Analysis of multivariables during porcine liver digestion to improve hepatocyte yield and viability for use in bioartificial liver support systems

In order to achieve optimal BALSS Function, preparation of porcine hepatocy tes with high yield, viability, and P450 activity is known to be important. To date hepatocyte yields have varied from 0.58 x 10(10) to 3.45 x 10(10) and viabilities from 75% to 95% within and between laboratories, even when using the same digestion methods and procedures, indicating that hepatocyte isolation during porcine liver digestion is not fully optimized. The aim o f this work was to identify the critical parameters affecting cell recovery during porcine liver harvesting by investigating 21 variables involved in the process, including pig body and liver weight, different digestion Limes of perfusates, DW, a range of concentrations of sodium and chloride in EDT A, and collagenase perfusates. Univariate and multivariate analysis of a re trospective study (n = 23) revealed that low perfusate pH during the proces s of digestion had a positive effect on hepatocyte yield (p < 0.05), while high (relative) concentrations of sodium and chloride in the perfusates had significant negative effects on hepatocyte viability (both p < 0.05). Sodi um and chloride had narrow optimal ranges for achieving a >90% viability. T hese findings were then tested in a prospective study (n = 10) and further verified. High hepatocyte viabilities (91.8 +/- 1.6%, p = 0.036) and yields (2.56 +/- 0.48 x 10(10)) were achieved consistently, and P450IA1 activity was increased after sodium acid chloride concentrations and pH in the perfu sates were controlled. The physiological mechanism by which sodium and chlo ride affects: hepatocyte viability during porcine liver digestion is discus sed.