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BIOMIMESYS® Liver, a 3D cell culture model for maintaining and promoting hepatocytes functions for metabolism and toxicity studies

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BIOMIMESYS® range are hyaluronan based hydroscaffold developed to overcome the 2D flat culture limitations by recreating an in vivo-like physiology within the in vitro environment.
BIOMIMESYS®Liver scaffold is made of RGDS and galactosamine-grafted Hyaluronic acid, Adipic acid dihydrazide crosslinker and extracellular matrix (ECM) proteins (collagen type I and collagen type IV) to mimic liver-ECM composition.

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BIOMIMESYS® Liver, a 3D cell culture model for maintaining and promoting hepatocytes functions for metabolism and toxicity studies

  1. 1. Human HepatocytesHepG2 BIOMIMESYS® Liver has been tested on HepG2 and on cryopreserved Human Hepatocytes. It makes 3D cell culture easy and provides a robust in vitro and reliable model for metabolism & toxicological studies. To know more about BIOMIMESYS®Liver visit our website: www.biomimesys.com  +33(0) 769 999 137; hello@biomimesys.com BIOMIMESYS®Liver is ready to use and compatible with all analytical technologies • Available in a ready-to-use format (96 well plates) it enables the culture of hepatocytes under physiological conditions that are representative of the microenvironment found in liver tissue(2). • Hepatocytes are simply seeded on top of the hydroscaffold and placed in the incubator. • The media can be refreshed easily by pipetting. Being transparent makes it suitable for microscopy (immunofluorescence, bright field) and use in plate readers (OD, fluorescence & luminescence). • Thanks to its mechanical properties, the hydroscaffold can be easily handled with fine forceps and proteins, nucleic acid can be extracted by directly adding the lysis buffer to the hydrogel, due to its high porosity. BIOMIMESYS®Liver is physiological BIOMIMESYS® range are hyaluronan based hydroscaffold developed to overcome the 2D flat culture limitations by recreating an in vivo-like physiology within the in vitro environment. In comparison to in vivo decellularized liver tissue SEM observation of a human decellularized liver tissue (from Mazza et al. 2015(1) ) SEM observation of a BIOMIMESYS®Liver section, highlighting the collagen chain, (artificially coloured) Biomimetic structure References: (1) Decellularized human liver as a natural 3D-scaffold for liver bioengineering and transplantation. Mazza G. et al. Sci Rep.7; 5:13079, 2015. (2) Decellularization and cell seeding of whole liver biologic scaffolds composed of extracellular matrix. Faulk D. et al. J Clin Exp Hepatol. 5; 1:69-80, 2015. BIOMIMESYS®Liver hydroscaffold is made of RGDS and galactosamine-grafted Hyaluronic acid, Adipic acid dihydrazide crosslinker and extracellular matrix (ECM) proteins (collagen type I and collagen type IV) to mimic liver- ECM composition. 0 50 100 150 Day 7 Day 14 Day 21 Day 28 Diameter(µm) HepG2 spheroids sizes 100µm400µm 100µm Alive Dead Day 28  Viability and Growth  Albumin Secretion • HepG2 form spheroids with an average diameter of 80 to 100 µm after one month of cultivation. • The amount of albumin secreted by HepG2 is 20 to 30 times higher in BIOMIMESYS®Liver compared to 2D. • Cryopreserved human hepatocytes form small aggregates, 40 to 80 µm, which have a very good viability until day14.  Key genes expression levels  Toxicity study • Cryopreserved human hepatocytes grown in BIOMIMESYS®Liver represent model for routine drug testing in 96-well format. n=3 0 20 40 60 80 100 120 Day 7 Day 14 Diameter(µm) Human Hepatocytes spheroids sizes donor 1 donor 2 3D Day 7 2D 100µm 25µm 25µm Day 21 100µm MRP-2  Biliary Canaliculi Formation • MRP2 expression, its colocalization with actin and CFDA fluorescence confirms the presence of active biliary canaliculi in HepG2 during 3 weeks.  CYP activities • Basal and induced CYP1A1/A2, CYP2B6 and CYP3A4 activities are higher in BIOMIMESYS®Liver compared to 2D collagen sandwich. Basal activity 2D vs 3D Basal and induced activities in 3D 0 50 100 150 200 CYP1A1/A2 2D 3D Acetaminophen/100ngof DNA 0 500 1000 1500 2000 CYP3A4 6β-OH-Testosterone (nM)/100ngofDNA 0 20 40 60 80 CYP2B6 OH-Bupropion (nM)/100ngofDNA 0 1000 2000 3000 CYP1A1/A2 Basal Induced *** Acetaminophen/100ngof DNA 0 1000 2000 3000 4000 5000 CYP3A4 *** 6β-OH-Testosterone (nM)/100ngofDNA 0 100 200 300 400 500 600 CYP2B6 *** OH-Bupropion (nM)/100ngofDNA Day 7  Viability and Growth 100µm Day 7 100µm Day 14 400µm 100µm 100µm • HepG2 grown in BIOMIMESYS®Liver allow acute and chronic dose (3 doses) drug exposure. -10% 10% 30% 50% 70% 90% 110% 130% 0,01 0,2 4 80 Viability(ATPcontent) Chlorpromazine (µM) BIOMIMESYS®Hepatocyte -10% 10% 30% 50% 70% 90% 110% 130% 0,1 2 40 800 Amiodarone (µM) BIOMIMESYS®Liver, a 3D cell culture model for maintaining and promoting hepatocytes functions for metabolism and toxicity studies Day 14 400µm 25µm Day 21 Actin MRP2 Day 14 CFDA 100µm  Toxicity study Day 7 Day 8 Day 9 Day 10 1 dose 1st dose 2nd dose 3rd dose Viability Viability Treatment 1 dose : Day 7 - Viability analysis : Day 8 • Better expression of phase I & II enzymes and transporters genes in 3D Day 7 0 2 4 6 mRNAnormalizedby RPLPO CYP1A1 0 1 2 3 UGT1A1 0 2 4 6 NTCP 0 2 4 6 8 BSEP Patented process 0,0 0,5 1,0 1,5 2,0 2,5 Day 7 Day 14 Day 21 µgalbumin/100ngofDNA 2D BIOMIMESYS®Hepatocyte *** *** *** BIOMIMESYS®Liver -5% 15% 35% 55% 75% 95% 115% 135% 0,01 0,1 1 10 100 Viability Chlorpromazine (µM) BIOMIMESYS®Hepatocyte - Chronic BIOMIMESYS®Hepatocyte - Acute BIOMIMESYS®Liver - Chronic BIOMIMESYS®Liver - Acute BIOMIMESYS®Liver

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