Hepregen products are created using breakthrough “Cell Micropatterning Technology” in which proprietary patterns of different cell types are plated on industry-standard microwell plates providing co-cultures of cells with longer life and improved function. The technology has been proven in the Hepregen HepatoPac® products in where functional liver cells (“hepatocytes”) are micropatterned to create proprietary patterns of hepatocyte “islands” surrounded by supportive stromal cells. This technology replicates the physiological microenvironment of the liver and allows the hepatocytes to exhibit normal metabolic activity for over four weeks for both short and long-term toxicology and efficacy studies during pre-clinical drug discovery. Hepatocyte health, functionality and liver enzyme activity can be extensively characterized on the HepatoPac® platform.
From Liver to Micropatterned Hepatocyte Co-Cultures
Cell Micropatterning Technology was invented by Howard Hughes Medical Institute Investigator, Sangeeta Bhatia, at the Massachusetts Institute of Technology and is centered on the concept that “architecture is important.” Cryopreserved primary hepatocytes from a species-of-interest (human, rat, monkey or dog) are micropatterned within industry-standard plates creating hepatocyte islands surrounded by supportive stromal cells. This proprietary architecture is the key to HepatoPac’s® longevity and superior predictive power.
Image above: Micropatterned Hepatocyte island (10x magnification)
Architecture is Important
HepatoPac® co-cultures are different from sandwich cultures and regular or “random” co-cultures. They contain proprietary patterning of primary hepatocyte “islands” in a “sea” of stromal cells which replicates the physiological microenvironment of the liver. The architectural organization of HepatoPac® co-cultures has been empirically optimized to promote hepatocyte health and enable stable metabolic activity for weeks, rather than hours or days.
How Does HepatoPac® Mimic the Liver?
In the liver, polarized hepatocytes are in direct contact with the blood supply and the bile cannuliculi, and are arranged in hepatic plates that are only one to two cells thick. The micropatterned design of HepatoPac® co-cultures was designed specifically to simulate this microscale in vivo architecture within the wells of industry-standard microtiter plate, thereby creating a model of fully functional hepatic plates.
Micropatterning of primary hepatocyte co-cultures, rather than simple random co-cultures, promotes and stabilizes physiologically relevant metabolic activity. Hepatocyte-stromal coupling is known to promote cellular health, but more than that, the optimal ratio and dimensions of hepatocyte islands to stromal cells is what sets HepatoPac products apart from the competition. Metabolic activity reflective of physiologically relevant levels of all Phase 1 and 2 metabolizing enzymes, transporter proteins, and in vivo-matched physiologic function comprise a platform that provides the most accurate data in the industry for all applications.
Various cellular patterns were assessed to determine optimum heterotypic interactions and co-culture stability. Current HepatoPac products are micropatterned so that individual hepatocyte islands are 500 µm in diameter, are 1200 µm from island center to center and are at 60° angles relative to adjacent islands. Morphology of HepatoPac and random co-cultures (10x) are shown.
1. Microscale Culture of Human Liver Cells for Drug Development. Nat Biotechnol. Vol. 26(1), p. 120-126 (2008).
2. Assessment of MicroPatterned Hepatocyte Co-culture System to Generate Metabolites. Drug Metabolism and Disposition, Vol. 38(10), p 1900-1905 (2010).
The involvement of inflammation and its underlying mechanisms are becoming increasingly important in the early detection of drug-induced hepatotoxicity. At present there is a clear, unmet need for an in vitro model of basal and inflamed human liver that is capable of predicting clinical outcomes. HepatoMune™ tri-cultures are HepatoPac® co-cultures (micropatterned primary hepatocytes and fibroblasts) supplemented with primary Kupffer cells that represent a highly functional miniature human liver tissue that models inflammation-mediated hepatotoxicity within an industry-standard multi-well format. The HepatoMune tri-cultures are viable for at least 10 days while maintaining both liver cell and Kupffer cell functionality.
HepatoMune™ tri-cultures enable:
- Evaluation of inflammation-mediated hepatotoxicity
- Modeling healthy liver physiology and inflammation
- Retention of long-term cultures (10 days) for testing hepatotoxicity of compounds
- Monitoring of protein therapeutic-drug interactions
- Determination of mechanistic information
HepatoMune™ tri-culture were created using patented microfabrication tools and consist of primary hepatocytes arranged in optimized domains and surrounded by stromal fibroblast. Cryopreserved primary Kupffer cells are added on day 7 of culture. pHrodo and CD68 staining illustrates the presence of functional Kupffer cells on day 10 of culture.