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Albumin, fibrinogen and transferrin synthesis in isolated rat hepatocyte suspensions. A model for the study of plasma protein synthesis.

ABSTRACT: A system using hepatocyte suspensions in vitro was developed for studying the synthesis of albumin, fibrinogen and transferrin. Conditions for optimum survival of the hepatocyte and for synthesis of these plasma proteins were defined for this system. These conditions included the use of horse serum (17.5 percent, v/v, heat-inactivated), an enriched medium (Waymouth's MB 752/1), an O2 tension of between 18.7 times 10(3) and 26.7 times 10(3) Pa and constant stirring. Albumin, fibrinogen and transferrin synthesis rates were obtained of 0.32 p 0.094(10), 0.12 p 0.030(11) and 0.097 p 0.017(10) [mean p S.D. (n)]mg/h per g of hepatocytes respectively. These rates were maintained for the first 12h of study and synthesis continued at a diminished rate up to 48h. The synthesis of albumin was decreased in a medium containing less amino acids and glucose, but that of fibrinogen was substantially unaffected. ATP concentrations up to 12h and RNA/DNA ratios up to 24h were comparable with values in vivo. The ability to study cells up to 48h permitted us to find that the addition of a mixture of hormones consisting of glucagon, cortisol, tri-iodothyronine and growth hormone enhanced fibrinogen synthesis. Addition of insulin to the above mixture resulted in increased synthesis for albumin and transferrin but not for fibrinogen.

SUBMITTER: Jeejeebhoy KN

PROVIDER: S-EPMC1165284 | biostudies-other | 1975 Jan

REPOSITORIES: biostudies-other

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Project description:The possibility of using two different isotopomers, for the incorporation of isotopically labeled amino acids, was explored to enable longitudinal studies of de novo synthesis of two export liver proteins, albumin and fibrinogen. The agreement of the synthesis rates between the two different labels was evaluated along with the reproducibility of repeated experiments using different time intervals. Healthy volunteers were studied in a standardized fed state. Protocol A (n = 10) involved two measurements 48 hours apart. Protocol B (n = 6) involved three measurements at baseline and five hours and then seven days after the initial measurement. De novo synthesis of albumin and fibrinogen by the incorporation of D5-phenylalanine or D8-phenylalanine were measured using the flooding dose technique. Albumin and fibrinogen were isolated from plasma using standard techniques. Fractional and absolute synthesis rates were calculated. Repeated measurements employing the two isotoptomers showed good agreement for albumin fractional synthesis rate after 48 hours (p = 0.92) and after 7 days (p = 0.99), with a coefficient of variation of 5.9% when using the same isotopic label. For fibrinogen, the coefficient of variation for the fractional synthesis rate employing the same isotopic label was 16.6%. Repeated measurements after 48 hours and seven days showed less agreement although there was no statistical difference (P = 0.32 and P = 0.30 respectively). Repeated measurement after five hours showed a statistical significant difference for the fractional synthesis rate of fibrinogen (p = 0.008) but not for albumin (p = 0.12). Repeated measurements of albumin de novo synthesis more than 48 hours apart show acceptable agreement using either one or two different isotopic labels. For fibrinogen the larger intra-individual scatter necessitates larger study groups to detect changes in longitudinal studies. Repeated measurements within 48 hours need to be validated further.

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Albumin, fibrinogen and transferrin synthesis in isolated rat hepatocyte suspensions. A model for the study of plasma protein synthesis.

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