MCF-7 breast cancer cell line grown in agarose culture for study of COX-2 inhibitors in three-dimensional growth system

Kinder, David H.; Aulthouse, Amy

doi:10.1016/j.canlet.2003.10.030

Cited by 4 publications

(12 citation statements)

References 3 publications

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“…Using this phenotype as a marker of OA, we established a functional screening assay based on retroviral‐mediated gene transfer of OA chondrocyte cDNA library into normal chondrocytes and identifying genes that induce cluster formation. While cell cluster formation is unusual for chondrocytes, cell colony formation in agarose is also evident in many cancer cells (Hohn et al, 1992; Yeates and Powis, 1997; Kinder and Aulthouse, 2004), and could similarly be utilized as a functional gene expression response for drug discovery targets. Chondrocytes grown in agarose suspension culture have low proliferation rates, similar to normal cartilage (Benya and Shaffer, 1982; Glowacki et al, 1983), however, bFGF added to agarose cultures will induce colony formation (Kato et al, 1987).…”

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confidence: 99%

Chondrocyte cluster formation in agarose cultures as a functional assay to identify genes expressed in osteoarthritis

Quintavalla¹,

Kumar²,

Daouti³

et al. 2005

Journal Cellular Physiology

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Understanding altered gene expression in osteoarthritic cartilage can lead to new targets for drug intervention. We established a functional assay based on chondrocyte cluster formation, a phenotype associated with osteoarthritis (OA), to screen an OA cartilage gene library. Previous reports have demonstrated that normal chondrocytes grown in suspension culture maintain their chondrocytic phenotype, however, certain growth factors such as basic fibroblast growth factor (bFGF) will induce the cells to proliferate in tight clusters similar to those seen in osteoarthritic cartilage. In this study we validate that overexpression of bFGF by retrovirally transduced normal chondrocytes would similarly induce the proliferation of tight cell clusters. We then used this approach as a basis to set up a functional screen where an entire OA cartilage cDNA library was tranduced into normal chondrocytes to search for other genes that would also induce cluster formation. Seven potential genes were isolated from the OA gene library, including BPOZ, IL-17 receptor C, NADH ubiquinone oxidoreductase, COMP, Soluble carrier 16 (MCT 3), C1r, and bFGF itself. None of the identified genes were upregulated by bFGF, however, all of them upregulated the expression of bFGF suggesting a common pathway. Although cluster formation is not considered to be destructive in OA cartilage, it is consistent with the disease and could yield answers to the altered phenotype. Further studies are needed to elucidate how these genes are linked to the disease state.

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mentioning

confidence: 99%

Chondrocyte cluster formation in agarose cultures as a functional assay to identify genes expressed in osteoarthritis

Quintavalla¹,

Kumar²,

Daouti³

et al. 2005

Journal Cellular Physiology

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“…This system has proven effective at providing a permissive and physiologically relevant environment for anchorage-independent cells including chondrocytes and cancer cells. 2,3,20 Full characterization of this model is novel for the 3T3-L1 preadipocytes and includes evaluation of change in morphology and non-chemically-induced spontaneous differentiation. This study demonstrates the effectiveness of this model for use by staining with Oil Red O, sectioning, and IHC.…”

Section: Discussionmentioning

confidence: 99%

“…The average diameter of a micro-lipid droplet was calculated per quadrant with an overall average of 0.5 μm. Spherical volume was calculated using the equation 4/3πr 3 where the radius was calculated from the measured micro-lipid droplet diameter. Once the lipid droplet volume was calculated, it was multiplied by the quadrant droplet number to obtain a total lipid volume for that quadrant.…”

Section: Lipid Quantitation Using Oil Red O Stainmentioning

confidence: 99%

“…Numerous studies have shown that 3D environments allow more conservative treatment doses than the monolayer parallel experiment. [1][2][3] Similarly, 3D environments are more permissible spatially, allowing changes in shape and morphology that sometimes results in changes in cell signaling and gene expression profiles. [4][5][6][7][8][9] Several 3D culture techniques have been used to study adipogenesis.…”

Section: Introductionmentioning

confidence: 99%

“…19 This 3D agarose system, in which single cells are suspended in agarose, has been used successfully in the past to provide a permissive environment for anchorage independent cells including chondrocytes and cancer cells. 2,3,20 Based on previous research by author Aulthouse and collaborators involving 3D environment-induced differentiation, we sought to determine whether this same 3D agarose cell culture environment would allow for spontaneous environment-driven differentiation of the preadipocytes to maturing adipocytes.…”

Section: Introductionmentioning

confidence: 99%

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Part 1: A Novel Model for Three-Dimensional Culture of 3T3-L1 Preadipocytes Stimulates Spontaneous Cell Differentiation Independent of Chemical Induction Typically Required in Monolayer

Aulthouse

Freeh

Newstead

et al. 2019

Nutr Metab Insights

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Differences in monolayer and three-dimensional (3D) culture systems have been recognized for several years. Despite the recognized importance of 3D systems, low cost and convenience of monolayer culture are still readily used for metabolic and nutritional studies. Here, we present part 1 of a 2-part series that will highlight (1) a novel and cost-effective model for culturing 3T3-L1 preadipocytes in 3D agarose as well as (2) an initial study showing the successful use of this 3D model for experimental analysis of these cells treated with cinnamon extract while suspended in agarose. In part 1, we provide a full characterization of the model system for the 3T3-L1 cells that demonstrate the functionality and convenience of this system. Importantly, we note spontaneous differentiation to adipocytes while cultured under these methods, independent of chemical induction. We present a 2.5-week time course with rounded cells forming vacuoles as early as 24 hours and accumulation of lipid detectable by Oil Red O stain at 0.5 weeks. Serum selection, lipid volume determination, and cell size are characterized. We conclusively demonstrate adipogenesis based on a peroxisome proliferator-activated receptor γ (PPARγ) detection using immunohistochemistry (IHC) of sections from these 3D cultures. Methods, materials and recommendations are described as well as proposed benefits to the use of this culture system for 3T3-L1 cells.

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Evaluation of Hydrazide-hydrazone and 4-thiazolidinone Derivatives of Etodolac as Potential Anticancer Agents in Leukemia Cells

Tiber,

Averbek,

Sevinç

et al. 2024

LDDD

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Background: Nonsteroidal anti-inflammatory drugs (NSAIDs), which are commonly used for their anti-inflammatory and analgesic properties, have also been found to prevent cancer. (±)(R,S) Etodolac is an NSAID that belongs to the class of cyclooxygenase-2 inhibitors. Various derivatives of etodolac are synthesized to boost its anti-proliferative action and lessen its potential negative effects. In our earlier studies, some novel derivatives of etodolac exhibited stronger cytotoxic effects on prostate cell lines and had similar effects on leukemia cells in pre-screening experiments. Objective: Using the K562 leukemia cell line as a model, we sought to investigate the anti-cancer properties of a hydrazide-hydrozone derivative (SGK-205) and a 4-thiazolidinone derivative of etodolac (SGK-216). Materials and Methods: In the current investigation, SGK-205 and SGK-216 compounds were administered to K562 cells for 24 and 48 hours at concentrations of 10, 25, 50, 75 and 100 μM. Cell viability was assessed using the MTT test, and apoptosis by Annexin V-PI staining and mitochondrial membrane potential assays, together with mRNA expressions of apoptotic proteins. The levels of the proteins, HER2 and COX2, were also examined to evaluate COX2 inhibitory capacity. Results: In K562 cells, there was a definite dose-dependent response to SGK-205 and SGK-216 compounds. Results from MTT viability tests, together with mitochondrial membrane potential measurements and Annexin V-PI staining, revealed that SGK-216 and SGK-205 significantly outperformed etodolac in terms of their apoptotic and anti-proliferative activities. The concentration range of 10-20 M for both chemicals was sufficient to start biological responses. Apoptosis was also investigated through the expressions of pro- and anti-apoptotic proteins. Additionally, gene expression research demonstrated SGK-205 to be a beneficial substitute to etodolac in lowering COX-2 and human epidermal growth factor receptor-2 (HER2) expression. Conclusion: Our data indicated both derivatives to have higher anti-proliferative and apoptotic effects compared to etodolac. An overall assessment highlighting apoptotic induction potential, acceptable toxicity levels, a consistent dose-response relationship, and COX2 inhibitory actions, in particular, indicated SGK-205 as a viable novel therapeutic.

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MCF-7 breast cancer cell line grown in agarose culture for study of COX-2 inhibitors in three-dimensional growth system

Cited by 4 publications

References 3 publications

Chondrocyte cluster formation in agarose cultures as a functional assay to identify genes expressed in osteoarthritis

Chondrocyte cluster formation in agarose cultures as a functional assay to identify genes expressed in osteoarthritis

Part 1: A Novel Model for Three-Dimensional Culture of 3T3-L1 Preadipocytes Stimulates Spontaneous Cell Differentiation Independent of Chemical Induction Typically Required in Monolayer

Evaluation of Hydrazide-hydrazone and 4-thiazolidinone Derivatives of Etodolac as Potential Anticancer Agents in Leukemia Cells

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