This study explored the effect and underlying mechanism of Stellera chamaejasme extract(SCE) on multidrug resistance of breast cancer. The chemotherapy-sensitive breast cancer cell line MCF-7 and adriamycin(ADR)-resistant cell line MCF-7/ADR were used as experimental subjects. MTT assay was used to detect cell proliferation activity. Pi staining was used to detect the cell cycle. 4',6-Diamidino-2-phenylindole, dihydrochloride(DAPI) staining and flow cytometry were used to detect apoptosis. Dansylcadaverine(MDC) staining and GFP-LC3B-Mcherry adenovirus transfection were used to detect autophagy. The protein expression of Bcl-2, Bax, caspase-9, caspase-3, LC3B, p62, and Beclin-1 was detected by Western blot. The results showed that SCE could significantly inhibit the proliferation of both sensitive and resistant breast cancer cell lines. The drug resistance factor was 0.53, which was significantly lower than 59 of ADR. Meanwhile, the proportion of sensitive/resistant cells in the G_0/G_1 phase increased significantly after SCE treatment. In addition, DAPI staining showed that a series of apoptosis phenomena such as nuclear pyknosis, staining deepening, and nuclear fragmentation appeared in sensitive/resistant cell lines after SCE administration. Moreover, the results of flow cytometry double staining showed that the proportion of apoptotic cells in sensitive/resistant cell lines increased significantly after SCE administration. Besides, Western blot showed that the protein expression levels of caspase-3, caspase-9, and Bcl-2 significantly decreased and the expression level of Bax protein significantly increased in both breast cancer cell lines after SCE administration. Furthermore, SCE could also increase the positive fluorescent spots after MDC staining and yellow fluorescent spots after GFP-LC3B-mcherry transfection, and up-regulate the expression levels of autophagy-related proteins LC3B-Ⅱ, p62, and Beclin-1 in breast cancer cells. In summary, SCE may play the role of anti-multidrug resistance by blocking the cell cycle of breast cancer multidrug-resistant cells, blocking autophagy flow, and ultimately interfering with the apoptosis resistance of drug-resistant cells.
Humans ; Female ; Breast Neoplasms/metabolism* ; MCF-7 Cells ; Caspase 3/metabolism* ; Caspase 9/metabolism* ; Beclin-1/pharmacology* ; Apoptosis ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Cell Line, Tumor ; Drug Resistance, Neoplasm ; Cell Proliferation

OBJECTIVETo investigate the effect of lonidamine on apoptosis of human breast carcinoma cells MCF-7 and the possible mechanisms.

METHODSMTT assay and colony-forming assay were used to evaluate the growth inhibition induced by lonidamine in breast cancer MCF-7 cells. PI/Annexin-V staining was used to detect the apoptotic cells. The ATP levels in the cells were detected using an ATP assay kit. The expression of glucose regulated protein 78 (GRP78), inhibitor of apoptosis protein (cIAP1) and caspase-8 were analyzed with Western blotting.

RESULTSMTT assay and colony-forming assay showed that 50-250 mmol/L lonidamine caused a time- and concentration-dependent inhibition of MCF-7 cell proliferation. Exposure to increased concentrations of lonidamine resulted in significantly increased apoptosis rate in MCF-7 cells. In MCF-7 cells treated with 50, 150 and 250 mmol/L lonidamine for 5 h, the intracellular ATP levels were lowered to 80.67%, 62.78% and 30.73% of the control level, respectively. Western blotting showed that lonidamine up-regulated the expression of GRP78, down-regulated the expression of cIAP1 and promoted caspase-8 activation as the treatment time extended.

CONCLUSIONLonidamine can inhibit the proliferation and induce apoptosis in MCF-7 cells, and these effects are probably mediated by reducing ATP level, inducing endoplasmic reticulum stress response, down-regulating cIAP1, and promoting caspase-8 activation in the cells.

Apoptosis ; drug effects ; Breast Neoplasms ; metabolism ; pathology ; Caspase 8 ; metabolism ; Cell Proliferation ; Down-Regulation ; Endoplasmic Reticulum Stress ; Heat-Shock Proteins ; metabolism ; Humans ; Indazoles ; pharmacology ; Inhibitor of Apoptosis Proteins ; metabolism ; MCF-7 Cells ; drug effects ; Ubiquitin-Protein Ligases ; metabolism ; Up-Regulation

Mcl-1 and Bcl-xL, key anti-apoptotic proteins of the Bcl-2 family, have attracted attention as important molecules in the cell survival and drug resistance. In this study, we investigated whether inhibition of Bcl-xL influences cell growth and apoptosis against simultaneous treatment of resveratrol and clofarabine in the human malignant mesothelioma H-2452 cells. Resveratrol and clofarabine decreased Mcl-1 protein levels but had little effect on Bcl-xL levels. In the presence of two compounds, any detectable change in the Mcl-1 mRNA levels was not observed in RT-PCR analysis, whereas pretreatment with the proteasome inhibitor MG132 led to its accumulation to levels far above basal levels. The knockdown of Bcl-xL inhibited cell proliferation with cell accumulation at G2/M phase and the appearance of sub-G0/G1 peak in DNA flow cytometric assay. The suppression of cell growth was accompanied by an increase in the caspase-3/7 activity with the resultant cleavages of procaspase-3 and its substrate poly (ADP-ribose) polymerase, and increased percentage of apoptotic propensities in annexin V binding assay. Collectively, our data represent that the efficacy of resveratrol and clofarabine for apoptosis induction was substantially enhanced by Bcl-xL-lowering strategy in which the simultaneous targeting of Mcl-1 and Bcl-xL could be a more effective strategy for treating malignant mesothelioma.
Adenine Nucleotides/*pharmacology ; Antimetabolites, Antineoplastic/*pharmacology ; Apoptosis/*drug effects ; Arabinonucleosides/*pharmacology ; Caspase 3/metabolism ; Caspase 7/metabolism ; Cell Line, Tumor ; Cell Proliferation/drug effects ; G2 Phase Cell Cycle Checkpoints/drug effects ; Gene Knockdown Techniques ; Humans ; Leupeptins/pharmacology ; Lung Neoplasms/metabolism/pathology ; M Phase Cell Cycle Checkpoints/drug effects ; Mesothelioma/metabolism/pathology ; Myeloid Cell Leukemia Sequence 1 Protein/antagonists & inhibitors/genetics/metabolism ; RNA Interference ; RNA, Messenger/metabolism ; RNA, Small Interfering/metabolism ; Stilbenes/*pharmacology ; bcl-X Protein/antagonists & inhibitors/*genetics/*metabolism

OBJECTIVETo evaluate apoptotic effects of cisplatin and cordycepin as single agent or in combination with cytotoxicity in oral cancer cells.

METHODSThe influences of cisplatin (2.5 μg/mL) and/or cordycepin treatment (10 or 100 μmol/L) to human OC3 oral cancer cell line were investigated by morphological observation for cell death appearance, methylthiazoletetrazolium (MTT) assay for cell viability, flow cytometry assay for cell apoptosis, and Western blotting for apoptotic protein expressions.

RESULTSData demonstrated that co-administration of cisplatin (2.5 μg/mL) and cordycepin (10 or 100 μmol/L) resulted in the enhancement of OC3 cell apoptosis compared to cisplatin or cordycepin alone treatment (24 h), respectively (P <0.05). In flow cytometry assay, percentage of cells arrested at subG1 phase with co-treatment of cordycepin and cisplatin (30%) was significantly higher than cisplatin (5%) or cordycepin (12%) alone group (P <0.05), confirming a synergistically apoptotic effect of cordycepin and cisplatin. In cellular mechanism study, co-treatment of cordycepin and cisplatin induced more stress-activated protein kinase/Jun terminal kinase (JNK), the expressions of caspase-7, and the cleavage of poly ADP-ribose polymerase (PARP) as compared to cisplatin or cordycepin alone treatment (P <0.05).

CONCLUSIONCisplatin and cordycepin possess synergistically apoptotic effect through the activation of JNK/caspase-7/PARP pathway in human OC3 oral cancer cell line.

Apoptosis ; drug effects ; Caspase 7 ; metabolism ; Cell Count ; Cell Line, Tumor ; Cell Shape ; drug effects ; Cell Survival ; drug effects ; Cisplatin ; pharmacology ; Deoxyadenosines ; pharmacology ; Drug Synergism ; G1 Phase ; drug effects ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Mouth Neoplasms ; pathology ; Phosphorylation ; drug effects ; Poly(ADP-ribose) Polymerases ; metabolism

This study aimed to investigate the synergistic effect of lidamycin (LDM) and rituximab on human B cell lymphoma Ramos cells. Cell proliferation was measured using MTS assay, cell apoptosis was analyzed by Annexin V-FITC/PI assay, the expression of apoptosis related proteins was analyzed by Western blotting, and the in vivo lymphoma inhibition was verified using BALB/c mice inoculated via tail vein using Ramos cells which stably expressed pEGFP-N1 plasmid. The results showed that, after the pretreatment with rituximab for 48 h, rituximab and LDM showed significantly synergistic effects on cell proliferation. Cells in combined treatment group had a higher apoptosis rate than that in LDM treatment group. Compared with the LDM treatment group, the expression of apoptosis-related proteins such as Cleaved caspase-3, Cleaved caspase-7, Cleaved caspase-9 and Cleaved PARP in combined treatment groups increased, and expression of cIAP-2 and Bcl-2 decreased. The result of in vivo experiment showed that, in the combined treatment group, the survival time of BALB/c mice was significantly longer than the mice in control group and LDM treatment group, and the degree of tumor accumulation and metastasis to lymph nodes and spleen was lower.
Aminoglycosides ; pharmacology ; Animals ; Antibiotics, Antineoplastic ; pharmacology ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Caspase 7 ; metabolism ; Caspase 9 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Drug Synergism ; Enediynes ; pharmacology ; Humans ; Inhibitor of Apoptosis Proteins ; metabolism ; Lymphoma, B-Cell ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Metastasis ; Neoplasm Transplantation ; Poly(ADP-ribose) Polymerases ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Random Allocation ; Rituximab ; pharmacology

Adiponectin, an adipokine predominantly secreted from adipose tissue, exhibits diverse biological responses, including metabolism of glucose and lipid, and apoptosis in cancer cells. Recently, adiponectin has been shown to modulate autophagy as well. While emerging evidence has demonstrated that autophagy plays a role in the modulation of proliferation and apoptosis of cancer cells, the role of autophagy in apoptosis of cancer cell caused by adiponectin has not been explored. In the present study, we demonstrated that globular adiponectin (gAcrp) induces both apoptosis and autophagy in human hepatoma cell line (HepG2 cells) and breast cancer cells (MCF-7), as evidenced by increase in caspase-3 activity, Bax, microtubule-associated protein light chain 3-II (LC3 II) protein levels, and autophagosome formation. Interestingly, gene silencing of LC3B, an autophagy marker, significantly enhanced gAcrp-induced apoptosis in both HepG2 and MCF-7 cell lines, whereas induction of autophagy by rapamycin, an mTOR inhibitor, significantly prevented gAcrp-induced apoptosis in hepatoma cells HepG2. Furthermore, modulation of autophagy produced similar effects on gAcrp-induced Bax expression in HepG2 cells. These results implicate that induction of autophagy plays a regulatory role in adiponectin-induced apoptosis of cancer cells, and thus inhibition of autophagy would be a novel promising target to enhance the efficiency of cancer cell apoptosis by adiponectin.
Adipokines ; Adiponectin ; Adipose Tissue ; Apoptosis* ; Autophagy* ; Breast Neoplasms ; Carcinoma, Hepatocellular ; Caspase 3 ; Cell Line ; Gene Silencing ; Glucose ; Hep G2 Cells ; Humans ; MCF-7 Cells ; Metabolism ; Sirolimus

Fucoidan is an active component of seaweed, which inhibits proliferation and induces apoptosis of several tumor cells while the detailed mechanisms underlying this process are still not clear. In this study, the effect of Fucoidan on the proliferation and apoptosis of human breast cancer MCF-7 cells and the molecular mechanism of Fucoidan action were investigated. Viable cell number of MCF-7 cells was decreased by Fucoidan treatment in a dose-dependent manner as measured by MTT assay. Fucoidan treatment resulted in G1 phase arrest of MCF-7 cells as revealed by flow cytometry, which was associated with the decrease in the gene expression of cyclin D1 and CDK-4. Annexin V/PI staining results showed that the number of apoptotic cells was associated with regulation of cytochrome C, caspase-8, Bax and Bcl-2 at transcriptional and translational levels. Both morphologic observation and Hoechst 33258 assay results confirmed the pro-apoptotic effect of Fucoidan. Meanwhile, the ROS production was also increased by Fucoidan treatment, which suggested that Fucoidan induced oxidative damage in MCF-7 cells. The results of present study demonstrated that Fucoidan could induce G1 phase arrest and apoptosis in MCF-7 cells through regulating the cell cycle and apoptosis-related genes or proteins expression, and ROS generation is also involved in these processes.
Antineoplastic Agents ; chemistry ; pharmacology ; Apoptosis ; drug effects ; genetics ; Blotting, Western ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Caspase 8 ; genetics ; metabolism ; Caspases ; genetics ; metabolism ; Cell Proliferation ; drug effects ; Cell Size ; drug effects ; Cyclin D1 ; genetics ; metabolism ; Cyclin-Dependent Kinase 4 ; genetics ; metabolism ; Cytochromes c ; genetics ; metabolism ; Dose-Response Relationship, Drug ; Fucus ; chemistry ; G1 Phase Cell Cycle Checkpoints ; drug effects ; genetics ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; MCF-7 Cells ; Microscopy, Fluorescence ; Molecular Structure ; Polysaccharides ; chemistry ; pharmacology ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; Reactive Oxygen Species ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; drug effects ; bcl-2-Associated X Protein ; genetics ; metabolism

OBJECTIVETo explore the inhibition of Jumi (traditional Chinese medicine) extraction on the growth of human cervical cancer cell line HeLa.

METHODSNude mouse model of human cervical cancer HeLa cell transplantation was established. The nude mice bearing cancer were randomly divided into control group and Jumi treated groups with different concentration (0.001, 0.002, 0.005, 0.01 mg/ml). The growth of cervical cancer cell in experimental mice were measured. Cultured HeLa cells were incubated in culture media with or without Jumi extract for 48 hours. Cell proliferation rate, cell apoptosis, caspase-3/7 and caspase-6 activity were determined by MTT colorimetric assay, flow cytometry analysis and spectrophotometric detection, respectively.

RESULTSWith the increase of the concentration of Jumi extract, tumor-bearing mice tumor inhibition rate gradually increased. The proliferation of cultured HeLa cells were significantly inhibited by Jumi extract in a dose-dependent manner. IC50 was 0.004 mg/ml. Apoptosis rates in the cells treated with Jumi extract were higher than those of the control group. Compared with the control group, except for lower Jumi treated group (0.001 mg/ml), caspase-3/7 and caspase-6 activity were significantly increased in the all Jumi treated groups.

CONCLUSIONJumi extract can inhibit the proliferation of human cervical cancer cell line HeLa in vitro in a dose-dependent manner and promote cell apoptosis through caspase-3, caspase-7 and caspase-6 pathway.

Animals ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Caspase 6 ; metabolism ; Caspase 7 ; metabolism ; Cell Proliferation ; drug effects ; Chrysanthemum ; Female ; HeLa Cells ; Humans ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Plant Extracts ; pharmacology ; Xenograft Model Antitumor Assays

Recent years, the incidence and mortality of prostate cancer have increased dramatically in China. At earlier stages, most diagnosed prostate cancers are responsive to androgen depletion treatment, yet, nearly all patients will eventually progress to metastatic androgen-independent prostate cancer (AIPC), which still has no effective therapeutic method or drug to deal with. 11'-Deoxyverticillin A (C42) belongs to the family of epipolythiodioxopiperazines (ETPs), an interesting class of fungal toxins that inhibit farnesyl transferase. Compounds holding such a property have been explored as putative anticancer agents. In this study, using PC3M cells, an AIPC cell line, we investigated the effect of the compound on apoptosis and explored the underlying mechanism. It revealed that C42 markedly enhanced the activity of caspase-3/7 and increased the accumulation of the cleaved PARP, all of which are the markers of apoptosis. It also revealed that C42 either decreased cell viability or inhibited the growth of PC3M cells. Moreover, we observed that the loss of cell viability and cell growth inhibition induced by C42 were both time- and dosage dependent. Taken together, we indicated that C42 can induce caspase-dependent apoptosis in AIPC cells, and the results presented here will broaden our knowledge about the molecular mechanisms by which C42 exerts its anticancer activity, and future work in this direction may provide valuable information in the development of these compounds into effective cancer therapeutic strategies against androgen-independent prostate cancer.
Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Caspase 7 ; metabolism ; Cell Line, Tumor ; Disulfides ; pharmacology ; Farnesyltranstransferase ; antagonists & inhibitors ; Humans ; Male ; Mycotoxins ; pharmacology ; Piperazines ; pharmacology ; Prostatic Neoplasms ; pathology

OBJECTIVETo observe the effect of NL-608 (a nutlin analog) on apoptosis induction in human breast cancer MCF-7 cells in vitro, and investigate the relevant molecular mechanism.

METHODSThe effect of NL-608 on proliferation of MCF-7 cells was determined by MTT assay. The apoptosis in MCF-7 cells was determined by flow cytometry with annexin V-FITC and PI. The activity of caspase 3, caspase 8 and caspase 9 was determined with caspase activity assay kit and Western blot, and the proteins of Fas and FasL were determined by Western blot.

RESULTSNL-608 showed a dose-dependent inhibitory effect on the proliferation of MCF-7 cells. It induced apoptosis in MCF-7 cells in a dose-dependent manner. The activity of caspase 3 and caspase 8 in MCF-7 cells was increased with the increasing concentration of NL-608, but caspase 9 had no changes. The proteins of Fas and FasL were increased in a dose-dependent manner.

CONCLUSIONNL-608 induces apoptosis in MCF-7 cells in vitro through inducing caspase 3 activity and death receptor-mediated signal pathway.

Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Caspase 8 ; metabolism ; Caspase 9 ; metabolism ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Fas Ligand Protein ; metabolism ; Humans ; Imidazoles ; pharmacology ; MCF-7 Cells ; Piperazines ; pharmacology ; fas Receptor ; metabolism