Regulation of secretion of a-2 macroglobulin by hepatic lipocytes in relation to hepatic fibrosis
Regulation of secretion of a-2 macroglobulin by hepatic lipocytes in relation to hepatic fibrosis
Alpha-2 macroglobulin is a non-specific protease inhibitor, released predominantly by hepatocytes but also by lipocytes, and is an acute phase protein in the rat. Hepatic lipocytes are the main source of extracellular matrix (ECM) components in both normal and fibrotic liver and also release matrix metalloproteinases. 72 kDa type IV Collagenase/Gelatinase secreted by activated lipocytes degrades normal basement membrane-like matrix in space of Disse. Degradation of this matrix leads to hepatocellular dysfunction and further lipocyte activation, thus promoting liver injury and fibrosis. Matrix metalloproteinases are inhibited by α-2 macroglobulin and other specific inhibitors (eg, TIMP-1). The study investigated release of α-2 macroglobulin by hepatic lipocytes in order to understand its role in matrix remodelling.
Lipocytes were prepared by pronase/collagenase digestion followed by density gradient centrifugation and centrifugal elutriation. α-2 macroglobulin released into serum-free media was assayed by a newly developed ELISA with an assay range between 10-200 ng/ml and also by immunostaining. 72 kDa type IV C/G was assayed both by gelatin substrate SDS-PAGE and ^14C gelatin degradation assay of total activity.
These studies confirmed secretion of α-2 macroglobulin by rat lipocytes which increases as the cells are activated by culture on plastic (2.8 ng/ml on D-5 to 16.2 ng/ml on D-9). Release was also increased by exposure of lipocytes to IL-6 (up to 2.2 fold) and Kupffer cell conditioned media (KCCM -2 fold), but not IL-1, TGF-β or TNF-α. These effects of IL-6 and KCCM mirrored those seen with hepatocytes but the absolute levels and magnitudes of response were much lower. This may be explained by a biological need for a pericellular action only. Dexamethasone further enhanced stimulation by IL-6 and KCCM. None of these cytokines or KCCM had an effect on release of lipocyte 72 kDa type IV C/G activity, but dexamethasone alone or in combination with KCCM caused significant suppression. With human lipocytes IL-1 inhibited α-2 macroglobulin release but stimulated release of type IV C/G activity, also indicating differential regulation.
University of Southampton
1993
Choudhury, Ali Kawser
(1993)
Regulation of secretion of a-2 macroglobulin by hepatic lipocytes in relation to hepatic fibrosis.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
Alpha-2 macroglobulin is a non-specific protease inhibitor, released predominantly by hepatocytes but also by lipocytes, and is an acute phase protein in the rat. Hepatic lipocytes are the main source of extracellular matrix (ECM) components in both normal and fibrotic liver and also release matrix metalloproteinases. 72 kDa type IV Collagenase/Gelatinase secreted by activated lipocytes degrades normal basement membrane-like matrix in space of Disse. Degradation of this matrix leads to hepatocellular dysfunction and further lipocyte activation, thus promoting liver injury and fibrosis. Matrix metalloproteinases are inhibited by α-2 macroglobulin and other specific inhibitors (eg, TIMP-1). The study investigated release of α-2 macroglobulin by hepatic lipocytes in order to understand its role in matrix remodelling.
Lipocytes were prepared by pronase/collagenase digestion followed by density gradient centrifugation and centrifugal elutriation. α-2 macroglobulin released into serum-free media was assayed by a newly developed ELISA with an assay range between 10-200 ng/ml and also by immunostaining. 72 kDa type IV C/G was assayed both by gelatin substrate SDS-PAGE and ^14C gelatin degradation assay of total activity.
These studies confirmed secretion of α-2 macroglobulin by rat lipocytes which increases as the cells are activated by culture on plastic (2.8 ng/ml on D-5 to 16.2 ng/ml on D-9). Release was also increased by exposure of lipocytes to IL-6 (up to 2.2 fold) and Kupffer cell conditioned media (KCCM -2 fold), but not IL-1, TGF-β or TNF-α. These effects of IL-6 and KCCM mirrored those seen with hepatocytes but the absolute levels and magnitudes of response were much lower. This may be explained by a biological need for a pericellular action only. Dexamethasone further enhanced stimulation by IL-6 and KCCM. None of these cytokines or KCCM had an effect on release of lipocyte 72 kDa type IV C/G activity, but dexamethasone alone or in combination with KCCM caused significant suppression. With human lipocytes IL-1 inhibited α-2 macroglobulin release but stimulated release of type IV C/G activity, also indicating differential regulation.
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Published date: 1993
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Local EPrints ID: 462218
URI: http://eprints.soton.ac.uk/id/eprint/462218
PURE UUID: 1e45d96c-307f-4b49-a9af-42ea58dd4b0e
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Date deposited: 04 Jul 2022 19:04
Last modified: 04 Jul 2022 19:04
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Author:
Ali Kawser Choudhury
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