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Enhance your epithelial barriers assay using Alvetex

By Frederique Tholozan, PhD

Barrier assays are an important tool for in vitro drug development and drug safety testing. For example, they can be used to assess the absorption of a drug candidate through the intestinal wall1 or the corrosion of chemical compounds on skin2. In this article we discuss how Alvetex can be used to increase the physiological relevance of your in vitro barrier assays. 

What are epithelial barrier assays?

Barrier assays have a long history and a whole range has been developed3 to respond to the needs of small academic labs all the way to large pharmaceutical toxicity departments. But where small-scale studies can use in vivo animal studies or ex vivo human tissue for the greatest degree of physiological relevance, large-scale studies have to rely on assays of much lesser complexity, such as Caco2 monolayers or PAMPA5 for intestinal permeability, to streamline workflows, keep costs down or comply with regulations to decrease animal testing6.

alvetex-caco2-ccd18-int-mucosaCaco2 cells co-cultured with CCD-18co fibroblasts in Alvetex Scaffold, forming a full-thickness model of intestinal mucosa.

Figure 1: Caco2 cells co-cultured with CCD-18co fibroblasts in Alvetex Scaffold, forming a full-thickness model of intestinal mucosa.

What are Bio-Engineered Tissue Models?

Enter bio-engineered 3D tissues models7. 3D cell culture scaffolds restore the Z-dimension which is missing in flat-surface cell culture and allows for the building of mucosal and sub-mucosal compartments in a single model. As a result, the epithelial layer can benefit from paracellular signalling from fibroblasts grown in 3D, with endogenously-deposited ECM. The epithelial cells mature better and their morphology and functionality end up being much closer to in vivo compared to conventional culture on flat surfaces.

Physiologically-Relevant Barrier Assays

At REPROCELL, our Alvetex 3D polyHIPE substrate has been proven to help in the building of such physiologically-relevant barrier assays using primary human cells for a number of applications:

  • Invasiveness of early-stage melanoma in a full-thickness skin model8
  • Barrier integrity against SDS challenge in a full-thickness skin model9
  • Infection of Influenza viral particles in a full-thickness airway epithelium model10

Alvetex is a great substrate for building barrier assays:

  • 200 microns thickness — mimics in vivo passive diffusion metrics
  • large pore-based structure — encourages cell migration, multilayering and true three-dimensionality
  • insert formats — compatible with 6-well and 12-well plates
  • inertness and chemical composition — precludes any exogenous signalling

Unconventional Applications

In addition, Alvetex has been used for more unconventional applications of barrier assays, such as the activation of immune cells after their migration through an epithelium11. Actual migration through an epithelium was required, so traditional paracrine co-culture models would have been inadequate, but using the barrier assay improved the functionality of the cells-of-interest prior to the main test of their efficacy. It is a clever solution to the problem of getting more physiologically-relevant cells without having to alter the set-up of an already-established assay system.

Increasing the complexity of established assays is always a challenge, but the rewards gained from greater physiological relevance are worth the effort, both in terms of better disease understanding and to avoid costly drug failures at late pipeline stages.

Find out more about Alvetex 3D cell culture

Discover the science: how Alvetex enables cells maintain their in vivo morphology, behaviour and responsiveness within an in vitro model system. Alvetex enables cells to  freely interact with their neighbours in 3D cell culture — so that they function in a more physiologically relevant manner. Download your Alvetex Brochure today.

Editors note: This blog was first published in July 2019 and has since been updated for accuracy and clarity.

Further reading

  1. Balimane, Praveen & Han, Yong-Hae & Chong, Saeho. (2006). Current industrial practices of assessing permeability and P-glycoprotein interaction. The AAPS journal. 8. E1-13.
  2. Macfarlane, M., Jones, P., Goebel, C., Dufour, E., Rowland, J., Araki, D., Costabel-Farkas, M., Hewitt, N. J., Hibatallah, J., Kirst, A., McNamee, P., Schellauf, F., & Scheel, J. (2009). A tiered approach to the use of alternatives to animal testing for the safety assessment of cosmetics: skin irritation. Regulatory toxicology and pharmacology : RTP54(2), 188–196.
  3. Sarmento, & Albuquerque, J. (2016). Concepts and models for drug permeability studies : cell and tissue based in vitro culture models. Woodhead Publishing.
  4. Balimane, P. V., Han, Y. H., & Chong, S. (2006). Current industrial practices of assessing permeability and P-glycoprotein interaction. The AAPS journal8(1), E1–E13.
  5. Balimane, P. V., Han, Y. H., & Chong, S. (2006). Current industrial practices of assessing permeability and P-glycoprotein interaction. The AAPS journal8(1), E1–E13.
  6. Gordon, Sarah & Daneshian et al (2015). Non-Animal Models of Epithelial Barriers (Skin, Intestine and Lung) in Research, Industrial Applications and Regulatory Toxicology. ALTEX. 32. 327-378.
  7. Torras, N., García-Díaz, M., Fernández-Majada, V., & Martínez, E. (2018). Mimicking Epithelial Tissues in Three-Dimensional Cell Culture Models. Frontiers in bioengineering and biotechnology6, 197.
  8. David S. Hill, Neil D.P. Robinson, Matthew P. Caley, Mei Chen, Edel A. O'Toole, Jane L. Armstrong, Stefan Przyborski, Penny E. Lovat; A Novel Fully Humanized 3D Skin Equivalent to Model Early Melanoma Invasion. Mol Cancer Ther 1 November 2015; 14 (11): 2665–2673.
  9. Roger, M., Fullard, N., Costello, L., Bradbury, S., Markiewicz, E., O'Reilly, S., Darling, N., Ritchie, P., Määttä, A., Karakesisoglou, I., Nelson, G., von Zglinicki, T., Dicolandrea, T., Isfort, R., Bascom, C. and Przyborski, S. (2019), Bioengineering the microanatomy of human skin. J. Anat., 234: 438-455.
  10. Marrazzo P, Maccari S, Taddei A, Bevan L, Telford J, et al. (2016) 3D Reconstruction of the Human Airway Mucosa In Vitro as an Experimental Model to Study NTHi Infections. PLOS ONE 11(4): e0153985.
  11. Grunwell, J.R., Giacalone, V.D., Stephenson, S. et al. Neutrophil Dysfunction in the Airways of Children with Acute Respiratory Failure Due to Lower Respiratory Tract Viral and Bacterial Coinfections. Sci Rep 9, 2874 (2019).

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