Studies on cellular engraftment and hepatocytic differentiation in liver injury and repair

BACKGROUND: In recent years, considerable attention has been directed towards the use of cellular therapies in the management of acute and chronic liver failure. The majority of clinical studies have utilised hepatocyte infusions, although there is limited understanding of the factors which limit the engraftment of cells in the face of liver injury. The use of stem cells as sources of hepatocytes which can contribute to liver repair has been examined in rodent models, but the extrapolation of these findings to human stem cells is unclear.AIM: In this thesis the factors which regulate the adhesion and survival of hepatocytes in the face of acute liver injury environment are examined. In addition, factors which regulate the differentiation of human stem cells towards hepatocytes are examined in vitro and in vivo.MATERIALS AND METHODS: Human hepatoblastoma (HepG2) cells were used as a model of human hepatocytes to study the effect of serum from patients with acute liver failure. Various laboratory assays were used to determine effects on adhesion, cell necrosis/apoptosis, integrin expression (flow cytometry) and integrin activation. Human cord blood was used as a source of human stem cells for both in vitro experiments and the in vivo work with the NOD-SCID miceRESULTS: Paracetamol-induced liver injury results in the marked up regulation of collagen on hepatic sinusoids. Adhesion of HepG2 cells to Collagen after exposure to fulminant serum was reduced within only a few hours. Apoptosis occurred approximately 24-48 hours after incubation and is associated with caspase3 activation. Furthermore, fulminant serum reduces the adhesive capabilities of HepG2 cells by a rapid down-regulation of their Pi-integrin activity. Loss of cellular adhesion and subsequent apoptosis can be reversed by treatment ofHepG2 cells with the stimulatory mAb TS2/16. Human cord blood could not be directed in vitro down the hepatocytic lineage under any of the different combinations of cytokines/matrix. In vivo however infused human cord blood cells are capable of engrafting into NOD-SCID mouse liver and differentiating down the hepatocytic lineage without fusion to host hepatocytesCONCLUSION: In this thesis, mechanisms regulating the engraftment and survival of HepG2 cells during exposure to fulminant serum were identified. Human stem cells were also demonstrated in vivo (but not in vitro) to differentiate into hepatocytes within the NOD-SCID mouse liver with no evidence of cellular fusion

Monitoring the dead as an ecosystem indicator

Dead animal biomass (carrion) is present in all terrestrial ecosystems, and its consumption, decomposition, and dispersal can have measurable effects on vertebrates, invertebrates, microbes, parasites, plants, and soil. But despite the number of studies examining the influence of carrion on food webs, there has been no attempt to identify how general ecological processes around carrion might be used as an ecosystem indicator. We suggest that knowledge of scavenging and decomposition rates, scavenger diversity, abundance, and behavior around carrion, along with assessments of vegetation, soil, microbe, and parasite presence, can be used individually or in combination to understand food web dynamics. Monitoring carrion could also assist comparisons of ecosystem processes among terrestrial landscapes and biomes. Although there is outstanding research needed to fully integrate carrion ecology and monitoring into ecosystem management, we see great potential in using carrion as an ecosystem indicator of an intact and functional food web. © 2021 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd. **Please note that there are multiple authors for this article therefore only the name of the first 5 including Federation University Australia affiliate “Philip Barton" is provided in this record*

Mesenchymal stromal cell therapy in liver disease: opportunities and lessons to be learnt?

End-stage liver disease is responsible for 30,000 deaths per year in the United States alone, and it is continuing to increase every year. With liver transplantation the only curative treatment currently available, new therapies are in great demand. Mesenchymal stem cells (MSC) offer an opportunity to both treat liver inflammatory damage, as well as reverse some of the changes that occur following chronic liver injury. With the ability to regulate both the innate and adaptive immune system, as well as both inhibit and promote apoptosis of effector inflammatory cells, there are numerous therapeutic opportunities for MSC in acute and chronic liver disease. This article critically appraises the potential therapeutic roles of MSC in liver disease, as well as the barriers to their adoption into clinical practice

Clinical effectiveness of cell therapies in patients with chronic liver disease and acute-on-chronic liver failure: a systematic review protocol

PRISMA-P (Preferred Reporting Items for Systematic review and Meta-Analysis Protocols) 2015 checklist: recommended items to address in a systematic review protocol*. (DOC 82 kb

Development of an invasively monitored porcine model of acetaminophen-induced acute liver failure

Background: The development of effective therapies for acute liver failure (ALF) is limited by our knowledge of the pathophysiology of this condition, and the lack of suitable large animal models of acetaminophen toxicity. Our aim was to develop a reproducible invasively-monitored porcine model of acetaminophen-induced ALF. Method: 35kg pigs were maintained under general anaesthesia and invasively monitored. Control pigs received a saline infusion, whereas ALF pigs received acetaminophen intravenously for 12 hours to maintain blood concentrations between 200-300 mg/l. Animals surviving 28 hours were euthanased. Results: Cytochrome p450 levels in phenobarbital pre-treated animals were significantly higher than non pre-treated animals (300 vs 100 pmol/mg protein). Control pigs (n=4) survived 28-hour anaesthesia without incident. Of nine pigs that received acetaminophen, four survived 20 hours and two survived 28 hours. Injured animals developed hypotension (mean arterial pressure; 40.8+/-5.9 vs 59+/-2.0 mmHg), increased cardiac output (7.26+/-1.86 vs 3.30+/-0.40 l/min) and decreased systemic vascular resistance (8.48+/-2.75 vs 16.2+/-1.76 mPa/s/m3). Dyspnoea developed as liver injury progressed and the increased pulmonary vascular resistance (636+/-95 vs 301+/-26.9 mPa/s/m3) observed may reflect the development of respiratory distress syndrome. Liver damage was confirmed by deterioration in pH (7.23+/-0.05 vs 7.45+/-0.02) and prothrombin time (36+/-2 vs 8.9+/-0.3 seconds) compared with controls. Factor V and VII levels were reduced to 9.3 and 15.5% of starting values in injured animals. A marked increase in serum AST (471.5+/-210 vs 42+/-8.14) coincided with a marked reduction in serum albumin (11.5+/-1.71 vs 25+/-1 g/dL) in injured animals. Animals displayed evidence of renal impairment; mean creatinine levels 280.2+/-36.5 vs 131.6+/-9.33 mumol/l. Liver histology revealed evidence of severe centrilobular necrosis with coagulative necrosis. Marked renal tubular necrosis was also seen. Methaemoglobin levels did not rise >5%. Intracranial hypertension was not seen (ICP monitoring), but there was biochemical evidence of encephalopathy by the reduction of Fischer's ratio from 5.6 +/- 1.1 to 0.45 +/- 0.06. Conclusion: We have developed a reproducible large animal model of acetaminophen-induced liver failure, which allows in-depth investigation of the pathophysiological basis of this condition. Furthermore, this represents an important large animal model for testing artificial liver support systems