Acellular Solution Research Articles

The Role of Adipose Tissue Mesenchymal Stem Cells in Colonic Anastomosis Healing in Inflammatory Bowel Disease: Experimental Study in Rats.

(1) Background: A surgical operation on an inflamed bowel is, diachronically, a challenge for the surgeon, especially for patients with inflammatory bowel disease. Adipose tissue-derived mesenchymal stromal cells are already in use in clinical settings for their anti-inflammatory properties. The rationale of the current study was to use AdMSCs in high-risk anastomoses to monitor if they attenuate inflammation and prevent anastomotic leak. (2) Methods: a total of 4 groups of rats were subjected to a surgical transection of the large intestine and primary anastomosis. In two groups, DSS 5% was administered for 7 days prior to the procedure, to induce acute intestinal inflammation. After the anastomosis, 5 × 106 autologous AdMSCs or an acellular solution was injected locally. Macroscopic evaluation, bursting pressure, hydroxyproline, and inflammatory cytokine expression were the parameters measured on the 8th post-operative day. (3) Results: Significantly less intra-abdominal complications, higher bursting pressures, and a decrease in pro-inflammatory markers were found in the groups that received AdMSCs. No difference in VEGF expression was observed on the 8th post-operative day. (4) Conclusions: AdMSCs attenuate inflammation in cases of acutely inflamed anastomosis.

Determination of Optical Properties and Photodynamic Threshold of Lung Tissue for Treatment Planning of In Vivo Lung Perfusion Assisted Photodynamic Therapy

BackgroundIsolated lung metastases in sarcoma and colorectal cancer patients are inadequately treated with current standard therapies. In Vivo Lung Perfusion, a novel platform, could overcome limitations to photodynamic therapy treatment volumes by using low cellular perfusate, removing blood, theoretically allowing greater light penetration. To develop personalized photodynamic therapy protocols requires in silico light propagation simulations based on optical properties and maximal permissible photodynamic threshold dose of lung tissue. This study presents quantification of optical properties for two perfusates and the photodynamic threshold for 5-ALA and Chlorin e6. MethodsPorcine and human lungs were placed on Ex Vivo Lung Perfusion, and perfused with acellular solution or blood. Isotropic diffusers were placed within bronchi and on lung surface for light transmission measurements, from which absorption and light scattering properties were calculated at multiple wavelengths. Separately, pigs were injected with 5-ALA or Chlorin e6, and lung tissue was irradiated at increasing doses. Resultant lesion sizes were measured by CT and histology to quantify the photodynamic threshold. ResultsLow cellular perfusate reduced the tissue absorption coefficient significantly, increasing penetration depth of light by 3.3 mm and treatment volumes 3-fold. The photodynamic threshold for lung exposed to 5-ALA was consistent with other malignancies. Chlorin e6 levels were undetectable in lung tissue and did not demonstrate photodynamic-induced necrosis. ConclusionsLight penetration with low cellular perfusate is significantly greater and could enable treatments for diffuse disease. This data aids photodynamic treatment planning and will guide clinical translation of photodynamic therapy protocols in the lung, especially during lung perfusion.

Addition of Dialysis to Ex-Vivo Lung Perfusion Maintains Homeostasis and Stability of Donor Lungs: A Proof of Concept Study

<h3>Purpose</h3> Ex-vivo lung perfusion (EVLP) maintains marginal donor lungs at body temperature with ventilation and circulating perfusate, allowing for functional assessment prior to transplantation. Prolonged EVLP would allow for advanced time-dependent therapies for donor lung repair and reconditioning. We hypothesized that the addition of a dialysis machine to the EVLP circuit (dEVLP) would maintain homeostasis of the donor lung and prolong EVLP duration. <h3>Methods</h3> Porcine donor lungs (n=6) were extracted and placed on the EVLP platform for 36 hours or until termination criteria (dynamic compliance < 15ml/cmH₂O) was reached. Lungs were perfused with an acellular solution and closed atrium, according to the Toronto protocol. A dialysis machine was incorporated into the EVLP circuit with a custom-designed dialysate to continuously dialyze perfusate using continuous venovenous hemodialysis (Fig1A). Physiological function, electrolytes and inflammatory mediators in EVLP perfusate were measured hourly. In this proof of concept study, dialysis cases were compared to historical controls with similar protocol (n=5). <h3>Results</h3> EVLP was prolonged in the dialysis group with a mean duration of EVLP reaching 30.83±0.57 h in the dEVLP group compared to 19.6±2.61 h in the historical control group (Fig 1D). Percent lung survival at 24h of perfusion was 100% in the dEVLP group, while only 20% was seen in historical controls (Fig 1B). Dialysis successfully prevented an increase in electrolyte levels (Fig 1C), maintained glucose and lactate levels at baseline (1C), and maintained physiological parameters (Fig 1D). Both lungs which survived to 36h of EVLP presented excellent lung function based on assessment by EVLP. <h3>Conclusion</h3> Dialysis may preserve lung function and length of EVLP by maintaining homeostasis of the lung. Following the results of this encouraging proof of concept study, a formal blinded experiment is underway.

Safety of continuous 12-hour delivery of antimicrobial doses of inhaled nitric oxide during ex vivo lung perfusion

IntroductionHigh-dose nitric oxide (NO) has been shown effective against a variety of micro-organisms in vitro, including common bacteria found in donor organs. However, clinical obstacles related to its implementation in vivo are the formation of methemoglobin and the accumulation of toxic nitrogen compounds. Ex vivo lung perfusion (EVLP) is a platform that allows for organ maintenance with an acellular perfusion solution, thus overcoming these limitations. The present study explores the safety of continuous high-dose inhaled (iNO) during EVLP for an extended period of 12 hours. MethodsLungs procured from Yorkshire pigs were randomized into control (standard ventilation) and treatment (standard ventilation + 200 ppm iNO) groups, then perfused with an acellular solution for 12 hours (n = 4/group). Lung physiology and biological markers were evaluated. ResultsAfter 12 hours of either standard EVLP or EVLP + 200 ppm iNO, we did not notice any significant physiologic difference between the groups: pulmonary oxygenation (P = .586), peak airway pressures (P = .998), and dynamic (P = .997) and static (P = .908) lung compliances. In addition, no significant differences were seen among proinflammatory cytokines measured in perfusate and lung tissue. Importantly, most common toxic compounds were kept at safe levels throughout the treatment course. ConclusionsHigh-dose inhaled NO delivered continuously over 12 hours appears to be safe without inducing any significant pulmonary inflammation or deterioration in lung function. These findings support further efficacy studies to explore the use of iNO for the treatment of infections in donor lungs during EVLP.

Normothermic kidney perfusion: An overview of protocols and strategies.

Normothermic machine perfusion (NMP) technologies are emerging as an important adjunct in organ preservation and transplantation. NMP can enable the reduction or avoidance of cold ischemia and allows for pretransplant measurement of function and metabolic status to assess the suitability of the organ for transplantation. The key requirement of NMP is to provide an environment that is protective to the organ, ensures optimal oxygen delivery and supports metabolic function. Red blood cell-based solutions, artificial hemoglobin solutions, and acellular solutions have all been utilized in NMP. However, there is no clear consensus on perfusion protocols. A period of NMP after hypothermic preservation is the most commonly used strategy. As an alternative, several groups have developed and tested the feasibility of more prolonged periods of NMP. There are only a few reports of the application of NMP in clinical kidney transplantation and each uses different approach and conditions. This review details the rationale for NMP protocols considering duration of NMP and different perfusate compositions in experimental and clinical models. We also include a discussion on the mechanistic action of NMP, comparison of subnormothermic and hypothermic conditions, the different logistical approaches and future requirements.

Inflammatory Response after Immediate versus Delayed Ex-Vivo Lung Perfusion in a Porcine Cardiac Arrest Donation Model

Ex Vivo Lung Perfusion (EVLP) is a promising tool for evaluation and reconditioning in marginal donor lungs. However, the optimal protocol is discussed controversially. It is our hypothesis that different EVLP protocols and different perfusion solutions result in different cytokine patterns during EVLP. Following cardiac arrest and 30 min of no-touch warm ischemia, pig lungs were harvested, flushed and stored in cold preservation solutions. Different groups were processed as immediate long EVLP (I-EVLP, 12 h), delayed short EVLP (4 h) after 9 hours of cold storage (D-EVLP). Lungs were perfused with different acellular solutions: Steen solution or modified HTK solution containing either dextran(CD) or dextran and albumin (CDA). Pulmonary gas exchange, biochemical and structural parameters were recorded hourly. Cytokine / chemokine levels were analyzed in perfusates at baseline and after 1 and 4 h of EVLP. ∆ pO2 (pulmonary venous - pulmonary arterialpO2) was higher in D-EVLP group, especially at h 1: I-EVLP: 259 ± 132, D-EVLP: 291 ± 96 and h 2: I-EVLP: 228 ± 125, D-EVLP: 252 ± 116 (n.s.). Compared to D-EVLP, lungs that underwent EVLP with CD and CDA showed significantly higher oxygen capacity (∆ pO2 CD: 402.79 +30.33 mmHg, CDA: 414.86 + 9.77 mmHg). Within 4 h ELVP, concentrations of IFN-γ, TNF-a, IL-1α/β, IL-10, CXCL8 (IL-8) and IL-18 increased significantly (all p<0.05) in the four groups. Of note, no significant differences could be detected between I- vs. D-EVLP lungs and CD vs. CDA solutions. In a non-heart beating porcine lung donor model with relevant ischemic tissue damage, lungs were perfused with normothermic EVLP. Even after 9 hours cold static preservation (CSP) and 4 hours EVLP lung, function is comparable to direct EVLP. The modified HTK solution with dextran and dextran/ albumin had significantly better oxygenation. Cytokines and chemokines including the cell death marker IL-18 increase during 4 h of EVLP but a delay before EVLP may not dramatically worsen the inflammatory response.

Twenty-Four-Hour Ex Vivo Perfusion with Acellular Solution Enables Successful Replantation of Porcine Forelimbs.

A critical barrier to successful limb replantation and allotransplantation is the maximum allowable limb ischemia time of 4 to 6 hours. The current gold standard is to preserve amputated limbs on an ice slurry. Experimental machine perfusion has yielded promising results as an alternative. In particular, hypothermic acellular perfusion has enabled preservation of amputated limbs for up to 12 hours thus far. Amputated forelimbs of Yorkshire pigs were preserved on static cold storage at 4°C for 4 hours (static cold storage group) or perfused at 8°C for 24 hours (perfusion group) with oxygenated modified STEEN Solution perfusate before replantation. Animals were followed up for 7 days after replantation. Eight animals underwent replantation (cold storage group, n = 4; perfusion group, n = 4). Seventy-five and 100 percent of animals in the static cold storage and perfusion groups survived for 7 days, respectively. Glycogen and adenosine triphosphate remained stable throughout perfusion. Heart and respiratory rate after replantation were increased in the static cold storage group. There was increased damage in muscle biopsy specimens obtained from animals in the static cold storage group after 7 days when compared with those from animals in the perfusion group. Hypothermic acellular ex vivo perfusion of limbs for up to 24 hours enables tissue preservation comparable to that obtained with conventional static cold storage for 4 hours and may reduce muscle damage and systemic reactions on limb replantation. Translation to human limbs may help improve limb replantation and allotransplantation outcomes.

Differential Gene Expression Profiling of Reperfusion During Ex Vivo Lung Perfusion and Lung Transplantation in Humans

Ischemia reperfusion (IR) injury is an unavoidable consequence of lung transplantation. Reperfusion of lung grafts induces physical stress and biological responses to recipient blood. Ex vivo lung perfusion (EVLP) can be considered to be a “modified reperfusion” that simulates the physical aspects of IR, by perfusing the lung with an acellular solution. By comparing the responses of implanted lungs versus those put on normothermic EVLP, we can distinguish the physical and biological responses to IR. We can also identify responses exclusive to EVLP.

Selective Blockade of the Metabotropic Glutamate Receptor mGluR5 Protects Mouse Livers in In Vitro and Ex Vivo Models of Ischemia Reperfusion Injury.

2-Methyl-6-(phenylethynyl)pyridine (MPEP), a negative allosteric modulator of the metabotropic glutamate receptor (mGluR) 5, protects hepatocytes from ischemic injury. In astrocytes and microglia, MPEP depletes ATP. These findings seem to be self-contradictory, since ATP depletion is a fundamental stressor in ischemia. This study attempted to reconstruct the mechanism of MPEP-mediated ATP depletion and the consequences of ATP depletion on protection against ischemic injury. We compared the effects of MPEP and other mGluR5 negative modulators on ATP concentration when measured in rat hepatocytes and acellular solutions. We also evaluated the effects of mGluR5 blockade on viability in rat hepatocytes exposed to hypoxia. Furthermore, we studied the effects of MPEP treatment on mouse livers subjected to cold ischemia and warm ischemia reperfusion. We found that MPEP and 3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pyridine (MTEP) deplete ATP in hepatocytes and acellular solutions, unlike fenobam. This finding suggests that mGluR5s may not be involved, contrary to previous reports. MPEP, as well as MTEP and fenobam, improved hypoxic hepatocyte viability, suggesting that protection against ischemic injury is independent of ATP depletion. Significantly, MPEP protected mouse livers in two different ex vivo models of ischemia reperfusion injury, suggesting its possible protective deployment in the treatment of hepatic inflammatory conditions.

Biomimetic heterogenous elastic tissue development

There is an unmet need for artificial tissue to address current limitations with donor organs and problems with donor site morbidity. Despite the success with sophisticated tissue engineering endeavours, which employ cells as building blocks, they are limited to dedicated labs suitable for cell culture, with associated high costs and long tissue maturation times before available for clinical use. Direct 3D printing presents rapid, bespoke, acellular solutions for skull and bone repair or replacement, and can potentially address the need for elastic tissue, which is a major constituent of smooth muscle, cartilage, ligaments and connective tissue that support organs. Thermoplastic polyurethanes are one of the most versatile elastomeric polymers. Their segmented block copolymeric nature, comprising of hard and soft segments allows for an almost limitless potential to control physical properties and mechanical behaviour. Here we show direct 3D printing of biocompatible thermoplastic polyurethanes with Fused Deposition Modelling, with a view to presenting cell independent in-situ tissue substitutes. This method can expeditiously and economically produce heterogenous, biomimetic elastic tissue substitutes with controlled porosity to potentially facilitate vascularisation. The flexibility of this application is shown here with tubular constructs as exemplars. We demonstrate how these 3D printed constructs can be post-processed to incorporate bioactive molecules. This efficacious strategy, when combined with the privileges of digital healthcare, can be used to produce bespoke elastic tissue substitutes in-situ, independent of extensive cell culture and may be developed as a point-of-care therapy approach.

Acellular Hypothermic Extracorporeal Perfusion Extends Allowable Ischemia Time in a Porcine Whole Limb Replantation Model.

One of the major challenges in traumatic amputation is the need to keep ischemia time brief (4 to 6 hours) to avoid ischemic damage and enable successful replantation. The current inability to meet this challenge often leads to traumatic limb loss, which has a considerable detrimental impact on the quality of life of patients. The authors' team built a portable extracorporeal membrane oxygenator device for the perfusion of amputated extremities with oxygenated acellular solution under controlled parameters. The authors amputated forelimbs of Yorkshire pigs, perfused them ex vivo with acellular Perfadex solution for 12 hours at 10°C in their device, and subsequently replanted them into the host animal. The authors used limbs stored on ice slurry for 4 hours before replantation as their control group. Clinical observation and histopathologic evaluation both demonstrated that there was less morbidity and less tissue damage to the cells during preservation and after replantation in the perfusion group compared with the standard of care. Significant differences in blood markers of muscle damage and tissue cytokine levels underscored these findings. The authors demonstrated the feasibility and superiority of ex vivo hypothermic oxygenated machine perfusion for preservation of amputated limbs over conventional static cold storage and herewith a substantial extension of the allowable ischemia time for replantation after traumatic amputation. This approach could also be applied to the field of transplantation, expanding the potential pool of viable donor vascularized composite allografts.

Intervertebral disc response to stem cell treatment is conditioned by disc state and cell carrier: An ex vivo study

SummaryIn vitro and in vivo studies evidenced that mesenchymal stem cells (MSCs) contribute to intervertebral disc (IVD) regeneration by differentiation towards the disc phenotype and matrix synthesis and/or by paracrine signalling to endogenous cells, thereby promoting a healthier disc phenotype in degenerative discs. The aim of this study was to investigate IVD response to human MSC (hMSC) treatment based on the disc degenerative state and hMSC carrier. Bovine caudal IVDs with endplates were cultured in a bioreactor under simulated physiological (0.1 Hz load and sufficient glucose) or degenerative (10 Hz load and limited glucose) conditions for 7 days. Discs were partially nucleotomised, restored with hMSCs in either fibrin gel or saline solution and cultured under physiological conditions for 7 days. Controls included fibrin and saline without hMSCs. Cell viability, histology, disc height, and gene expression analyses were performed to evaluate regeneration. hMSCs in fibrin were viable and homogenously distributed following 7 days of culture under dynamic loading in partially nucleotomised discs. IVD response to hMSCs was conditioned by both disc degenerative state and hMSC carrier. The effect of the regenerative treatment was stronger on simulated-degenerative discs than on simulated-physiological discs. hMSCs in fibrin induced a superior anabolic response in degenerative IVDs compared with fibrin alone, thus suggesting an added value of the cellular therapy compared with an acellular solution. When comparing fibrin and saline as a hMSC carrier, a significantly higher anabolic response was observed in IVDs treated with hMSCs in fibrin. Moreover, it was found that the degenerative state of the disc influenced hMSC differentiation. Indeed, a significantly higher expression of specific discogenic markers (ACAN and CA12) was observed in hMSCs implanted into physiological discs than in those implanted into degenerative discs. In conclusion, host disc cells and donor hMSC response depend on the degenerative state of the host disc and carrier used for hMSC delivery, and these two aspects need to be considered for a successful translation of hMSC therapies for the treatment of IVD degeneration.

(121) - Comparison of Two Strategies for Experimental Ex-Vivo Lung Perfusion - Cellular Solution and Open Atrium versus Acellular Solution and Closed Atrium

(121) - Comparison of Two Strategies for Experimental Ex-Vivo Lung Perfusion - Cellular Solution and Open Atrium versus Acellular Solution and Closed Atrium

Comparison between cellular and acellular perfusates for ex vivo lung perfusion in a porcine model

Ex vivo lung perfusion with acellular solutions is an established technique for assessing marginal donor lungs. We evaluated the utility of a blood-based lung preservation fluid as an alternative perfusate. Donor lungs from 50-kg donation after cardiac death pigs (n = 24) were randomized into 3 groups: acellular, commercial blood-based, and Papworth-Blood. Physiologic function was evaluated using conventional markers of pulmonary vascular resistance, pulmonary compliance, lactate excretion, partial pressure of oxygen/fraction of inspired oxygen, and wet-to-dry ratios. The immunologic profile was assessed by fluorescence-activated cell sorting analysis of bronchoalveolar lavage and cells entrapped in the leucocyte filter. Cytokines were quantified using a commercial platform. No significant difference was noted in pulmonary vascular resistance (p = 0.26), compliance (p = 0.12), partial pressure of oxygen/fraction of inspired oxygen (p = 0.06) and wet-to-dry ratios (p = 0.26) between groups. There was no difference between the percentages of lymphocytes (p = 0.51), macrophages (p = 0.87), monocytes (p = 0.68), and dendritic cells (p = 0.65) in the leukocyte filters. Interleukin (IL)-1β (p = 0.36), IL-6 (p = 0.08), IL-8 (p = 0.64), and IL-18 (p = 0.14) were elevated in all groups. In bronchoalveolar lavage, IL-8 was significantly higher in the acellular group (p = 0.04). Electron microscopy cell characteristics were similar among the groups. This study demonstrated no significant difference in the physiologic, immunologic, or ultrastructural parameters between lungs perfused with cellular or acellular solutions. The Papworth-Blood solution is a potential alternative perfusate for ex vivo lung perfusion.

(727) - The Impact of Perfusate Oxygenation During Ex Vivo Lung Perfusion on Post-transplant Outcomes: Deoxygenated vs. Fully-Oxygenated

Purpose: The metabolic circumstances in lung grafts on ex vivo lung perfu- sion (EVLP) are compromised as compared with those in lungs in vivo as described in previous studies. While the current EVLP protocol suggests using deoxygenated perfusate with acellular solution, little evidence supports that is the optimal condition. This study aimed to evaluate the quality of lung grafts in the different modes of oxygenated perfusate during EVLP and to define their impact on posttransplant functional outcomes. Methods: Using rat heart-lung blocks after 1 hr cold preservation, the EVLP sys- tem was established with Steen solution at 37°C for 4 hrs. We compared the results of four different modes of perfusate O2 during EVLP: 6%, 40%, 60% and 100%. The evaluation included lung graft function, compliance (Cdyn), and vascular resistance (PVR) as well as circulating glucose, lactate levels and other biochemi-

Bioactivity and cytotoxicity of glass and glass–ceramics based on the 3CaO·P2O5–SiO2–MgO system

The mechanical strength of bioactive glasses can be improved by controlled crystallization, turning its use as bulk bone implants viable. However, crystallization may affect the bioactivity of the material. The aim of this study was to develop glass-ceramics of the nominal composition (wt%) 52.75(3CaO·P₂O₅)-30SiO₂-17.25MgO, with different crystallized fractions and to evaluate their in vitro cytotoxicity and bioactivity. Specimens were heat-treated at 700, 775 and 975 °C, for 4 h. The major crystalline phase identified was whitlockite, an Mg-substituted tricalcium phosphate. The evaluation of the cytotoxicity was carried out by the neutral red uptake methodology. Ionic exchanges with the simulated body fluid SBF-K9 acellular solution during the in vitro bioactivity tests highlight the differences in terms of chemical reactivity between the glass and the glass-ceramics. The effect of crystallinity on the rates of hydroxycarbonate apatite (HCA) formation was followed by Fourier transformed infrared spectroscopy. Although all glass-ceramics can be considered bioactive, the glass-ceramic heat-treated at 775 °C (V775-4) presented the most interesting result, because the onset for HCA formation is at about 24 h and after 7 days the HCA layer dominates completely the spectrum. This occurs probably due to the presence of the whitlockite phase (3(Ca,Mg)O·P₂O₅). All samples were considered not cytotoxic.

The effect of retrograde autologous priming on intraoperative blood product transfusion in coronary artery bypass grafting

Retrograde autologous priming (RAP) of the cardiopulmonary bypass (CPB) circuit could reduce the degree of haemodilution associated with priming with acellular solutions. However, there is no strong evidence to prove that the practice of RAP reduced intraoperative packed red cell (PRC) or blood product transfusion. To evaluate the effect of RAP on intraoperative PRC usage in coronary artery bypass grafting (CABG). This study is a prospective, observational study on patients who underwent first-time, isolated CABG using CPB between April 2012 and July 2012. Two groups of patients were identified: 1. Non-RAP group (n=128) and 2. RAP group (n=73). The primary outcome for the study was the amount of PRC and blood product usage between the induction of anaesthesia and the cessation of CPB. Use of PRC and blood products in the operating room was comparable in both groups. Univariate logistic regression showed that RAP was not an independent predictor of PRC or blood product transfusion (p=0.43). Multivariate logistic regression showed that CPB time, preoperative haemoglobin (Hb) levels and creatinine clearance were independent predictors of blood product transfusion. Practising RAP with mean volumes of 300 ml does not necessarily reduce PRC and other blood product transfusion requirements during CABG. In our practice, RAP was performed, aiming at displacing CPB circuit prime volume with which the perfusionist felt comfortable and dictated by haemodynamic parameters prior to commencing CPB. We presume this is the case in many units around the world. This practice, in our opinion, is not enough to achieve the benefits of RAP, if any, in the form of a reduction of packed red cell transfusion requirements. The true advantages of RAP in cardiac surgery need to be studied in a prospective, randomized, controlled trial.

Editorial Comment: Growing experience with ex vivo lung perfusion: many ways leading to the same goal

Ex vivo lung perfusion (EVLP) has gained enormous interest, and a substantial number of lung transplant centres are using EVLP to increase the donor pool. The most widespread approach currently is the optimization and second evaluation of initially unacceptable donor organs with reversible impairments or donors after circulatory death. In this issue, the Sahlgrenska group presents an update of their experience with 5 additional cases [1] compared with the intial report from 2012 [2]. The results of this paper confirm the satisfying outcome achieved with the use of EVLP lungs. During the additional 10 months reported in this paper, 20% of all transplanted lungs were evaluated with EVLP. This represents a substantial increase in the transplant activity compared with if these lungs had all been otherwise rejected. The 100% acceptance rate after EVLP (with the exception of two single lungs while still using the contralateral side) suggests a careful preselection of EVLP cases. There are several different technical EVLP solutions available on the market, and there are distinct differences in the way EVLP is performed and in the interpretation of results obtained during EVLP. To accept a lung after EVLP, the Sahlgrenska group relies on a single measurement of venous pO2 and functional parameters if the initial values are fine. Due to variabilities in gas flow to the deoxygenator and differences in the oxygen content in the inflow, the pO2 difference between the arterial inflow and venous outflow (ΔpO2) is used by the majority of centres to judge oxygenation on EVLP. Using venous pO2 as the main parameter blinds out this potentially valuable additional information and might be insufficient to completely assess the oxygenation capacity; however, no direct comparison between these two parameters has been performed in the clinical setting. The largest EVLP experience so far has been reported by centres using acellular Steen solution [3–6] with consistent results regarding the recipient outcome. In contrast to this, the Sahlgrenska group advocates the use of red cells in addition to the Steen solution with a haematocrit of 10–15%. So far, it remains unclear if the initial oxygenation measurements obtained on EVLP directly correspond with the values obtained in the donor. Since absolute oxygen values in an acellular solution are not equal to the values obtained in the donor, the development of functional parameters and oxygenation levels during EVLP are additional major parameters in deciding about the acceptance of a lung. If the addition of red cells enhances the direct comparability to the donor is unclear and the interpretation of a single measurement of the venous pO2 seems doubtful. If the first measurement is already satisfying and the authors speculate on direct comparability with the donor, there is a high likelihood that the increase in pO2 is rather due to a full recruitment than a true improvement. In this aspect, EVLP could be seen as a rather time-consuming and sophisticated method for what could have been easily achieved by further optimization in the donor. No detailed figures on the comparability between donor and recipient values are provided in the current paper. Even though the clinical results are convincing, there are still questions remaining on the exact interpretation of the results that might receive answers once higher patient numbers have been recruited. It is very exciting to see the different ways in which EVLP is developing and to compare the advantageous aspects of individual approaches. EVLP has the potential to completely alter our current practice of organ preservation, serve as a platform for a multitude of therapeutic interventions and improve outcome after lung transplantation.

A New Lung Preservation Technique through Bronchial Artery Perfusion on Ex-Vivo Lung

Purpose Airway hypoxia due to microvascular loss may be a predisposing factor for subsequent fibrotic remodeling in allografts, resulting in suboptimal transplant outcomes in lung transplantation. Ex-vivo lung perfusion (EVLP) is a novel strategy for lung allograft preservation that keeps the organs at physiological protective conditions. Modifying EVLP strategy, we have developed a new lung preservation technique focusing on airway protection through bronchial artery perfusion. Methods and Materials Using heart-lung blocks harvested from Lewis rats, we have developed a model where bronchial circulation (using another cannula from descending aorta, retrogradely) was synchronously perfused in addition to pulmonary circulation with acellular Steen solution on EVLP (the scheme of the model is shown in Fig 1 ). Rat lungs after 1 hour cold preservation, perfused and ventilated on EVLP for 4 hours were evaluated and lungs with bronchial artery perfusion were compared with those without bronchial perfusion on EVLP. The studies were performed based on our rat EVLP protocol that has proven to be reliable with at least 4 hours of duration with stable results. Results At 4 hours on EVLP, bronchial artery perfusion was associated with significantly better preserved oxygenation and better compliance in lung allografts through reduced glucose consumption and increased ATP level as compared to the control group. Conclusions Bronchial artery perfusion on EVLP may be a new lung preservation strategy that breaks through the current major limitation in hypoxic transplant airways after lung transplantation.

The Importance of Repeated Measurements To Assess Transplant Suitability in Clinical Ex Vivo Lung Perfusion (EVLP)

Purpose Normothermic acellular EVLP is a safe and feasible option to evaluate and recondition marginal donor lungs. A ΔPo2 of 350 mmHg between venous and arterial solution is used as a cutoff value for acceptance. The validity of oxygenation values obtained in an acellular perfusion solution remain however unclear. Methods and Materials An analysis of our prospective clinical EVLP database since 3/2010 was performed to compare oxygenation values during clinical EVLP to values obtained in the brain dead donor as well as differences in functional parameters between accepted and rejected lungs assessed during EVLP. Results From 3/2010-10/2012 20 EVLPs were performed (15 lungs transplanted, 5 rejected). The first pvO2 (FiO2=1.0) in the venous solution was significantly higher in both, accepted lungs (median: 429 mmHg, range 258-523) and rejected lungs (median: 417mmHg, range 304–474) compared to the respective last donor blood gases (median accepted 215 mmHg, range 133–330, p Conclusions Oxygenation values obtained during EVLP are not directly comparable to donor values. pvO2, ΔPo2, compliance as well as AWP showed a significantly different development in accepted lungs compared to rejected lungs. These findings underline that the decision for transplant suitability during EVLP should not be based on a single assessment but the trend in all functional parameter should be taken into account.