scholarly journals The cytotoxicity and apoptotic effects of verbascoside on breast cancer 4T1 cell line

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Atena Daneshforouz ◽  
Samad Nazemi ◽  
Omid Gholami ◽  
Marzieh Kafami ◽  
Bahareh Amin
Keyword(s):  
Breast Cancer ◽  
Flow Cytometry ◽  
Mtt Assay ◽  
Caspase 3 ◽  
Mrna Level ◽  
Annexin V ◽  
Underlying Mechanism ◽  
Mammary Tumor Cell ◽  
Mouse Mammary Tumor ◽  
Apoptotic Effects

Abstract Background Despite significant advancements in breast cancer therapy, novel drugs with lower side effects are still being demanded. In this regard, we investigated the anti-cancer features of verbascoside in 4 T1 mouse mammary tumor cell. Methods First, MTT assay was performed with various concentrations (ranging between 5 to 200 μM) of verbascoside and IC50 was calculated. Then the expression of Bax, Bcl-2, and caspase-3 was evaluated in treated 4 T1 cells. In addition, we investigated the expression of TLR4, MyD88, and NF-κB to ascertain the underlying mechanism of the anti-proliferative feature of verbascoside. Also, flow cytometry followed by double PI and Annexin V was conducted to confirm the apoptosis-inducing effect of verbascoside. Results Our results from MTT assay showed verbascoside inhibits proliferation of 4 T1 cancer cells (IC50 117 μM) while is safe for normal HEK293T cells. By qRT-PCR, we observed that verbascoside treatment (100, 117 and, 130 μM) increases the expression of caspase-3 and Bax while reduces the expression of Bcl-2. Also, verbascoside (100, 117 and, 130 μM) increased the expression of TLR4 only at 130 μM dose and the expression of MyD88 whereas reduced the expression of NF-κB at mRNA level. Flow cytometry analysis also confirmed verbascoside induces apoptosis in 4 T1 cells at 117 μM. Conclusion Taken together, our data showed verbascoside is a safe natural compound for normal cells while has apoptosis-inducing feature through TLR4 axis on 4 T1 cells.

Combination of Histone Deacetylase Inhibitor with Cu(II) 5,5- diethylbarbiturate Complex Induces Apoptosis in Breast Cancer Stem Cells: A Promising Novel Approach

2020 ◽  
Vol 21 ◽  
Author(s):  
Merve Erkisa ◽  
Nazlihan Aztopal ◽  
Elif Erturk ◽  
Engin Ulukaya ◽  
Veysel T. Yilmaz ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Oxidative Stress ◽  
Stem Cells ◽  
Cancer Stem Cells ◽  
Histone Deacetylases ◽  
Caspase 3 ◽  
Cancer Cell Line ◽  
Annexin V ◽  
Tumor Formation ◽  
Dependent Manner

Background: Cancer stem cells (CSC) are subpopulation within the tumor that acts a part in the initiation, progression, recurrence, resistance to drugs and metastasis of cancer. It is well known that epigenetic changes lead to tumor formation in cancer stem cells and show drug resistance. Epigenetic modulators and /or their combination with different agents have been used in cancer therapy. Objective: In our study we scope out the effects of combination of a histone deacetylases inhibitor, valproic acid (VPA), and Cu(II) complex [Cu(barb-κN)(barb-κ2N,O)(phen-κN,N’)]·H2O] on cytotoxicity/apoptosis in a stem-cell enriched population (MCF-7s) obtained from parental breast cancer cell line (MCF-7). Methods: Viability of the cells was measured by the ATP assay. Apoptosis was elucidated via the assessment of caspase-cleaved cytokeratin 18 (M30 ELISA) and a group of flow cytometry analysis (caspase 3/7 activity, phosphatidylserine translocation by annexin V-FITC assay, DNA damage and oxidative stress) and 2ˈ,7ˈ–dichlorofluorescein diacetate staining. Results: The VPA combined with Cu(II) complex showed anti proliferative activity on MCF-7s cells in a dose- and time-dependently. Treatment with combination of 2.5 mM VPA and 3.12 μM Cu(II) complex induces oxidative stress in a time-dependent manner, as well as apoptosis that is evidenced by the increase in caspase 3/7 activity, positive annexin-V-FITC, and increase in M30 levels. Conclusion: The results suggest that the combination therapy induces apoptosis following increased oxidative stress, thereby making it a possible promising therapeutic strategy that further analysis is required.

scholarly journals Graphene-modified CePO4 nanorods effectively treat breast cancer-induced bone metastases and regulate macrophage polarization to improve osteo-inductive ability

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Yu-Wei Ge ◽  
Xiao-Liang Liu ◽  
De-gang Yu ◽  
Zhen-An Zhu ◽  
Qin-Fei Ke ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Flow Cytometry ◽  
Bone Metastases ◽  
Bone Regeneration ◽  
Caspase 3 ◽  
Macrophage Polarization ◽  
Tunel Staining ◽  
Alizarin Red ◽  
Arginase 1 ◽  
Local Bone

Abstract Background Breast cancer bone metastasis has become one of the most common complications; however, it may cause cancer recurrence and bone nonunion, as well as local bone defects. Methods Herein, In vitro, we verified the effect of bioscaffold materials on cell proliferation and apoptosis through a CCK8 trial, staining of live/dead cells, and flow cytometry. We used immunofluorescence technology and flow cytometry to verify whether bioscaffold materials regulate macrophage polarization, and we used ALP staining, alizarin red staining and PCR to verify whether bioscaffold material promotes bone regeneration. In vivo, we once again studied the effect of bioscaffold materials on tumors by measuring tumor volume in mice, Tunel staining, and caspase-3 immunofluorescence. We also constructed a mouse skull ultimate defect model to verify the effect on bone regeneration. Results Graphene oxide (GO) nanoparticles, hydrated CePO4 nanorods and bioactive chitosan (CS) are combined to form a bioactive multifunctional CePO4/CS/GO scaffold, with characteristics such as photothermal therapy to kill tumors, macrophage polarization to promote blood vessel formation, and induction of bone formation. CePO4/CS/GO scaffold activates the caspase-3 proteasein local tumor cells, thereby lysing the DNA between nucleosomes and causing apoptosis. On the one hand, the as-released Ce3+ ions promote M2 polarization of macrophages, which secretes vascular endothelial growth factor (VEGF) and Arginase-1 (Arg-1), which promotes angiogenesis. On the other hand, the as-released Ce3+ ions also activated the BMP-2/Smad signaling pathway which facilitated bone tissue regeneration. Conclusion The multifunctional CePO4/CS/GO scaffolds may become a promising platform for therapy of breast cancer bone metastases.

The Effect of Sirtuin 2 (Sirt2) Overexpressing Bone Marrow Mesenchymal Stem Cells on the Growth of Human Epidermal Growth Factor Receptor 2 (Her-2) Breast Cancer Cells and Its Mechanism

2021 ◽  
Vol 11 (4) ◽  
pp. 778-785
Author(s):  
Xiaolin Chen ◽  
Yan Wang ◽  
Sunlu Jiang
Keyword(s):  
Breast Cancer ◽  
Flow Cytometry ◽  
Nk Cells ◽  
Tumor Cells ◽  
Cancer Cell ◽  
Breast Cancer Cell ◽  
Caspase 3 ◽  
Tumor Bearing ◽  
Her 2 ◽  
Ifn Γ

Our study investigates the effect of high expression of Sirt2 in MSCs (MSCs-Sirt2) on Her-2 breast cancer cell proliferation. A mouse subcutaneous xenograft tumor model was established and MSCssirt2 analysis was performed on nude mice. TUNEL staining, flow cytometry, western-blot, real-time PCR and immunohistochemistry were used to detect cancer cell apoptosis. The number of NK cells infiltrated by flow cytometry detected the tumor tissue of tumor-bearing mice, and its killing activity on tumor-bearing mice was detected by isotope labeling and release method. The levels of TNF-α, IFN-γ, IL-8, IL-6 and IL-10 were detected by ELISA. Caspase-3 level was decreased in the MSCs group (P <0.01) while increased in the MSCs-sirt2 group (P <0.001). However, PCNA expression showed an opposite profile in the Her-2 group and MSCs-sirt2 group compared to Caspase-3 level (P <0.01). The tumor volume and weight in the MSCs-sirt2 group was significantly reduced (P < 0.01), while increased in the MSCs group significantly (P < 0.05). The number of Ki-67-positive tumor cells in MSCs-sirt2 group was significantly reduced (P <0.01) and increased in MSCs group (P < 0.001) with oppositive number of TUNEL-positive tumor cells in the MSCs-sirt2 group and MSCs group (P <0.01). IFN-γ level showed an upward trend (P <0.001). The NK cell toxicity of MSCs-Sirt2 group was significantly higher (P <0.001). MSCs-Sirt2 has an inhibitory effect on Her-2 breast cancer cell growth by enhancing the local inflammatory response of NK cells.

scholarly journals PCDHGB7 Increases Chemosensitivity to Carboplatin by Inhibiting HSPA9 via Inducing Apoptosis in Breast Cancer

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Siqi Hou ◽  
Ming Shan ◽  
Chunyang Gao ◽  
Xinxin Feng ◽  
Yongheng Yang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cancer Progression ◽  
Poor Prognosis ◽  
Caspase 3 ◽  
Triple Negative ◽  
Chemotherapy Resistance ◽  
Underlying Mechanism ◽  
High Recurrence Rate ◽  
Tnbc Cell ◽  
Resistance To Chemotherapy

Breast cancer is one of the most serious cancers worldwide, and chemotherapy resistance frequently drives cancer progression. Triple-negative breast cancer (TNBC) has a high recurrence rate and poor prognosis given its resistance to chemotherapy. In our previous study, we found a remarkable abnormal methylation modification of the PCDHGB7 gene in breast cancer. However, the roles of PCDHGB7 in the progression and treatment of breast cancer are unclear. In this study, we examined the effects of PCDHGB7 on the sensitivity of TNBC cells to carboplatin and investigated the underlying mechanism. By knocking down and overexpressing PCDHGB7 in HS578T and BT549 cells, we confirmed that PCDHGB7 increases TNBC cell chemosensitivity to carboplatin. Mechanistically, we found that PCDHGB7 negatively regulates the expression of HSPA9, uplifting its inhibition on P53 translocation and caspase-3 activation. Thus, we demonstrated that PCDHGB7 increases chemosensitivity of TNBC cells to carboplatin by inhibiting HSPA9 via inducing apoptosis. PCDHGB7 and HSPA9 represent potential therapeutic targets for chemosensitivity in breast cancer.

Angelica Polysaccharide and TPO Have a Protective Effect On Hcmv-Induced Apoptosis In Megakaryocytes

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 3553-3553
Author(s):  
Mo Yang ◽  
Jian Liang Chen ◽  
Jie yu Ye ◽  
Su yi Li ◽  
En yu Liang ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Protective Effect ◽  
Caspase 3 ◽  
Hcmv Infection ◽  
Annexin V ◽  
Angelica Sinensis ◽  
Control Group ◽  
Mock Control ◽  
Apoptotic Effect ◽  
Induced Apoptosis

Abstract Human cytomegalovirus (hCMV) infection is often associated with thrombocytopenia. Megakaryocytes may be one of the major sites of hCMV infection, then inducing this cell apoptosis. Angelica Sinensis (Danggui) is an important ingredient of many commonly used herbal Medicine for promoting blood production. Our previous study has showed that the hematopoietic effect of Angelica Sinensis is related to its constituent, angelica polysaccharide (APS) (Yang M et al, J Ethnopharma, 2009). This present study investigated the anti-apoptotic effect of APS and TPO on hCMV-induced apoptosis in megakaryocytes. Human bone marrow mononuclear cells (MNC) or megakaryocytic cell line CHRF-288-11 and hCMV AD169 strain were co-cultured in this study. hCMV significantly inhibited the formation of CFU-MK as shown in three different concentrations of viral infection groups (103, 104 and 105 pfu/ml), compared with blank control and mock control (n=10, P<0.05). hCMV also significantly inhibited the growth of CHRF cells in these three different concentrations after incubation for 3 days, which compared with control group (n=10, P<0.01). hCMV DNA and mRNA were also positively detected in CHRF cells and the cells of CFU-MK with IS-PCR and RT-PCR respectively, while it was negative in blank and mock control groups. We further studied the effect of APS and TPO on CFU-MK formation. Results showed that APS (50 ug/ml) like TPO (50 ng/ml) enhanced hCMV-reduced CFU-MK (P=0.05, n=6). CHRF cells were also analyzed by Annexin V/PI with flow cytometry at day 3 after infection with hCMV AD169. The percentage of apoptotic cells in group of 103 pfu/ml was 19.0 ± 2.0%; The group of 104 pfu/ml was 23.0 ± 1.5%; The group of 105 pfu/ml was 28.0 ± 3.0%. The control group was 2.0 ± 0.5%. The apoptotic cells were confirmed by morphologic observation. In addition, apoptotic signals from megakaryocytic surface, cytoplasma and mitochondria were detected in hCMV infected cells by flow cytometry with Caspase-3 and JC-1 assay. Compared to mock infection control at day 5, Annexin-V positive cells population increased by 58%; active caspase-3 signal increased by 120% in viable cell population; and cell population with damaged mitochondial membrane showed a 5-times increase. Moreover, the anti-apoptotic effect of APS and TPO on CHRF cells was also demonstrated by using Annexin-V assay. Our studies showed that hCMV induces the apoptosis in megakaryocytes via mitochondrial and caspase-3 signaling, and angelica polysaccharide (APS) like TPO has a protective effect on hCMV-induced apoptosis in these cells. Disclosures: No relevant conflicts of interest to declare.

scholarly journals In Vitro and Ex Vivo Evaluation of Smart Infra-Red Fluorescent Caspase-3 Probes for Molecular Imaging of Cardiovascular Apoptosis

2011 ◽  
Vol 2011 ◽  
pp. 1-13 ◽  
Author(s):  
Manuelle Debunne ◽  
Christophe Portal ◽  
Bruno Delest ◽  
Ebba Brakenhielm ◽  
Françoise Lallemand ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Endothelial Cells ◽  
Caspase 3 ◽  
Near Infrared ◽  
Ex Vivo ◽  
Annexin V ◽  
Fluorescent Signal ◽  
Infra Red ◽  
Infrared Probes

Purpose. The aim of this paper is to develop new optical bioprobes for the imaging of apoptosis. Procedure. We developed quenched near-infrared probes which become fluorescent upon cleavage by caspase-3, the key regulatory enzyme of apoptosis. Results. Probes were shown to be selectively cleaved by recombinant caspase-3. Apoptosis of cultured endothelial cells was associated with an increased fluorescent signal for the cleaved probes, which colocalized with caspase-3 and was reduced by the addition of a caspase-3 inhibitor. Flow cytometry demonstrated a similar profile between the cleaved probes and annexin V. Ex vivo experiments showed that sections of hearts obtained from mice treated with the proapoptotic drug doxorubicin displayed an increase in the fluorescent signal for the cleaved probes, which was reduced by a caspase-3 inhibitor. Conclusion. We demonstrated the capacity of these novel probes to detect apoptosis by optical imaging in vitro and ex vivo.

Acrylamide-derived cytotoxic, anti-proliferative, and apoptotic effects on A549 cells

2017 ◽  
Vol 37 (5) ◽  
pp. 468-474 ◽  
Author(s):  
S Kacar ◽  
D Vejselova ◽  
HM Kutlu ◽  
V Sahinturk
Keyword(s):  
Confocal Microscopy ◽  
Cell Lines ◽  
Mtt Assay ◽  
A549 Cells ◽  
Morphological Characteristics ◽  
Annexin V ◽  
Fluorescein Isothiocyanate ◽  
Morphological Alterations ◽  
Lung Adenocarcinoma Cell Line ◽  
Apoptotic Effects

Background: Acrylamide is a very common compound even reaching up to our daily foods. It has been studied in a wealth of cell lines on which it proved to have various toxic effects. Among these cell lines, human lung adenocarcinoma cell line (A549) is one of that on which acrylamide’s toxicity has not been studied well yet. Aim: We intended to determine the half maximal inhibitory concentration (IC50) dose of acrylamide and to investigate its cytotoxic, anti-proliferative and apoptotic effects on A549 cells. Methods: We determined the IC50 dose by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Then, the mode of cell death was evaluated by flow cytometry using Annexin-V fluorescein isothiocyanate (FITC)/propidium iodide (PI) staining. Next, we performed transmission electron microscopy (TEM) and confocal microscopy analyses for morphological alterations and apoptotic indices. Results: According to the MTT assay results, A549 cell viability decreases proportionally with increasing acrylamide concentrations and IC50 for A549 was 4.6 mM for 24 h. Annexin-V FITC/PI assay results indicated that acrylamide induces apoptosis in 64% of the A549 cells. TEM and confocal microscopy analyses showed nuclear condensations, fragmentations, cytoskeleton laceration, and membrane blebbing, which are morphological characteristics of apoptosis. Conclusion: Our research suggests that acrylamide causes cytotoxic, anti-proliferative, and apoptotic effects on A549 cells at 4.6 mM IC50 dose in 24 h.

scholarly journals Saikosaponin D Inhibits the Proliferation and Promotes the Apoptosis of Rat Hepatic Stellate Cells by Inducing Autophagosome Formation

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Hong Jiang ◽  
Jia Liu ◽  
Kun Zhang ◽  
Qingxin Zeng
Keyword(s):  
Flow Cytometry ◽  
Hepatic Stellate Cells ◽  
Caspase 3 ◽  
Stellate Cell ◽  
Annexin V ◽  
Stellate Cells ◽  
Double Staining ◽  
Autophagosome Formation ◽  
Proliferation And Apoptosis ◽  
Normal Rat

Objective. This study aimed to investigate the effects of saikosaponin D (SSd) on the proliferation and apoptosis of the HSC-T6 hepatic stellate cell line and determine the key pathway that mediates SSd’s function. Methods. Cell viability was detected using the CCK-8 kit. The EdU kit and flow cytometry were used to examine cell proliferation. The Annexin V-FITC/PI double staining kit and flow cytometry were used to examine cell apoptosis. Western blot analysis was performed to analyze the expression levels of LC3, Ki67, cleaved caspase 3, Bax, and Bcl2. Autophagosome formation was detected by LC3-GFP adenovirus transfection. Results. SSd inhibits the proliferation and promotes the apoptosis of acetaldehyde-activated HSC-T6 cells. SSd treatment increased the expression of cleaved caspase 3 and Bax but reduced that of Ki67 and Bcl2. The same concentration of SSd barely influenced the growth of normal rat liver BRL-3A cells. SSd upregulated LC3-II expression and induced autophagosome formation. Autophagy agonist rapamycin had the same effect as SSd and autophagy inhibitor 3-methyladenine could neutralize the effect of SSd in acetaldehyde-activated HSC-T6 cells. Conclusions. SSd could inhibit the proliferation and promote the apoptosis of HSC-T6 cells by inducing autophagosome formation.

scholarly journals Anti-bacterial and anti-viral nanchangmycin displays anti-myeloma activity by targeting Otub1 and c-Maf

2020 ◽  
Vol 11 (9) ◽  
Author(s):  
Yujia Xu ◽  
Tong Sun ◽  
Kun Zeng ◽  
Min Xu ◽  
Jinhao Chen ◽  
...  
Keyword(s):  
Cell Apoptosis ◽  
Caspase 3 ◽  
Plasma Cells ◽  
Mrna Level ◽  
Annexin V ◽  
Safety And Efficacy ◽  
In Vivo Assays ◽  
Subsequent Degradation ◽  
Factor C

Abstract As a deubiqutinase Otub1 stabilizes and promotes the oncogenic activity of the transcription factor c-Maf in multiple myeloma (MM), a malignancy of plasma cells. In the screen for bioactive inhibitors of the Otub1/c-Maf axis for MM treatment, nanchangmycin (Nam), a polyketide antibiotic, was identified to suppress c-Maf activity in the presence of Otub1. By suppressing Otub1, Nam induces c-Maf polyubiquitination and subsequent degradation in proteasomes but does not alter its mRNA level. Consistently, Nam downregulates the expression of CCND2, ARK5, and ITGB7, the downstream genes regulated by c-Maf, and promotes MM cell apoptosis as evidenced by PARP and Caspase-3 cleavage, as well as Annexin V staining. In line with the hypothesis, overexpression of Otub1 partly rescues Nam-induced MM cell apoptosis, and interestingly, when Otub1 is knocked down, Nam-decreased MM cell survival is also partly ablated, suggesting Otub1 is essential for Nam anti-MM activity. Nam also displays potent anti-MM activity synergistically with Doxorubicin or lenalidomide. In the in vivo assays, Nam almost completely suppresses the growth of MM xenografts in nude mice at low dosages but it shows no toxicity. Given its safety and efficacy, Nam has a potential for MM treatment by targeting the Otub1/c-Maf axis.

Study on the effects of c-erbB-2-specific antisense oligonucleotides on c-erbB-2 overexpressing breast cancer cells

2006 ◽  
Vol 24 (18_suppl) ◽  
pp. 10724-10724
Author(s):  
X. Liu ◽  
P. Fan ◽  
Z. Wu
Keyword(s):  
Breast Cancer ◽  
Flow Cytometry ◽  
Cell Proliferation ◽  
Mtt Assay ◽  
Cell Apoptosis ◽  
Antisense Oligonucleotides ◽  
Therapy Group ◽  
The Other ◽  
Antisense Odns ◽  
Breast Cancer Xenografts

10724 Background: Overexpression of c-erbB2 oncoprotein was always correlated with bad prognostic. In this study, we will research on the effects of c-erbB-2-specific antisense oligonucleotides (ODNs) on c-erbB-2 expression, cell proliferation and apoptosis of c-erbB-2 over-expressing breast cancer TM40D cells, and to further study the effects of c-erbB-2-specific oligonucleotides on breast cancer xenografts in Balb/c mice. Methods: Balb/c mouse derived breast cancer cell line TM40D was incubated with liposome-mediated ODNs for 4 h and cultured for another 72 h, then the effects of ODNs on c-erbB-2 expression, cell proliferation and activation of apoptosis were examined by western blot, MTT assay and flow cytometry. Twenty-six mice with breast cancer xenografts were randomized into three groups—9 in control group, 9 in lipsome group and 8 in therapy group, which were injected hypodermically with 0.1 ml serum-free RPMI-1640 culture-medium, lipsome (10 ug/ml) solution, and the mixture of lipsome solution (10 ug/ml) and anti-sense ODN (1 uM) weekly for consecutive 6 weeks, respectively. After the therapy, the incidence of skin ulcer was recorded, the lumps were removed and weighted, and part of them were used for Flow Cytometry. Results: Western Blotting showed treatment of TM40D cells with c-erbB-2-specific antisense ODNs resulted in inhibition of c-erbB-2 expression. The effects of antisense ODNs on c-erbB-2 protein levels correlated with their effects on cell proliferation. MTT Assay showed antisense ODNs inhibited cell growth by about 50%. Flow cytometry analysis revealed that antisense ODNs increased cell apoptosis by38.5%, compared with cultured cells group 9.13% and liposome group 9.29%. The weight of lumps in the therapy group was significantly lower than that of in the other two groups. Flow Cytometry showed that in the therapy group the ratio of G0/G1 cells in cell cycles was 87.18%, which is higher compared with the other two and proliferation index was lower. Conclusions: Antisense ODNs reduced c-erbB-2 expression, inhibited cell proliferation and induced cell apoptosis. Anti-sense ODNs can inhibit the growth of c-erbB2-overexpressing breast cancer xenogarfts in Balb/c mice. No significant financial relationships to disclose.

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