To effectively study diseases, we must first replicate their processes and symptoms outside the human body.
Understanding the mechanisms of disease development and progression has long been a goal of biomedical research. In humans, analysis of disease has been largely limited to relatively non-invasive tests, such as blood tests, small biopsies, and ECG and EEG. To more comprehensively understand human disease, researchers have instead turned to disease models.
Animals have been used as models to study physiology and disease since the 4th and 3rd centuries BCE. Between the mid-16th and late-17th centuries, researchers and physicians observed animals to document physiological revelations such as embryo formation and the systemic and pulmonary blood circuits. Physicians also tested surgical procedures on animals before employing them on human subjects.
Today, researchers can replicate many symptoms of disease, from behavioural to systemic to cellular, by breeding animals with specific mutations. Knock-out and knock-in mutations can produce or reduce diseases in animals by altering the function of disease-associated genes. For example, the p53 knockout mouse is a strain of mouse in which the p53 gene, normally responsible for suppressing excessive cell replication that would otherwise cause tumours, is non-functional. This allows researchers to investigate tumour formation and novel cancer drugs and therapies.
Most medical advances and treatments used today, from diabetes treatments to pain relief medications, were in part developed using animals. It is imperative that novel drugs and treatments are tested for safety in animals before they are trialled in humans.
But there are limitations on the use of animals for modelling human disease. Because animals have a different genetic make-up from humans, disease development and drug and therapy action may not be comparable. Compounds that are safe and effective in a mouse, for example, may be toxic and life threatening in humans. A tragic example of this was the development of the drug fialuridine, for the treatment of Hepatitis B. Fialuridine was tested in mice, rats and primates for safety and efficacy before being tested in humans. Out of 15 trial participants, five people died from liver failure and two people only survived after life-saving liver transplants.
The use of animals for research and drug development carries many hefty ethical questions: should animals be bred and kept in lab environments for research? And should animals be produced with disease phenotypes to aid scientific and medical research? The Australian code for the care and use of animals for scientific purposes provides guidelines and regulations for using animals in research. However, it is widely accepted in the scientific community that ‘lower’ animals such as mice and rats — as opposed to primates — are used experimentally and terminated readily and inconsequentially. For example, to maintain a specific genetic mouse linage, mice must be bred and kept in the lab environment until future use, with excess offspring culled if not utilised. These lab animals are often kept in small cages in sterile environments with limited enrichment, conditions that would be ethically questionable were they inflicted upon household pets.
Another widespread technique for modelling disease is to culture a range of cell types, allowing researchers to understand disease physiology on cellular and sub-cellular levels. Cell lines with gene knock-out or knock-in mechanisms, similar to animal models, allow the study of specific cell types and specific disease phenotypes. Cell function modulators — molecules that alter cell processes — are used to investigate the enhancement or blockage of distinct cellular functions.
In the last decade, the generation of stem cells from adult human cells has become routine. This allows scientists to make stem cells from small tissue samples, such as skin or blood, which can be subsequently differentiated into any cell type of interest to make human model organoids, a type of 3D cell culture. These methodologies allow scientists to recapitulate organ structures using cells of human origin, which helps overcome the limitations of 2D cell culture. Biopsies taken from patients with known disease mutations allow the generation of organoids using cells carrying the disease genotype, as opposed to induced mutations as in animal models. Three-dimensional cell culture permits cultured cells to grow and interact with their environment and other cells in all three directions. Compared to 2D or monolayer cell culture, researchers can study cell interactions in a more biologically and physiologically representative way.
Further, a range of organ-on-a-chip models are in development. These artificial organ models imitate the physiology, structure and functions of organs and improve on 3D cell culture systems by incorporating tissue-tissue interactions and molecular gradients.
The future is looking bright for these physiological models. “I think for most people, the goal is to replace animal testing and to carry out personalized medicine in a more effective way,” said Dinal Ingber, founding director of Harvard University’s Wyss Institute for Biologically Inspired Engineering, in an interview last year.
While monolayer and 3D cell culture can tell us much about cellular function and dysfunction, they cannot reproduce the systemic functions of physiology and disease provided by animal models. Additionally, drug screening trials in 2D and 3D cultures have yielded different results, highlighting the limitations of cell culture for investigating disease pathology, and necessitating more complex tissue and disease models, such as organs-on-a-chip.
Techniques for the non-animal modelling of disease are rapidly becoming more available and reliable, improving options for human-derived experimental models. 3D culture techniques, including organs-on-a-chip, are hugely valuable for understanding disease pathology and treatment action on a cellular and microenvironment level. However, for the moment, animal testing remains a vital part of the disease modelling and drug development processes. The future may see the advancement of organ models to replicate systemic features of human physiology; but for now, at least, that is a science-fiction pipe dream.
Edited by Andrew Katsis