Efrat Manaster, Ph.D
Sigma-Aldrich Research Technical Service, Jerusalem, Israel
Three-dimensional (3D) extracellular matrix-based cell culture systems are being used as research tools for understanding normal and disease systems, and for drug screening in vitro. The extracellular matrix (ECM) and its attachment factor components are discussed in this article in relation to their function in structural biology and their availability on the market for in vitro applications.
All animal cells are organized as highly complex specialized tissues. Whether the tissues are in a solid or liquid state, cells do not comprise the tissue alone, but are in constant close contact with the extracellular matrix environment. Cells secrete various types of proteins that create an intercalated mesh comprising a tissuespecialized ECM.
In general, the function of the ECM is important for:
Thus, the ECM plays a key role in tissue homeostasis, in normal, pathological, and malignant states.
In general, the ECM is comprised of insoluble collagen fibers and soluble proteins. Different ECM components share identical structure/function modules, are interconnected, and some possess the ability to associate with cell surface components, hence their designation as attachment factors. Thus, a constant functional association exists between the ECM and tissue cells.
It is clear from these descriptions why alterations of the ECM can result in changes in pathological states, such as enhancement of tumor cell metastatic ability. 2
As explained previously, the structure and function of ECM components create intricate networks of cells and the ECM, leading to constant cross talk between the inner and outer cellular environment. The contact areas between the plasma membrane and the ECM are called focal adhesions. The molecular composition of these structures varies between tissues, but in general, cell-surface integrin molecules associate with both intracellular cytoskeleton-associated proteins and with ECM components (Figure 1). The ECM is a functional unit in intracellular signaling.
Figure 1. A typical focal adhesion structure.
It has become clear that the ECM is not merely a physical scaffold for holding cells in place within tissue. The following are examples of the roles attachment factors play at the ECM-cell membrane junction in cellular function:
Ex-vivo/in-vitro research with primary or immortalized cells is undoubtedly a convenient, relatively cheap, and reliable way to acquire preliminary information about various biological functions. However, in vitro cultures are inferior to in vivo studies in many ways. One concern is that growing cells on two-dimensional (2D) substrata creates an artificial lower and upper surface polarity, an artifact reflected in many physiological properties. However, in vivo studies are expensive, slow, and often present difficulty in isolating a single studied process/mechanism.
Three-dimensional (3D) cell cultures developed over the past years have proven to be a bridge between 2D cultures and in vivo studies, thus combining the convenience of a controlled, relatively cheap, and rapid experimental environment with physiological reliability. Researcher can now have control over both cell and matrix content, which allows mimicking of different tissues and physiological conditions, while maintaining a tissue-like structure of cells in an ECM context. Cells can be diversely manipulated prior to cultivation in the 3D matrix. The matrices relative transparency also allows visualization of processes and structures.9 For these reasons, 3D cultures have been designated “Engineered Tissues”.
In general, 3D cell cultures contain primary or immortalized cells, naïve or manipulated, seeded either on, below, or within an ECM-based matrix. Specialized cell/tissue-derived ECM matrices can then be prepared because the ECM is secreted from cells in vivo.10
Major attachment factors and ECM components are also commercially available to aid in preparing specific engineered tissues such as the following:
Recent advances in 3D cell culture development have led to the realization that not only are engineered tissues a valuable tool for mimicking in vivo tissue conditions, but their physical structure and attachment factor composition are critical for this behavior.
Below are examples of how 3D tissue performance is dependent on ECM and attachment factor composition:
Research over recent years has led to optimal conditions for in vitro/ex vivo work with cell cultures. The growing dilemma of complications with in vivo protocols on one hand and in vitro derived artifacts on the other hand have brought about the need for a research platform that would be reliable, resemble in vivo conditions, and allow rapid research under controlled conditions. Due to excellent academic and clinical research, and the responsiveness of the life science industry, some state-ofthe- art models exist today for growing cells in three-dimensional engineered tissues. Although some tissue types are yet to be successfully reproduced in vitro and one cannot fully mimic the complex cell types and processes intercalated in vivo, engineered 3D tissues have potential in the clinical diagnosis and treatment fields.
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