Cold-to-warm machine perfusion of the liver: a novel circuit for an uninterrupted combined perfusion protocol

International audienceBackground: Ex-vivo perfusion of liver grafts is associated with promising results for the preservation of marginal grafts. Recent studies highlight the need for a combination of perfusion conditions, such as hypothermic followed by normothermic perfusion. While comprehensive machines dedicated to liver perfusion have been developed, these systems remain costly and poorly adaptable to perfusion condition switch, which requires a complete interruption of the perfusion process. Our team aimed at developing an adaptable and simple circuit for uninterrupted ex-vivo liver perfusion.Methods: Together with specialized bioengineers, we developed a highly adaptable circuit that can fit on already pre-existing extracorporeal oxygenation machines routinely used in cardiovascular surgery. This circuit, owing to its reservoir, allows any type of perfusion conditions without interrupting the perfusion process.Results: In a preliminary study, to assess the technical feasibility of liver perfusion using our circuit under different conditions, we performed 7 perfusions of discarded liver grafts. HOPE and DHOPE hypothermic perfusion could be performed, and a switch to normothermia was easily possible within seconds. From there, a dynamic perfusion sequence model was developed.Conclusion: This circuit may represent a simpler alternative or a new refinement to existing perfusion systems allowing uninterrupted combined perfusion protocols

Rapid or slow time to brain death? Impact on kidney graft injuries in an allotransplantation porcine model

International audienceThe use of donors deceased after brain death (DBD) with extended criteria in response to the shortage of grafts leads to the removal of more fragile kidneys. These grafts are at greater risk of not being grafted or delayed function. A better knowledge of the pathophysiology of DBDs would improve this situation. There is a difference between the results from animal models of DBD and the clinical data potentially explained by the kinetics of brain death induction. We compared the effect of the induction rate of brain death on the recovery of post-transplant renal function in a pig model of DBD followed by allografts in nephrectomized pigs. Resumption of early function post-transplant was better in the rapidly generated brain death group (RgBD) and graft fibrosis at three months less important. Two groups had identical oxidative stress intensity but a greater response to this oxidative stress by SIRT1, PGC1-α and NRF2 in the RgBD group. Modulation of mechanistic target of rapamycin (mTOR) stimulation by NRF2 would also regulate the survival/apoptosis balance of renal cells. For the first time we have shown that an allostatic response to oxidative stress can explain the impact of the rapidity of brain death induction on the quality of kidney transplants

Individual and Combined Impact of Oxygen and Oxygen Transporter Supplementation during Kidney Machine Preservation in a Porcine Preclinical Kidney Transplantation Model

Marginal kidney graft preservation in machine perfusion (MP) is well-established. However, this method requires improvement in order to mitigate oxidative stress during ischemia-reperfusion, by using oxygenation or an O2 carrier with anti-oxidant capacities (hemoglobin of the marine worm; M101). In our preclinical porcine (pig related) model, kidneys were submitted to 1h-warm ischemia, followed by 23 h hypothermic preservation in Waves® MP before auto-transplantation. Four groups were studied: W (MP without 100%-O2), W-O2 (MP with 100%-O2; also called hyperoxia), W-M101 (MP without 100%-O2 + M101 2 g/L), W-O2 + M101 (MP with 100%-O2 + M101 2 g/L) (n = 6/group). Results: Kidneys preserved in the W-M101 group showed lower resistance, compared to our W group. During the first week post-transplantation, W-O2 and W-M101 groups showed a lower blood creatinine and better glomerular filtration rate. KIM-1 and IL-18 blood levels were lower in the W-M101 group, while blood levels of AST and NGAL were lower in groups with 100% O2. Three months after transplantation, fractional excretion of sodium and the proteinuria/creatinuria ratio remained higher in the W group, creatininemia was lower in the W-M101 group, and kidney fibrosis was lower in M101 groups. We concluded that supplementation with M101 associated with or without 100% O2 improved the Waves® MP effect upon kidney recovery and late graft outcome

Inhibition of complement improves graft outcome in a pig model of kidney autotransplantation

International audienceBackground: Ischemia reperfusion injury (IRI) induced immune response is a critical issue in transplantation. Complement and contact system activation are among its key mechanisms. Study design: We investigated the benefits of pre-reperfusion treatment with recombinant human C1INH (rhC1INH), inhibitor of both complement and contact activation, in a pig model of kidney autotransplantation, subjecting the organ to 60 min warm ischemia prior to 24 h static preservation to maximize damage. Results: Serum creatinine measurement showed that treated animals recovered glomerular function quicker than the Vehicle group. However, no difference was observed in tubular function recovery, and elevated level of urinary NGal (Neutrophil gelatinase-associated lipocalin) and plasma AST (Aspartate Aminotransferase) were detected, indicating that treatment did not influence IRI-mediated tubular cell necrosis. Regarding chronic graft outcome, rhC1INH significantly prevented fibrosis development and improved function. Immunohistochemistry and western blot showed decreased invasion by macrophages and T lymphocytes, and reduction of epithelial to mesenchymal transition. We determined the effect of treatment on complement activation with immunofluorescence analyses at 30 min post reperfusion, showing an inhibition of C4d deposition and MBL staining in treated animals. Conclusions: In this model, the inhibition of complement activation by rhC1INH at reperfusion, while not completely counteracting IRI, limited immune system activation, significantly improving graft outcome on the short and long term

Defining the optimal duration for normothermic regional perfusion in the kidney donor: A porcine preclinical study

International audienceKidneys from donation after circulatory death (DCD) are highly sensitive to ischemia‐reperfusion injury and thus require careful reconditioning, such as normothermic regional perfusion (NRP). However, the optimal NRP protocol remains to be characterized. NRP was modeled in a DCD porcine model (30 minutes of cardiac arrest) for 2, 4, or 6 hours compared to a control group (No‐NRP); kidneys were machine‐preserved and allotransplanted. NRP appeared to permit recovery from warm ischemia, possibly due to an increased expression of HIF1α‐dependent survival pathway. At 2 hours, blood levels of ischemic injury biomarkers increased: creatinine, lactate/pyruvate ratio, LDH, AST, NGAL, KIM‐1, CD40 ligand, and soluble‐tissue‐factor. All these markers then decreased with time; however, AST, NGAL, and KIM‐1 increased again at 6 hours. Hemoglobin and platelets decreased at 6 hours, after which the procedure became difficult to maintain. Regarding inflammation, active tissue‐factor, cleaved PAR‐2 and MCP‐1 increased by 4‐6 hours, but not TNF‐α and iNOS. Compared to No‐NRP, NRP kidneys showed lower resistance during hypothermic machine perfusion (HMP), likely associated with pe‐NRP eNOS activation. Kidneys transplanted after 4 and 6 hours of NRP showed better function and outcome, compared to No‐NRP. In conclusion, our results confirm the mechanistic benefits of NRP and highlight 4 hours as its optimal duration, after which injury markers appear

Preventing acute kidney injury during transplantation: the application of novel oxygen carriers

International audienceIntroduction: Delayed graft function (DGF) has a significant impact on kidney transplantation outcome. One of the underlying pivotal mechanisms is organ preservation and associated hypothermia and biochemical alteration. Area covered: This paper focuses on organ preservation and its clinical consequences and describes 1. A comprehensive presentation of the pathophysiological mechanism involved in delayed graft function development; 2. The impact on endothelial cells and microvasculature integrity and the consequences on transplanted organ outcome; 3. The reassessment of dynamic organ preservation motivated by the growing use of extended criteria donors and the interest in the potential of normothermia; 4. The role of oxygenation during dynamic preservation; and 5. Novel oxygen carriers and their proof of concept in transplantation, among which M101 (HEMO(2)life (R)) is currently the most extensively investigated. Expert opinion: Metabolic disturbances and imbalance of oxygen supply during preservation highlight the importance of providing oxygen. Normothermia, permitted by recent advances in machine perfusion technology, appears to be the leading edge of preservation technology. Several oxygen transporters are compatible with normothermia; however, only M101 also demonstrates compatibility with standard hypothermic preservation

MOESM1 of Inhibition of complement improves graft outcome in a pig model of kidney autotransplantation

Additional file 1: Table 1. Power calculations for the different statistical analyses. Calculations were performed using Anastat ( http://www.anastats.fr/ ) with an alpha threashold of 0.05. Table 2. Immune cell counts

MOESM2 of Inhibition of complement improves graft outcome in a pig model of kidney autotransplantation

Additional file 2: Figure 1. Early outcome: tubular function. Evolution of the fraction of excreted sodium in the urine in the first week post-transplant. Serial blood and urine samples were collected from transplanted pigs and processed for biochemical analysis. Shown are means¹SD, n=7