Project description:Full title: Three-dimensional culture of AIDS-NHL cells influences gene expression related to B-cell development, proliferation and survival The AIDS-NHL-derived cell line, UMCL01-101, representing diffuse large B-cell lymphoma of immunoblastic morphology (AIDS-IBL), was grown in conventional, static suspension culture or three-dimensionally (3D) in the Rotating Wall Vessel (RWV) bioreactor. The objective was to assess the impact on gene expression of growth as a three-dimensional tissue assembly. Global gene expression analysis was performed on UMCL01-101 cells grown under either condition using Affymetrix microarray. UMCL01-101 cells were cultured in the Rotating Wall Vessel bioreactor to form 3D assemblies, or in conventional suspension culture, for 15 days. RNA was prepared from triplicate samples under each growth condition and submitted for microarray analysis.
Project description:Full title: Three-dimensional culture of AIDS-NHL cells influences gene expression related to B-cell development, proliferation and survival The AIDS-NHL-derived cell line, UMCL01-101, representing diffuse large B-cell lymphoma of immunoblastic morphology (AIDS-IBL), was grown in conventional, static suspension culture or three-dimensionally (3D) in the Rotating Wall Vessel (RWV) bioreactor. The objective was to assess the impact on gene expression of growth as a three-dimensional tissue assembly. Global gene expression analysis was performed on UMCL01-101 cells grown under either condition using Affymetrix microarray.
Project description:HCV proliferation is closely related to three-dimentional cellular condition. In case of blood-borne (bb) HCV culture in HuS-E2 cells, bbHCV was reproduced only from 3D-cultured cells in hollow fibers. Thus, in order to identify novel factors which support HCV proliferation under three-dimentional condition, we compared gene expression profile between 2D- and 3D-cultured HuS-E/2 cells with 3D-gene Human oligo chip 25k (Toray, Tokyo, Japan).
Project description:The generation of cell-based three dimensional (3D) nerve tissue is an attractive subject to improve graft survival and integration into host tissue for neural tissue regeneration or to model biological events in stem cell differentiation. However, although 3D organotypic culture strategy has been well established for 3D nerve tissue formation of pluripotent stem cells to study underlying biology in nerve development, cell-based nerve tissue have not been developed using human postnatal stem cells with therapeutic potential of stem cell-based therapy. Here, we established culture strategy for the generation of in vitro cell-based 3D nerve tissue from postnatal stem cells from apical papilla (SCAPs) of tooth, which is known to be originated from neural crest-derived ectomesenchyme. A stem cell population capable of differentiating into neural cell lineages was generated during the ex-vivo expansion of SCAPs in the presence of EGF and bFGF, and SCAPs differentiated into neural cells showing neural cell lineage-related molecular and gene expression profiles with morphological change under neural inductive culture condition. Moreover, we showed the first evidence that 3D cell-based nerve-like tissue with axon and myelin structure could be generated from SCAPs via 3D organotypic culture using an integrated bioprocess composing of polyethylene glycol (PEG) microwell-mediated cell spheroid formation and subsequent dynamic culture in a high aspect ratio vessel (HARV) bioreactor. In conclusion, our study propose that the culture strategy provide a novel approach to develop in vitro engineered nerve tissue using SCAPs and a foundation to study biological events in neural differentiation of postnatal stem cells.
Project description:Brain microenvironment plays an important role in neurodevelopment and function, where extracellular matrix (ECM) components and soluble factors modulate cellular features, as migration, proliferation survival and neuronal function. Disruption of microenvironment’s homeostasis is often related to pathological conditions. Here, we addressed the microenvironment remodeling occurring during in vitro differentiation of human neural stem cells (NSC) in a three-dimensional (3D) culture system. Proteome and transcriptome dynamics revealed significant changes namely at cell membrane and ECM composition during 3D differentiation, diverging significantly from the profile of monolayer cultures (2D). Structural proteoglycans typically found in brain ECM were enriched during 3D differentiation, while 2D cultures presented increased levels of basement membrane constituents (e.g., laminins, collagens and fibrillins). Moreover, higher expression levels of synaptic machinery and ion transport machinery constituents observed for 3D cultures, both at mRNA and protein levels, suggested a higher degree of neuronal maturation and organization relative to 2D differentiation. This work demonstrated that neural cellular and extracellular features can be recapitulated in the presented 3D neural cell model, highlighting its value to address molecular defects in cell-ECM interactions associated with neurological disorders. <html><head>Associated GEO dataset is available at</head><body><a href="https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi">GSE102139</a></body></html>
Project description:3D cell culture models are recognized for representing the physiological microenvironment and exhibiting higher concordance with in vivo conditions, when compared to a conventional 2D cell culture model. However, cells grown in 3D cultures are likely to exhibit slower growth than those in 2D cultures. We found that addition of a novel small molecule named GA-017 to culture media promotes the cell proliferation particularly under 3D conditions. Gene microarrays were used to observe the global gene expression in Skov3 cells cultured under 3D condition with DMSO or GA-017 and identified distinct classes of up or down-regulated genes.
Project description:Obvious advantages of 3D cell culture model are the cell morphology better reflecting tissue cell morphology, the formation of zones of i) active proliferation, ii) quiescent viable cell zone and iii) necrotic zone, as well as formation of nutrition, oxygen and drug gradients better reflecting cellular environment in tissue. Nevertheless the 3D cultures are a model still not resembling full complexity of tumor tissue environment in vivo. Few obvious limitations of 3D cell cultures as cancer research model are the lack of vasculature, host immune response and other cell-cell interactions that occur between cancer and stromal cells in tumors. Recognized advantages and limitations of the 3D cell culture models, however do not suggest directly the areas of cancer biology where 3D models could be applied with highest success. Hence detailed analysis at the molecular level of 2D/3D cell cultures and tumors in vivo are needed to unlock the power of 3D cell culture model. In order to elucidate which biological pathways of cancer cells in tumors are best resembled by the 3D cell culture model we have analyzed whole genome gene expression changes in mouse LLC1 cell line when cultured in 2D or laminin rich ECM 3D system. RNA was isolated 48h after growing in two different cell culture systems.
Project description:3D in vitro culture models of cancer cells in extracellular matrix (ECM) have been developed to investigate drug targeting and resistance or, alternatively, mechanisms of invasion, however models allowing analysis of shared pathways mediating invasion and therapy resistance are lacking. To evaluate therapy response associated with cancer cell invasion, we here used 3D invasion culture of tumor spheroids in 3D fibrillar collagen and applied Ethanol-Ethyl cinnamate (EtOH-ECi) based optical clearing to detect both spheroid core and invasion zone by subcellular-resolved 3D microscopy. When subjected to a single dose of irradiation (4 Gy), we detected preferential cell survival in the invasion zone, compared to the spheroid core. By physical separation of core and invasion zone we identified differentially regulated genes between preferentially engaged in invading cells controlling cell division, repair and survival. This imaging-based 3D invasion culture may be useful for analysis of complex therapy-response patterns in cancer cells in drug discovery and invasion-associated resistance development.
Project description:Human bone marrow mesenchymal stromal cells (MSCs) are conventionally cultured as adherent monolayers on tissue culture plastic. MSCs can also be cultured as 3D cell aggregates (spheroids). Optimised 3D conditions (60,000 MSCs cultured as a spheroid for 5 days) inhibited MSC proliferation and induced cell shrinkage in the absence of cell death. Primary human MSCs isolated from 2 donors were cultured under both monolayer (2D MSCs) and optimised 3D (3D MSCs) conditions. High quality RNA was isolated from all samples, and global gene expression analysis was performed in duplicate (using Agilent SurePrint G3 Human Gene Expression 8x60K v2 Microarrays) to identify gene expression changes in 3D compared to 2D MSC cultures.