In VitroCell. Dev.Biol.26:67-74,January1990 9 1990TissueCultureAssociation 0883-8364/90 $01.50+0.00

C U L T U R E O F H U M A N H E P A T O C Y T E S F R O M SMALL S U R G I C A L L I V E R B I O P S I E S . B I O C H E M I C A L C H A R A C T E R I Z A T I O N AND C O M P A R I S O N W I T H I N V l V O

M. JOSl~ GOMEZ-LECH6N, PILAR LOPEZ, TERESA DONATO, ANGEL MONTOYA, AMPARO LARRAURi, PATRICIA GIMENEZ, RAMON TRULLENQUE, RICARDO FABRA, ANnJOSI~V. CASTELL Centro de Investigact~n, Hospital La Fe, INSALUD. Avda Campanar 21, 46009-Valencia, Spain, and Servicio de Cirugi~ A. Hospital General de Valencia, Camino Tres Cruces s/n., 4600 Valencia, Spain (R. T., R. F.)

{Received28 July 1989; accepted 10 August 1989)

SUMMARY High yields of human hepatocytes Cup to 23 )< liP viable cells/g) were obtained from small surgical liver biopsies {1 to 3 g) by a two-step collagenase microperfusion method. Cell viability was about 95%, attachment efficiency of hepatocytes seeded on fibronectin-coated plates was 80% within 1 h after plating, and cells survived for about 2 wk in serum-free Ham's F12 containing 0.2% bovine serum albumin, 10-8 M insulin, and 10-8 M dexamethasone. To evaluate the metabolism of human hepatocytes in serum-free conditions, we measured their most characteristic biochemical functions and compared them to those reported for human liver. Mter 24 h in culture, glycogen content was 1250 -t- 177 nmol glucose/mg cell protein and remained stable for several days. Gluconeogenesis from lactate in hormone-free media was (3.50 ___ 0.17 nmol glucose.mg-'.min-q similar to that reported for human liver. Insulin at 10-8 M activated glycolysis ()

Culture of human hepatocytes from small surgical liver biopsies. Biochemical characterization and comparison with in vivo.

High yields of human hepatocytes (up to 23 X 10(6) viable cells/g) were obtained from small surgical liver biopsies (1 to 3 g) by a two-step collagena...
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