P.N. Newsome

Isolation of Primary Human Hepatocytes from Normal and Diseased Liver Tissue: A One Hundred Liver Experience

Successful and consistent isolation of primary human hepatocytes remains a challenge for both cell-based therapeutics/transplantation and laboratory research. Several centres around the world have extensive experience in the isolation of human hepatocytes from non-diseased livers obtained from donor liver surplus to surgical requirement or at hepatic resection for tumours. These livers are an important but limited source of cells for therapy or research. The capacity to isolate cells from diseased liver tissue removed at transplantation would substantially increase availability of cells for research. However no studies comparing the outcome of human hepatocytes isolation from diseased and non-diseased livers presently exist. Here we report our experience isolating human hepatocytes from organ donors, non-diseased resected liver and cirrhotic tissue. We report the cell yields and functional qualities of cells isolated from the different types of liver and demonstrate that a single rigorous protocol allows the routine harvest of good quality primary hepatocytes from the most commonly accessible human liver tissue samples.

Advanced non-alcoholic fatty liver disease and adipose tissue fibrosis in patients with Alström syndrome.

Alström syndrome (AS) is a recessive monogenic syndrome characterized by obesity, extreme insulin resistance and multi‐organ fibrosis. Despite phenotypically being high risk of non‐alcoholic fatty liver disease (NAFLD), there is a lack of data on the extent of fibrosis in the liver and its close links to adipose in patients with AS. Our aim was to characterize the hepatic and adipose phenotype in patients with AS.

N-acetylcysteine and liberase improve success of hepatocyte isolation and viability of hepatocytes isolated from normal and diseased liver

Abstract Background Successful hepatocyte isolation is crucial for development of cellular transplantation and biochemical research. Most researchers isolate hepatocytes from surplus donor tissue or normal tissue removed during resection of liver tumours. However, most tissue available for research is from explanted diseased liver and donor tissue rejected for transplant. We previously described our experience of hepatocyte isolation using liberase from such livers with a success rate of 51% and median viability of 40%. Liberase is a highly purified collagenase that has been shown to improve the viability of isolated porcine hepatocytes. N-acetylcysteine (NAC) has been shown to improve the viability of human hepatocytes isolated from steatotic donor tissue. The aim of this study was to determine the effect of both reagents in combination on the outcome of hepatocyte isolation from normal and diseased liver. Methods Hepatocytes were isolated from 30 consecutive liver specimens as previously described (old protocol). A further 30 consecutive liver specimens were perfused with buffer containing NAC and standard collagenase substituted by liberase (new protocol). Success was defined as maintenance of cell adhesion and morphology for 48 h and/or their successful use in laboratory studies. Mann-Whitney tests were used to compare results. Fishers exact test was used for categorical data. Findings Baseline factors were similar for both groups. The delay to tissue processing was slightly less in the new protocol group (median 2 h vs 4 h, p=0·007). The success rate improved from 40% (12/30) with the old protocol to 70% (21/30) with the new protocol (p=0·037), and the median viable cell yield increased from 7·3 × 10 4 to 28·3 × 10 4 cells per g tissue (p=0·003). After multivariable analysis adjusting for the difference in time delay, the success rate (p=0·014) and viable cell yield per g tissue (p=0·001) remained significantly improved. Interpretation NAC and liberase greatly improve the success of hepatocyte isolation and result in a significantly higher viable cell yield. Use of these agents may improve the availability of hepatocytes for transplantation as well as laboratory research. Funding UK Medical Research Council.

Identifying patients with NASH that are non‐responders to therapy

Alessandra Mangia, M.D. Gregory J. Dore, M.D., Ph.D. Andrew R. Lloyd, M.D., Ph.D. Jacob George, M.D., Ph.D. Mohammed Eslam, M.D., Ph.D. Storr Liver Centre Westmead Millennium Institute and Westmead Hospital University of Sydney Sydney, Australia Division of Hepatology Ospedale Casa Sollievo della Sofferenza, IRCCS San Giovanni Rotondo, Italy Kirby Institute The University of New South Wales Sydney, NSW, Australia School of Medical Sciences, University of New South Wales, Sydney, Australia