Long-term production of major coagulation factors and inhibitors by primary human hepatocytes in vitro: perspectives for clinical application.
OBJECTIVE
Patients with coagulation factor disorders require lifelong symptomatic treatment. This is associated with limited efficacy and transmission risks. From a clinical point of view, hepatocyte transplantation offers a rational alternative but is currently being hampered by lack of functional stability of engrafted cells. It was the aim of our study to devise culture conditions providing stable cell polarity, attachment and growth factor stimulation to improve longevity and coagulation factor production.


METHODS
Human hepatocytes (HC) were plated on different extracellular matrices, inside collagen gel or Matrigel. HC were grown inside growth factor-enriched serum-free medium (SFM) or exposed to media switching from differentiation (DM) to dedifferentiation (DeDM).


RESULTS
Over more than 30 days in vitro human HC synthesized coagulation factors (factors VII, VIII, IX, fibrinogen) and coagulation inhibitors (antithrombin III, protein C). Protein synthesis was augmented when HC were grown inside a 3D collagen type I matrix, while Matrigel showed no additional benefit. Soluble growth factors improved coagulation factor production when applied in SFM or in sequential DM/DeDM. Coagulation factor levels ranged from 3% to 12% in the first week to 2.5-5% after 4 weeks, reaching biologically relevant levels.


CONCLUSIONS
Preserved synthesis and secretion of coagulation factors in balanced proportion by human HC in this model may offer new perspectives for HC transplantation in coagulation defects of patients.
Boost
Kim A
KA
Department of Anesthesiology, Intensive Care and Pain Therapy, Johann Wolfgang Goethe-University, Frankfurt am Main, Germany.
Auth
Marcus K H
MK
Woitaschek
Dirk
D
Kim
Hyun-Soo
HS
Hilgard
Philip
P
Nadalin
Silvio
S
Blaheta
Roman A
RA
D019774:Blood Coagulation Factor Inhibitors / Q000378:metabolism*; D001779:Blood Coagulation Factors / Q000378:metabolism*; D002448:Cell Adhesion; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D016764:Cell Polarity; D002470:Cell Survival; D017690:Cell Transplantation / Q000379:methods*; D002478:Cells, Cultured; D020147:Coagulation Protein Disorders / Q000378:metabolism / Q000601:surgery*; D003094:Collagen / Q000378:metabolism; D024042:Collagen Type I / Q000378:metabolism; D003470:Culture Media / Q000737:chemistry; D004338:Drug Combinations; D022781:Hepatocytes / Q000378:metabolism* / Q000473:pathology / Q000637:transplantation; D006801:Humans; D007797:Laminin / Q000378:metabolism; D011509:Proteoglycans / Q000378:metabolism; D013997:Time Factors
D019774:Blood Coagulation Factor Inhibitors; D001779:Blood Coagulation Factors; D024042:Collagen Type I; D003470:Culture Media; D004338:Drug Combinations; D007797:Laminin; D011509:Proteoglycans; C059485:matrigel; D003094:Collagen