Rotary culture promotes the proliferation of MCF-7 cells encapsulated in three-dimensional collagen–alginate hydrogels via activation of the ERK1/2-MAPK pathway

2012 ◽  
Vol 7 (1) ◽  
pp. 015003 ◽  
Author(s):  
Hongxia Zheng ◽  
Weiming Tian ◽  
Hongji Yan ◽  
Lei Yue ◽  
Yao Zhang ◽  
...  
Keyword(s):  
Mapk Pathway ◽  
Three Dimensional ◽  
Alginate Hydrogels ◽  
Rotary Culture ◽  
Mcf 7

scholarly journals Dihydrochalcone Derivative Induces Breast Cancer Cell Apoptosis via Intrinsic, Extrinsic, and ER Stress Pathways but Abolishes EGFR/MAPK Pathway

2019 ◽  
Vol 2019 ◽  
pp. 1-18 ◽  
Author(s):  
Wasitta Rachakhom ◽  
Patompong Khaw-on ◽  
Wilart Pompimon ◽  
Ratana Banjerdpongchai
Keyword(s):  
Breast Cancer ◽  
Er Stress ◽  
Cell Lines ◽  
Cell Apoptosis ◽  
Mapk Pathway ◽  
Annexin V ◽  
Mapk Signaling ◽  
Dependent Manner ◽  
Induced Apoptosis ◽  
Mcf 7

Dihydrochalcone derivatives are active compounds that have been purified from the Thai medicinal plant Cyathostemma argenteum. The objectives of this study were to investigate the effects of two dihydrochalcone derivatives on human breast cancer MDA-MB-231 and MCF-7 cell proliferation and to study the relevant mechanisms involved. The two dihydrochalcone derivatives are 4′,6′-dihydroxy-2′,4-dimethoxy-5′-(2″-hydroxybenzyl)dihydrochalcone (compound 1) and calomelanone (2′,6′-dihydroxy-4,4′-dimethoxydihydrochalcone, compound 2), both of which induced cytotoxicity toward both cell lines in a dose-dependent manner by using MTT assay. Treatment with both derivatives induced apoptosis as determined by annexin V-FITC/propidium iodide employing flow cytometry. The reduction of mitochondrial transmembrane potential (staining with 3,3′-dihexyloxacarbocyanine iodide, DiOC6, employing a flow cytometer) was established in the compound 1-treated cells. Compound 1 induced caspase-3, caspase-8, and caspase-9 activities in both cell lines, as has been determined by specific colorimetric substrates and a spectrophotometric microplate reader which indicated the involvement of both the extrinsic and intrinsic pathways. Calcium ion levels in mitochondrial and cytosolic compartments increased in compound 1-treated cells as detected by Rhod-2AM and Fluo-3AM intensity, respectively, indicating the involvement of the endoplasmic reticulum (ER) stress pathway. Compound 1 induced cell cycle arrest via enhanced atm and atr expressions and by upregulating proapoptotic proteins, namely, Bim, Bad, and tBid. Moreover, compound 1 significantly inhibited the EGFR/MAPK signaling pathway. In conclusion, compound 1 induced MDA-MB-231 and MCF-7 cell apoptosis via intrinsic, extrinsic, and ER stress pathways, whereas it ameliorated the EGFR/MAPK pathway in the MCF-7 cell line. Consequently, it is believed that compound 1 could be effectively developed for cancer treatments.

scholarly journals Jaagsiekte Sheep Retrovirus Transformation in Madin-Darby Canine Kidney Epithelial Cell Three-Dimensional Culture

2010 ◽  
Vol 84 (10) ◽  
pp. 5379-5390 ◽  
Author(s):  
Chassidy Johnson ◽  
Kiah Sanders ◽  
Hung Fan
Keyword(s):  
Epithelial Cell ◽  
Mdck Cells ◽  
Mapk Pathway ◽  
Three Dimensional ◽  
Bronchioloalveolar Carcinoma ◽  
Envelope Gene ◽  
Madin Darby Canine Kidney ◽  
Jaagsiekte Sheep Retrovirus ◽  
Darby Canine Kidney ◽  
Canine Kidney

ABSTRACT Jaagsiekte sheep retrovirus (JSRV) is the causative agent of a contagious lung cancer in sheep that shares similarities with human bronchioloalveolar carcinoma (BAC). JSRV is unique because the envelope gene (env) is the oncogene, as it can transform cells in culture and induce tumors in animals. The phosphatidylinositol 3-kinase (PI3K)-Akt-mTOR and H/N-Ras-MEK-mitogen-activated protein kinase (MAPK) pathways have been shown to be critical for Env transformation. However, the question still remains of how disruption of these pathways relates to tumor formation. To address this, JSRV Env transformation was studied in the context of epithelial structure, using the polarized Madin-Darby canine kidney (MDCK) epithelial cell three-dimensional (3-D) culture system. The results indicated that JSRV Env-transformed MDCK cells were larger and had full or multiple lumens, in contrast to the single lumens observed in controls. The altered phenotype was largely mediated by an increase in proliferation, in addition to overcoming the proliferative suppression signal. JSRV Env was not found to disrupt polarity or tight junctions or to inhibit lumen apoptosis. The PI3K-Akt-mTOR pathway was important for Env transformation in MDCK cells, although the mechanisms of action differed in 3-D and monolayer cultures. PI3K-dependent signaling to mTOR occurred in monolayers, while PI3K-independent signaling to mTOR occurred in 3-D culture. In contrast, the H/N-Ras-MEK-MAPK pathway was found to be inhibitory to transformation in both normal and transformed MDCK cells in 3-D culture. However, in monolayer culture, inhibition of MEK reverted the transformed phenotype, suggesting a different mechanism(s) of action in monolayer versus 3-D culture.

99 DIFFERENTIAL GENE REGULATION OF STEROIDOGENIC TRANSCRIPTS AND ESTRADIOL PRODUCTION FOLLOWING IN VITRO PIG EMBRYO ELONGATION IN ALGINATE HYDROGEL THREE-DIMENSIONAL MATRIX

2012 ◽  
Vol 24 (1) ◽  
pp. 162
Author(s):  
J. R. Miles ◽  
C. N. Sargus ◽  
S. A. Plautz ◽  
J. L. Vallet ◽  
A. K. Pannier
Keyword(s):  
Equal Opportunity ◽  
Culture Media ◽  
Morphological Changes ◽  
Three Dimensional ◽  
Differential Gene Regulation ◽  
Alginate Hydrogels ◽  
The Media ◽  
And Control

Between Day 10 and 12 of gestation, the pig embryo elongates from a sphere to a long thin, filament. During this time, the embryo increases the production of oestrogen via an increase in steroidogenic transcripts, which is critical for maternal recognition of pregnancy. To date, attempts to elongate porcine embryos in vitro have been unsuccessful. Therefore, the objective of this study was to utilise alginate hydrogels to establish a culture system that promotes in vitro embryo elongation with a corresponding increase in steroidogenic transcripts and oestradiol production. In 3 replicate collections, White crossbred gilts (n = 15) were bred at Day 0 of the oestrous cycle. At Day 9 of gestation, reproductive tracts were collected and flushed with RPMI-1640 containing antibiotics. Embryos were recovered, grouped according to size and washed with RPMI-1640 containing antibiotics and 10% fetal bovine serum (FBS). Embryos were randomly assigned to be encapsulated using a double encapsulation technique (0.7% sodium alginate and 1.5% calcium chloride solution) or used as controls. Encapsulated and control embryos were cultured for 96 h in CO2 -pretreated RPMI-1640 containing antibiotics and 10% FBS at 38°C, 5% CO2 in air and 100% humidity. Every 24 h, the embryos were imaged and half of the media was replaced. The removed media was stored at –20°C and used to assess oestradiol levels by radioimmunoassay. At the end of culture, a subset of encapsulated and control embryos were snap frozen and used to assess the expression level of steroidogenic transcripts (STAR, CYP11 and CYP19) using quantitative PCR. All data were analysed using general linear model (GLM) procedures for ANOVA. Cell survival, assessed by blastocyst fragmentation and confirmed by live/dead staining in representative embryos, was greater (P = 0.01) for encapsulated embryos (60.1 ± 4.8%) compared with controls (33.3 ± 4.8%). Of encapsulated embryos, 27% had some morphological change (minor flattening and tubal formation) and 14% had significant morphological changes (considerable flattening and tubal formation elongating through the gel), consistent with in vivo embryo elongation. In contrast, the control embryos had no morphological changes observed and remained spherical during culture. The expression levels of STAR, CYP11 and CYP19 were significantly (P < 0.05) greater in encapsulated embryos compared with control embryos. Furthermore, a significant (P < 0.01) time-dependent increase in oestradiol levels in the culture media of encapsulated embryos was identified compared with controls and culture media alone. These results illustrate that cultured pig embryos encapsulated in alginate hydrogels undergo limited morphological changes with increased expression of steroidogenic transcripts and oestrogen production. †USDA is an equal opportunity provider and employer.

P38-β/SAPK-inhibiting and apoptosis-inducing activities of (E)-4-chloro-2-((3-ethoxy-2-hydroxybenzylidene) amino)phenol

2020 ◽  
Vol 39 (10) ◽  
pp. 1374-1389
Author(s):  
O Karaosmanoğlu
Keyword(s):  
Protein Kinase ◽  
Mapk Pathway ◽  
Neutral Red ◽  
Mitogen Activated Protein Kinase ◽  
Breast Cancer Patients ◽  
Trastuzumab Resistance ◽  
Qrt Pcr ◽  
Amino Phenol ◽  
Induced Apoptosis ◽  
Mcf 7

The present study has three purposes; first evaluating cytotoxicity of (E)-4-chloro-2-((3-ethoxy-2-hydroxybenzylidene)amino)phenol (ACES), second deciphering ACES-mediated cellular death mechanism, and third estimating ACES-mediated alterations in the expressions of mitogen-activated protein kinase (MAPK) pathway-related genes. Neutral red uptake assay, cell cycle analysis, mitochondrial membrane potential (MMP), reactive oxygen species (ROS) measurements, caspase 3/7 and 9 activations, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) were implemented. IC50 values of ACES-treated five cells were around 4–6 µg/mL. However, Caco-2 and Huh-7 cells were found to be twofold resistant and fivefold sensitive with IC50 values of 11 µg/mL and 0.93 µg/mL, respectively. In this study, it was initially reported that ACES exhibits selective cytotoxicity to Huh-7 cells. In addition, ACES induced apoptosis by nuclear fragmentation, MMP disruption, and intracellular ROS elevation in MCF-7 cells. qRT-PCR experiment indicated the expressions of 30 genes including ATF2, CREB1, MYC, NFATC4 (NFAT3), CCNA1, CCNB1, CCND2, CDK2, CDKN1A (p21CIP1), CDKN1C (p57KIP2), CDKN2A (p16INK4a), CDKN2B (p15INK4b), DLK1, NRAS, CDC42, PAK1, MAP4K1 (HPK1), MAP3K3 (MEKK3), MAP2K3 (MEK3), MAP2K6 (MEK6), MOS, MAPK1 (ERK2), MAPK8 (JNK1), MAPK10 (JNK3), MAPK11 (p38-β), LAMTOR3 (MP1), MAPK8IP2 (JIP-1), PRDX6 (AOP2), COL1A1, and HSPA5 (Grp78) were downregulated at least 1.5-fold. Moreover, ACES effectively inhibited expressions of genes that code for elements of p38-β/stress-activated protein kinase (SAPK) pathway. ACES has the potential to be used for the reversal of trastuzumab resistance in breast cancer patients by inhibiting p38/SAPK pathway in MCF-7 cells. Therefore, with the selective cytotoxic, apoptosis-inducing, and p38-β/SAPK-inhibiting activities, ACES can be utilized for developing a novel anticancer drug.

Oestrogen Receptor-Mediated Modulation of the EGFR/MAPK Pathway in Tamoxifen-Resistant MCF-7 Cells

2003 ◽  
Vol 81 (1) ◽  
pp. 81-93 ◽  
Author(s):  
Iain R. Hutcheson ◽  
Janice M. Knowlden ◽  
Tracie-Ann Madden ◽  
Denise Barrow ◽  
Julia M.W. Gee ◽  
...  
Keyword(s):  
Oestrogen Receptor ◽  
Mapk Pathway ◽  
Tamoxifen Resistant ◽  
Mcf 7

scholarly journals Anti-tumor activity of plant extracts against human breast cancer cells are different in monolayer and three-dimensional cell culture screening models: A comparison on 34 extracts

2020 ◽  
Vol 7 (3) ◽  
pp. 3667-3677
Author(s):  
Nhan Lu-Chinh Phan ◽  
Khuong Duy Pham ◽  
Mai Thi-Thanh Nguyen ◽  
Ngoc Kim Phan ◽  
Kiet Dinh Truong ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Culture ◽  
Cancer Cells ◽  
Cell Lines ◽  
Plant Extracts ◽  
Breast Cancer Cells ◽  
Three Dimensional ◽  
Culture Model ◽  
Anti Tumor Activity ◽  
Mcf 7

Introduction: The monolayer cell culture model is a popular model for screening anti-tumor activity of plant extracts. However, almost the extracts selected for screening in this model have failed in subsequent animal models. Therefore, there is only about 5 % of candidates from the original thousands of drugs that are screened which ultimately reach clinical trial. This study aimed to compare the differences in anti-tumor activity of 34 plant extracts against breast cancer cells in 2 models of monolayer cell culture (2D) and in three-dimensional (3D) cell culture. Methods: Four breast cancer cell lines (MCF-7, CD44+CD24- MCF-7, VN9, and CD44+CD24- VN9) were used to generate the 2D and 3D models (the 3D model was developed by culturing breast cancer cells in matrigel). The extracts were got from the plant extract library that prepared in the previous study. The anti-tumor activity was evaluated via half inhibitory concentrations( IC50 values). Results: Of the 34 extracts, E12, E7, E5 and E6 of them had an effect on MCF-7, CD44+CD24- MCF-7, VN9 and CD44+CD24- VN9 cells, respectively. The results indicated 10 potentially strong candidates for future drug development targeting hypoxic areas in breast cancer. Conclusion: The 3D culture model exhibited higher resistance to extracts than the 2D culture model. The CD44+CD24- cell population of both VN9 and MCF-7 cell lines showed higher drug resistance than the original cell lines (VN9 and MCF-7).  

scholarly journals Synergistic Effects of Controlled-Released BMP-2 and VEGF from nHAC/PLGAs Scaffold on Osteogenesis

2018 ◽  
Vol 2018 ◽  
pp. 1-12 ◽  
Author(s):  
Ting Wang ◽  
Shu Guo ◽  
Hua Zhang
Keyword(s):  
Bone Defect ◽  
Nuclear Translocation ◽  
Mapk Pathway ◽  
Synergistic Effects ◽  
Three Dimensional ◽  
Bone Morphogenetic Protein 2 ◽  
Related Factors ◽  
P38 Inhibitor ◽  
Growth Space ◽  
Scaffold Materials

Tissue engineering bones take great advantages in massive bone defect repairing; under the induction of growth factors, seed cells differentiate into osteoblasts, and the scaffold materials gradually degrade and are replaced with neogenetic bones, which simulates the actual pathophysiological process of bone regeneration. However, mechanism research is required and further developed to instruct elements selection and optimization. In the present study, we prepared vascular endothelial growth factor/bone morphogenetic protein-2- nanohydroxyapatite/collagen (VEGF/ BMP-2- nHAC/ PLGAs) scaffolds and inoculated mouse MC3T3-E1 preosteoblasts to detect osteogenic indexes and activation of related signaling pathways. The hypothesis is to create a three-dimensional environment that simulates bone defect repairing, and p38 mitogen-activated kinase (p38) inhibitor was applied and osterix shRNA was transferred into mouse MC3T3-E1 preosteoblasts to further investigate the molecular mechanism of crosstalk between BMP-2 and VEGF. Our results demonstrated the following: (1) BMP-2 and VEGF were sustainably released from PLGAs microspheres. (2) nHAC/PLGAs scaffold occupied a three-dimensional porous structure and has excellent physical properties. (3) MC3T3-E1 cells proliferated and differentiated well in the scaffold. (4) Osteogenic differentiation related factors expression of VEGF/BMP-2 loaded scaffold was obviously higher than that of other groups; p38 inhibitor SB203580 decreased the nucleus/cytoplasm ratio of osterix expression. To conclude, the active artificial bone we prepared could provide a favorable growth space for MC3T3-E1 cells, and osteogenesis and maturation reinforced by simultaneous VEGF and BMP-2 treatment may be mainly through the activation of the p38 MAPK pathway to promote nuclear translocation of osterix protein.

scholarly journals lncMat2B regulated by severe hypoxia induces cisplatin resistance by increasing DNA damage repair and tumor-initiating population in breast cancer cells

2020 ◽  
Vol 41 (11) ◽  
pp. 1485-1497 ◽  
Author(s):  
Alfredo García-Venzor ◽  
Edna Ayerim Mandujano-Tinoco ◽  
Araceli Ruiz-Silvestre ◽  
José Manuel Sánchez ◽  
Floria Lizarraga ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Dna Damage ◽  
Cancer Cells ◽  
Breast Cancer Cells ◽  
Ectopic Expression ◽  
Three Dimensional ◽  
Severe Hypoxia ◽  
Loss Of Function ◽  
Mcf 7

Abstract Multicellular tumor spheroids (MCTSs) constitute a three-dimensional culture system that recapitulates the in vivo tumor microenvironment. Tumor cells cultured as MCTSs present antineoplastic resistance due to the effect of microenvironmental signals acting upon them. In this work, we evaluated the biological function of a new microenvironment-regulated long non-coding RNA, lncMat2B, in breast cancer. In MCTSs, the expression of lncMat2B presented an increase and a zonal heterogeneity, as it was expressed principally in quiescent cells of hypoxic regions of the MCTSs. As expected, functional assays supported the role of severe hypoxia in the regulation of lncMat2B. Moreover, gain- and loss-of-function assays using a transcriptional silencing CRISPR/Cas9 system and gBlock revealed that lncMAT2B regulates the tumor-initiating phenotype. Interestingly, lncMat2B is overexpressed in a cisplatin-resistant MCF-7 cell line, and its ectopic expression in wild type MCF-7 cells increased survival to cisplatin exposure by reducing DNA damage and reactive oxygen species accumulation. lncMAT2B is a possible link between severe hypoxia, tumor-initiating phenotype and drug resistance in breast cancer cells.

Arsenic Disulfide Combined with L-Buthionine-(S, R)-Sulfoximine Induces Synergistic Antitumor Effects in Two-Dimensional and Three-Dimensional Models of MCF-7 Breast Carcinoma Cells

2019 ◽  
Vol 47 (05) ◽  
pp. 1149-1170 ◽  
Author(s):  
Yuxue Zhao ◽  
Sachiko Tanaka ◽  
Bo Yuan ◽  
Kentaro Sugiyama ◽  
Kenji Onda ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Three Dimensional ◽  
Two Dimensional ◽  
Antitumor Effects ◽  
Anticancer Efficacy ◽  
Three Dimensional Models ◽  
Novel Strategy ◽  
2D And 3D ◽  
Combinatorial Treatment ◽  
Mcf 7

Three-dimensionally (3D) cultured tumor cells (spheroids) exhibit more resistance to therapeutic agents than the cells cultured in traditional two-dimensional (2D) system (monolayers). We previously demonstrated that arsenic disulfide (As2[Formula: see text] exerted significant anticancer efficacies in both 2D- and 3D-cultured MCF-7 cells, whereas 3D spheroids were shown to be resistant to the As2S2 treatment. L-buthionine-(S, R)-sulfoximine (BSO), an inhibitor of glutathione (GSH) synthesis, has been regarded to be a potent candidate for combinatorial treatment due to its GSH modulation function. In the present study, we introduced BSO in combination with As2S2 at a low concentration to investigate the possible enhancing anticancer efficacy by the combinatorial treatment on 2D- and 3D-cultured MCF-7 cells. Our results presented for the first time that the combination of As2S2 and BSO exerted potent anticancer synergism in both MCF-7 monolayers and spheroids. The [Formula: see text] values of As2S2 in combinatorial treatment were significantly lower than those in treatment of As2S2 alone in both 2D- and 3D-cultured MCF-7 cells ([Formula: see text], respectively). In addition, augmented induction of apoptosis and enhanced cell cycle arrest along with the regulation of apoptosis- and cell cycle-related proteins, as well as synergistic inhibitions of PI3K/Akt signals, were also observed following co-treatment of As2S2 and BSO. Notably, the combinatorial treatment significantly decreased the cellular GSH levels in both 2D- and 3D-cultured MCF-7 cells in comparison with each agent alone ([Formula: see text] in each). Our results suggest that the combinatorial treatment with As2S2 and BSO could be a promising novel strategy to reverse arsenic resistance in human breast cancer.

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