Caspase-dependent Mitochondrial Pathway Research Articles

Effect of co-infection with Newcastle disease virus on Mycoplasma gallisepticum pathogenesis in vivo and in vitro

The co-infection of Newcastle disease virus (NDV) and Mycoplasma gallisepticum (MG) has a detrimental effect on chicken production performance, exerts a deleterious impact on poultry production performance, resulting in substantial economic losses. However, the exact impact and underlying mechanisms remain ambiguous. In this study, co-infection models were established both in vivo and in vitro. Through these models, it was found that the co-infection facilitated the replication of MG and NDV, as well as MG induced pathogenesis. The administration of lentogenic NDV resulted in the suppression of the innate immune response in vivo. At cellular level, co-infection promoted MG induced apoptosis through caspase-dependent mitochondrial endogenous pathway and suppressed the inflammatory secretion. This research contributes novel insights in co-infection.

Organophosphorus Flame Retardant TPP-Induced Human Corneal Epithelial Cell Apoptosis through Caspase-Dependent Mitochondrial Pathway.

A widely used organophosphate flame retardant (OPFR), triphenyl phosphate (TPP), is frequently detected in various environmental media and humans. However, there is little known on the human corneal epithelium of health risk when exposed to TPP. In this study, human normal corneal epithelial cells (HCECs) were used to investigate the cell viability, morphology, apoptosis, and mitochondrial membrane potential after they were exposed to TPP, as well as their underlying molecular mechanisms. We found that TPP decreased cell viability in a concentration-dependent manner, with a half maximal inhibitory concentration (IC50) of 220 μM. Furthermore, TPP significantly induced HCEC apoptosis, decreased mitochondrial membrane potential in a dose-dependent manner, and changed the mRNA levels of the apoptosis biomarker genes (Cyt c, Caspase-9, Caspase-3, Bcl-2, and Bax). The results showed that TPP induced cytotoxicity in HCECs, eventually leading to apoptosis and changes in mitochondrial membrane potential. In addition, the caspase-dependent mitochondrial pathways may be involved in TPP-induced HCEC apoptosis. This study provides a reference for the human corneal toxicity of TPP, indicating that the risks of OPFR to human health cannot be ignored.

Synergistic inhibitory actions of resveratrol, epigallocatechin-3-gallate, and diallyl trisulfide against skin cancer cell line A431 through mitochondrial caspase dependent pathway: a combinational drug approach.

The harmful effect of chemotherapeutic side effects has paid a way to discover a novel with curative way for skin cancer treatment. Skin cancer prevention is more viable with the use of combination of bioactive agents than using of single bioactive compounds. Present work was demonstrated to evaluate the interaction of Resveratrol (Res), Epigallocatechin-3-gallate (EGCG), and diallyl trisulfide (DATS) with each other as a binary combination on A431 cells. Nuclear fragmentation analysis of combination of bioactive agents using DAPI analysis, detection of apoptosis, analysis of cell cycle, ROS assay, antimigration assays, and western blotting were implemented to study the combination of bioactive compounds on A431 cell line. Among the selected combination EGCG + DATS had a synergetic effect reducing cellular migration, increased intercellular reactive oxygen species generation, condensation, cell phagocytosis induced by phosphatidylserine externalization, rise in sub-G1 DNA content, and S-phase were cell cycle arrest. The combinations EGCG + DATS induced apoptotic proteins in A431 cells by upregulation of proapoptotic Bax and Bad proteins, a downmodulation of anti-apoptotic proteins Bcl2 and caspases (caspase-3, and -9) activity got triggered by intrinsic pathway. The combination of EGCG + DATS showed good anticancer potential against A431 skin cancer cell line via the mitochondrial caspase dependent pathway with very strong synergism. This finding will help to produce a novel combination/chemoprevention using dietary bioactive agents (EGCG + DATS) for the treatment of skin cancer after clinical trial.

Anti-tumor effect and mechanism of the total biflavonoid extract from S doederleinii on human cervical cancer cells in vitro and in vivo

In this study, the therapeutic effect and possible mechanism of the total biflavonoid extract of Selaginella doederleinii Hieron (SDTBE) against cervical cancer were originally investigated in vitro and in vivo. First, the inhibition of SDTBE on proliferation of cervical cancer HeLa cells was evaluated, followed by morphological observation with AO/EB staining, Annexin V/PI assay, and autophagic flux monitoring to evaluate the possible effect of SDTBE on cell apoptosis and autophagy. Cell cycle, as well as mitochondrial membrane potential (ΔѰm), was detected with flow cytometry. Further, the apoptosis related protein expression and the autophagy related gene LC3 mRNA transcription level were analyzed by Western blot (WB) and real-time quantitative polymerase chain reaction (RT-qPCR), respectively. Finally, the anti-cervical cancer effect of the SDTBE was also validated in vivo in HeLa cells grafts mice. As results, SDTBE inhibited HeLa cells proliferation with the IC50 values of 49.05 ± 6.76 and 44.14 ± 4.75 μg/mL for 48 and 72 h treatment, respectively. The extract caused mitochondrial ΔѰ loss, induced cell apoptosis by upregulating Bax, downregulating Bcl-2, activating Caspase-9 and Caspase-3, promoting cell autophagy and blocking the cell cycle in G0/G1 phase. Furthermore, 100, 200, and 300 mg/kg SDTBE suppressed the growth of HeLa cells xenografts in mice with the mean inhibition rates, 25.3 %, 57.5 % and 62.9 %, respectively, and the change of apoptosis related proteins and microvascular density was confirmed in xenografts by immunohistochemistry analysis. The results show that SDTBE possesses anti-cervical cancer effect, and the mechanism involves in activating Caspase-dependent mitochondrial apoptosis pathway.

Rhein suppresses African swine fever virus replication in vitro via activating the caspase-dependent mitochondrial apoptosis pathway

African swine fever (ASF) is a virulent infectious diseases of pigs caused by the African swine fever virus (ASFV) that can spread widely and cause high fatality rates. Currently, there is no effective way to treat the disease, and there is no effective vaccine to prevent it. Rhein, an anthraquinone compound extracted from many traditional Chinese medicines, exhibits anti-inflammatory, anti-tumor, and anti-viral activities. However, the anti-viral effects of rhein on ASFV remain unclear. Therefore, this study aimed to investigate the anti-ASFV activity of rhein in porcine alveolar macrophages (PAMs) and the underlying mechanisms. In this study, we confirmed that rhein inhibits ASFV replication significantly in a dose-dependent manner in vitro. Moreover, rhein could alter the susceptibility of PAMs to ASFV and promoted the production of superoxide in the mitochondria, which induced the loss of mitochondrial membrane potential, leading to the activation of caspase-9, caspase-3, and apoptosis. Mito-TEMPO, a mitochondria-targeted antioxidant, blocked rhein-induced mitochondrial superoxide generation and loss of mitochondrial membrane potential, prevented caspase-9 and caspase-3 activation, alleviated apoptosis, and suppressed the anti-ASFV activity of rhein. Altogether, our results suggested that rhein could play an anti-ASFV role by inducing apoptosis through the activation of the caspase-dependent mitochondrial apoptotic pathway and may provide a novel compound for developing anti-ASFV drugs.

Cis-vaccinic acid, Dibutyl-Cyanamide and 9, 12-Octadecadienoic acid from pistachio (Pistachia vera) seed coat (PSCE) induced nuclear damage and cytotoxicity in human colon cancer HT-115 cells

Current strategy for colorectal cancer (CRC) therapy like regular intake of non-steroidal anti-inflammatory drugs (NSAIDs) may reduce the risk of developing CRC but they induce the regression of adenomas. Even the CRC management technique include colectomy, have not yet been proven to reduce mortality. The present study aimed to reduce the susceptibility of CRC via controlling cellular oxidative stress, inflammation and caspase depended apoptosis in HT-115 human colorectal cancer cells using pistachio seed coat extract as nutritional therapy. Increasing concentrations of pistachio seed coat ethyl acetate extract (PSCE) were applied to HT-115 cells, which were incubated for 24 h and 48 h. The IC50 values were 14 mg/dl after 24 h, and and 7.5 mg/dl after 48 h. Acridine orange/ ethidium bromide staining confirmed the presence of 32% early apoptotic, 27% pre-apoptotic, 12% apoptotic, and 6% necrotic cells after 48 h. PSCE at 14 mg/dl significantly increased the antioxidant capacity via the expression of CYP1A and GSK3β, and decreased inflammatory agents via decreasing NF-κb, TNF-α, COX-2 and PGE-2 expression three-fold after 48 h. The expression of the tumor suppression related genes p53 and mdm2, and the apoptosis related genes Bax, Bcl-2, Caspase 3, p21, and PCNA levels increased one-fold, and levels of mdm2, Bcl-2 and PCNA decreased after PME treatment of 48 h. PSCE effectively controlled colon cancer cell proliferation via the caspase-dependent mitochondrial mediated apoptotic pathway.
 Keywords: Pistachia vera, cytotoxicity, human colon cancer, Cis-Vaccininc acid, apoptosis

Non-thermal plasma inhibited the growth and aflatoxins production of Aspergillus flavus, degraded aflatoxin B1 and its potential mechanisms

Aspergillus flavus (A. flavus) and its toxic metabolites, aflatoxins, are widely distributed in agricultural products and foods, posing a grave threat to food safety and public health. This study investigated the decontamination effects and mechanism of non-thermal plasma (NTP) on A. flavus and aflatoxins. NTP primarily generated reactive oxygen and nitrogen species (RONS), effectively inactivating A. flavus spores via the Weibull + Tail model. The results of ultrastructural observation and membrane integrity revealed that fungal inactivation by NTP was mainly attributed to intracellular destruction rather than the direct damage of cell membranes. Furthermore, NTP induced the accumulation of intracellular ROS and NO, disrupting anti-oxidative and energy metabolic systems, thereby inducing fungal apoptosis via a caspase-dependent mitochondrial pathway, which is responsible for A. flavus spore inactivation. In addition, the results showed that NTP reduced aflatoxin B1 (AFB1) production by regulating the expression of key genes in the aflatoxin biosynthetic pathway. Moreover, NTP effectively degraded AFB1 by disintegrating the furan ring, modifying the methoxy group, and forming the double bond in cyclopentanone, thereby decreasing the toxicity of AFB1. Hence, NTP exhibits exceptional decontamination effects against A. flavus and AFB1 via fungal inactivation, aflatoxin biosynthesis suppression, and AFB1 degradation, making it a viable alternative strategy for agricultural disinfection.

Effect of the extra-nuclear cation on the cytotoxicity and mechanism of action of pyridine-2,6-dicarboxylate Ga(III) complexes

Two Ga(III) complexes (C1) and (C2) were prepared by the one-pot reaction of pyridine-2,6-dicarboxylic acid and aminopyridine derivatives with gallium(III) nitrate octahydrate. The compounds were characterized by single-crystal X-ray diffraction. The distorted octahedral geometry was confirmed by crystallographic data for both complexes. The study of the in vitro cytotoxicity of the compounds showed that the presence of different extra-nuclear cations can affect the cytotoxicity of the same anionic complexes. The most significant antiproliferative activity was observed for C1 (IC50 = 0.69 μM, MAE = 73.96%) and C2 (IC50 = 3.78 μM, MAE = 60.35%) (where MAE represents the maximal antiproliferative effect) against A431 cell line. The mechanistic study evidenced the same pathway for the death of A431 cells treated with the complexes, although the results for C2 were obtained at approximately five times the concentration of C1. According to the study, both complexes induced cell cycle arrest in G2/M phase in A431 cells by upregulating the levels of p21, p27, p-cdc25C, and p-cdc2 and downregulating the levels of cdc25C, cdc2, and cyclin B1. In addition, apoptosis via a caspase-dependent mitochondrial pathway was confirmed by a decrease in Bcl-2 family proteins and an increase in the expression of procaspase-9 and 3. Also, the complexes induced autophagic cell death by activating the RAGE /PI3KC3/Beclin 1 pathway in A431 cells. Data availabilityCCDC 874052 and 874055 contain the supplementary crystallographic data for C1 and C2, respectively. These data can be obtained free of charge via http://www.ccdc.cam.ac.uk/services/structures?pid=ccdc:874052,874055&sid=CCDCManual, or from the Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge CB2 1EZ, UK; fax: (+44) 1223–336–033; or e-mail: deposit@ccdc.cam.ac.uk.

Anticancer Mechanism of Parthenolide Analogue PTLA-1 on Adriamycin Resistant Human Chronic Myeloid Leukemia Cell Line K562/ADR

Objective: To investigate the anticancer effect and mechanism of parthenolide analogue (PTLA-1) on adriamycin resistant human chronic myeloid leukemia cell line K562/ADR cells. Methods K562/ADR cells were cultured in vitro. The inhibitory effects of PTLA-1 on cell proliferation were detected by MTT assay, and the effects of PTLA-1 on K562/ADR cell cycle, apoptosis, mitochondrial membrane potential (ΔΨm) and reactive oxygen species were detected by flow cytometry. The effects of PTLA-1 on K562/ADR cell cycle and apoptosis related proteins expressions were detected by Western blot. Results PTLA-1 had higher inhibitory activity on K562/ADR cells than parthenolide. It inhibited the expressions of Cyclin B1/D1 and CDK1/2/4/6, resulting in the arrest of K562/ADR cells in G0/G1 phase; PTLA-1 induced apoptosis of K562/ADR cells in mitochondrial pathway. PTLA-1 could induce mitochondrial membrane potential (ΔΨm) and down regulate the expressions of reactive oxygen species, Bcl-2 and p-Akt proteins, and up regulate the expressions of Bax, cyto c, caspase 9/3 proteins. The chromatin of K562/ADR cells contracted and apoptotic bodies appeared. Conclusion Parthenolide analogue PTLA-1 could inhibit cell proliferation by causing K562/ADR cells to block in G0/G1 phase. Secondly, it could induce K562/ADR cells to apoptosis through caspase dependent mitochondrial pathway, which was related to Akt signal pathway. It had certain anticancer therapeutic potential for chronic myeloid leukemia.

Bioactive chemical constituents, in vitro anti-proliferative activity and in vivo toxicity of the extract of Curcuma singularis Gagnep rhizomes

Ethnopharmacological relevanceCurcuma singularis Gagnep is a Vietnamese medicinal plant which has been commonly used as a medicinal remedy in traditional and folk medicines for improving health as well as for treating some diseases, like rheumatoid arthritis, kidney failure. However, pharmacological effects, including anti-cancer activity and the safety of this plant has not been fully investigated. Aim of the studyThis study aimed to investigate the in vitro anti-growth activity of an extract derived from Curcuma singularis rhizome extract (CSE) against cell lines as well as determine its phytochemical composition. The other goal of our study was to assess the safety of CSE in rats. Materials and methodsThe main constituents in the extract were identified and quantitatively analyzed. The in vitro cytotoxicity of CSE was evaluated in several cancer and normal cell lines. The apoptotic activity of CSE and the expression of the apoptosis-related genes were investigated in AGS cells to clarify the underlying molecular mechanisms. The in vivo toxicity of CSE was assessed via acute and subacute oral studies on Sprague-Dawley rats, respectively according to the guidelines 425 and 407 of the Organization for Economic Cooperation and Development (OECD). The drug-related toxicity signs, mortality, body and organ weights were recoreded during the experimental period. In addition, the selected hematological and biochemical parameters, and histological alterations were determined at the end of the subacute toxicity test. ResultsGermacrone, ar-turmerone, and curcumol were three sesquiterpene components found in the extract. CSE showed cytotoxic effects in different cancer cells, but had minimal effects on normal cells. Apoptosis in AGS cells was caused by CSE in a concentration-dependent pattern through increase of Bax/Bcl-2 ratio, and release of cytochrome c, which leads to activation of caspase-3/-7, caspase-9, as well as cleavage of PARP. In the acute toxicity test, no signs of toxicity and no mortality were recorded in rats at both doses of 1000 and 5000 mg/kg. In the subacute toxicity study, CSE showed no drug-related adverse effects on water and food consumption, body and organ weights. CSE at a dose of 1000 mg/kg slightly increased WBC and platelet values in female rats, while it increased WBC values in male rats in all tested doses. The decrease of total cholesterol and triglyceride levels were found in female rats treated CSE at doses of 250 or 500 mg/kg. In addition, the increase of serum ALT and AST levels in rats treated at the dose of 1000 mg/kg were noted. No significant changes in histopathological structures of kidneys, spleen, heart and lungs, except liver tissue with minor modifications was found. ConclusionsOur findings indicated that CSE exhibited in vitro anti-proliferative effects on AGS cells by mainly activating the caspase-dependent mitochondrial apoptotic pathway. CSE also showed in vivo toxicity signals at the dose of 1000 mg/kg with proven minor hepatic injuries, which should be avoided the high dose for prolonged use. Curcuma singularis rhizomes may be used as a chemotherapeutic agent for the treatment of gastric cancer with in vitro anti-cancer investigation and in vivo biological safety evaluation.

Cadmium-induced oxidative stress in Meretrix meretrix gills leads to mitochondria-mediated apoptosis.

Cadmium (Cd) is one of the most important marine environmental pollutants that can cause oxidative damage and apoptosis in living organisms, and mitochondria are the key cell organelles affected by Cd toxicity. In this study, we investigated the effect of Cd on the mitochondria in the gill cells of the clam Meretrix meretrix and the underlying mechanism of mitochondria-mediated apoptosis following exposure to the metal. Exposure of the clams to artificial seawater containing 1.5, 3, 6 and 12 mg L-1 Cd2+ led to swollen mitochondria compared with the untreated clams. The mitochondria also became vacuolated at the higher Cd2+ concentrations. Biochemical assays showed that monoamine oxidase (MAO) activity and mitochondrial membrane potential (Δψm) increased at 1.5 mg L-1 Cd2+, but decreased at higher Cd2+ concentrations, while the activities of malate dehydrogenase (MDH) and cytochrome oxidase (CCO) and the scavenging capacities of anti-superoxide anion (ASA) and anti-hydroxy radical (AHR) all decreased with increasing Cd2+ concentrations. Significant increases in the levels of malondialdehyde (MDA) and H2O2 as well as in the activity levels of caspase-3, -8, and -9 were also observed in the Cd2+-treated clams. The results implied that Cd might induce apoptosis in M. meretrix via the mitochondrial caspase-dependent pathway.

Berberine-photodynamic therapy sensitizes melanoma cells to cisplatin-induced apoptosis through ROS-mediated P38 MAPK pathways

The clinical use of cisplatin are limited due to its drug resistance. Thus, it is urgent to find effective combination therapy that sensitizes tumor cells to this drug. The combined chemo-photodynamic therapy could increase anti-tumor efficacy while also reduce the side effects of cisplatin. Berberine is an isoquinoline alkaloid, which has been reported to show high photosensitizing activity.In this study, we have examined the effect of a combination of cisplatin and berberine-PDT in cisplatin-resistant melanoma cells. The cytotoxic effects of berberine-PDT alone or in combination with cisplatin were tested by MTT assays. We then examined the subcellular localization of berberine with confocal fluorescence microscopy. The percentage of apoptotic cells, the mitochondrial membrane potential (Δψm) and reactive oxygen species (ROS) generation assessed using flow cytometry analysis. Western blotting used in this study to determine the expression levels of MAPK signaling pathways and apoptosis-related proteins. Experimental data revealed that the mode of cell death is the caspase-dependent mitochondrial apoptotic pathways. Excessive accumulation of ROS played a key role in this process, which is confirmed by alleviation of cytotoxicity upon pretreatment with NAC. Furthermore, we found that the combined treatment activated MAPK signaling pathway. The inhibition of p38 MAPK by pretreating with SB203580 block the combined treatment-induced apoptotic cell death.In conclusion, berberine-PDT could be used as a chemo-sensitizer by promoting cell death through activation of a ROS/p38/caspase cascade.

Toosendanin Shows Potent Efficacy Against Human Ovarian Cancer through Caspase-Dependent Mitochondrial Apoptotic Pathway

Toosendanin (TSN) is a triterpenoid extracted from the bark or fruits of Melia toosendan Sieb et Zucc, which is a traditional Chinese medicine and mainly grows in China and India. TSN has been verified to possess antitumor activities on various human cancers, whereas the effects of TSN on ovarian cancer (OC) has not been reported yet. Here, TSN was shown to significantly inhibit proliferation of SKOV3 and OVCAR3 cell lines in a dose- and time-dependent manner. Treatment of OC cells with TSN resulted in colony formation reduction, S and G2/M phase arrest, cell apoptosis, and dramatic decrease in mitochondrial membrane potential. Furthermore, TSN suppressed invasion and migration of OC cells. Research on molecular mechanism indicated that the above efficacy of TSN was associated with decreased expression of survivin, PARP-1, Bcl-2, Bcl-xl, caspase-3, caspase-9, MMP-2 and MMP-9 and increased expression of cleaved PARP-1, Bax, cleaved caspase-3 and cleaved caspase-9. Finally, in vivo results showed that TSN suppressed OC xenograft tumor growth by inducing apoptosis and regulating the related protein expression levels of SKOV3 cells in transplanted tumors. Taken together, our data provide new insights into TSN as a potentially effective reagent against human OC through caspase-dependent mitochondrial apoptotic pathway.

Green Biosynthesized Silver Nanoparticles With Aqueous Extracts of Ginkgo Biloba Induce Apoptosis via Mitochondrial Pathway in Cervical Cancer Cells.

Biosynthetic silver nanoparticles (AgNPs), specifically formed using medicinal plant extracts, have recently exhibited a remarkable therapeutic effect due to their anticancer potential. Here, we synthesized AgNPs using an aqueous extract of Ginkgo biloba leaves and evaluated its activity against cervical cancer (CCa) and the related molecular mechanisms. The physiochemical properties of the AgNPs were measured by ultraviolet-visible spectrophotometry, nanometre particle size analyzer and transmission electron microscopy. The AgNPs effects on cell proliferation and apoptosis were investigated through MTT, MTS, and colony formation assay; Hoechst 33258 staining; and flow cytometry. The intracellular ROS and oxidative stress levels were assessed using the appropriate commercial kits. Apoptosis-related protein levels were determined by western blotting. We prepared a series of different sized ginkgo extract synthesized AgNPs (GB-AgNPs), and the smallest mean particle size was 40.2 ± 1.2 nm with low polydispersity (0.091 ± 0.011), zeta potential values showed -34.56 mV. Compared to the controls, the GB-AgNP treatment inhibited the cell proliferation and induced the apoptosis of HeLa and SiHa cells. In addition, GB-AgNP treatment led to markedly increased levels of intracellular ROS, the release of cytochrome c (Cyt C) from mitochondria into the cytosol and the cleavage of caspase -9 and -3 in both CCa cell lines. Moreover, NAC, an ROS scavenger, eliminated the effect of GB-AgNPs on the HeLa and SiHa cells. This study reveals that GB-AgNPs suppresses cancer cell proliferation and induces apoptosis by upregulating intracellular ROS generation and inducing the activation of the caspase-dependent mitochondrial apoptotic pathway in CCa cells. Thus, GB-AgNPs may be a potential alternative drug for CCa therapy.

GPR4 Knockout Improves the Neurotoxin-Induced, Caspase-Dependent Mitochondrial Apoptosis of the Dopaminergic Neuronal Cell.

In Parkinson’s disease, mitochondrial oxidative stress-mediated apoptosis is a major cause of dopaminergic neuronal loss in the substantia nigra (SN). G protein-coupled receptor 4 (GPR4), previously recognised as an orphan G protein coupled-receptor (GPCR), has recently been claimed as a member of the group of proton-activated GPCRs. Its activity in neuronal apoptosis, however, remains undefined. In this study, we investigated the role of GPR4 in the 1-methyl-4-phenylpyridinium ion (MPP+) and hydrogen peroxide (H2O2)-treated apoptotic cell death of stably GPR4-overexpressing and stably GPR4-knockout human neuroblastoma SH-SY5Y cells. In GPR4-OE cells, MPP+ and H2O2 were found to significantly increase the expression levels of both mRNA and proteins of the pro-apoptotic Bcl-2-associated X protein (Bax) genes, while they decreased the anti-apoptotic B-cell lymphoma 2 (Bcl-2) genes. In addition, MPP+ treatment activated Caspase-3, leading to the cleavage of poly (ADP-ribose) polymerase (PARP) and decreasing the mitochondrial membrane potential (ΔΨm) in GPR4-OE cells. In contrast, H2O2 treatment significantly increased the intracellular calcium ions (Ca2+) and reactive oxygen species (ROS) in GPR4-OE cells. Further, chemical inhibition by NE52-QQ57, a selective antagonist of GPR4, and knockout of GPR4 by clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 decreased the Bax/Bcl-2 ratio and ROS generation, and stabilised the ΔΨm, thus protecting the SH-SY5Y cells from MPP+- or H2O2-induced apoptotic cell death. Moreover, the knockout of GPR4 decreased the proteolytic degradation of phosphatidylinositol biphosphate (PIP2) and subsequent release of the endoplasmic reticulum (ER)-stored Ca2+ in the cytosol. Our results suggest that the pharmacological inhibition or genetic deletion of GPR4 improves the neurotoxin-induced caspase-dependent mitochondrial apoptotic pathway, possibly through the modulation of PIP2 degradation-mediated calcium signalling. Therefore, GPR4 presents a potential therapeutic target for neurodegenerative disorders such as Parkinson’s disease.

Palladium (II) Complex Enhances ROS-Dependent Apoptotic Effects via Autophagy Inhibition and Disruption of Multiple Signaling Pathways in Colorectal Cancer Cells.

Inhibition of autophagy is reported to be a therapeutically effective strategy in overcoming resistance that is a deadly outcome in cancer. One of the most common reasons for chemo-resistance to treatment is the patients with tumors exhibiting a KRAS mutation, which occurs in approximately 40% of colorectal cancer patients. Hence, we assessed whether a Palladium (Pd)(II) complex is a promising anticancer complex, compared to 5-fluorouracil in KRAS wt HT-29 and KRAS mutant HCT-15 cells. HCT-15 and HT-29 cells were used for colorectal cancer and Chloroquine (CQ) was used as an inhibitor of autophagy. In this context, cells were treated with Pd(II) complex and 5-FU in combination with CQ for 48h and cell viability was measured by SRB assay. Cell death mode was examined with M30 and M65 ELISA assays, using annexin V/propidium iodide. Autophagy was determined by Acridine Orange (AO) staining. Furthermore, the expressions of various autophagy and apoptosis-related proteins were evaluated with Western blotting. Luminex assay and the level of Reactive Oxygen Species (ROS) were examined. Cell viability was found to decrease in a dose-dependent manner and CQ enhanced cytotoxic effect in Pd(II) and 5-FU treated cells in colorectal cancer cells. Our data showed that inhibition of autophagic flux significantly increased intrinsic apoptosis through the activation of ROS. We showed that combinatorial treatment with CQ induced apoptosis via the caspase-dependent mitochondrial pathway. Luminex analysis revealed that the combination resulted in a down-regulation of NF-κB/AKT/CREB signaling pathways in both cell lines, however, decreased Erk1/2 protein expression was only observed after treatment with CQ combination in HCT-15 cells. We suggest that the inhibition of autophagy along with Pd(II) and 5-FU treatment has a synergistic effect on KRAS-mutant colorectal cancer cells. Autophagy inhibition by CQ promotes apoptosis via blockade of the NF-κB/AKT/CREB and activation of ROS.

Inhibition of mitotic kinase Mps1 promotes cell death in neuroblastoma

Neuroblastoma is the most common paediatric cancer type. Patients diagnosed with high-risk neuroblastoma have poor prognosis and occasionally tumours relapse. As a result, novel treatment strategies are needed for relapse and refractory neuroblastoma patients. Here, we found that high expression of Mps1 kinase (mitotic kinase Monopolar Spindle 1) was associated with relapse-free neuroblastoma patient outcomes and poor overall survival. Silencing and inhibition of Mps1 in neuroblastoma or PDX-derived cells promoted cell apoptosis via the caspase-dependent mitochondrial apoptotic pathway. The mechanism of cell death upon Mps1 inhibition was dependent on the polyploidization/aneuploidization of the cells before undergoing mitotic catastrophe. Furthermore, tumour growth retardation was confirmed in a xenograft mouse model after Mps1-inhibitor treatment. Altogether, these results suggest that Mps1 expression and inhibition can be considered as a novel prognostic marker as well as a therapeutic strategy for the treatment of high-risk neuroblastoma patients.

3-Bromopyruvate potentiates TRAIL-induced apoptosis in human colon cancer cells through a reactive oxygen species- and caspase-dependent mitochondrial pathway.

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a promising anticancer cytokine with minimal toxicity towards normal cells. Nevertheless, most primary cancers are often intrinsically TRAIL-resistant or can acquire resistance after TRAIL therapy. This study aimed to investigate the inhibitory effect of co-treatment of 3-bromopyruvate (3-BP) as a potent anticancer agent with TRAIL on colon cancer cells (HT-29). The results of present study indicated that combined treatment with 3-BP and TRAIL inhibited the proliferation of HT-29 cells to a greater extent (88.4%) compared with 3-BP (54%) or TRAIL (11%) treatment alone. In contrast, the combination of 3-BP and TRAIL had no significant inhibitory effect on the proliferation of normal cells (HEK-293) (8.4%). At a cellular mechanistic level, the present study showed that 3-BP sensitized human colon cancer cells to TRAIL-induced apoptosis via reactive oxygen species generation, upregulation of Bax, downregulation of Bcl-2 and survivin, release of cytochrome c into the cytosol, and activation of caspase-3. In normal cells, 3-BP, TRAIL, or combination of both had no significant effect on the reactive oxygen species levels, release of cytochrome c, and caspase-3 activity. Therefore, the combination of 3-BP and TRAIL can be a promising therapeutic strategy for treatment of colon cancer.

Xanthone derivatives from the leaves of Garcinia oligantha

Nine new and unique xanthone derivatives, including one novel hybrid monoterpene-tetrahydroxanthone (1), three dihydro-xanthone derivatives (2-4), and five skeleton-rearranged xanthone derivatives (5–9), were obtained from a 95% EtOH extract of Garcinia oligantha leaves by a LC-MS-guided fractionation procedure. The structures of the new compounds were elucidated by analysis of their 1D and 2D NMR and MS data. The relative configurations of 2 and 8 were determined via X-ray crystallographic data analysis, while the absolute configurations of 1–2, 5–9 were assigned based on a comparison of calculated and experimental ECD and/or OR data. In SRB, PI-exclusion and Hoechst staining assays, 6 showed strong cytotoxic activities which could dose-dependently induce Taxol-insensitive quiescent LNCaP cell death. Additionally, a preliminary mechanism investigation using immunoblotting and Caspase-3 activity assay, indicated that 6 induced quiescent LNCaP cell death potentially through caspase-dependent mitochondrial apoptosis pathway.

The methyl-triclosan induced caspase-dependent mitochondrial apoptosis in HepG2 cells mediated through oxidative stress

Methyl-triclosan (MTCS) is a dominant transformation product of triclosan (TCS), which has been widely used as an effective antimicrobial ingredient with increasing concentrations in the environment. MTCS shows higher persistence in environment than its parent chemical TCS. The toxic effects of MTCS and toxicological mechanism are not well understood up to now. This study investigated the cytotoxic effects of MTCS in HepG2 cells in terms of cell viability, apoptosis induction, ROS production, GSH/GSSG levels, Mitochondrial Membrane Potential (MMP) reduction, LDH release, glucose uptake and ATP production. Moreover, the related gene transcripts were measured with RT-qPCR assay. Cytotoxic experiments in HepG2 cells revealed that MTCS exposure at micromol per liter levels had toxic effects as evidenced by decreased cell survival, elevated cell apoptosis, reduced MMP and increased LDH release. These toxic effects were associated with increased ROS production and reduced GSH/GSSG ratio. Meanwhile, elevated glucose uptake and ATP production indicated that MTCS induced membrane damages resulted not from a typical mitochondrial uncoupler, but from oxidative stress. Analysis of gene transcripts showed that MTCS exposure induced mRNA expressions alterations associated with oxidative stress response, energy production, cell cycle regulation and cell apoptosis. In general, the caspase-dependent mitochondrial apoptosis pathway might play a role in MTCS induced cytotoxicity in HepG2 cells.