Uwe Gottmann

Effects of Donor Pretreatment With Dopamine on Graft Function After Kidney Transplantation A Randomized Controlled Trial

CONTEXT Kidney graft function after transplantation can be improved through pharmacological donor pretreatment to limit organ injury from cold preservation. OBJECTIVE To determine whether pretreatment of brain-dead donors with low-dose dopamine improves early graft function in human renal transplant recipients. DESIGN, SETTING, AND PATIENTS Randomized, open-label, multicenter, parallel-group trial of 264 deceased heart-beating donors and 487 subsequent renal transplants performed at 60 European centers between March 2004 and August 2007 (final follow-up, December 31, 2008). Eligible donors were stable under low-dose norepinephrine with a normal serum creatinine concentration on admission. INTERVENTIONS Donors were randomized to receive low-dose dopamine (4 mug/kg/min). MAIN OUTCOME MEASURES Dialysis requirement during first week after transplantation. RESULTS Dopamine was infused for a median of 344 minutes (IQR, 215 minutes). Dialysis was significantly reduced in recipients of a dopamine-treated graft. Fewer recipients in the treatment group needed multiple dialyses (56/227; 24.7%; 95% CI, 19.0%-30.3%; vs 92/260; 35.4%; 95% CI, 29.5%-41.2%; P = .01). The need for multiple dialyses posttransplant was associated with allograft failure after 3 years (HR, 3.61; 95% CI, 2.39-5.45; P < .001), whereas a single dialysis was not (HR, 0.67; 95% CI, 0.21-2.18; P = .51). Besides donor dopamine (OR, 0.54; 95% CI, 0.35-0.83; P = .005), cold ischemic time (OR, 1.07; 95% CI, 1.02-1.11 per hour; P = .001), donor age (OR, 1.03; 95% CI, 1.01-1.05 per year; P < .001), and recipient body weight (OR, 1.02; 95% CI, 1.01-1.04 per kg; P = .009) were independent explanatory variables in a multiple logistic regression model. Dopamine resulted in significant but clinically meaningless increases in the donors systolic blood pressure (3.8 mm Hg; 95% CI, 0.7-6.9 mm Hg; P = .02) and urine production before surgical recovery of the kidneys (29 mL; 95% CI, 7-51 mL; P = .009) but had no influence on outcome. CONCLUSION Donor pretreatment with low-dose dopamine reduces the need for dialysis after kidney transplantation. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00115115.

Prevention of Cold‐Preservation Injury of Cultured Endothelial Cells by Catecholamines and Related Compounds

The present study was conducted to dissect the underlying mechanisms by which catecholamines protect cells against preservation injury.

Hypothermic injury: the mitochondrial calcium, ATP and ROS love-hate triangle out of balance.

Background/Aims: Catecholamines prevent hypothermic cell death which accounts for severe tissue damage and impaired allograft function after prolonged organ preservation. Here, we identified cellular processes which govern hypothermia-mediated cell death in endothelial cells and how they are influenced by dopamine. Methods: Lactate dehydrogenase assay, intracellular ATP, reactive oxygen species and reduced thio-group measurement, intracellular calcium measurement and mitochondrial calcium staining were performed in the study. Results: Intracellular ATP was almost completely depleted within 12 hrs of hypothermic preservation in untreated human umbilical vein endothelial cells (HUVEC), while dopamine pre-treatment significantly delayed ATP depletion. 4 hrs after hypothermia a redox imbalance was observed in untreated cells, which increased with the duration of hypothermia. The redox imbalance was primarily caused by depletion of SH reduction equivalents and was significantly inhibited by dopamine. In addition, hypothermia-induced Ca2+ influx and mitochondrial Ca2+ accumulation were both prevented by dopamine. The protective effect of dopamine was abrogated by ionomycin and sodium azide and partly by oligomycin and CCCP. Conclusions: Our data demonstrated that loss of intracellular ATP, generation of a redox imbalance and accumulation of intracellular Ca2+ underlie cold preservation injury. Dopamine improves the redox balance, prevents intracellular Ca2+ accumulation and delays ATP depletion.

Effects of Donor Pre-Treatment With Dopamine on Survival After Heart Transplantation A Cohort Study of Heart Transplant Recipients Nested in a Randomized Controlled Multicenter Trial

OBJECTIVES We determined the outcome of cardiac allografts from multiorgan donors enrolled in a randomized trial of donor pre-treatment with dopamine. BACKGROUND Treatment of the brain-dead donor with low-dose dopamine improves immediate graft function after kidney transplantation. METHODS A cohort study of 93 heart transplants from 21 European centers was undertaken between March 2004 and August 2007. We assessed post-transplant left ventricular function (LVF), requirement of a left ventricular assist device (LVAD) or biventricular assist device (BVAD), need for hemofiltration, acute rejection, and survival of recipients of a dopamine-treated versus untreated graft. RESULTS Donor dopamine was associated with improved survival 3 years after transplantation (87.0% vs. 67.8%, p = 0.03). Fewer recipients of a pre-treated graft required hemofiltration after transplant (21.7% vs. 40.4%, p = 0.05). Impaired LVF (15.2% vs. 21.3%, p = 0.59), requirement of a LVAD (4.4% vs. 10.6%, p = 0.44), and biopsy-proven acute rejection (19.6% vs. 14.9%, p = 0.59) were not statistically different between groups. Post-transplant impaired LVF (hazard ratio [HR]: 4.95; 95% confidence interval [CI]: 2.08 to 11.79; p < 0.001), requirement of LVAD (HR: 6.65; 95% CI: 2.40 to 18.45; p < 0.001), and hemofiltration (HR: 2.83; 95% CI: 1.20 to 6.69; p = 0.02) were predictive of death. The survival benefit remained (HR: 0.33; 95% CI: 0.12 to 0.89; p = 0.03) after adjustment for various risks affecting mortality, including pre-transplant LVAD/BVAD, inotropic support, and impaired kidney function. CONCLUSIONS Treatment of brain-dead donors with dopamine of 4 μg/kg/min will not harm cardiac allografts but appears to improve the clinical course of the heart allograft recipient. (Prospective Randomized Trial to Evaluate the Efficacy of Donor Preconditioning With Dopamine on Initial Graft Function After Kidney Transplantation; NCT00115115).

Dopamine treatment in brain-dead rats mediates anti-inflammatory effects: the role of hemodynamic stabilization and D-receptor stimulation

Brain death (BD) is associated with profound inflammation in end‐organs. Dopamine (DA) treatment reduces this inflammatory response, but the underlying mechanisms remain thus far largely unknown. In this study, we investigated if the anti‐inflammatory effect of DA was related to hemodynamic stabilization and by which receptors it was mediated. BD was induced in F344 donor rats. DA was given either before BD for 24 h or after BD induction during a definite time. Adrenergic or D‐receptor blockers were administered to inhibit the receptor stimulation mediated by DA. Hemodynamic changes were recorded and kidneys were harvested after 6 h of BD. Mean arterial pressure was completely normalized by DA treatment. DA pretreatment before BD induction and treatment during BD both significantly inhibited the monocyte infiltration. The anti‐inflammatory as well as its blood pressure stabilizing effect was abrogated by concomitant application of adrenergic receptor blockers. In contrast, concomitant application of D‐receptor blockers only abrogated the anti‐inflammatory effect, but did not affect blood pressure stabilization. In contrast, pergolide and adrenergic receptor blockers completely normalized the blood pressure, but did not affect renal inflammation. Hence, DA might reduce BD‐induced monocyte infiltration possibly by hemodynamic stabilization, D‐receptor activation, or a combination of both.

Atorvastatin donor pretreatment prevents ischemia/ reperfusion injury in renal transplantation in rats : Possible role for aldose-reductase inhibition

Background. The aim of the present study was to evaluate the effect of donor pretreatment with atorvastatin on ischemia/reperfusion (I/R) injury in renal transplantation in rats. Methods. Donor rats were pretreated orally with atorvastatin or vehicle 2 days prior to explantation. Kidneys were stored for 24 hr at 4°C in University of Wisconsin solution and transplanted into isogeneic or allogeneic recipients. Results. Donor treatment with atorvastatin improved initial graft function, reduced renal inflammation, and the number of TUNEL-positive cells in renal tissue after prolonged cold storage and isogeneic transplantation. In the allogeneic transplantation model, donor treatment with atorvastatin reduced renal inflammation in grafts harvested after 5 days, but no improvement of long-term graft survival (24 weeks) could be observed. A genome wide gene expression profile of donor kidneys from atorvastatin treated or vehicle treated rats revealed a fivefold downregulation of aldose reductase in all atorvastatin treated animals (P<0.01). Donor treatment with an aldose-reductase inhibitor improved kidney function and reduced renal inflammation after prolonged cold storage and isogeneic transplantation. Conclusion. Our data suggest that downregulation of aldose reductase in renal tissue might underlie the protective effect of donor atorvastatin treatment. Donor pretreatment with a statin or an aldose reductase inhibitor could offer a new treatment strategy to prevent transplantation associated tissue injury.

Hypothermia-induced loss of endothelial barrier function is restored after dopamine pretreatment: role of p42/p44 activation.

Background. Donor dopamine usage is associated with improved immediate graft function after renal transplantation. Although prolonged cold preservation results in an increased vascular permeability, the present study was conducted to examine in vitro and in vivo if dopamine pretreatment influences endothelial barrier function under such conditions. Methods. To assess cold preservation injury in vitro and in vivo, cultured human umbilical vein endothelial cells (HUVEC) and Lewis donor rats were pretreated with dopamine or isotonic saline prior to cold storage. Injury was determined by lactate dehydrogenase (LDH) release, histology, and functional analysis. Results. In vitro cold storage resulted in intercellular gap formation in both untreated and dopamine pretreated HUVEC. In the latter monolayer integrity was completely restored upon rewarming and paracellular transport of fluorescein isothiocyanate-dextran was significantly reduced. In dopamine treated HUVEC, intercellular gap formation was independent of cell death and was associated with redistribution of junctional proteins and condensation of cytoskeleton proteins. In untreated HUVEC proteolysis and cell death were clearly evident after hypothermia. Closing of intercellular gaps was dependent on p42/p44 activation. Regeneration of adenosine triphosphate was only observed in dopamine pretreated cells. Only in dopamine treated Lewis renal allografts subjected to cold storage, activation of p42/p44 occurred upon rewarming. These grafts had a better renal function and displayed less inflammatory cells five days after transplantation. Conclusion. Our study demonstrates beneficial effects of dopamine treatment on cold storage induced endothelial barrier disturbances. This may contribute to the positive effects of catecholamines on immediate graft function of renal allografts in men.

Donor dopamine treatment in brain dead rats is associated with an improvement in renal function early after transplantation and a reduction in renal inflammation

Brain death (BD) is associated with tissue inflammation. As dopamine treatment of BD donor rats reduces renal monocyte infiltration, we tested if this treatment affects renal function and inflammation in recipients. BD was induced in F344 rats and was maintained for 6 h in all experiments. Dopamine was given for 6 (DA6) or 3 h (DA3) from the onset of BD. Ventilated non‐BD (NBD) and BD animals served as controls. Kidneys were transplanted into bilaterally nephrectomized Lewis recipients. Serum creatinine (s‐crea) was measured and leukocyte infiltration was assessed 10 days after transplantation. One day after transplantation, s‐crea was significantly reduced in recipients who received a renal allograft from dopamine treated BD or from NBD rats compared to BD vehicle (P < 0.05). Ten days after transplantation, the number of infiltrating monocytes was significantly lower in grafts obtained from dopamine treated and from NBD rats (P < 0.05). A reduced infiltration in these grafts was confirmed by Banff 97 classification. Cytokine‐induced neutrophil‐chemoattractant 1 and interleukin (IL)‐6 mRNA expression were reduced in DA rats compared to BD controls. No difference for macrophage chemoattractant protein 1 and IL‐10 were found. These findings may explain the salutary effect of donor dopamine treatment in renal transplantation.

Influence of hypersulfated and low molecular weight heparins on ischemia/reperfusion: injury and allograft rejection in rat kidneys

The aim of the study was to evaluate the effect of the hypersulfated nonanticoagulant heparin derivative LU 51198 (LU) and of the low molecular weight heparin reviparin (REVI) on ischemia/reperfusion (I/R) injury, acute rejection (AR) and chronic allograft nephropathy (CAN) in rats. Organs were harvested 5 days after 60 min of renal I/R injury. For investigation of AR and CAN we used the allogeneic Fisher–Lewis model. Kidneys were harvested at one respectively 32 weeks after transplantation. Rats were treated with either vehicle, LU or REVI. After I/R injury, treatment with REVI or LU reduced infiltration with MHC II and R73‐positive cells, whereas only REVI reduced ED1‐positive cells and expression of monocyte chemoattractant protein‐1. There was no effect of REVI and LU on acute allograft rejection. Treatment with LU or REVI reduced glomerular infiltration with ED1 and MHCII‐positive cells and renal expression of transforming growth factor‐beta 32 weeks after transplantation. Only REVI treatment reduced albuminuria, interstitial infiltration and histological signs of CAN. LU, and in a more potent manner REVI, reduce signs of CAN and renal inflammation after I/R injury. Chemically modified heparins without anticoagulatory effects may offer a new treatment option in preventing I/R injury and CAN in human kidney transplantation.

Atorvastatin interferes with activation of human CD4+ T cells via inhibition of small guanosine triphosphatase (GTPase) activity and caspase‐independent apoptosis

Although a beneficial effect of hydroxy‐methylglutaryl coenzyme A (HMG‐CoA) reductase inhibitors, i.e. statins, on cell‐mediated immunity has been suggested in vivo and in vitro, little is known about the molecular and biochemical events by which statins inhibit T cell proliferation. To address this question, we investigated the effects of atorvastatin (AT) on intracellular cytokine production, T cell activation markers, cell cycle progression and apoptosis in human CD4+ T cells. AT did not influence intracellular cytokine production after short‐term stimulation of whole blood with phorbol myristate acetate (PMA)/ionomycin or superantigen (SEB). In contrast, AT influenced CD45RA to RO switching dose‐dependently, as well as CD25 expression, and caused cell cycle arrest in the G1 phase after long‐term T cell stimulation. This occurred in conjunction with a reduced expression of cyclin‐dependent kinases 2 and 4 and p21wav1/cip1 and was paralleled by an increased protein expression of p27kip1. In addition to G1 arrest, increased apoptosis was observed in AT‐treated cells. In line with this, the expression of Bcl‐xl and pBad were decreased by AT. Apoptosis was independent of caspases 3 and 9 activation. The inhibitory effect of AT on T cell proliferation could be overcome by addition of mevalonic acid or geranylgeranyl pyrophosphate, but not by farnesyl pyrophosphate or squalen, suggesting reduced protein prenylation. Activation of Rho, Rac and Ras were strongly reduced in AT‐treated T cells, suggesting that impaired geranylation of these molecules might underlie the inhibitory effect of AT on T cell proliferation.