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M J Arthur

Publications and source records attributed to M J Arthur.

At least 55 records · Page 3Linked to original sources
PubMed1995

Role of Ito cells in the degradation of matrix in liver.

Liver fibrosis is a dynamic process caused by changes in not only the synthesis of matrix proteins but also their degradation. Current evidence indicates that Ito cells, when activated to a myofibroblastic phenotype, play a very active role in regulating matrix degradation in liver. This is mediated via their ability to synthesize and release several members of the matrix metalloproteinase family, a class of enzymes which are responsible for degradation of matrix proteins in the extracellular space. Activated Ito cells have been demonstrated to release prostromelysin, progelatinase A and the pro-enzyme form of interstitial collagenase. In addition, these cells can express appropriate systems for cleaving pro-metalloproteinases to active forms (e.g. the plasminogen activator system, urokinase) as well as specific tissue inhibitors of the activated metalloproteinases (TIMP). In the early phases of liver injury, enzymes with the ability to degrade components of normal liver matrix are expressed (stromelysin and gelatinase A). In contrast, in the fibrotic phase of liver injury, during which fibrillar collagens accumulate, there is little (if any) expression of interstitial collagenase but marked expression of TIMP. These findings suggest that metalloproteinase and their inhibitors play a significant role in liver injury and fibrosis.

Animals↗
PubMed1995

Kupffer cell-derived 95-kd type IV collagenase/gelatinase B: characterization and expression in cultured cells.

Release of 92-kd type IV collagenase/gelatinase, also known as gelatinase B, by inflammatory and tumor cells is increasingly recognized and is believed to facilitate cellular migration across basement membranes. It has been implicated in the pathogenesis of many diseases, but little is known of its cellular origin(s) and function in liver. In this study we have demonstrated synthesis and release of gelatinase B by human and rat Kupffer cells in primary culture. Northern analysis of RNA extracted from Kupffer cells stimulated with phorbol ester demonstrated a 2.8 kb transcript for gelatinase B. Immunoblotting and zymography of serum-free Kupffer cell-conditioned media demonstrated extracellular release of immunoreactive enzyme and gelatinase activity, Mr 92,000 (95,000 from rat cells). The organomercurial 4-aminophenyl mercuric acetate (APMA) activated the enzyme in vitro, indicating secretion primarily as a proenzyme. Stimulation of Kupffer cells by phorbol ester markedly induced gelatinase B release, which was inhibited by cycloheximide. In contrast, cycloheximide had no effect on constitutive secretion in culture, suggesting that there is some intracellular storage. Kupffer cell-derived gelatinase B was also partially purified and characterized. After separation by gelatin sepharose and gel filtration chromatogrpahy, gelatin-degrading activities of 95, 88, 75, and 65 kd were detected, the three lower-molecular-weight species probably representing activated forms. Enzyme activity was inhibited by ethyl-enediaminetetra-acetic acid (EDTA), but not by serine- and thiol-protease inhibitors, and was restored by zinc. Activity was also inhibited by tissue inhibitor of metalloproteinase-1 (TIMP-1) and alpha-2 macroglobulin. The partially purified enzyme rapidly degraded denatured collagens (gelatin) as well as native types III, IV, and V collagens, but had no activity against casein, types I and VI collagens.

Animals↗
PubMed1995

Collagenases and liver fibrosis.

Hepatic lipocytes which are activated to a myofibroblastic phenotype synthesize many of the metalloproteinases and their inhibitors, particularly TIMP-1. The available evidence suggests that this enzyme/inhibitor system for regulating matrix degradation is important in liver in two respects; (i) degradation of the normal liver matrix by the gelatinases (A and B) and stromelysin and the role this has in the pathogenesis of liver injury, and (ii) failure of matrix degradation consequent upon the relative expression of interstitial collagenase and TIMP-1 by hepatic lipocytes and the role this has in the progression of liver fibrosis. Recent progress in this field provides a clear indication that liver fibrosis is dynamic, involving a balance between matrix synthesis and regulated matrix degradation. These observations offer opportunities for the development of new therapeutic strategies in the management of liver fibrosis.

Animals↗
PubMed1994

Degradation of matrix proteins in liver fibrosis.

Hepatic fibrosis occurs as a consequence of net accumulation of matrix proteins (particularly collagen types I and III) in liver. Current concepts of the pathogenesis of liver fibrosis place major emphasis on the activation of hepatic lipocytes (fat-storing or Ito cells) to a myofibroblast-like phenotype with a consequent increase in their synthesis of matrix proteins. While this is an important factor, there is increasing evidence to indicate that liver fibrosis is a dynamic pathologic process in which altered matrix degradation may also play a significant role. Extracellular degradation of matrix proteins is regulated by a family of enzymes called the matrix metalloproteinases, which is subdivided into three groups; collagenases which degrade interstitial collagens (types I, II and III), type IV collagenases/gelatinases which degrade basement membrane (type IV) collagen and gelatins and stromelysins which degrade a broad range of substrates including proteoglycans, laminin, gelatins and fibronectin. The extracellular activity of these enzymes is regulated by several mechanisms which include alterations in gene transcription and proenzyme synthesis, cleavage of secreted proenzymes to active forms, and specific inhibition of activated forms by tissue inhibitor(s) of metalloproteinases (TIMPs). In liver, current evidence indicates that activated hepatic lipocytes and Kupffer cells play a central role in synthesis of matrix metalloproteinases. Under defined conditions they synthesize interstitial collagenase, 72 kDa and 95 kDa type IV collagenase/gelatinase and possibly stromelysin. Moreover, lipocytes also contribute to regulation of the extracellular activity of these enzymes by secretion of TIMP-1 and alpha 2-macroglobulin.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Differential cellular synthesis of insulin-like growth factor binding protein-1 (IGFBP-1) and IGFBP-3 within human liver.

The two major insulin-like growth factor-binding protein (IGFBP) species in the human circulation, IGFBP-1 and IGFBP-3, are synthesized in large amounts by liver. To determine which hepatic cell populations in human liver were responsible for the synthesis and release of these IGFBPs, we 1) performed in situ hybridization with specific complementary RNA (RNA) probes for human IGFBP-1 or-3 or performed immunohistochemical analysis to reveal the sites of messenger RNA (mRNA) presence and peptide translation, respectively, in sections of normal liver derived from organ donors; and 2) examined the release of IGFBP species by Western ligand and immunoblots of medium conditioned by isolated cultures of hepatocytes, lipocytes, and Kupffer cells. In situ hybridization showed that IGFBP-1 mRNA was distributed widely among the parenchymal cell population, which also showed immunohistochemical staining for IGFBP-1 peptide. Conversely, IGFBP-3 mRNA and immunoreactive peptide were mainly localized to Kupffer cells, which were positively identified by immunoreactivity with antiserum against the glycoprotein marker, CD68. Isolated hepatocytes released two species of IGFBP of 28 and 30-32 kilodaltons, which were recognized immunologically as IGFBP-1. Isolated Kupffer cells released only a 43- to 46-kilodalton IGFBP immunologically recognized as IGFBP-3. Lipocyte cultures released no detectable IGFBP species. The results suggest that IGFBP-1 and IGFBP-3 are derived from separate cell populations in human liver.

Adolescent↗
PubMed1992

Secretion of 72 kDa type IV collagenase/gelatinase by cultured human lipocytes. Analysis of gene expression, protein synthesis and proteinase activity.

The matrix metalloproteinases play an important role in matrix degradation, but there is limited information about this family of enzymes in either normal or diseased human liver. In this study, we have examined the synthesis of a 72 kDa type IV collagenase/gelatinase by human hepatic lipocytes in primary culture. Hepatic lipocytes were isolated from wedges of normal human donor liver by Pronase/collagenase perfusion, purified by density-gradient centrifugation, and established in primary culture on uncoated plastic. By Northern-blot analysis, the total RNA extracted from cultured human lipocytes was found to contain 3.4 kb mRNA for the 72 kDa type IV collagenase/gelatinase. Low levels of expression of this mRNA were observed in freshly isolated lipocytes but expression increased with the duration of lipocyte culture. Using anti-human 72 kDa type IV collagenase/gelatinase IgG, synthesized enzyme was immunolocalized to monensin-treated human lipocyte cultures. De novo synthesis and secretion of 72 kDa type IV collagenase/gelatinase were confirmed by immunoprecipitation of radiolabelled enzyme from medium obtained from [35S]methionine-treated cells. Activity of the secreted enzyme was demonstrated by gelatin-zymography and by degradation of soluble, radiolabelled [14C]gelatin. The enzyme was released both in active and latent pro-enzyme forms and its inhibition profile was that of a metalloproteinase. These studies indicate that cultured human hepatic lipocytes express the gene for the 72 kDa type IV collagenase/gelatinase, and secrete this enzyme, particularly in prolonged primary culture. As this enzyme exhibits degradative activity against basement membrane collagen, its release by activated hepatic lipocytes in the space of Disse could lead to disruption of the normal subendothelial liver matrix. It is suggested that this enzyme may have an important role in human liver injury and fibrosis.

Adipose Tissue↗
PubMed1992

Reversible inhibition of albumin production by rat hepatocytes maintained on a laminin-rich gel (Engelbreth-Holm-Swarm) in response to secretory products of Kupffer cells and cytokines.

Decreased albumin synthesis by hepatocytes in liver injury is thought to occur in response to Kupffer cell-derived acute-phase cytokines. In this study we used hepatocytes maintained in a differentiated phenotype, by culture on a laminin-rich gel substratum (Engelbreth-Holm-Swarm matrix), to investigate the effects of Kupffer cell-conditioned medium and purified cytokines (interleukin-1, interleukin-6 and tumor necrosis factor-alpha) on albumin synthesis. Kupffer cell-conditioned medium caused a reversible decrease in albumin synthesis to 64.7% of control (p less than 0.01, Wilcoxon's rank sum test, n = 11) on day 2. Repeated doses caused further dose-dependent reversible responses. The same result was obtained when protease inhibitors (alpha 1-antitrypsin and alpha 2-macroglobulin) were added to Kupffer cell-conditioned medium (n = 3), thus eliminating the potential effect of matrix degradation. Pure interleukin-1, interleukin-6 and tumor necrosis factor-alpha also inhibited albumin synthesis (p less than 0.05, Wilcoxon's rank sum test, n = 5), interleukin-6 having the greatest effect. After exposure to interleukin-1 (30 U.ml-1) and tumor necrosis factor-alpha (300 U.ml-1), decreased albumin synthesis was followed by a rebound increase (n = 3). Our results support the hypothesis that reduced albumin synthesis in the acute-phase response is modulated by cytokines released from Kupffer cells. Moreover, our results suggest that hepatocytes may exhibit a compensatory increase in albumin synthesis after cytokine withdrawal. These findings may be of physiological importance in the recovery from injury and the acute-phase response in vivo.

Albumins↗
PubMed1992

Familial and idiopathic colonic varices: an unusual cause of lower gastrointestinal haemorrhage.

A patient is described presenting with an acute lower gastrointestinal haemorrhage as a result of extensive colonic varices. Further investigation revealed that there were no oesophageal varices or splenomegaly. Liver biopsy showed grade II fatty change only, with no other specific or significant pathological features. Transhepatic portography showed a raised portal pressure (20 mm/Hg) but the portal system was patent throughout. There was an abnormal leash of vessels in the caecum thought to represent a variceal plexus. This patient was diagnosed as having idiopathic colonic varices. This case is discussed together with nine other reports of idiopathic colonic varices from the published literature. Four of these reports describe idiopathic colonic varices in more than one member of the same family. Possible modes of inheritance, aetiology of variceal change, natural history, and prognosis are discussed.

Colon↗
PubMed1992

Human hepatic lipocytes synthesize tissue inhibitor of metalloproteinases-1. Implications for regulation of matrix degradation in liver.

Hepatic lipocytes play a central role in the pathogenesis of liver fibrosis, both via production of extracellular matrix proteins and through secretion of matrix metalloproteinases. In this study, we have characterized lipocyte expression and release of tissue inhibitor of metalloproteinases-1 (TIMP-1), an important inhibitor of metalloproteinase activity, whose role in liver has not previously been examined. TIMP-1 was immunolocalized to human lipocytes, and secretion of TIMP-1 was confirmed by ELISA of culture media; (mean +/- SD) 159 +/- 79 ng of TIMP-1/10(6) cells per 24 h. Evidence for functional inhibitory activity of released TIMP-1 was obtained by (a) reverse zymography that demonstrated a single inhibitor band, M(r) 28 kD, that co-migrated with a TIMP-1-positive control sample; and (b) unmasking of inhibited gelatinase activity in lipocyte medium by separating it from TIMP-1 using gelatin sepharose chromatography; gelatinase activity in chromatographed medium increased more than 20-fold, compared with unfractionated medium, and could be reinhibited by adding back fractions that contained inhibitor. By Northern analysis, freshly isolated human lipocytes exhibited low levels of mRNA expression for TIMP-1, but this increased markedly relative to beta-actin expression with lipocyte activation during cell culture. We conclude that human hepatic lipocytes synthesize TIMP-1, a potent metalloproteinase inhibitor, and that TIMP-1 expression increases with lipocyte activation. These data indicate that hepatic lipocytes can regulate matrix degradation in the liver, and suggest that expression of TIMP-1 by activated lipocytes may contribute to the progression of liver fibrosis.

Adipose Tissue↗
PubMed1989

Lipocytes from normal rat liver release a neutral metalloproteinase that degrades basement membrane (type IV) collagen.

We report a proteinase that degrades basement-membrane (type IV) collagen and is produced by the liver. Its cellular source is lipocytes (fat-storing or Ito cells). Lipocytes were isolated from normal rat liver and established in primary culture. The cells synthesize and secrete a neutral proteinase, which by gelatin-substrate gel electrophoresis and gel filtration chromatography, has a molecular mass of 65,000 D. The enzyme is secreted in latent form and is activated by p-aminophenylmercuric acetate but not by trypsin. Enzyme activity in the presence of EDTA is restored selectively by zinc and is unaffected by serine-protease inhibitors. In assays with radiolabeled soluble substrates, it degrades native type IV (basement membrane) collagen but not interstitial collagen types I or V and exhibits no activity against laminin or casein. At temperatures causing partial denaturation of soluble collagen in vitro, it rapidly degrades types I and V. Thus, it is both a type IV collagenase and gelatinase. The enzyme may play a role in initiating breakdown of the subendothelial matrix in the Disse space as well as augmenting the effects of collagenases that attack native interstitial collagen.

Animals↗
PubMed1989

Activation of cultured rat hepatic lipocytes by Kupffer cell conditioned medium. Direct enhancement of matrix synthesis and stimulation of cell proliferation via induction of platelet-derived growth factor receptors.

Hepatic lipocytes appear to be central to the pathogenesis of hepatic fibrosis, undergoing activation during inflammation to a matrix-producing, proliferative cell type. We have studied the activation process in culture by examining the response of lipocytes to conditioned medium from hepatic macrophages (Kupffer cells). Lipocytes exposed to Kupffer cell medium (KCM) exhibited cellular and nuclear enlargement associated with up to a threefold increase in collagen and total protein synthesis per cell. Cell proliferation was also stimulated as measured by [3H]thymidine incorporation and direct cell counting. The latter effect was serum dependent and inhibited by antibodies to platelet-derived growth factor (PDGF). Proliferation could be stimulated by recombinant PDGF, but only after preincubation of cells with KCM. These findings suggested that KCM was eliciting expression of the PDGF receptor in lipocytes, and this was confirmed by immunoblot analysis with antibodies to the PDGF receptor. DNA synthesis in lipocytes exposed to KCM occurred at 48 h, which reflected the time required for PDGF receptor expression (24 h) plus initiation of [3H]thymidine incorporation (24 h). These results indicate that KCM has multiple stimulatory effects on cultured lipocytes similar to activation of these cells observed in vivo.

Animals↗
PubMed1988

IFAT detection of IgG specific to toxoplasma in thoracic fluids from aborted lambs: evaluation on routine diagnostic submissions.

Thoracic fluids from 171 aborted lambs from 55 flocks were examined by the indirect fluorescent antibody test for the presence of IgG specific to Toxoplasma gondii. The technique was shown to be both sensitive and specific for the diagnosis of toxoplasma abortion at a titre of 1/256, when compared with diagnoses made in the flocks and in the small number of individual fetuses in which the pathology of the cotyledons and the serology of the ewe were known.

Abortion, Veterinary↗
PubMed1988

Effect of endotoxin on release of reactive oxygen intermediates by rat hepatic macrophages.

Reactive oxygen intermediates released by activated hepatic macrophages have been implicated in the pathogenesis of a rat model of liver injury induced by sequential administration of Corynebacterium parvum and endotoxin. In this model, C. parvum causes extensive infiltration of the liver with activated macrophages, but severe liver injury occurs only after subsequent exposure to endotoxin. We have therefore investigated the effects of endotoxin on the release of reactive oxygen intermediates by C. parvum-activated hepatic macrophages. After in vitro exposure to zymosan, opsonized zymosan, or phorbol myristate acetate, hepatic macrophages isolated from C. parvum- and endotoxin-treated rats demonstrated significantly (1.5-2-fold) increased release of superoxide and oxidation of [1-14C]glucose via the hexose monophosphate shunt compared with hepatic macrophages isolated from C. parvum- and saline-treated control rats. These results indicate that endotoxin enhances the state of activation of hepatic macrophages already partially activated by C. parvum. We suggest that the increased release of reactive oxygen intermediates by these cells promotes liver injury in this model.

Animals↗