Earlier charcoal haemoperfusion in fulminant hepatic failure

Protein adsorption on polymeric biomaterials I. Adsorption isotherms

A technique has been developed by the authors that allows hepatocyte attachment on collagen-coated microcarriers resulting in prolonged hepatocyte viability and function both in vivo and in vitro. Rat hepatocytes were obtained by portal vein collagenase perfusion. Intraperitoneally transplanted microcarrier-attached normal hepatocytes into congeneic Gunn rats were functioning 3-4 weeks later, as shown by the presence and persistence of conjugated bilirubin in recipient bile, sustained decrease in serum bilirubin, uptake of Tc99m-DESIDA, and morphologic criteria. Intraperitoneal transplantation of normal microcarrier-attached hepatocytes into genetically albumin deficient rats (NAR) resulted in marked increase in plasma albumin levels (6 days without and 21 days with Cyclosporin A immunosuppression). Microcarrier-attached hepatocytes transplanted after 2 weeks of storage at -80 C into congeneic Gunn rats were viable and functional as assessed by criteria outlined above. An extracorporeal liver perfusion system was developed using the microcarrier-attached hepatocytes that was capable of synthesizing and conjugating bilirubin and synthesizing liver-specific proteins.

New method of hepatocyte transplantation and extracorporeal liver support.

Combination of detoxifying liver support systems with liver cell bioreactors may have additional benefits for the treatment of liver failure due to the replacement of known and unknown metabolic activities of the liver. However, the problem of side effects and possible risks caused by the use of animal hepatocytes or hepatoma cells remains unsolved which underlines the need of a safety barrier between the patients blood and the extracorporeal bioreactor. Passive filters do not meet the requirements of such membranes, because in liver failure desired and undesired molecules in the patients blood share similar physicochemical properties. That challenges the development of biologically designed separation membranes. A hybrid membrane is formed by implementation of transport proteins into a highly permeable hollow fiber. The transport of free solutes and albumin bound toxins is tested in vitro in comparison with conventional high flux membranes. The transport characteristics for tightly albumin bound toxins are significantly improved for the hybrid membrane. The transport of albumin bound toxins across the membrane is not associated with albumin. The selectivity of the transport is evaluated in vivo. No significant loss of middle molecular weight hormones attached to other carrier proteins was observed. Neither transport of immunologically relevant proteins across the membrane nor loss of valuable proteins was measured. Also in vivo, a significant reduction of protein bound toxins and a transport of metabolically relevant solutes, like amino acids, was shown. The presented hybrid membrane may be used like an "intelligent membrane" as a safety barrier between the patients blood and cell devices.

A carrier-mediated transport of toxins in a hybrid membrane. safety barrier between a patients blood and a bioartificial liver

Liver diseases are of major concern as they now account for millions of deaths annually. As a result of the increased incidence of liver disease, many patients die on the transplant waiting list, before a donor organ becomes available. To meet the huge demand for donor liver, alternative approaches using liver tissue engineering principles are being actively pursued. Even though adult hepatocytes, the primary cells of the liver are most preferred for tissue engineering of liver, their limited availability, isolation from diseased organs, lack of in vitro propagation and deterioration of function acts as a major drawback to their use. Various approaches have been taken to prevent the functional deterioration of hepatocytes including the provision of an adequate extracellular matrix and co-culture with non-parenchymal cells of liver. Great progress has also been made to differentiate human stem cells to hepatocytes and to use them for liver tissue engineering applications. This review provides an overview of recent challenges, issues and cell sources with regard to liver tissue engineering.

https://onlinelibrary.wiley.com/doi/pdf/10.1111/liv.12134

Liver tissue engineering and cell sources: issues and challenges

Platelets adhere to most foreign surfaces. As a result, polymers and albumin have been suggested as possible coatings to improve the blood compatibility of such surfaces. Amberlite XAD-7 has a high affinity for human serum albumin (HSA) and several protein-bound toxic metabolites. In the present study it is shown that HSA can be tightly bound to Amberlite XAD-7 without the use of chemical coupling agents. Optimal binding was achieved at pH 5.0 and the adsorption conforms to Langmuir's isotherm. Theoretical analysis of data and absence of a residual surface coating visible with scanning electron microscopy suggest a monolayer of albumin. The amount of HSA eluted from the resin under severe flow conditions was negligible (approximately 1%). Furthermore, in stirred batch studies with human plasma, the HSA coating did not decrease the adsorptive capacity of XAD-7 for bile acids and bilirubin.

Albumin-coated Amberlite XAD-7 resin for hemoperfusion in acute liver failure. Part I: adsorption studies.

Albumin-coated Amberlite XAD-7 has been previously shown to be blood compatible in in vitro hemoperfusion experiments with human blood. In this study, the preliminary results are Reported on single hemoperfusions with albumin-coated XAD-7 resin in four patients with acute liver failure. The mean platelet count was 116 ± SE 16.3% of the initial arterial value and the mean white cell count was 96 ± SE 6.5% of initial at the end of four hours of hemoperfusion. Removal of bilirubin, phenols and substances in the middle molecular weight range by the resin was demonstrated.



These preliminary results suggest albumin-coated Amberlite XAD-7 resin to be blood compatible and capable of removing protein-bound and middle molecular weight substances from patients with acute liver failure. Further clinical evaluation of repeated resin hemoperfusion is required to determine whether this treatment will be beneficial to patient survival.

Albumin-coated Amberlite XAD-7 resin for hemoperfusion in acute liver failure. Part II: in vivo evaluation.

In the present study the conditions leading to tight binding of human serum albumin to the Amberlite XAD-7 resin without the use of chemical coupling agents have been defined. Optimal binding (10.97 mm/kg dry XAD-7) was achieved at pH 5.0 and adsorption conformed to a Langmuir isotherm. Theoretical analysis of the data suggest adsorption of a monolayer of albumin which is supported by the absence of visual surface coating on scanning electron micrographs. Binding of human serum albumin was reduced when two chemical coupling agents, glutaraldehyde and carbodiimide were included in the human serum albumin solution, the elution of adsorbed HSA from the resins under severe flow conditions was reduced, and the amount eluted was minimal in all instances.

Adsorption of human serum albumin to amberlite XAD‐7 resin

In previous studies severe side effects, including irreversible hypotension, platelet losses and rises in Swank screen filtration pressure, have been recorded during the treatment by charcoal haemoperfusion of patients with fulminant hepatic failure. It has also been demonstrated that rises in Swank screen filtration pressure in an in vitro test circuit are due to the presence of platelet aggregates. In this study Prostacyclin was infused at a constant rate, to give a blood concentration of 5 ng/ml, into the in vitro circuit just prior to the charcoal column. Rises in Swank screen filtration pressure were prevented, and it was also shown that the giving of a bolus dose of Prostacyclin (400 ng/ml blood concentration) reversed rises in Swank screen filtration pressure that have already occurred. On the basis of these findings, Prostacyclin would be expected to be of considerable clinical value in the prevention of adverse platelet reactions to charcoal haemoperfusion when carried out in patients with fulminant hepatic failure.

The use of prostacyclin, PGI2, in the prevention of platelet aggregation during charcoal haemoperfusion.

We have developed an in vitro haemoperfusion circuit, which closely simulates the conditions encountered in a clinical haemoperfusion, for testing the blood compatibility of new adsorbent materials. Fresh heparinised human blood is perfused for 2 hours over two model columns arranged in parallel closed circuits. The present study was performed to investigate the blood compatibility of human serum albumin-coated Amberlite XAD-7 resin. Mean platelet losses over 2 hours were greater following perfusion of the uncoated resin (44 +/- SE 5.6%) than the albumin-coated resin (17 +/- SE 2.2%) (p less than 0.01). Mean white cell losses were similar (55%) for both resins. No rises in Swank screen filtration pressure, which detects the presence of cellular aggregates in blood, were observed. Coating of XAD-7 resin with HSA therefore improves its blood compatibility with respect to platelet losses and we thus intend to use this preparation in a clinical trial of resin haemoperfusion in fulminant hepatic failure.

H. Y. Ton

Papers

Albumin-coated Amberlite XAD-7 resin for hemoperfusion in acute liver failure. Part I: adsorption studies.

Albumin-coated Amberlite XAD-7 resin for hemoperfusion in acute liver failure. Part II: in vivo evaluation.

Adsorption of human serum albumin to amberlite XAD‐7 resin

The use of prostacyclin, PGI2, in the prevention of platelet aggregation during charcoal haemoperfusion.

The use of an in vitro haemoperfusion circuit to evaluate the blood compatibility of albumin-coated Amberlite XAD-7 resin.