB-231- and triple negative BT-20 breast cancer cell lines as well as the A549 lung cancer cell line were used. The minimal compound- and radiation doses able to induce apoptosis were determined using annexin-V and cell cycle progression markers. These doses (cell line dependent) were used to pre-sensitize the cancer cells 24 hours prior to 6 gray (Gy) radiation. Experiments were conducted on samples exposed to the individual- as well as the combination treatment conditions in order to determine whether the combination treatment yielded an additive cell death response. Morphological studies included light-, fluorescence- and transmission electron microscopy. Apoptosis induction was determined by flow cytometry employing annexin V, cell cycle analysis, B-cell lymphoma 2 (Bcl-2) signalling, as well as reactive oxygen species (ROS) production. Clonogenic studies were performed by allowing colony formation for 10 days post radiation. Deoxyribonucleic acid (DNA) damage was quantified via γ-H2AX foci and micronuclei quantification. Amplification of the p53 signalling pathway was determined by western blot. Results indicated that exposing breast- and lung cancer cells to nanomolar concentrations of these analogues 24 hours prior to γ-radiation induced more cell death than the compound- and radiation treatments alone. Hypercondensed chromatin, decreased cell density, a damaged cytoskeleton and an increase in apoptotic body formation were observed in cells exposed to the combination treatment condition. An increased number of cells present in the sub-G1 phase as well as increased annexin-V staining, elevation of ROS formation and decreased Bcl-2 signalling confirmed the additive effect of the combination treatment. In addition, colony formation decreased significantly. p53 signalling pathways were significantly amplified in cells exposed to the analogues 24 hours prior to radiation, as was the amount of DNA damage. In conclusion, our results indicated that pre-treatment of breast- and lung cancer cells with low doses of 2-ME analogues sensitized breast- and lung cancer cells to γ-radiation and induced apoptosis more so than the individual treatments alone. Future studies will focus on the effect of the combination treatment on non-malignant cellular counterparts.

Keywords: cancer, microtubule dynamics, radiation therapy, radiosensitization

Procedia PDF Downloads 179

Authors: Mohammad M. Aria, Fatma Öz, Yashar Esmaeilian, Marco Carofiglio, Valentina Cauda, Özlem Yalçın

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Photoelectrical stimulation of cells with semiconductor organic polymers have been shown promising applications in neuroprosthetics such as retinal prosthesis. Photoelectrical stimulation of the cell membranes can be induced through a photo-electric charge separation mechanism in the semiconductor materials, and it can alter intracellular calcium level through both stimulation of voltage-gated ion channels and increase of intracellular reactive oxygen species (ROS) level. On the other hand, targeting voltage-gated ion channels in cancer cells to induce cell apoptosis through calcium signaling alternation is an effective mechanism which has been explained before. In this regard, remote control of the voltage-gated ion channels aimed to alter intracellular calcium by using photo-active organic polymers can be novel technology in cancer therapy. In this study, we used P (ITO/Indium thin oxide)/P3HT(poly(3-hexylthiophene-2,5-diyl)) and PN (ITO/ZnO/P3HT) photovoltaic junctions to stimulate MDA-MB-231 breast cancer cells. We showed that the photo-stimulation of breast cancer cells through photo capacitive current generated by the photovoltaic junctions are able to excite the cells and alternate intracellular calcium based on the calcium imaging (at 8mW/cm² green light intensity and 10-50 ms light durations), which has been reported already to safety stimulate neurons. The control group did not undergo light treatment and was cultured in T-75 flasks. We detected 20-30% cell death for ITO/P3HT and 51-60% cell death for ITO/ZnO/P3HT samples in the light treated MDA-MB-231 cell group. Western blot analysis demonstrated poly(ADP-ribose) polymerase (PARP) activated cell death in the light treated group. Furthermore, Annexin V and PI fluorescent staining indicated both apoptosis and necrosis in treated cells. In conclusion, our findings revealed that the photoelectrical stimulation of cells (through long time overstimulation) can induce cell death in cancer cells.

Keywords: Ca²⁺ signaling, cancer therapy, electrically excitable cells, photoelectrical stimulation, voltage-gated ion channels

Procedia PDF Downloads 148

Authors: Dhruv J. Limaye, Hanga Galfalvy, Cheick A. Sissoko, Yung-yu Huang, Chunanning Tang, Ying Liu, Shu-Chi Hsiung, Andrew J. Dwork, Gorazd B. Rosoklija, Victoria Arango, Lewis Brown, J. John Mann, Maura Boldrini

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Memory and emotion require hippocampal cell viability and connectivity and are disrupted in major depressive disorder (MDD). Applying shotgun proteomics and stereological quantification of neural progenitor cells (NPCs), intermediate neural progenitors (INPs), and mature granule neurons (GNs), to postmortem human hippocampus, identified differentially expressed proteins (DEPs), and fewer NPCs, INPs and GNs, in untreated MDD (uMDD) compared with non-psychiatric controls (CTRL) and antidepressant-treated MDD (MDDT). DEPs lower in uMDD vs. CTRL promote mitosis, differentiation, and prevent apoptosis. DEPs higher in uMDD vs. CTRL inhibit the cell cycle, and regulate cell adhesion, neurite outgrowth, and DNA repair. DEPs lower in MDDT vs. uMDD block cell proliferation. We observe group-specific correlations between numbers of NPCs, INPs, and GNs and an abundance of proteins regulating mitosis, differentiation, and apoptosis. Altered protein expression underlies hippocampus cellular and volume loss in uMDD, supports a trophic effect of antidepressants, and offers new treatment targets.

Keywords: proteomics, hippocampus, depression, mitosis, migration, differentiation, mitochondria, apoptosis, antidepressants, human brain

Procedia PDF Downloads 69

Authors: Min-Chien Su, Tzu-Hsuan Hsu, Guan-Xuan Wu, Shyh-Ming Kuo

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Lung cancer is one of the clinically challenging malignant diseases worldwide. For efficient therapeutics in cancer, combination therapy has developed to acquire a better outcome. PLK-1 was one of the major factors affecting cell mitosis in cancer cells, its inhibitor Bi6727 was proven effective in treating several different cancers namely oral cancer, colon cancer and lung cancer. Despite its low toxicity toward normal cells compared to traditional chemotherapy, it is still yet to be evaluated in detail. Livistona Chinensis (LC) is a Chinese herb that used as a traditional prescription to treat lung cancer. Due to the uncertainty of the efficacy of LC, we utilized a water extraction method to extract the Livistona Chinensis and then lyophilized into powder for further study. In this study we investigated the antiproliferation activities of Bi6727 and LC extracts (LCE) on A549 non-small lung cancer cells. The IC50 of Bi6727 and LCE on A549 are 60 nM and 0.8 mg/mL, respectively. The fluorescent staining images shown nucleolus damage in cells treated with Bi6727 and mitochondrial damage after treated with LCE. A549 cells treated with Bi6727 and LCE showed increased expression of Bax, Caspase-3 and Caspase-9 proteins from Western blot assay. LCE also inhibited A549 cells growth keeping cells at G2-M phase from cell cycle assay. Apoptosis assay results showed that LCE induced late apoptosis of A549 cells. JC-1 assay showed that the mitochondria damaged at the LCE concentration of 0.4 mg/mL. In our preliminary anti-proliferation test of combined LCE and Bi-6727 on A549 cells, we found a dramatically decrease in proliferation after treated with LCE first for 24-h and then Bi-6727 for extra 24-h. This was an important finding regarding synergistic anti-proliferation effect of these drugs, However, the usage, the application sequence of LCE and Bi-6727 on A549 cells and their related mechanisms still need to be evaluated. In summary, the drugs exerted anti-proliferation effect on A549 cells independently. We hopefully combine the usage of these two drugs will bring a different and potential outcome in treating lung cancer.

Keywords: anti-proliferation, A549, Livistona Chinensis fruit extracts, PLK-1 inhibitor

Procedia PDF Downloads 112

Authors: Xingxin Wu, Fenli Shao, Tao Tan, Yang Tan, Yang Sun, Qiang Xu

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Psoriasis is a chronic inflammatory skin disease that affects about 2% of the world's population. IL-23 signaling plays a key role in the pathogenesis of psoriasis. Control of IL-23 release by small molecule compounds during developing psoriasis has not been well established. Here, we show that compound 1, a small molecule nature product, protected mice from imiquimod-induced psoriasis with improved skin lesions, reduced skin thickness, and reduced IL-23 mRNA expression in the skin tissue. FACS results showed compound 1 reduced the number of dendritic cells in the skin. Interestingly, compound 1 was not able to ameliorate IL-23-induced psoriasis-like skin inflammation in mice. Further, compound 1 inhibited MyD88-dependent IL-23 mRNA expression induced by LPS, CpG and imiquimod in BMDC cells, but not MyD88-independent CD80 and CD86 expression induced by LPS. The methods included real-time PCR, western blot, H & E staining, FACS and ELISA et al. In conclusion, compound 1 regulates MyD88-dependent signaling to control IL-23 release in dendritic cells, which improves imiquimod-induced psoriasis.

Keywords: dendritic cells, IL-23, toll-like receptor signaling, psoriasis

Procedia PDF Downloads 614

Authors: Noraziah Nordin, Najihah Mohd Hashim, Nazia Abdul Majid, Mashitoh Abdul Rahman, Hamed Karimian, Hapipah Mohd Ali

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Enicosanthellum pulchrum belongs to family Annonaceae is also known as family of 'mempisang' in Malaysia. Liriodenine was isolated by prep-HPLC method. This method was first technique used for the isolation of this compound. The structure of the liriodenine was elucidated by 1D and 2D spectroscopy techniques. Liriodenine was tested on ovarian cancer cells line (CAOV-3) for MTT, AO/PI and cytotoxicity 3 assays. The MTT assay was performed to determine the cytotoxicity effect of lirodenine on CAOV-3 cells. The morphological changes on CAOV-3 cells were observed by AO/PI assay for the early and late stage of apoptosis, as well as necrosis. Meanwhile, the measurement of cell loss, nuclear morphology, DNA content, cell membrane permeability, mitochondrial membrane potential changes and cytochrome c release from mitochondria were detected through cytotoxicity 3 assay. The IC50 results showed liriodenine inhibits the growth of CAOV-3 cells after 24 h of treatment at 10.25 ± 1.06 µg/mL. After 48 and 72 h of treatments, the IC50 values were decreased to 7.65 ± 0:07 and 6.35 ± 1.62 µg/mL, respectively. The morphology changes can be seen on CAOV-3 with a production of cell membrane blebbing, cromatin condensation and apoptotic bodies with increasing time of treatment from 24 to 72 h. Evaluation of cytotoxicity 3 on CAOV-3 cells after treated with liriodenine, resulting loss of mitochondrial membrane potential and release of cytochrome c from mitochondria. The results demonstrated the capability of liriodenine as a promising anticancer agent, particularly on human ovarian cancer.

Keywords: Enicosanthellum pulchrum, ovarian cancer, apoptosis, cytotoxicity

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