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The Journal of Thoracic and... Aug 2022Ex vivo lung perfusion has emerged as a novel technique to safely preserve lungs before transplantation. Recent studies have demonstrated an accumulation of...
OBJECTIVE
Ex vivo lung perfusion has emerged as a novel technique to safely preserve lungs before transplantation. Recent studies have demonstrated an accumulation of inflammatory molecules in the perfusate during ex vivo lung perfusion. These proinflammatory molecules, including damage-associated molecular patterns and inflammatory cytokines, may contribute to acute and chronic allograft dysfunction. At present, ex vivo lung perfusion is performed clinically at normothermic temperature (37°C). The effect of lowering temperature to the subnormothermic range during ex vivo lung perfusion has not been reported. In this study, we hypothesized that lower ex vivo lung perfusion temperature will lead to a reduction in allograft inflammation and result in improved post-transplant graft function.
METHODS
Lewis rat heart-lung blocs underwent 4 hours of ex vivo lung perfusion in 3 temperature groups: 37°C (MP37), 30°C (MP30), and 25°C (MP25). In the control group, lung grafts were preserved by static cold storage before transplantation. After ex vivo lung perfusion or static cold storage, the left lung was transplanted for 2 hours before the animal was killed. Sera and tissue were collected and analyzed.
RESULTS
There were no differences in partial pressure of arterial oxygenation to fraction of inspired oxygen ratios during 4 hours of ex vivo lung perfusion between temperature groups. Tumor necrosis factor α significantly increased in the MP37 group during ex vivo lung perfusion, whereas this was not seen at lower temperatures. Extracellular DNA and high-mobility group box 1 perfusate concentrations increased significantly during ex vivo lung perfusion in all groups, but the rate of increase was diminished at lower temperature. Two hours post-transplant, there were no significant differences in partial pressure of arterial oxygenation to fraction of inspired oxygen ratios of the lung graft or serum damage-associated molecular pattern levels among groups. On histologic grading after transplantation, greater injury was observed in the MP30 and MP37 groups, but not MP25, when compared with static cold storage.
CONCLUSIONS
Subnormothermic ex vivo lung perfusion at 25°C reduces the production of inflammatory mediators during ex vivo lung perfusion and is associated with reduced histologic graft injury after transplantation.
Topics: Animals; Inflammation; Lung; Lung Transplantation; Organ Preservation; Oxygen; Perfusion; Rats; Rats, Inbred Lew; Reperfusion Injury
PubMed: 33640121
DOI: 10.1016/j.jtcvs.2021.01.066 -
Journal of Visualized Experiments : JoVE Sep 2023This protocol presents an optimized erythrocytes-free NEVLP system using mouse livers. Ex vivo preservation of mouse livers was achieved by employing modified cannulas...
This protocol presents an optimized erythrocytes-free NEVLP system using mouse livers. Ex vivo preservation of mouse livers was achieved by employing modified cannulas and techniques adapted from conventional commercial ex vivo perfusion equipment. The system was utilized to evaluate the preservation outcomes following 12 h of perfusion. C57BL/6J mice served as liver donors, and the livers were explanted by cannulating the portal vein (PV) and bile duct (BD), and subsequently flushing the organ with warm (37 °C) heparinized saline. Then, the explanted livers were transferred to the perfusion chamber and subjected to normothermic oxygenated machine perfusion (NEVLP). Inlet and outlet perfusate samples were collected at 3 h intervals for perfusate analysis. Upon completion of the perfusion, liver samples were obtained for histological analysis, with morphological integrity assessed using modified Suzuki-Score through Hematoxylin-Eosin (HE) staining. The optimization experiments yielded the following findings: (1) mice weighing over 30 g were deemed more suitable for the experiment due to the larger size of their bile duct (BD). (2) a 2 Fr (outer diameter = 0.66 mm) polyurethane cannula was better suited for cannulating the portal vein (PV) when compared to a polypropylene cannula. This was attributed to the polyurethane material's enhanced grip, resulting in reduced catheter slippage during the transfer from the body to the organ chamber. (3) for cannulation of the bile duct (BD), a 1 Fr (outer diameter = 0.33 mm) polyurethane cannula was found to be more effective compared to the polypropylene UT - 03 (outer diameter = 0.30 mm) cannula. With this optimized protocol, mouse livers were successfully preserved for a duration of 12 h without significant impact on the histological structure. Hematoxylin-Eosin (HE) staining revealed a well-preserved morphological architecture of the liver, characterized by predominantly viable hepatocytes with clearly visible nuclei and mild dilation of hepatic sinusoids.
Topics: Mice; Animals; Eosine Yellowish-(YS); Hematoxylin; Polypropylenes; Polyurethanes; Liver Transplantation; Organ Preservation; Mice, Inbred C57BL; Liver; Perfusion
PubMed: 37811934
DOI: 10.3791/65363 -
Transplant International : Official... 2023Pancreas transplantation is the only curative treatment for patients with complicated diabetes, and organ shortage is a common and increasing problem. Strategies to...
Pancreas transplantation is the only curative treatment for patients with complicated diabetes, and organ shortage is a common and increasing problem. Strategies to expand the donor pool are needed, and normothermic perfusion of the pancreas has the potential to test and repair grafts before implantation. Between January 2021 and April 2022, six human pancreases, declined for transplantation or islet isolation, were perfused using a previously established method by our group. All 6 cases were successfully perfused for 4 h, with minimal edema. The mean age of the donors was 44.16 ± 13.8 years. Five grafts were obtained from neurological death donors, and one was obtained from a donation after cardiac death. The mean glucose and lactate levels decreased throughout perfusion and insulin levels increased. All 6 grafts were metabolically active during perfusion and histopathology showed minimal tissue injury and no edema. Human normothermic perfusion of the pancreas is feasible and safe and has the potential to expand the donor pool. Future studies will focus on tests and biomarkers for the assessment of grafts.
Topics: Humans; Adult; Middle Aged; Organ Preservation; Feasibility Studies; Perfusion; Tissue Donors; Pancreas; Allografts
PubMed: 37252614
DOI: 10.3389/ti.2023.10936 -
Critical Care (London, England) Jun 2021In acute respiratory distress syndrome (ARDS), non-ventilated perfused regions coexist with non-perfused ventilated regions within lungs. The number of unmatched regions... (Observational Study)
Observational Study
BACKGROUND
In acute respiratory distress syndrome (ARDS), non-ventilated perfused regions coexist with non-perfused ventilated regions within lungs. The number of unmatched regions might reflect ARDS severity and affect the risk of ventilation-induced lung injury. Despite pathophysiological relevance, unmatched ventilation and perfusion are not routinely assessed at the bedside. The aims of this study were to quantify unmatched ventilation and perfusion at the bedside by electrical impedance tomography (EIT) investigating their association with mortality in patients with ARDS and to explore the effects of positive end-expiratory pressure (PEEP) on unmatched ventilation and perfusion in subgroups of patients with different ARDS severity based on PaO/FiO and compliance.
METHODS
Prospective observational study in 50 patients with mild (36%), moderate (46%), and severe (18%) ARDS under clinical ventilation settings. EIT was applied to measure the regional distribution of ventilation and perfusion using central venous bolus of saline 5% during end-inspiratory pause. We defined unmatched units as the percentage of only ventilated units plus the percentage of only perfused units.
RESULTS
Percentage of unmatched units was significantly higher in non-survivors compared to survivors (32[27-47]% vs. 21[17-27]%, p < 0.001). Percentage of unmatched units was an independent predictor of mortality (OR 1.22, 95% CI 1.07-1.39, p = 0.004) with an area under the ROC curve of 0.88 (95% CI 0.79-0.97, p < 0.001). The percentage of ventilation to the ventral region of the lung was higher than the percentage of ventilation to the dorsal region (32 [27-38]% vs. 18 [13-21]%, p < 0.001), while the opposite was true for perfusion (28 [22-38]% vs. 36 [32-44]%, p < 0.001). Higher percentage of only perfused units was correlated with lower dorsal ventilation (r = - 0.486, p < 0.001) and with lower PaO/FiO ratio (r = - 0.293, p = 0.039).
CONCLUSIONS
EIT allows bedside assessment of unmatched ventilation and perfusion in mechanically ventilated patients with ARDS. Measurement of unmatched units could identify patients at higher risk of death and could guide personalized treatment.
Topics: Adult; Aged; Electric Impedance; Female; Humans; Italy; Male; Middle Aged; Perfusion; Prognosis; Prospective Studies; Respiration, Artificial; Respiratory Distress Syndrome; Simplified Acute Physiology Score
PubMed: 34082795
DOI: 10.1186/s13054-021-03615-4 -
Annals of Surgery Nov 2023To provide mechanistic insight into key biological alterations in donation after circulatory death kidneys during continuous pefusion we performed mass spectrometry... (Randomized Controlled Trial)
Randomized Controlled Trial
OBJECTIVE
To provide mechanistic insight into key biological alterations in donation after circulatory death kidneys during continuous pefusion we performed mass spectrometry profiling of perfusate samples collected during a phase 3 randomized double-blind paired clinical trial of hypothermic machine perfusion with and without oxygen (COMPARE).
BACKGROUND
Despite the clinical benefits of novel perfusion technologies aiming to better preserve donor organs, biological processes that may be altered during perfusion have remained largely unexplored. The collection of serial perfusate samples during the COMPARE clinical trial provided a unique resource to study perfusate proteomic profiles, with the hypothesis that in-depth profiling may reveal biologically meaningful information on how donor kidneys benefit from this intervention.
METHODS
Multiplexed liquid chromatography-tandem mass spectrometry was used to obtain a proteome profile of 210 perfusate samples. Partial least squares discriminant analysis and multivariate analysis involving clinical and perfusion parameters were used to identify associations between profiles and clinical outcomes.
RESULTS
Identification and quantitation of 1716 proteins indicated that proteins released during perfusion originate from the kidney tissue and blood, with blood-based proteins being the majority. Data show that the overall hypothermic machine perfusion duration is associated with increasing levels of a subgroup of proteins. Notably, high-density lipoprotein and complement cascade proteins are associated with 12-month outcomes, and blood-derived proteins are enriched in the perfusate of kidneys that developed acute rejection.
CONCLUSIONS
Perfusate profiling by mass spectrometry was informative and revealed proteomic changes that are biologically meaningful and, in part, explain the clinical observations of the COMPARE trial.
Topics: Humans; Kidney Transplantation; Proteome; Proteomics; Organ Preservation; Kidney; Perfusion; Tissue Donors
PubMed: 37503631
DOI: 10.1097/SLA.0000000000006046 -
Transplantation Aug 2022Ischemia-reperfusion injury remains a primary concern in upper extremity transplantation. Ex vivo normothermic perfusion (EVNP) enables near-physiological organ...
BACKGROUND
Ischemia-reperfusion injury remains a primary concern in upper extremity transplantation. Ex vivo normothermic perfusion (EVNP) enables near-physiological organ preservation, avoiding the deleterious effects of hypoxia and cooling. We investigated the effectiveness of human limb EVNP compared with static cold storage (SCS).
METHODS
Twenty human upper extremities were procured. Ten were perfused at 38 °C with an oxygenated red blood cell-based solution, and contralateral limbs served as SCS control (4 °C). EVNP was terminated with systolic arterial pressure ≥115 mm Hg, compartment fullness, or a 20% decline in oxygen saturation. Weight, contractility, compartment pressure, tissue oxygen saturation, and uptake rates were assessed. Perfusate fluid dynamics, gases, electrolytes, and metabolites were measured. Myocyte injury scores and liquid chromatography-mass spectrometry analysis were performed.
RESULTS
EVNP duration was 41.6 ± 9.4 h. Vascular resistance averaged 173.0 ± 29.4 mm Hg × min/L. Weight change and compartment pressures were 0.4 ± 12.2% ( P = 0.21) and 21.7 ± 15.58 mm Hg ( P = 0.003), respectively. Arterial and venous carbon dioxide partial pressure, oxygen saturation, and pH were 509.5 ± 91.4 mm Hg, 15.7 ± 30.2 mm Hg, 87.4 ± 11.4%, and 7.3 ± 0.2, respectively. Oxygen uptake rates averaged 5.7 ± 2.8 mL/min/g. Lactate reached 20 mmol/L after 15 (interquartile range = 6) h. Limb contractility was preserved for 30.5 (interquartile range = 15.8) h ( P < 0.001) and negatively correlated with perfusate potassium (ρ = -0.7, P < 0.001). Endpoint myocyte injury scores were 28.9 ± 11.5% (EVNP) and 90.2 ± 11.8% (SCS) ( P < 0.001). A significant increase in taurine ( P = 0.002) and decrease in tryptophan ( P = 0.002) were detected. Infrared thermography and indocyanine green angiography confirmed the presence of peripheral perfusion.
CONCLUSIONS
EVNP can overcome the limitations of cold preservation by extending preservation times, enabling limb quality assessment, and allowing limb reconditioning before transplantation.
Topics: Extracorporeal Circulation; Humans; Organ Preservation; Organ Preservation Solutions; Perfusion; Upper Extremity
PubMed: 35152257
DOI: 10.1097/TP.0000000000004045 -
Critical Care (London, England) May 2020Microcirculatory perfusion disturbances are associated with increased morbidity and mortality in patients undergoing cardiac surgery with cardiopulmonary bypass (CPB)....
BACKGROUND
Microcirculatory perfusion disturbances are associated with increased morbidity and mortality in patients undergoing cardiac surgery with cardiopulmonary bypass (CPB). Technological advancements made it possible to monitor sublingual microcirculatory perfusion over time. The goal of this review is to provide an overview of the course of alterations in sublingual microcirculatory perfusion following CPB. The secondary goal is to identify which parameter of sublingual microcirculatory perfusion is most profoundly affected by CPB.
METHODS
PubMed and Embase databases were systematically searched according to PRISMA guidelines and as registered in PROSPERO. Studies that reported sublingual microcirculatory perfusion measurements before and after onset of CPB in adult patients undergoing cardiac surgery were included. The primary outcome was sublingual microcirculatory perfusion, represented by functional capillary density (FCD), perfused vessel density (PVD), total vessel density (TVD), proportion of perfused vessels (PPV), and microvascular flow index (MFI).
RESULTS
The search identified 277 studies, of which 19 fulfilled all eligibility criteria. Initiation of CPB had a profound effect on FCD, PVD, or PPV. Seventeen studies (89%) reported one or more of these parameters, and in 11 of those studies (65%), there was a significant decrease in these parameters during cardiac surgery; the other 6 studies (35%) reported no effect. In 29% of the studies, FCD, PVD, or PPV normalized by the end of cardiac surgery, and in 24% percent of the studies, this effect lasted at least 24 h. There was no clear effect of CPB on TVD and a mixed effect on MFI.
CONCLUSION
CPB during cardiac surgery impaired sublingual microcirculatory perfusion as reflected by reduced FCD, PVD, and PPV. Four studies reported this effect at least 24 h after surgery. Further research is warranted to conclude on the duration of CPB-induced microcirculatory perfusion disturbances and the relationship with clinical outcome.
TRIAL REGISTRATION
PROSPERO, CRD42019127798.
Topics: Adult; Cardiopulmonary Bypass; Female; Humans; Male; Microcirculation; Middle Aged; Perfusion; Postoperative Complications
PubMed: 32404120
DOI: 10.1186/s13054-020-02948-w -
Journal of Cardiovascular Medicine... Aug 2022Selective antegrade cerebral perfusion technique is a method of cerebral protection used worldwide during aortic arch surgery. This study was designed to identify a...
AIMS
Selective antegrade cerebral perfusion technique is a method of cerebral protection used worldwide during aortic arch surgery. This study was designed to identify a potential correlation between perfusion flows and the development of postoperative transient neurological dysfunctions.
METHODS
From January 2015 to May 2020, 175 patients underwent elective surgical replacement of the aortic arch using selective antegrade cerebral perfusion at the Cardiac Surgery Unit of Sant'Orsola Hospital in Bologna. Considering that patients who developed a permanent neurological dysfunction and those who died before a possible evaluation of neurological status were excluded, the study population included 160 patients. The perfusion flows were collected and analyzed. Univariate and multivariate analyses were performed to identify the statistical risk factors involved in the onset of transient neurological dysfunctions.
RESULTS
The study population was divided into two groups: 138 patients (86.3%) without and 22 (13.8%) with postoperative transient neurological complications. Among the intra-operative parameters collected in the study, the univariate analysis showed that the indexed medium perfusion flow of selective antegrade cerebral perfusion was significantly lower in the transient neurological dysfunctions group (11.63 ± 2.41 ml/kg/min vs 12.62 ± 2.39 ml/kg/min, P -value = 0.03). The multivariate logistic regression analysis showed that the female gender ( P = 0.004, OR = 4.816, IC = 1.636-14.174) was predictor of transient neurological dysfunctions.
CONCLUSION
The results of the study showed that lower perfusion flows seem to be related to a higher probability of developing transient neurological dysfunctions. However, the analysis of a wider population is required to confirm these preliminary data.
Topics: Aorta, Thoracic; Cerebrovascular Circulation; Female; Humans; Perfusion; Postoperative Complications; Retrospective Studies; Risk Factors; Treatment Outcome
PubMed: 35904991
DOI: 10.2459/JCM.0000000000001340 -
Analytical Chemistry Oct 2022Targeted delivery and labeling of single living cells in heterogeneous cell populations are of great importance to understand the molecular biology and physiological...
Targeted delivery and labeling of single living cells in heterogeneous cell populations are of great importance to understand the molecular biology and physiological functions of individual cells. However, it remains challenging to perfuse fluorescence markers into single living cells with high spatial and temporal resolution without interfering neighboring cells. Here, we report a single cell perfusion and fluorescence labeling strategy based on nanoscale glass nanopipettes. With the nanoscale tip hole of 100 nm, the use of nanopipettes allows special perfusion and high-resolution fluorescence labeling of different subcellular regions in single cells of interest. The dynamic of various fluorescent probes has been studied to exemplify the feasibility of nanopipette-dependent targeted delivery. According to experimental results, the cytoplasm labeling of Sulfo-Cyanine5 and fluorescein isothiocyanate is mainly based on the Brownian movement due to the dyes themselves and does not have a targeting ability, while the nucleus labeling of 4',6-diamidino-2-phenylindole (DAPI) is originated from the adsorption between DAPI and DNA in the nucleus. From the finite element simulation, the precise manipulation of intracellular delivery is realized by controlling the electro-osmotic flow inside the nanopipettes, and the different delivery modes between nontargeting dyes and nucleus-targeting dyes were compared, showcasing the valuable ability of nanopipette-based method for the analysis of specially defined subcellular regions and the potential applications for single cell surgery, subcellular manipulation, and gene delivery.
Topics: DNA; Fluoresceins; Fluorescent Dyes; Isothiocyanates; Nanotechnology; Perfusion
PubMed: 36162134
DOI: 10.1021/acs.analchem.2c02537 -
Current Opinion in Organ Transplantation Dec 2023A major hurdle hindering more widespread application of reconstructive transplantation is the very limited cold ischemia time (CIT) of vascularized composite allografts... (Review)
Review
PURPOSE OF REVIEW
A major hurdle hindering more widespread application of reconstructive transplantation is the very limited cold ischemia time (CIT) of vascularized composite allografts (VCAs). In this review, we discuss cutting edge machine perfusion protocols and preservation strategies to overcome this limitation.
RECENT FINDINGS
Several preclinical machine perfusion studies have demonstrated the multifactorial utility of this technology to extend preservation windows, assess graft viability prior to transplantation and salvage damaged tissue, yet there are currently no clinically approved machine perfusion protocols for reconstructive transplantation. Thus, machine perfusion remains an open challenge in VCA due to the complexity of the various tissue types. In addition, multiple other promising avenues to prolong preservation of composite allografts have emerged. These include cryopreservation, high subzero preservation, vitrification and nanowarming. Despite several studies demonstrating extended preservation windows, there are several limitations that must be overcome prior to clinical translation. As both machine perfusion and subzero preservation protocols have rapidly advanced in the past few years, special consideration should be given to their potential complementary utilization.
SUMMARY
Current and emerging machine perfusion and preservation technologies in VCA have great promise to transform the field of reconstructive transplantation, as every extra hour of CIT helps ease the complexities of the peri-transplant workflow. Amongst the many advantages, longer preservation windows may allow for elective procedures, improved matching, establishment of novel immunomodulatory protocols and global transport of grafts, ultimately enabling us the ability to offer this life changing procedure to more patients.
Topics: Humans; Composite Tissue Allografts; Organ Preservation; Perfusion; Transplantation, Homologous; Liver Transplantation
PubMed: 37823760
DOI: 10.1097/MOT.0000000000001107