The role of hepatic macrophages in the regulation of hepatocyte function
The role of hepatic macrophages in the regulation of hepatocyte function
The secretory products of Kupffer cells and inflammatory hepatic macrophages and their effects on normal hepatocyte function have been investigated in this thesis. The influence of extracellular matrix on these effects was also studied. A cell culture system was developed for these studies. Hepatic macrophages were assessed for purity, viability, function (Hexose monophosphate shunt activity (HMPS), protein symthesis) and activation (HMPS) in cell culture. Hepatocytes were assessed for viability and function (protein synthesis, albumin secretion) and modulation of function by hormones (dexamethasone) and extra cellular matrix (EHS-laminin rich gel extracted from the Engelbreth Holm Sward tumour) in cell culture. The assays used and final culture system were optimised in order to investigate the effects of hepatic macrophages on hepatocytes. Media collected from normal Kupffer cell cultures on plastic substrata caused depression of hepatocyte albumin production in cell culture, (77% of control, p< 30.01, n= 6) but did not effect leucine incorporation into hepatocyte protein (97% of control, n= 6). Soluble factors that were greater than 12kD also elicited these effects (n= 3-5). Activation of hepatic macrophages `in vivo' via the C.parvum model of liver injury enhanced their ability to diminish hepatocyte albumin production via soluble factors in cell culture (53% of control, p< 0.05, n= 6). Kupffer cell conditioned medium (KCCM) also diminished albumin production of hepatocytes that were maintained in a well differentiated state on EHS matrix. (extracellular matrix analogue) (74.2% of control, p< 0.001, n= 22). These hepatocytes were normally able to sustain albumin production for 22 days in cell culture. (mean 30.1μg m^-1 24hrs^-1 ± 3.39, n= 22, on day 11 of culture). Kupffer cell products mediated these effects via mechanisms that were independent of disruption of hepatocyte matrix interactions. Similar depressive effects occurred with hepatocytes subjected to KCCM with added protease inhibitors of alpha 1 antitrypsin and alpha 2 macroglobulin (n= 3). Therefore it was hypothesised that specific cytokines which are produced by Kupffer cells caused these effects on EHS cultured hepatocytes. The dynamics of hepatocyte albumin production in response to human recombination IL-1, IL-6, TNF-alpha and KCCM were therefore subsequently studied with the following results:- IL-6, IL-1, TNF-alpha and KCCM reversibly inhibited hepatocyte albumin production on EHS (p< 0.05 n= 5) (Wilcoxon). IL-6 produced the greatest initial depressive effects (p< 0.05 n= 5) (Wilcoxon). Preliminary data showed that EHS cultured hepatocytes demonstrated a `rebound increase' in albumin production after subjection to low doses of TNF-alpha and/or IL-1. In conclusion:- Hepatocyte albumin production is increasingly depressed by hepatic macrophages with an enhanced state of activation. Differentiated hepatocytes can regain normal rates of albumin production after subjection to Kupffer cell soluble products and cytokines. This latter observation was possible by the use of the EHS matrix as a model culture substratum.
University of Southampton
Kowalski Saunders, Piotr Josef Wladislaw
1990
Kowalski Saunders, Piotr Josef Wladislaw
Kowalski Saunders, Piotr Josef Wladislaw
(1990)
The role of hepatic macrophages in the regulation of hepatocyte function.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
The secretory products of Kupffer cells and inflammatory hepatic macrophages and their effects on normal hepatocyte function have been investigated in this thesis. The influence of extracellular matrix on these effects was also studied. A cell culture system was developed for these studies. Hepatic macrophages were assessed for purity, viability, function (Hexose monophosphate shunt activity (HMPS), protein symthesis) and activation (HMPS) in cell culture. Hepatocytes were assessed for viability and function (protein synthesis, albumin secretion) and modulation of function by hormones (dexamethasone) and extra cellular matrix (EHS-laminin rich gel extracted from the Engelbreth Holm Sward tumour) in cell culture. The assays used and final culture system were optimised in order to investigate the effects of hepatic macrophages on hepatocytes. Media collected from normal Kupffer cell cultures on plastic substrata caused depression of hepatocyte albumin production in cell culture, (77% of control, p< 30.01, n= 6) but did not effect leucine incorporation into hepatocyte protein (97% of control, n= 6). Soluble factors that were greater than 12kD also elicited these effects (n= 3-5). Activation of hepatic macrophages `in vivo' via the C.parvum model of liver injury enhanced their ability to diminish hepatocyte albumin production via soluble factors in cell culture (53% of control, p< 0.05, n= 6). Kupffer cell conditioned medium (KCCM) also diminished albumin production of hepatocytes that were maintained in a well differentiated state on EHS matrix. (extracellular matrix analogue) (74.2% of control, p< 0.001, n= 22). These hepatocytes were normally able to sustain albumin production for 22 days in cell culture. (mean 30.1μg m^-1 24hrs^-1 ± 3.39, n= 22, on day 11 of culture). Kupffer cell products mediated these effects via mechanisms that were independent of disruption of hepatocyte matrix interactions. Similar depressive effects occurred with hepatocytes subjected to KCCM with added protease inhibitors of alpha 1 antitrypsin and alpha 2 macroglobulin (n= 3). Therefore it was hypothesised that specific cytokines which are produced by Kupffer cells caused these effects on EHS cultured hepatocytes. The dynamics of hepatocyte albumin production in response to human recombination IL-1, IL-6, TNF-alpha and KCCM were therefore subsequently studied with the following results:- IL-6, IL-1, TNF-alpha and KCCM reversibly inhibited hepatocyte albumin production on EHS (p< 0.05 n= 5) (Wilcoxon). IL-6 produced the greatest initial depressive effects (p< 0.05 n= 5) (Wilcoxon). Preliminary data showed that EHS cultured hepatocytes demonstrated a `rebound increase' in albumin production after subjection to low doses of TNF-alpha and/or IL-1. In conclusion:- Hepatocyte albumin production is increasingly depressed by hepatic macrophages with an enhanced state of activation. Differentiated hepatocytes can regain normal rates of albumin production after subjection to Kupffer cell soluble products and cytokines. This latter observation was possible by the use of the EHS matrix as a model culture substratum.
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Published date: 1990
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Local EPrints ID: 460529
URI: http://eprints.soton.ac.uk/id/eprint/460529
PURE UUID: 1c1b67a2-d593-4e93-a9ed-6a40da82035e
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Date deposited: 04 Jul 2022 18:24
Last modified: 04 Jul 2022 18:24
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Author:
Piotr Josef Wladislaw Kowalski Saunders
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