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Alginate foam-based three-dimensional culture to investigate drug sensitivity in primary leukaemia cells.

ABSTRACT: The development of assays for evaluating the sensitivity of leukaemia cells to anti-cancer agents is becoming an important aspect of personalized medicine. Conventional cell cultures lack the three-dimensional (3D) structure of the bone marrow (BM), the extracellular matrix and stromal components which are crucial for the growth and survival of leukaemia stem cells. To accurately predict the sensitivity of the leukaemia cells in an in vitro assay a culturing system containing the essential components of BM is required. In this study, we developed a porous calcium alginate foam-based scaffold to be used for 3D culture. The new 3D culture was shown to be cell compatible as it supported the proliferation of both normal haematopoietic and leukaemia cells. Our cell differential assay for myeloid markers showed that the porous foam-based 3D culture enhanced myeloid differentiation in both leukaemia and normal haematopoietic cells compared to two-dimensional culture. The foam-based scaffold reduced the sensitivity of the leukaemia cells to the tested antileukaemia agents in K562 and HL60 leukaemia cell line model and also primary myeloid leukaemia cells. This observation supports the application of calcium alginate foams as scaffold components of the 3D cultures for investigation of sensitivity to antileukaemia agents in primary myeloid cells.

SUBMITTER: Karimpoor M

PROVIDER: S-EPMC5938583 | biostudies-literature | 2018 Apr

REPOSITORIES: biostudies-literature

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Alginate foam-based three-dimensional culture to investigate drug sensitivity in primary leukaemia cells.

Karimpoor Mahroo M Yebra-Fernandez Eva E Parhizkar Maryam M Orlu Mine M Craig Duncan D Khorashad Jamshid S JS Edirisinghe Mohan M

Journal of the Royal Society, Interface 20180401 141

The development of assays for evaluating the sensitivity of leukaemia cells to anti-cancer agents is becoming an important aspect of personalized medicine. Conventional cell cultures lack the three-dimensional (3D) structure of the bone marrow (BM), the extracellular matrix and stromal components which are crucial for the growth and survival of leukaemia stem cells. To accurately predict the sensitivity of the leukaemia cells in an <i>in vitro</i> assay a culturing system containing the essentia ...[more]

PMID: 29695605

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Project description:Defining the final size and geometry of engineered tissues through precise control of the scalar and vector components of tissue growth is a necessary benchmark for regenerative medicine, but it has proved to be a significant challenge for tissue engineers. The growth plate cartilage that promotes elongation of the long bones is a good model system for studying morphogenetic mechanisms because cartilage is composed of a single cell type, the chondrocyte; chondrocytes are readily maintained in culture; and growth trajectory is predominately in a single vector. In this cartilage, growth is generated via a differentiation program that is spatially and temporally regulated by an interconnected network composed of long- and short-range signaling mechanisms that together result in the formation of functionally distinct cellular zones. To facilitate investigation of the mechanisms underlying anisotropic growth, we developed an in vitro model of the growth plate cartilage by using neonatal mouse growth plate chondrocytes encapsulated in alginate hydrogel beads. In bead cultures, encapsulated chondrocytes showed high viability, cartilage matrix deposition, low levels of chondrocyte hypertrophy, and a progressive increase in cell proliferation over 7 days in culture. Exogenous factors were used to test functionality of the parathyroid-related protein-Indian hedgehog (PTHrP-IHH) signaling interaction, which is a crucial feedback loop for regulation of growth. Consistent with in vivo observations, exogenous PTHrP stimulated cell proliferation and inhibited hypertrophy, whereas IHH signaling stimulated chondrocyte hypertrophy. Importantly, the treatment of alginate bead cultures with IHH or thyroxine resulted in formation of a discrete domain of hypertrophic cells that mimics tissue architecture of native growth plate cartilage. Together, these studies are the first demonstration of a tunable in vitro system to model the signaling network interactions that are required to induce zonal architecture in growth plate chondrocytes, which could also potentially be used to grow cartilage cultures of specific geometries to meet personalized patient needs.

Project description:BackgroundAlginate matrix 3-dimensional culture offers the opportunity for the development and maturation of human secondary follicles in vitro. However, alginate may not be the most suitable culture system for human primordial/primary follicles in vitro. Thus, the innovation of alginate matrix 3-dimensional culture systems for human primordial/primary follicles could hold promise as an ideal approach to restoring fertility.MethodsWe extracted primordial/primary follicles from ovarian tissues collected from patients with non-ovarian benign gynecological conditions. Fibroblasts were isolated from dermal tissue from 1 male patient who had undergone posthectomy. The isolated human follicles were randomly divided into 2 groups and encapsulated within fibroblast-alginate-hydrogels or alginate hydrogels. The survival and growth of human primordial/primary follicles were measured after 21 days of in vitro culture.ResultsThe dermal fibroblasts in alginate hydrogel microcapsules were round in shape, and were distributed as uniform clouds on the surface and gaps of the alginate. After 21 days of culture, the survival rate of follicles in the fibroblast-alginate group was higher than that of the alginate group (P<0.05). The diameter of follicles in the fibroblast-alginate group and the alginate group after 21 days of culture was 152.80±13.64 and 129.14±9.95 μm, respectively (P<0.05). After 21-day culture, the mean cpm (log-converted) for 3H-thymidine incorporated by granulosa cells in the fibroblast-alginate and alginate groups was 6.87±0.24 and 4.63±0.38, respectively (P<0.05). After 21 days of culture, the messenger RNA expression levels of growth differentiation factor 9 (GDF9) and bone morphogenetic protein 15 (BMP15) were significantly higher in oocytes in fibroblast-alginate hydrogels than in those in alginate hydrogels (P<0.05).ConclusionsHuman fibroblasts are beneficial to the development of human follicles in 3-dimensional culture alginate gel systems over a long period of time. More studies are required to investigate the molecular biological mechanisms of human fibroblasts that promote follicle growth in vitro.

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Alginate foam-based three-dimensional culture to investigate drug sensitivity in primary leukaemia cells.

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Alginate foam-based three-dimensional culture to investigate drug sensitivity in primary leukaemia cells.

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