scholarly journals The effects and mechanism of peiminine-induced apoptosis in human hepatocellular carcinoma HepG2 cells

2018 ◽  
Author(s):  
Xu Chao ◽  
Guoquan Wang ◽  
Yuping Tang ◽  
Changhu Dong ◽  
Hong Li ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Flow Cytometry ◽  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Hepg2 Cells ◽  
Mitochondrial Membrane ◽  
Molecular Mechanisms ◽  
Human Hepatocellular Carcinoma ◽  
Nuclear Staining ◽  
Induced Apoptosis

AbstractPeiminine is a compound that is isolated fromBolbostemma paniculatum(Maxim) Franquet (Cucurbitaceae family), which has demonstrated antitumor activities. Its precise molecular mechanisms underlying antitumor activity remain elusive. In this study, peiminine-induced apoptosis towards human hepatocellular carcinoma and its molecular mechanisms were investigated. MTT assay was employed to assess anticancer effects of peiminine at concentrations of 2, 4, 6, 8, 10, 12, and 14 μg/ml after 24, 48, or 72 h. Nuclear staining and flow cytometry were carried out to further assess apoptosis. Mitochondrial membrane potential evaluation and Western blot analysis were performed to investigate the mechanism of peiminine-induced apoptosis. Peiminine reduced the viability of HepG2 cells in a time- and dose-dependent manner and had an IC50of 4.58 μg/mL at 24h. Flow cytometry assessment indicated that peiminine markedly increased the cell number of apoptotic cells and the mitochondrial membrane potential dose-dependently in HepG2 cells. The results of Western blotting showed the expression of Bcl-2, procaspase-3, procaspase-8, procaspase-9, and PARP1decreased in HepG2 cells treated with peiminine, while the expression of Bax, caspase-3, caspase-8, caspase-9, and cleaved PARP1increased. The result suggest taht peiminine can induce apoptosis in human hepatocellular carcinoma HepG2 cells through both extrinsic and intrinsic apoptotic pathways.

scholarly journals Autoregulatory Feedback Mechanism of P38MAPK/Caspase-8 in Photodynamic Therapy-Hydrophilic/Lipophilic Tetra-α-(4-carboxyphenoxy) Phthalocyanine Zinc-Induced Apoptosis of Human Hepatocellular Carcinoma Bel-7402 Cells

2014 ◽  
Vol 2014 ◽  
pp. 1-9 ◽  
Author(s):  
Yu Wang ◽  
Chunhui Xia ◽  
Wei Chen ◽  
Yuhang Chen ◽  
Yiyi Wang ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Photodynamic Therapy ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Caspase 3 ◽  
Human Hepatocellular Carcinoma ◽  
Caspase 8 ◽  
Induced Apoptosis

Photodynamic therapy (PDT) is a novel and promising antitumor treatment. Our previous study showed that hydrophilic/lipophilic tetra-α-(4-carboxyphenoxy) phthalocyanine zinc- (TαPcZn-) mediated PDT (TαPcZn-PDT) inhibits the proliferation of human hepatocellular carcinoma Bel-7402 cells by triggering apoptosis and arresting cell cycle. However, mechanisms of TαPcZn-PDT-induced apoptosis of Bel-7402 cells have not been fully clarified. In the present study, therefore, effect of TαPcZn-PDT on apoptosis, P38MAPK, p-P38MAPK, Caspase-8, Caspase-3, Bcl-2, Bid, Cytochrome c, and mitochondria membrane potential in Bel-7402 cells without or with P38MAPK inhibitor SB203580 or Caspase-8 inhibitor Ac-IEFD-CHO was investigated by haematoxylin and eosin (HE) staining assay, flow cytometry analysis of annexin V-FITC/propidium iodide (PI) double staining cells and 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolylcarbocyanine iodide (JC-1), and immunoblot assay. We found that TαPcZn-PDT resulted in apoptosis induction, activation of P38MAPK, Caspase-8, Caspase-3, and Bid, downregulation of Bcl-2, release of Cytochrome c from mitochondria, and disruption of mitochondrial membrane potential in TαPcZn-PDT-treated Bel-7402 cells. In contrast, SB203580 or Ac-IEFD-CHO attenuated induction of apoptosis, activation of P38MAPK, Caspase-8, Caspase-3, and Bid, downregulation of Bcl-2, release of Cytochrome c from mitochondria, and disruption of mitochondrial membrane potential in TαPcZn-PDT-treated Bel-7402 cells. Taken together, we conclude that Caspase-3, Bcl-2, Bid, and mitochondria are involved in autoregulatory feedback of P38MAPK/Caspase-8 during TαPcZn-PDT-induced apoptosis of Bel-7402 cells.

scholarly journals The Short Isoform of DNAJB6 Protects against 1-Methyl-4-phenylpridinium Ion-Induced Apoptosis in LN18 Cells via Inhibiting Both ROS Formation and Mitochondrial Membrane Potential Loss

2017 ◽  
Vol 2017 ◽  
pp. 1-12 ◽  
Author(s):  
Yeon-Mi Hong ◽  
Yohan Hong ◽  
Yeong-Gon Choi ◽  
Sujung Yeo ◽  
Soo Hee Jin ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Molecular Mechanisms ◽  
Mitochondrial Pathway ◽  
Caspase 9 ◽  
Ros Formation ◽  
Mitochondrial Membrane Potential Loss ◽  
Induced Apoptosis ◽  
Shock Proteins

In a previous study, we found that the short isoform of DNAJB6 (DNAJB6(S)) had been decreased in the striatum of a mouse model of Parkinson’s disease (PD) induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). DNAJB6, one of the heat shock proteins, has been implicated in the pathogenesis of PD. In this study, we explored the cytoprotective effect of DNAJB6(S) against 1-methyl-4-phenylpyridinium ion- (MPP+-) induced apoptosis and the underlying molecular mechanisms in cultured LN18 cells from astrocytic tumors. We observed that MPP+ significantly reduced the cell viability and induced apoptosis in LN18 glioblastoma cells. DNAJB6(S) protected LN18 cells against MPP+-induced apoptosis not only by suppressing Bax cleavage but also by inhibiting a series of apoptotic events including loss of mitochondrial membrane potential, increase in intracellular reactive oxygen species, and activation of caspase-9. These observations suggest that the cytoprotective effects of DNAJB6(S) may be mediated, at least in part, by the mitochondrial pathway of apoptosis.

scholarly journals Chitopentaose inhibits hepatocellular carcinoma by inducing mitochondrial mediated apoptosis and suppressing protective autophagy

2021 ◽  
Vol 8 (1) ◽  
Author(s):  
Chunfeng Zhu ◽  
Mengyao Zhao ◽  
Liqiang Fan ◽  
Xuni Cao ◽  
Quanming Xia ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Membrane Potential ◽  
Cell Viability ◽  
Mtt Assay ◽  
Hepg2 Cells ◽  
Mitochondrial Membrane ◽  
Molecular Mechanisms ◽  
Degree Of Polymerization ◽  
Intrinsic Pathway ◽  
Autophagy Flux

AbstractHepatocellular carcinoma (HCC) is one of the most prevalent and deadliest cancers. In this study, the anti-tumor effect of singular degree of polymerization (DP) chitooligosaccharides (COS) (DP 2–5) and the underlay molecular mechanisms were investigated on HCC cell line HepG2. MTT assay showed that (GlcN)5 have the best anti-proliferation effect among the different DP of COS (DP2-5). Furthermore, the administration of (GlcN)5 could decrease mitochondrial membrane potential, release cytochrome c into cytoplasm, activate the cleavage of Caspases9/3, thus inducing mitochondrial-mediated apoptosis in HepG2 cells (accounting for 24.57 ± 2.25%). In addition, (GlcN)5 treatment could increase the accumulation of autophagosomes. Further investigation showed that (GlcN)5 suppressed protective autophagy at the fusion of autophagosomes and lysosomes. Moreover, the inhibition of protective autophagy flux by (GlcN)5 could further decrease cell viability and increase the apoptosis rate. Our findings suggested that (GlcN)5 suppressed HepG2 proliferation through inducing apoptosis via the intrinsic pathway and impairing cell-protective autophagy. COS might have the potential to be an agent for lowering the risk of HCC.

Cucurbitacin IIb from Ibervillea sonorae Induces Apoptosis and Cell Cycle Arrest via STAT3 Inhibition

2020 ◽  
Vol 20 (10) ◽  
pp. 1188-1196
Author(s):  
Heriberto Torres-Moreno ◽  
Maria C. Marcotullio ◽  
Carlos Velazquez ◽  
Víctor M. Arenas-Luna ◽  
Salomón Hernández-Gutiérrez ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Flow Cytometry ◽  
Dna Damage ◽  
Membrane Potential ◽  
Cell Cycle Arrest ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Molecular Mechanisms ◽  
Cyclin B1 ◽  
Cycle Arrest

Background: Cucurbitacin IIb (CIIb) from Ibervillea sonorae has a high capacity to suppress cancer cell proliferation and induce apoptosis. This study investigated the molecular mechanisms related to the antiproliferative and apoptosis induction capacity of CIIb in HeLa cells. Materials and Methods: The cell viability and anti-proliferative effect of CIIb were evaluated by using the trypan blue exclusion assay. The effect of CIIb on the mitochondrial membrane potential was determined by flow cytometry using JC-1. The activity of caspase-3 and caspase-9 was evaluated by flow cytometry using commercial kits. The effect of CIIb on the cell cycle was investigated using Fluorescence-Activated Cell Sorting (FACS) analysis. Western blot analysis was used to evaluate both the inhibitory effect of CIIb on the STAT3 signaling pathway and cyclin –B1, and DNA damage by the comet assay. Results: CIIb triggers disruption of the mitochondrial membrane potential (Δψm) and consequently activated the caspases -3 and -9, as a result of the activation of the intrinsic pathway of the apoptosis. Likewise, the CIIbinduced cell cycle was arrested in S and G2/M after 24h of treatment. CIIb also reduced the expression of STAT3 and cyclin –B1. Finally, CIIb produced an antiproliferative effect at 48 and 72 h, inducing DNA damage. Conclusion: These results demonstrate CIIb-induced apoptosis and cell cycle arrest in HeLa through the inhibition of STAT3.

scholarly journals Osthole Induces Apoptosis, Secondary Necrosis and Mitophagy Via NQO1-Mediated ROS Overproduction in HeLa Cells

2020 ◽  
Author(s):  
Mengjie Huangfu ◽  
Juan Wang ◽  
Dan Yu ◽  
Jianli Qin ◽  
Xiao Guan ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Flow Cytometry ◽  
Membrane Potential ◽  
Cancer Cells ◽  
Mitochondrial Membrane Potential ◽  
Hela Cells ◽  
Mitochondrial Membrane ◽  
Oxygen Species ◽  
Secondary Necrosis ◽  
Induced Apoptosis

Abstract Background: Osthole is a natural coumarin which has been proved to inhibit growth of cancer cells by inducing cancer cells death, while its mechanism of anticancer remains unclearly. In our study, we found that osthole activated multiple forms of cell death including apoptosis, secondary necrosis and mitophagy in receptor interacting protein kinase (RIP) 3-deficient cervical cancer HeLa cells. Methods: Cell viability was detected by MTT assay. Cell membrane integrity was detected by LDH release assay and PI staining. Cell apoptosis and necrosis were detected by flow cytometry assay. Reactive oxygen species (ROS) was detected by DCFH-DA staining and mitochondrial membrane potential (MMP) was detected by JC-1 staining using flow cytometry. The expression of proteins was detected by western blotting assay and proteomics. Xenograft tumor model was used to evaluate the effect of osthole in vivo.Results: Our study showed osthole caused HeLa cells apoptosis and secondary necrosis, which is a phenomenon of the apoptotic cells’ plasma membrane breakdown. And when Hela cells pretreatment with Z-DEVD-FMK, an irreversible caspase-3 inhibitor, not only inhibited osthole-induced apoptosis but also necrosis. Moreover, we found that Z-DEVD-FMK reversed the effect of osthole on the induction of cleaved the N-terminal fragment of GSDME in Hela cells. Furthermore, inhibition of NAD (P) H: quinone oxidoreductase 1 (NQO1) by osthole induced the overproduction of reactive oxygen species (ROS). ROS inhibitor N-Acetyl-L-cysteine (NAC) not only reduced osthole-induced apoptosis, but also reversed its effect on the necrotic induction and the GSDME N-terminal generation. It was shown that osthole decreased mitochondrial membrane potential (MMP) and increased the expression of PTEN-induced putative kinase 1 (PINK1) and Parkin, which indicated that the activation of mitophagy induced by osthole. Meanwhile, as well as apoptosis and secondary necrosis, mitophagy was also restrained by NAC. Conclusions: In conclusion, all these data suggested that osthole induced apoptosis, secondary necrosis and mitophagy via NQO1-mediated ROS overproduction.

Platelets apoptosis in rats after global brain ischemia

2018 ◽  
pp. 27-35
Author(s):  
А.А. Соколовская ◽  
Э.Д. Вирюс ◽  
В.В. Александрин ◽  
А.С. Роткина ◽  
К.А. Никифорова ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Ischemic Stroke ◽  
Membrane Potential ◽  
Mitochondrial Membrane Potential ◽  
Brain Ischemia ◽  
Mitochondrial Membrane ◽  
Male Rats ◽  
Global Brain Ischemia ◽  
Platelet Apoptosis ◽  
Global Brain

Цель исследования. Ишемические повреждения головного мозга, являются одной из наиболее частой причин инвалидности и смертности во всем мире. Недавно была установлена роль апоптоза тромбоцитов в патофизиологии инсульта, однако его механизмы до сих пор остаются невыясненными. Несмотря на различные экспериментальные модели, направленные на мониторинг апоптоза тромбоцитов, результаты, относительно изучения и выявления апоптоза тромбоцитов при ишемии головного мозга у крыс, весьма немногочисленны. Цель исследования - анализ апоптоза тромбоцитов с помощью метода проточной цитофлуориметрии на модели глобальной ишемии мозга у крыс. Методика. В экспериментах использовано 6 крыс-самцов Вистар в возрасте от 5 до 6 мес., разделенных на 2 группы: интактный контроль (К) и глобальная ишемия головного мозга. Модель глобальной ишемии головного мозга у крыс воспроизводилась путём билатеральной окклюзии общих сонных артерий на фоне гипотензии. Уровень системного артериального давления снижали посредством кровопотери до 40-45 мм рт. ст. Суспензию тромбоцитов крыс получали методом гельфильтрации с использованием сефарозы 2B. Для анализа экстернализации фосфатидилсерина (ФС) тромбоциты крыс инкубировали с Аннексином V-PE в связывающем буфере. Для оценки митохондриального мембранного потенциала (ММП) тромбоциты инкубировали с катионным красителем JC-1. После инкубации образцы немедленно анализировали на проточном цитофлуориметре FACSCalibur (Becton Dickinson, США). Результаты. Согласно полученным данным, экстернализация ФС на тромбоцитах крыс, перенесших инсульт, была значительно выше (53,45 ± 4,21%), чем в контрольной группе крыс (5,27 ± 2,40%). Данный эффект подтверждается выраженной деполяризацией митохондриальных мембран (DYm). После экспериментальной ишемии мозга почти 40% тромбоцитов было деполяризовано. Заключение. Использованный в работе подбор методов и маркеров обеспечивает понимание механизмов апоптоза тромбоцитов как в экспериментальных, так и в клинических условиях. Полученные данные позволяют сделать заключение, что апоптоз тромбоцитов является одним из факторов развития глобальной ишемии головного мозга у крыс. Результаты могут быть использованы для понимания механизмов, участвующих в развитии ишемического повреждения, что, в свою очередь, может быть использовано при разработке новых терапевтических стратегий. Aim. Stroke is one of the most common causes of disability and mortality worldwide. Multiple experimental models of stroke have focused on monitoring of platelet apoptosis. However, studies on and detection of platelet apoptosis in rats with ischemic stroke are very scarce. We investigated platelet apoptosis in rats with global brain ischemia using flow cytometry. Methods. Experiments were carried out on healthy, adult Wistar male rats weighing 300-350 g. The rats were divided into the following 2 groups: intact rats and rats with global brain ischemia. Global brain ischemia was induced by two-vessel (2-VO) carotid occlusion in combination with hypotension. Systemic blood pressure was reduced by 40-45 mm Hg by inducing haemorrhage. Platelets were isolated by gel filtration on Sepharose 2B. For evaluation of phosphatidylserine (PS) externalization, platelets were incubated with Annexin V-PE and analyzed on FACSCalibur (BD Biosciences). Mitochondrial membrane potential (DY) was measured during platelets apoptosis using JC-1, a mitochondrial membrane potential indicator. Platelets were analyzed by flow cytometry immediately after the incubation. Results. PS externalization on platelets was significantly greater after global brain ischemia (53.45 ± 4.21%) than in the control group (5.27 ± 2.40%). Pronounced depolarization of mitochondrial membrane potential (DYm) confirmed this finding. In the rat group with experimental brain ischemia, almost 40% (35.24 ± 5.21%) of platelets were depolarized. Conclusion. Our results provide insight into mechanisms involved in platelet apoptosis during ischemic stroke and can be used in further development of new therapeutic strategies.

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