Hip fractures in nonagenarians: perioperative mortality and survival.

The nonagenarian population is increasing yet there is little known about their survival and mortality rates following hip fractures. The aim was to review and evaluate all the nonagenarians who underwent surgery for intra- or extracapsular hip fracture at the Trauma Unit of the Department of Surgery at the Academic Medical Center (AMC) in Amsterdam in the past 20 years. In this retrospective study, 155 nonagenarians sustaining hip fractures between 1982 and 2001 were included and reviewed. In this population there were 128 women (83%) and 27 men (17%). Mortality in women was higher than in men. In men more intracapsular fractures (70%) were diagnosed compared to extracapsular fractures. The opposite, 38% intracapsular and 62% extracapsular fractures were found in women (p=0.002). A trend in higher perioperative mortality for extracapsular fractures and increasing ASA-classification was observed. Mean survival for the whole population, men and women was 756, 544 and 787 days, respectively. This study shows that ASA-II nonagenarians with hip fractures have comparable life expectancy compared to nonagenarians without a hip fracture. Shortening of hospital stay after hip fracture surgery reduces the costs and will benefit the patient.

Subnormothermic preservation maintains viability and function in a porcine hepatocyte culture model simulating bioreactor transport.

Bioartificial liver (BAL) systems have been developed to bridge patients with acute liver failure (ALF) to liver transplantation or liver regeneration. Clinical application of BAL systems is dependent on the supportive quality of cells used and direct availability of the whole system. Reliable transport of BAL systems from the laboratory to remote treatment centers is therefore inevitable. Subsequently, preservation conditions play a crucial role during transport of a BAL, with temperature being one of the most determining factors. In this study, we assessed the effect of subnormothermic preservation on freshly isolated porcine hepatocytes cultured in monolayer under oxygenation. Additionally, the effect of the University of Wisconsin (UW) preservation solution was compared with Williams' E (WE) culture medium at 4 degrees C. The control group was cultured for 3 days at 37 degrees C, whereas the transport groups were cultured at 4 degrees C, 15 degrees C, 21 degrees C, or 28 degrees C for 24 h at day 2. All groups were tested each day for cell damage and hepatic functions. Subnormothermic culture (i.e., 15 degrees C to 28 degrees C) for a period of 24 h did not reduce any hepatic function and did not increase cellular damage. In contrast, culture of hepatocytes in WE medium and preservation in UW solution at 4 degrees C significantly reduced hepatic function. In conclusion, freshly isolated porcine hepatocytes can be preserved for 24 h at subnormothermic temperatures as low as 15 degrees C. Future research will focus on the implementation of the AMC-BAL in an oxygenated culture medium perfusion system for transport between the laboratory and the hospital.

Liver support therapy: an overview of the AMC-bioartificial liver research.

Acute liver failure (ALF) is a disease with a mortality of 60-90% depending on the cause. Only high-urgency liver transplantation is able to increase survival compared to standard intensive care therapy. Liver transplantation is hampered by the increasing shortage of organ donors, resulting in a high incidence of patients with ALF dying on the transplantation waiting list. Amongst a variety of liver assist therapies, bioartificial liver (BAL) therapy is marked as the most promising solution to bridge ALF patients to liver transplantation or to liver regeneration, since several BAL systems showed significant improvement of survival time in experimental animals with irreversible ALF. One of these systems has been developed at the Academic Medical Center in Amsterdam, The Netherlands - the AMC-BAL. This overview describes the development of the AMC-BAL based on porcine hepatocytes which was started 10 years ago. Positive results of in-vitro functionality and in vivo safety and efficacy led to a successful phase I study in 12 ALF patients in Italy. However, xenotransplantation legislation in many European countries prohibits the use of porcine hepatocytes in clinically applied BAL systems. The future of the BAL, therefore, resides in the development of a human-derived hepatocyte cell line as biocomponent of BAL systems.

Assessment and improvement of liver specific function of the AMC-bioartificial liver.

UNLABELLED: The variety of methods for measuring bioactive mass and functionality of bioartificial livers (BAL) is confusing and prevents accurate comparison of reported data. Here we present a comparison of different hepatocyte quantification methods and propose that estimation of cell pellet volume after centrifugation generates a reliable, useful and fast method. In addition a correlation is made between several function tests performed in 26 bioreactors to assess their predictive value. The ammonia eliminating capacity was found to be most predictive for other liver functions, except for lidocaine elimination as a measure of mixed function oxidase activity, which should therefore be determined separately. The oxygen consumption test proved to be an easy and predictive parameter as well. The first generation of our BAL system needed further development to assure optimal treatment of acute liver failure (ALF) patients. Changes in the porcine hepatocyte isolation method and bioreactor loading as well as changes in bioreactor configuration, including use of different materials, resulted in a significantly improved level and maintenance of in vitro BAL function. A fourfold increase in ammonia eliminating capacity, which is only reduced to 75% after seven days of culturing, offers promising prospects for further clinical application. CONCLUSION: The current second generation of our BAL and improvement of hepatocyte isolation and testing protocols have led to a significant increase in the level as well as the maintenance of hepatocyte specific function in our BAL. Finally, consensus on definition of the bioactive mass to be loaded in the bioreactor and insight in the variation and reliability of the functional and metabolic parameters enhances comparison of the different types of bioartificial livers presented in literature.

Large animal models of fulminant hepatic failure in artificial and bioartificial liver support research.

Among the large range of organs involved in the field of tissue engineering (skin, blood vessels, cartilage, etc.) the liver has been given broad attention in the last decade. Liver support systems encompassing artificial and bioartificial systems are applied to treat patients with fulminant hepatic failure (FHF) as a bridge to orthotopic liver transplantation or to liver regeneration. To test safety, technical applicability and therapeutic effect of liver support systems, reliable animal models are needed. Due to the complexity of FHF many diverse attempts have been made to develop an adequate animal model to study liver failure, liver regeneration and liver support systems. In this paper an overview is given of the different models and their advantages and disadvantages are discussed. Suggestions are made for the most suitable large animal model to test liver support systems.

Blood coagulation in anhepatic pigs: effects of treatment with the AMC-bioartificial liver.

The function of a newly devised bioartificial liver (AMC-BAL) based on viable, freshly isolated porcine hepatocytes has been evaluated in anhepatic pigs. The aim of this study was to assess the contribution of BAL treatment on blood coagulation parameters. Pigs were anesthetized and a total hepatectomy was performed (n = 15). The infrahepatic caval vein and the portal vein were connected to the subdiaphragmatic caval vein using a three-way prosthesis. Animals received standard intensive care (control, n= 5), treatment with an empty BAL (device control, n= 5) or with a cell-loaded BAL (BAL-treatment, n= 5) for a period of 24 h starting 24 h after hepatectomy. Coagulation parameters studied concerned prothrombin time (PT), platelet count, the procoagulant system (factors (F)II, FV, FVII, FVIII and fibrinogen), anticoagulant system (AT III), fibrinolytic system (t-PA, PAI-1) as well as markers of coagulation factor activation (TAT complexes, prothrombin fragment F1 + 2). FII, FV, FVII, AT III and fibrinogen rapidly decreased after total hepatectomy in pigs in accordance with the anhepatic state of the animals. FVIII levels were not influenced by the hepatectomy. A mild drop in platelet count was seen in all groups. Treatment of anhepatic pigs with the cell-loaded BAL did not restore PT or clotting factor levels. TAT and F1 + 2 complexes, however, were significantly increased in this group. Levels of t-PA and PAI-1 were not influenced by cell-loaded BAL treatment. Treatment of anhepatic pigs with the AMC-BAL based on freshly isolated porcine hepatocytes does not result in an improved coagulation state due to extensive consumption of clotting factors. However, increased levels of TAT complexes and prothrombin fragments F1 + 2 during treatment of anhepatic pigs indicate synthesis and direct activation of coagulation factors, leading to thrombin generation. This demonstrates that this bioartificial liver is capable of synthesizing coagulation factors.

Phase I clinical trial with the AMC-bioartificial liver.

UNLABELLED: Recently a bio-artificial liver (BAL) system has been developed at the Academic Medical Center (AMC) of Amsterdam to bridge patients with acute liver failure (ALF) to orthotopic liver transplantation (OLT). After successful testing of the AMC-BAL in rodents and pigs with ALF, a phase I study in ALF patients waiting for (OLT) was started in Italy. We present the safety outcome of the first 7 patients aged 21-56 years with coma grade III or IV The total AMC-BAL treatment time ranged from 8 to 35 hours. Three patients received 2 treatments with two different BAL's within three days. Six of the 7 patients were successfully bridged to OLT. One patient showed improved liver function after two treatments and did not need OLT. No severe adverse events of the BAL treatment were noted. CONCLUSION: Treatment of ALF patients with the AMC-BAL is a safe and feasible technique to bridge the waiting time for an adequate liver-graft.

Treatment of acute liver failure in pigs reduces hepatocyte function in a bioartificial liver support system.

Several different types of bioartificial liver (BAL) support systems have been developed to bridge patients suffering from acute liver failure (ALF) to transplantation or liver regeneration. In this study we assessed the effects of ALF plasma on hepatocyte function in the BAL system that has been developed in our center. Pigs (40-60 kg) were anaesthetised and a total hepatectomy was performed. Cells were isolated from the resected livers and were transferred to the bioreactor of the BAL system. Twenty hours after cell isolation, hepatocytes in the BAL were tested for cell viability and functional activity by using a recirculating test medium in which assessment of LDH leakage, ammonia clearance, urea synthesis, 7-ethoxycoumarin O-deethylase (ECOD) activity and pseudocholine esterase production was performed. Subsequently, two groups were studied. In one group (I, n=5), the cell-loaded bioreactor was used to treat the donor pig, rendered anhepatic, for 24 hours. In the second group (II, n=5) the bioreactor was cultured for 24 h and served as a control. After 24 hours treatment or culturing, the cell viability count and functional activity tests were repeated. The results show that hepatocytes in the BAL remained viable after 24 h treatment of anhepatic pigs, as shown by the LDH release and pseudocholine esterase production. However, metabolic functions such as ammonia clearance, ECOD and urea synthesis were reduced after 24 h exposure of hepatocytes to autologous ALF plasma, whereas these functions were unaltered after 24 h culturing of the cells in the bioreactor.

The migratory response to platelet-derived growth factor of smooth muscle cells isolated from synthetic vascular grafts in a canine model.

OBJECTIVE: Previous studies on smooth muscle cells (SMCs) harvested from implanted synthetic grafts demonstrate increased production of platelet-derived growth factor (PDGF) but decreased proliferative response compared with aortic SMCs. The purpose of this study was to determine the migratory response of graft versus aortic SMCs. METHODS: Thoracoabdominal grafts were implanted in beagles. The SMCs were harvested from the graft and infrarenal aorta. Migration was determined with the use of a razor-scrape assay and computerized image analysis. RESULTS: The mean distance migrated and the number of cells that migrated were greater in graft SMCs at baseline (185 +/- 18 micrometer and 108 +/- 17 cells) compared with aortic cells (110 +/- 10 micrometer and 42 +/- 5 cells)(P <.05). Baseline differences persisted after treatment with antibodies to PDGF. The addition of PDGF (10 ng/mL) resulted in increased migration in both graft (229 +/- 23 micrometer and 146 +/- 20 cells) and aortic SMCs (130 +/- 9 micrometer and 70 +/- 5 cells) compared with baseline (P <.05). The relative increase in response to PDGF was similar between the two groups (P = not significant). CONCLUSIONS: Graft SMCs differ phenotypically from aortic SMCs; they exhibit increased basal migration that is independent of autocrine stimulation by PDGF. In contrast to their blunted proliferative response, graft SMCs have a similar migratory response to PDGF compared with aortic SMCs.

The proliferative response to platelet-derived growth factor of smooth muscle cells isolated from synthetic vascular grafts in a canine model.

OBJECTIVE: Previous studies on graft healing have shown increased platelet-derived growth factor (PDGF) production in graft segments versus native aortic segments. The purpose of this study was to characterize the proliferative response of graft smooth muscle cells (SMCs) to PDGF. METHODS: Thoracoabdominal grafts were implanted in beagles. SMCs were harvested from the graft and the proximal and distal aortas. Basal proliferation was assessed with growth curves in primary culture. The proliferative response to PDGF then was compared with [3H]thymidine uptake studies and cell counts. Finally, PDGF receptors were characterized with radio-labeled ligand binding assays. RESULTS: The growth curves showed that the graft SMCs entered log-phase growth 2 days earlier than did the aortic SMCs. Stimulation of quiescent early-passage graft SMCs with PDGF (10 ng/mL) resulted in a 1.7 +/- 0.1-fold increase in [3H]thymidine incorporation, which was significantly less than that of the SMCs from both the proximal aorta (11.8 +/- 3.0) and the distal aorta (10. 2 +/- 1.9; P <.5). Similarly, the 1.1 +/- 0.1-fold increase in graft SMC cell number was significantly less than the increases for both proximal (2.8 +/- 0.5) and distal (2.9 +/- 0.8) aortic SMCs (P <.5). Binding studies on quiescent first-passage cells showed fewer PDGF receptors available for binding in the graft SMCs (185 +/- 70 fmol/million cells) as compared with both the proximal (419 +/- 147 fmol/million cells) and the distal (387 +/- 112 fmol/million cells) aortas (P <.5). Binding affinity was similar for the three groups. CONCLUSION: Graft SMCs exist in a chronic proliferative state but exhibit a decreased proliferative response to PDGF and have fewer receptors available for binding PDGF than do aortic SMCs in vitro.