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Three-dimensional organotypic culture: experimental models of mammalian biology and disease

Key Points

  • Three-dimensional (3D) culture protocols have been developed for diverse tissues, organs and disease states.

  • 3D culture enables imaging of mammalian organogenesis at the cellular level.

  • Genetic manipulation within 3D cultures can resolve the cellular and molecular basis of tissue-level phenotypes.

  • 3D culture enables the independent evaluation of how distinct features of the microenvironment regulate organogenesis and disease.

  • Induced pluripotent stem (iPS) cell-derived 3D cultures enable the generation and study of tissues derived from the somatic cells of a patient.

  • 3D culture is a natural point of integration for fundamental, translational and clinical research.

Abstract

Mammalian organs are challenging to study as they are fairly inaccessible to experimental manipulation and optical observation. Recent advances in three-dimensional (3D) culture techniques, coupled with the ability to independently manipulate genetic and microenvironmental factors, have enabled the real-time study of mammalian tissues. These systems have been used to visualize the cellular basis of epithelial morphogenesis, to test the roles of specific genes in regulating cell behaviours within epithelial tissues and to elucidate the contribution of microenvironmental factors to normal and disease processes. Collectively, these novel models can be used to answer fundamental biological questions and generate replacement human tissues, and they enable testing of novel therapeutic approaches, often using patient-derived cells.

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Figure 1: Cellular inputs to organotypic cultures.
Figure 2: The major categories of cell culture.
Figure 3: The cellular basis of epithelial tube elongation.
Figure 4: Genetic regulation of cell behaviours in mammalian tissues.
Figure 5: The role of the microenvironment in regulating epithelial function.

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Acknowledgements

The authors apologize to the many scientists whose outstanding work could not be cited owing to space limitations. A.J.E. and E.R.S. were supported by a Research Scholar Grant (RSG-12-141-01-CSM) from the American Cancer Society. A.J.E. was also supported in part by funds from the National Institutes of Health National Cancer Institute (NIH–NCI) (U01 CA155758), by a Jerome L. Greene Foundation Discovery Project, by a grant from the Mary Kay Ash Foundation (036-13), by funds from the Cindy Rosencrans Fund for Triple Negative Breast Cancer Research and by a grant from the Breast Cancer Research Foundation.

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Glossary

Epithelium

A type of animal tissue derived from ectoderm or endoderm that lines all cavities and body surfaces and consists of one or more layers of polarized, tightly connected cells.

Connective tissue

A type of animal tissue derived from mesenchyme that provides structural and nutritional support and connectivity among other tissues; it consists of individual cells, ground substance and fibres.

Extracellular matrix

(ECM). The non-cellular component of tissues that provides both structural support and signalling cues to cells; it is composed of a network of proteins such as collagen, fibronectin and laminin.

Mesenchyme

Loosely organized, undifferentiated cells derived from embryonic mesoderm that give rise to the connective tissues of the body and the lymphatic and circulatory systems.

Induced pluripotent stem cells

(iPS cells). Adult somatic cells that are genetically reprogrammed to generate embryonic-like pluripotent stem cells.

Basement membrane

An organized thin layer of extracellular matrix proteins that separates the epithelium from the surrounding connective tissue.

Stromal cells

The connective tissue cells of an organ (for example, fibroblasts), which support the function of the parenchymal cells of the organ.

Tissue stem cells

Adult stem cells that can give rise to some or all of the specialized cells of the tissue or organ from which they originate.

Matrigel

A gelatinous basement membrane matrix derived from Engelbreth–Holm–Swarm mouse sarcoma cells; it promotes cell differentiation and models the in vivo microenvironment of many tissues.

Self-organization

In tissues, the spontaneous formation of a highly ordered structure from a population of cells in the absence of pre-patterns.

Stratified epithelium

An epithelium that is composed of two or more layers of cells; it is often found in locations that require increased protection, such as exterior body surfaces.

Microfluidic systems

Devices that comprise submillimetre channels, pumps and valves that enable controlled, reproducible analysis of small samples of cells in nanolitre or picolitre volumes.

Asymmetric divisions

Cell divisions that result in two daughter cells with different fates, such as localization into distinct epithelial cell layers with unequal inheritance of polarity proteins.

Cre–lox

A site-specific recombination tool that uses the enzyme Cre recombinase to induce deletions, translocations or inversions in segments of genomic DNA that are flanked by loxP sites.

CRISPR–Cas9

(Clustered, regularly interspaced short palindromic repeats–CRISPR-associated protein 9). A genome-editing tool that uses the microbial RNA-guided Cas9 nuclease to make targeted changes in the DNA of eukaryotic cells.

Hypoplasia

Abnormal tissue or organ development owing to a deficient number of cells.

Hyperplasia

Abnormal tissue or organ development owing to an excess number of cells.

Tumour xenografts

Human tumours that are implanted into immunocompromised animal hosts.

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Shamir, E., Ewald, A. Three-dimensional organotypic culture: experimental models of mammalian biology and disease. Nat Rev Mol Cell Biol 15, 647–664 (2014). https://doi.org/10.1038/nrm3873

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