Flow in human tissue models
Go with the flow: Modelling human tissues using microfluidics
Microfluidics is the science of controlling fluids in tiny channels. It’s a rapidly developing field of research which opens an enormous range of possibilities for diagnostics and lab-on-a-chip. Most recently, microfluidic technology is applied for the development of organ-on-a-chip devices that allows for physiologically relevant 3D tissue cultures.
Why is flow essential for biological tissue functioning?
Blood vessels and microvasculature
Diffusion of nutrients and oxygen is limited to short distances (several micrometers). Humans and larger animals in general therefore need the cardiovascular system to transport nutrients and oxygen to all cells of the body. This is achieved by an extensive network of capillaries through every tissue, which ensures cells of proper nourishment and waste removal. Blood vessels transport oxygen, ions, glucose, and other metabolites.
Flow drives molecules across cell membranes
The flow of fluids through our body maintains local concentration differences on which most forms of transport rely. Oxygen and carbon dioxide cross cell membranes by simple molecular diffusion. This process does not require additional energy, since it is driven by a concentration gradient. Co-transport, by which ions and glucose molecules are transported into the cell, also relies on a concentration difference of one of the transported molecules. Other gradients are achieved by active transport of ions or metabolites across the cell membrane. The continuous refreshment of local fluids maintains the gradients between tissue and blood, which drives the uptake of oxygen and nutrients, and the release of waste products by the cells in the tissue.
Flow initiates cell polarity
Most tissues are characterized by coherent cellular polarization, by which the opposing sides of cells are defined by specific properties. Cell polarity is critical for tissue development and enables cells to carry out specialized functions, such as migration and molecular transport.
Many cell types are known to sense flow with their primary cilia. The presence of fluid flow contributes to the initiation of dynamic cytoskeleton rearrangements important for apical-basal and planar cell polarity in endothelial as well as epithelial cells. This process partly underlies the adherence between epithelial or endothelial cells by tight junctions, desmosomes, and adherens junctions, to form a surface or internal cavity.
Why do we need flow in human tissue models?
- Continuous supply of media — Microfluidics continuously provide oxygen and nutrients to the cells in culture, while waste products are removed or diluted in large volumes of culture media. This allows to establish and maintain 3D cell culture models with an in vivo like environment for research and drug evaluations.
- Higher liquid-to-cell ratio — In standard 2D culture, the media content changes considerably over time. Until the medium is refreshed, nutrients are depleted and (toxic) waste products accumulate, leading to suboptimal culture conditions. Fluctuations in the composition of culture media may affect the quality of your culture, since cells respond by metabolic adaptation. In a microfluidic system with media reservoirs, the medium is continuously refreshed and the liquid-to-cell ratio is higher. In this way, consumption of growth media by the cells does not drastically change the medium content, meaning cells and organ models grow in a more physiologically relevant way.
- In vivo-like diffusion time — When testing the effects of pharmacological compounds in standard 2D cultures, the monolayer is directly exposed to the substances. This is almost never the case in the human body. There, compounds are taken up, distributed through the vascular network, and diffuse into underlying tissues. 3D cultures can mimic the diffusion of compounds into tissues. Besides distribution, the microvascular network also limits diffusion of compounds and should therefore be taken into account. These aspects can be easily mimicked in microphysiological systems such as the OrganoPlate®.
- Polarised cells — Flow of fluids induces cell polarization in epithelial and endothelial tissues. Correct cell polarization is essential for proper functioning of a tissue, and therefore contributes to a physiologically relevant tissue model.
- The establishment of gradients in culture — A microfluidic device can be used to control chemical gradients in tissues, by adding compounds to a perfusion channel, adjacent to a channel containing cells and extracellular matrix. Such a gradient can for example be used to induce vessel sprouting when angiogenic factors are added.
- Vascularized tissue (co-)cultures — Microfluidic devices such as the OrganoPlate®, enable co-cultures with spatial organisation of specific cell types. In vitro tissue models can be combined with vasculature, to make them more physiologically relevant. Vascularized tissue co-cultures can be used for drug efficacy studies, transport studies, compound screens, and toxicity studies.
Methods to create flow in a microfluidic culture system
Gravity-driven perfusion does not require pumping equipment nor tubes. Gravity-driven perfusion is based on a continuously changing angle of tissue-chips, which initiates a flow of fluids through the microfluidic channels to the lowest position within the plate. The OrganoPlate® is the only organ-on-a-chip platform available that can maintain flow without the use of pumps. Placing the OrganoPlate® on a Perfusion Rocker™ in the incubator provides continuous medium flow through the microfluidic channels.
The traditional way to perfuse tissue cultures, is by using a pump system to drive flow in a microfluidic system. Pumps can be used to create a unidirectional flow of fluids through the system, and enables for high controllability of flow rate. However, pump-driven flow systems require you to manually connect tubing and a pump to every individual chip. This should be air- and fluid-tight and air bubble formation in the system should be prevented at all times. When culturing multiple perfused tissue-chips at the same time, this can take up a considerable amount of time and incubator space.
The choice between pump-driven or gravity driven perfusion depends on the application and the required throughput and ease-of-handling. For lower flow rates, which are in most cases sufficient to exert the functions of perfusion flow described in this article, gravity-driven flow is more cost-effective and easier to implement and maintain.
- Lizama, C. O., & Zovein, A. C. (2013). Polarizing pathways: balancing endothelial polarity, permeability, and lumen formation. Experimental cell research, 319(9), 1247-1254.
- Iruela-Arispe, M. L., & Davis, G. E. (2009). Cellular and molecular mechanisms of vascular lumen formation. Developmental cell, 16(2), 222-231.