Background: This study describes the pre‐clinical trials of an extracorporeal bioartificial liver support system (BALSS). It includes the biochemical changes which occur in the plasma and blood of pigs with devascularized livers when the plasma is treated in a BALSS, and the testing of the system for presence or absence of infective agents, pyrogens and for toxicity. Methods: Hepatic cells were prepared from littermate juvenile white landrace pigs with a double‐step collagenase digest technique. The cell preparations were incubated with collagen‐coated dextran microspheres (CDM) for 3 h and the medium was tested to determine cellular metabolic activity. Incubation continued for a further 20 h during which the hepatic cells attach to the CDM. The CDM‐attached cells were inoculated into a hollow fibre bioreactor which was part of an extracorporeal liver support system. Results: Hepatic cell content of the bioreactor was 6 × 109± 3 × 1018 cells, equivalent to those present in half a pig's liver. The system was tested in a controlled trial with the plasma of pigs with fulminant hepatic failure (FHF) due to devascularized livers. When plasma from FHF pigs was circulated through the device there was significantly less of an increase in the accumulation of ammonia, lactate and most amino acids when hepatic cells were included in the circuit compared with those in control experiments when they were excluded. Similar changes occurred in porcine blood. There were few infections diagnosed and an absence of pyrogens, endotoxins and toxicity in the bioreactor contents or in the terminating reservoir or animal blood samples. Conclusions: We believe that the results, demonstrating function of the porcine hepatic cells in the circuit, together with low risks, justify a clinical trial of use of the BALSS in Australia.
The new 3-(4,5-dimethylthiazol-2yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) and 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5[(phenylamino)carbonyl]-2H-tetrazolium hydroxide (XTT) calorimetric assays for assessing hepatocyte density, viability and proliferation were evaluated and compared with 3-(4,5-dimethlthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and tritiated thymidine (3HTdR) incorporation. OD values of MTS or XTT, which are metabolically reduced in viable cells to a watersoluble formazan product and can be read directly, had a good correlation coefficient with hepatocyte densities in a range of 2.5-40 x lo4 cells/ml (MTS r = 0.952; XTT r = 0.902) and with hepatocyte viability (MTS r = 0.974; XTT r = 0.975). At 0, 20, 40 and 60 hr cultures, the correlation coefficients with 3HTdR for assessing hepatocyte viability and proliferation (MTS r = 0.942-0.981; XTT r = 0.953-0.992) were excellent. In contrast with MTS and XTT, MTT OD values had a poor correlation coefficient with hepatocyte densities (r = 0.672), viability 0. = 0.622) and 3HTdR incorporation (r = 0.701-0.818 at 0, 20 and 40 hour cultures). This study shows that the MTS or XTT calorimetric assays for assessing hepatocyte density, viability and proliferation are more accurate, reliable and simpler than the widely used MTT assay. They avoid use of radioactive material as required for 3HTdR incorporation. Of the two, the MTS calorimetric assay is more sensitive than the XTT.
Patterns are pervasive in nature with many examples being found in both living and inanimate systems. While researchers recognize the importance of the behavior of individuals to the structure and shape of an aggregation, a major hurdle in describing aggregated organisms has been the difficulty of tracking the movement of individuals over time. Here we present an innovative application of an analytical technique derived from statistical mechanics (a subfield of physics) to describe the spatial distribution of grouped organisms. Radial distribution and pair-correlation functions are traditionally used by physicists to describe inert particle dynamics. This novel biological application allows one to infer the behavioral characteristics of individuals within a group based solely on the spatial distribution of the aggregate population. Additionally, the method allows one to determine the correlation length, the average maximum distance over which one individual may exert an influence on another member of the aggregation. The analytical technique presented here is also important in that it minimizes two problems that typically plague studies of grouped organisms: it eliminates the need to track the movements of individuals, and it partially takes into account the presence of occluded individuals. This technique also permits quantitative comparison between aggregations formed under various environmental and/or experimental conditions. Thus, this technique may be of value to resource managers, ecologists, and others working with grouped organisms (e.g., plankton swarms, schooling fish, flocking birds, or migratory mammals) who seek to gain information about factors influencing the structure and behavior of such groups.
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