Isolation of primary human hepatocytes & LiMAx-test

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Tissue Engineering (Part C: Methods) accepted our paper entitled "The predictive value of the LiMAx-test for the isolation of primary human hepatocytes".
Authors are R.D. Major, M. Kluge, M. Jara, M. Nösser, R. Horner, J. Gassner, B. Struecker, P. Tang, S. Lippert, A. Reutzel-Selke, D. Geisel, T. Denecke, M. Stockmann, J. Pratschke, I.M. Sauer, and N. Raschzok.

The need for primary human hepatocytes is constantly growing, for basic research as well as for therapeutic applications. However, the isolation outcome strongly depends on the quality of liver tissue, and we are still lacking a preoperative test that allows the prediction of the hepatocyte isolation outcome. Here we evaluated the “maximal liver function capacity test” (LiMAx) as predictive test for the quantitative and qualitative outcome of hepatocyte isolation. This test is already used in clinical routine to measure preoperative and to predict postoperative liver function.
The patient’s preoperative mean LiMAx was obtained from the patient records and preoperative CT and MRI images were used to calculate the whole liver volume in order to adjust the mean LiMAx. The outcome parameters of the hepatocyte isolation procedures were analyzed in correlation with the adjusted mean LiMAx.
Primary human hepatocytes were isolated from partial hepatectomies (n=64).
From these 64 hepatectomies we included 48 to our study and correlated their isolation outcome parameters with volume corrected LiMAx values. From a total of 11 hepatocyte isolation procedures, metabolic parameters (albumin, urea and aspartate aminotransferase) were assessed during the hepatocyte cultivation period of 5 days. The volume adjusted mean LiMAx showed a significant positive correlation with the total cell yield (p= 0.049;r= 0.242;n= 48). The correlations of volume adjusted LiMAx values with viable cell yield and cell viability did not reach statistical significance. A sub-group analysis of isolations from patients with colorectal metastasis revealed a significant correlation between volume adjusted mean LiMAx and total cell yield (p= 0.012;r= 0.488;n= 21) and viable cell yield (p=0.034;r=0.405;n=21). Whereas a sub-group analysis of isolations of patients with carcinoma of the biliary tree showed significant correlations of volume adjusted mean LiMAx with cell viability (r= 0.387;p= 0.046;n=20) and lacked significant correlations with total cell yield (r= - 0.060;p= 0.401;n=20) and viable cell yield (r= 0.012;p= 0.480;n=20). The volume-adjusted mean LiMAx did not show a significant correlation with any of the metabolic parameters. In conclusion, the LiMAx-test might be a useful tool to predict the quantitative outcome of hepatocyte isolation, as long as underlying liver disease is taken into consideration.

Cells isolated from diseased explanted livers

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The International Journal of Artificial Organs (official journal of the European Society for Artificial Organs [ESAO]) published our paper on Isolation, characterization and cold storage of cells isolated from diseased explanted livers. Authors are Belaschk E, Rohn S, Mukiibi R, Reutzel-Selke A, Tang P, Sawitzki B, Pratschke J, Sauer IM and Mogl MT.

Livers discarded after standard organ retrieval are commonly used as a cell source for hepatocyte transplantation. Due to the scarcity of organ donors, this leads to a shortage of suitable cells for transplantation. Here, the isolation of liver cells from diseased livers removed during liver transplantation is studied and compared to the isolation of cells from liver specimens obtained during partial liver resection.
Hepatocytes from 20 diseased explanted livers (Ex-group) were isolated, cultured and stored at 4°C for up to 48 hours, and compared to hepatocytes isolated from the normal liver tissue of 14 liver lobe resections (Rx-group). The nonparenchymal cell fraction (NPC) was analyzed by flow cytometry to identify potential liver progenitor cells, and OptiPrep™ (Sigma-Aldrich) density gradient centrifugation was used to enrich the progenitor cells for immediate transplantation.
There were no differences in viability, cell integrity and metabolic activity in cell culture and survival after cold storage when comparing the hepatocytes from the Rx-group and the Ex-group. In some cases, the latter group showed tendencies of increased resistance to isolation and storage procedures. The NPC of the Ex-group livers contained considerably more EpCAM+ and significantly more CD90+ cells than the Rx-group. Progenitor cell enrichment was not sufficient for clinical application.
Hepatocytes isolated from diseased explanted livers showed the essential characteristics of being adequate for cell transplantation. Increased numbers of liver progenitor cells can be isolated from diseased explanted livers. These results support the feasibility of using diseased explanted livers as a cell source for liver cell transplantation.

Int J Artif Organs. 2017 May 23:0. doi: 10.5301/ijao.5000594. [Epub ahead of print]