Gerhard Rakhorst

Improved kidney graft function after preservation using a novel hypothermic machine perfusion device

Objective:To study graft function and ischemia/reperfusion injury of porcine kidneys after preservation with the new Groningen Machine Perfusion (GMP) system versus static cold storage (CS). Introduction:The increasing proportion of marginal and nonheart beating donors necessitates better preservation methods to maintain adequate graft viability. Hypothermic machine preservation (HMP) is a promising alternative to static CS. We have therefore developed and tested an HMP device, which is portable and actively oxygenates the perfusate via an oxygenator. The aim of the present study was to examine the efficacy of the GMP system in a transplantation experiment. Materials and Methods:In a porcine autotransplantation model, kidneys were retrieved and either cold stored in University of Wisconsin CS for 20 hours at 4°C or subjected to HMP using University of Wisconsin machine perfusion at 4°C with 2 different pressure settings: 30/20 mm Hg or 60/40 mm Hg. Results:HMP at 30/20 mm Hg was found to better preserve the viability of kidneys reflected by improved cortical microcirculation, less damage to the proximal tubule, less damage mediated by reactive oxygen species, less proinflammatory cytokine expression, and better functional recovery after transplantation. However, high perfusion pressures (60/40 mm Hg) resulted in higher expression of von Willebrand factor and monocyte chemotactic peptide-1 in postpreservation biopsies and subsequent graft thrombosis in 2 kidneys. Conclusions:It is concluded that the GMP system improves kidney graft viability and perfusion pressures are critically important for outcome.

The value of machine perfusion perfusate biomarkers for predicting kidney transplant outcome

Background. Retrospective evidence suggests that lactate dehydrogenase, aspartate aminotransferase, total glutathione-S-transferase (GST), alanine-aminopeptidase, N-acetyl-&bgr;-d-glucosaminidase (NAG), and heart-type fatty acid binding protein (H-FABP) measured during kidney machine perfusion (MP) could have predictive value for posttransplant outcome. However, these data may be biased due to organ discard based on biomarker measurements, and previous analyses were not adjusted for likely confounding factors. No reliable prospective evidence has been available so far. Nevertheless, some centers already use these biomarkers to aid decisions on accepting or discarding a donor kidney. Methods. From 306 deceased-donor kidneys donated after brain death or controlled cardiac death and included in an international randomized controlled trial, these six biomarkers were measured in the MP perfusate. In this unselected prospective data set, we tested whether concentrations were associated with delayed graft function, primary nonfunction, and graft survival. Multivariate regression models investigated whether the biomarkers remained independent predictors when adjusted for relevant confounding factors. Results. GST, NAG, and H-FABP were independent predictors of delayed graft function but not of primary nonfunction and graft survival. Lactate dehydrogenase, aspartate aminotransferase, and alanine-aminopeptidase had no independent prognostic potential for any of the endpoints. Perfusate biomarker concentrations had no relevant correlation with cold ischemic time or renal vascular resistance on the pump. Conclusions. Increased GST, NAG, or H-FABP concentrations during MP are an indication to adjust posttransplant recipient management. However, this study shows for the first time that perfusate biomarker measurements should not lead to kidney discard.

Normothermic ex vivo lung perfusion of non-heart-beating donor lungs in pigs: from pretransplant function analysis towards a 6-h machine preservation

Donor shortage urges optimal use of all lungs available. Ex vivo lung perfusion (EVLP) is a method to evaluate lung function before implantation. EVLP was performed in pigs to evaluate lung function, using two different clinical non‐heart‐beating (NHS) donor protocols: flush perfusion and topical cooling after 1‐h warm ischaemia (n = 5 each). Secondly, we investigated whether EVLP can be used for 6 h ex vivo machine preservation (n = 4). In comparison with topical cooling, flush perfusion preserved lung function better during EVLP. During 6 h normothermic EVLP, gas exchange remained stable; however, the pulmonary artery pressure and ventilation pressure showed a significant increase. EVLP is a reliable method for evaluation of lung graft function. Flush perfusion with Perfadex is preferred above topical cooling in NHB lung donation. Six‐hour normothermic EVLP is feasible but should be further improved to make ex vivo machine preservation or treatment of lung grafts successful.

Hypothermic machine preservation in liver transplantation revisited: concepts and criteria in the new millennium.

To overcome the present shortage of liver donors by expansion of the existing donor pool and possibly lengthening of the storage time, hypothermic machine perfusion of the liver as a dynamic preservation method is revisited. The three most important aspects are defined to be the type of preservation solution, the characteristics of perfusion dynamics, and the oxygen supply. Reviewing hypothermic liver machine perfusion experiments, the University of Wisconsin machine preservation solution is the solution most used. It is also found that nothing conclusive can be said about the optimal perfusion characteristics, since either perfusion pressure or perfusion flow is reported. The best estimation is perfusion of the liver in a physiological manner, i.e. pulsatile arterial perfusion and continuous portal venous perfusion. The applied pressures could be chosen to be somewhat lower than physiological pressures to prevent possible endothelial cell damage. Oxygen supply is necessary to achieve optimal preservation of the liver. The minimal amount of partial oxygen pressure required is inversely related to the normalized flow. Incorporating these features in a system based on existing standard surgical and organ sharing procedures and which is able to work stand-alone for 24 h, weighing less than 23 kg, could successfully implement this technique into every day clinical practise.

Animal models for tracheal research.

Tracheal research covers two main areas of interest: tracheal reconstruction and tracheal fixation. Tracheal reconstructions are aimed at rearranging or replacing parts of the tracheal tissue using implantation and transplantation techniques. The indications for tracheal reconstruction are numerous: obstructing tracheal tumors, trauma, post-intubation tissue reactions, etc. Although in the past years much progress has been made, none of the new developed techniques have resulted in clinical application at large scale. Tissue engineering is believed to be the technique to provide a solution for reconstruction of tracheal defects. Although developing functional tracheal tissue from different cultured cell types is still a challenge. Tracheal fixation research is relatively new in the field and concentrates on solving fixation-related problems for laryngectomized patients. In prosthetic voice rehabilitation tracheo-esophageal silicon rubber speech valves and tracheostoma valves are used. This is often accompanied by many complications. The animal models used for tracheal research vary widely and in most publications proper scientific arguments for animal selection are never mentioned. It showed that the choice on animal models is a multi-factorial process in which non-scientific arguments tend to play a key role. The aim of this study is to provide biomaterials scientists with information about tracheal research and the animal models used.

Is red blood cell rheology preserved during routine blood bank storage

BACKGROUND: Red blood cell (RBC) units stored for more than 2 weeks at 4°C are currently considered of impaired quality. This opinion has primarily been based on altered RBC rheologic properties (i.e., enhanced aggregability, reduced deformability, and elevated endothelial cell interaction), during prolonged storage of nonleukoreduced RBC units. In this study, the rheologic properties and cell variables of leukoreduced RBC units, during routine blood bank storage in saline‐adenine‐glucose‐mannitol, were investigated.

Abnormal endothelium-dependent microvascular reactivity in recently preeclamptic women

OBJECTIVE: To assess endothelial function at the level of skin microvasculature, using iontophoretic administration of acetylcholine (endothelium-dependent vasodilator) and sodium nitroprusside (endothelium-independent vasodilator), in women who recently had a preeclamptic pregnancy. METHODS: Microvascular skin reactivity was assessed by laser Doppler perfusion monitoring and iontophoresis of acetylcholine (ACh) and sodium nitroprusside (SNP) in 25 women with a history of early onset preeclampsia and 23 women with previous uncomplicated pregnancies, all of whom were between 3 and 11 months postpartum. RESULTS: Mean (± standard error of the mean) ACh-mediated vasodilatation, expressed as a percentage increase in flux, was higher in women who recently had a preeclampsia than in controls (535 ± 46% versus 314 ± 29%, P < .001). In contrast, SNP-mediated vasodilatation was not significantly different (560 ± 71% versus 483 ± 69%, P = .4) in both groups. Linear regression analysis revealed that the difference in ACh-mediated vasodilatation was explained by preeclampsia (P = .004), whereas vascular risk factors such as maternal age, diastolic blood pressure, and family history of premature cardiovascular diseases had no significant effect. CONCLUSION: The increased ACh-mediated vasodilatation in the microcirculation of recently preeclamptic women indicates abnormal endothelial function. Furthermore, it may represent a compensatory response to an impaired vasodilatory response of the macrocirculation, thereby supporting the hypothesis of an underlying (micro)angiopathy. LEVEL OF EVIDENCE: II-2

Early events in kidney donation: progression of endothelial activation, oxidative stress and tubular injury after brain death.

Cerebral injury leading to brain death (BD) causes major physiologic derangements in potential organ donors, which may result in vascular‐endothelial activation and affect posttransplant graft function. We investigated the kinetic of pro‐coagulatory and pro‐inflammatory endothelial activation and the subsequent oxidative stress and renal tubular injury, early after BD declaration. BD was induced by slowly inflating a balloon‐catheter inserted in the extradural space over a period of 30 min. Rats (n = 30) were sacrificed 0.5, 1, 2 or 4 h after BD‐induction and compared with sham‐controls. This study demonstrates immediate pro‐coagulatory and pro‐inflammatory activation of vascular endothelium after BD in kidney donor rats, proportional with the duration of BD. E‐ and P‐Selectins, Aα/Bβ‐fibrinogen mRNA were abruptly and progressively up‐regulated from 0.5 h BD onwards; P‐Selectin membrane protein expression was increased; fibrinogen was primarily visualized in the peritubular capillaries. Plasma von Willebrand factor was significantly higher after 2 h and 4 h BD. Urine heart‐fatty‐acid‐binding‐protein and N‐acetyl‐glucosaminidase, used as new specific and sensitive markers of proximal and distal tubular damage, were found significantly increased after 0.5 h, with a maximum at 4 h. Unexpectedly, oxidative stress was detectable only late, after the installation of tubular injury, suggesting only a secondary role for hypoxia in triggering these injuries.

Medical technology assessment: The use of the analytic hierarchy process as a tool for multidisciplinary evaluation of medical devices

Most types of medical technology assessment are performed only after the technology has been developed. Consequently, they have only minor effects on changes in clinical practice. Our study introduces a new method of constructive medical technology assessment that can change the development and diffusion of a medical device to improve its later clinical effectiveness. The method, based on Saatys Analytic Hierarchy Process, quantitatively supports discussions between various parties involved in technological development and diffusion. We applied this method in comparing a new blood pump with two competitors based on technical, medical and social requirements. These discussions changed the evaluators’ perspectives, reduced diasagreements, and ended in a reliable evaluation of the pumps performance. On the basis of these results, adaptations were derived which improved the design and diffusion of the blood pump. This application shows the adequate potential of our method to steer technological development and diffusion of artificial organs.

Advanced glycation end-products and skin autofluorescence in end-stage renal disease: a review

Abstract Chronic kidney disease (CKD), especially in its end stage, is marked by extremely high cardiovascular rates of morbidity and mortality; hemodialysis patients have a five-fold shorter life expectancy than healthy subjects of the same age. In CKD the metabolic products that accumulate in the body are so-called uremic toxins. These include advanced glycation end-products (AGE). AGE levels are markedly increased in CKD patients not only because of impaired excretion but also because of increased production. AGE formation has initially been described as a non-enzymatic reaction between proteins and glucose in the so-called Maillard reaction, but they are also more rapidly formed during oxidative stress and subsequent formation of reactive carbonyl compounds like (methyl)glyoxal. AGE accumulate in tissue where they cross-link with proteins, e.g., collagen, inducing tissue stiffening of blood vessels and skin. They may also interact with receptor of AGE (RAGE) and other receptors, which lead to activation of intracellular transduction mechanisms resulting in cytokine release and further tissue damage in CKD. The accumulation of AGE in the skin can be measured non-invasively using autofluorescence. The skin autofluorescence is a strong marker of cardiovascular mortality in CKD. The focus of this review is on the role of tissue and plasma AGE, and of skin autofluorescence as a proxy of tissue AGE accumulation, in the increase in cardiovascular disease in end stage renal disease (ESRD). This review will also present the possibility of reducing the AGE accumulation in ESRD patients using the following five methods: 1) use of low AGE peritoneal dialysis solutions; 2) use of advanced hemodialysis techniques; 3) use of AGE reducing drugs; 4) optimizing the nutrition of hemodialysis patients; and 5) renal transplantation.