In Vitro Models and Alternatives to Animal Testing

Stem Cells

Many in vitro test systems and replacement methods are based on the use of stem cells. Stem cells are undifferentiated cells of a multicellular organism that are capable of giving rise to an infinite number of other cells of the same type and from which certain other cell types can arise through differentiation. With the help of stem cells, researchers can, among other aspects, study molecular mechanisms of the body and the diseases associated with them, build patient-specific disease models in the laboratory and thus develop specific therapeutic approaches e.g. by testing active substances in these models.

Embryonic and Adult Stem Cells

Embryonic (so-called pluripotent) stem cells can differentiate into all tissues of the body, whereas adult (multipotent) stem cells can only develop into specific tissue types. Although embryonic stem cells are of particular interest for scientific and biomedical research due to their differentiation potential, their use and especially their isolation is ethically highly problematic since a human embryo has to be destroyed for this procedure.

Although the use of adult stem cells is ethically far less problematic, these already specialized stem cells can only give rise to certain cell types of the human body (e.g. various cell types of the skin but not blood or nerve cells).

Induced Pluripotent Stem Cells (iPS)

Ethically far less problematic than the work with embryonic stem cells is the use of so-called induced pluripotent stem cells (iPS). Due to their differentiation properties, iPS cells are almost as versatile as embryonic stem cells. Through artificial reprogramming of non-pluripotent cells (so called somatic cells), iPS cells have almost as much of a medical potential as embryonic stem cells. Of particular medical interest is the fact that for instance patient-specific iPS cells can be generated from simple skin cells of the respective patient.

Spheroids and Organoids

Spheroids

Spheroids are simple 3D cell aggregates that are formed when cells become highly spatially confined during culture in the laboratory and, as a result, adhere to each other. Viewed over a longer period of time it can be observed that the cells in the spheroid interact with each other, surround themselves with their own tissue-specific extracellular matrix (a complex mixture of a wide variety of biomolecules) and henceforth also interact with the biomolecules contained therein. This natural microenvironment provides cells with important signaling molecules, growth factors and biological stimuli which are, amongst other factors, important for cell self-organization. Due to the spherical arrangement, a metabolic gradient is formed in the cell aggregate in which the concentration of nutrients, oxygen but also various metabolic products of the cells varies. In the outermost layer, the cells divide and are well supplied by the cell culture medium which surrounds them. The cells in the inner layer are in a dormant state due to the altered concentrations while the cells in the core of the spheroid die over time, forming a necrotic core. Spheroids behave more like a natural tissue than cells in a conventional cell culture dish due to the 3D arrangement. Due to the layers that form as well as due to the concentration gradients within them, the cell aggregates resemble solid tumors and are therefore often used as in vitro models in cancer and drug research.

Organoids

Organoids are formed when cells self-organize in a three-dimensional space, forming complex organ-like structures in contrast to simple spheroids. The tissue structures are much more suitable for many scientific studies than two-dimensional cell cultures or even spheroid cultures because their nature, composition and functionality are similar to the natural organ in the body. By using human (stem) cells, they are often far superior to animal experiments in terms of their informative value and thus represent a promising approach for the development of drugs, the development of disease models and for cancer research.

Organoids can be generated from pluripotent stem cells or organ precursor cells which are usually embedded in their own tissue-specific extracellular matrix and which provides the cells for instance with important signaling molecules, growth factors and biological stimuli which are important for the self-organization of the cells into the three-dimensional organ-like tissue structures.

Tissue Models

In order to be able to investigate for example the influence of new substances in consumer protection, the effects of UV radiation or the influence of new active substances in preclinical biomedical research on specific organs or tissues, a wide variety of 3D tissue models are developed in the laboratory. By using human cells, they are ethically far less critical than animal experiments and, moreover, often more informative due to better transferability.

The design of these human tissue models reflects not only the architecture and the tissue-specific cells and biomolecules used to construct them but also allows the respective functionalities of the individual layers to be mimicked. In addition to the above mentioned applications, in vitro models can also be used to study molecular processes during infection or to understand specific pathological changes in tissues.

Organ-on-Chip-Systems and Multi-Organ-Chips

Organ-on-chip systems are small three-dimensional polymer chips with tiny micrometer-scale channels and chambers in which tissue-specific cells, spheroids or organoids can be cultured under microphysiological conditions. This means that the cells, just like in the natural human tissue, are embedded in a tissue-specific extracellular matrix and thus integrated into the small chambers of the chips. With the help of special membranes, the blood vessels are reconstructed in the chip in which the cell culture medium can flow as a blood substitute. In this way and analogous to the human body, the cells are dynamically supplied with nutrients and oxygen, metabolic products are removed and the cells are exposed to fluid mechanical stimuli. Because the chips replicate the natural, 3D microenvironment of the cells in the tissue, the cells and tissues in the chip behave very similarly to how they would in the human body. By examining the cellular behavior, aspects like the effect of new drugs or the influence of external stimuli on the cells can be studied.

Organ-on-Chip systems thus combine the unique selling points of classical in vitro models (human cells and genes) and that of animal models (complex 3D tissues and blood circulation). These microphysiological in vitro test systems can be used to answer a wide variety of medical, biological, pharmacological and toxicological questions without having to use laboratory animals.

In-silico-Models

The term in silico model refers to highly complex mathematical computer simulations based on a large amount of pre-existing animal test and laboratory data which can be used to predict the effects of specific substances on the human organism. Such models are for exapmle used for the development of new drugs to determine the interaction of a substance with organ-specific cells and to identify potentially dangerous active substances at an early developmental stage. In this way it is possible to predict at an early stage how and by which organs a new active ingredient will most likely be absorbed and metabolized in the body.

In silico models are also developed for the study of diseases and to understand their underlying molecular mechanisms as well as the complex interactions. This also makes it possible to identify potential targets for new therapies or to optimize the experimental design of clinical trials.

Computer simulations can also replace animal experiments in further education and training, for example by allowing students to virtually experience and examine the structure of the human and animal body without having to use frogs or cattle eyes.