Combined Mesenchymal Stromal Cell Therapy and Extracorporeal Membrane Oxygenation in Acute Respiratory Distress Syndrome. A Randomized Controlled Trial in Sheep.

Rationale: Mesenchymal stromal cell (MSC) therapy is a promising intervention for acute respiratory distress syndrome (ARDS), although trials to date have not investigated its use alongside extracorporeal membrane oxygenation (ECMO). Recent preclinical studies have suggested that combining these interventions may attenuate the efficacy of ECMO.Objectives: To determine the safety and efficacy of MSC therapy in a model of ARDS and ECMO.Methods: ARDS was induced in 14 sheep, after which they were established on venovenous ECMO. Subsequently, they received either endobronchial induced pluripotent stem cell-derived human MSCs (hMSCs) (n = 7) or cell-free carrier vehicle (vehicle control; n = 7). During ECMO, a low Vt ventilation strategy was employed in addition to protocolized hemodynamic support. Animals were monitored and supported for 24 hours. Lung tissue, bronchoalveolar fluid, and plasma were analyzed, in addition to continuous respiratory and hemodynamic monitoring.Measurements and Main Results: The administration of hMSCs did not improve oxygenation (PaO2/FiO2 mean difference = -146 mm Hg; P = 0.076) or pulmonary function. However, histological evidence of lung injury (lung injury score mean difference = -0.07; P = 0.04) and BAL IL-8 were reduced. In addition, hMSC-treated animals had a significantly lower cumulative requirement for vasopressor. Despite endobronchial administration, animals treated with hMSCs had a significant elevation in transmembrane oxygenator pressure gradients. This was accompanied by more pulmonary artery thromboses and adherent hMSCs found on explanted oxygenator fibers.Conclusions: Endobronchial hMSC therapy in an ovine model of ARDS and ECMO can impair membrane oxygenator function and does not improve oxygenation. These data do not recommend the safe use of hMSCs during venovenous ECMO.

Shear-dependent platelet aggregation size.

Nonsurgical bleeding is the most frequent complication of left ventricular assist device (LVAD) support. Supraphysiologic shear rates generated in LVAD causes impaired platelet aggregation, which increases the risk of bleeding. The effect of shear rate on the formation size of platelet aggregates has never been reported experimentally, although platelet aggregation size can be considered to be directly relevant to bleeding complications. Therefore, this study investigated the impact of shear rate and exposure time on the formation size of platelet aggregates, which is vital in predicting bleeding in patients with an LVAD. Human platelet-poor plasma (containing von Willebrand factor, vWF) and fluorochrome-labeled platelets were subjected to a range of shear rates (0-10 000 s-1 ) for 0, 5, 10, and 15 minutes using a custom-built blood-shearing device. Formed sizes of platelet aggregates under a range of shear-controlled environment were visualized and measured using microscopy. The loss of high molecular weight (HMW) vWF multimers was quantified using gel electrophoresis and immunoblotting. An inhibition study was also performed to investigate the reduction in platelet aggregation size and HMW vWF multimers caused by either mechanical shear or enzymatic (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13-ADAMTS13, the von Willebrand factor protease) mechanism under low and high shear conditions (360 and 10 000 s-1 ). We found that the average size of platelet aggregates formed under physiological shear rates of 360-3000 s-1 (200-300 µm2 ) was significantly larger compared to those sheared at >6000 s-1 (50-100 µm2 ). Furthermore, HMW vWF multimers were reduced with increased shear rates. The inhibition study revealed that the reduction in platelet aggregation size and HWM vWF multimers were mainly associated with ADAMTS13. In conclusion, the threshold of shear rate must not exceed >6000 s-1 in order to maintain the optimal size of platelet aggregates to "plug off" the injury site and stop bleeding.

The effect of hyperoxia on inflammation and platelet responses in an ex vivo extracorporeal membrane oxygenation circuit.

Use of extracorporeal membrane oxygenation (ECMO) is expanding, however, it is still associated with significant morbidity and mortality. Activation of inflammatory and innate immune responses and hemostatic alterations contribute to complications. Hyperoxia may play a role in exacerbating these responses. Nine ex vivo ECMO circuits were tested using fresh healthy human whole blood, with two oxygen levels: 21% inspired fraction of oxygen (FiO2 ; mild hyperoxia; n = 5) and 100% FiO2 (severe hyperoxia; n = 4). Serial blood samples were taken for analysis of platelet aggregometry, leukocyte activation, inflammatory, and oxidative stress markers. ECMO resulted in reduced adenosine diphosphate- (P < .05) and thrombin receptor activating peptide-induced (P < .05) platelet aggregation, as well as increasing levels of the neutrophil activation marker, neutrophil elastase (P = .013). Additionally, levels of the inflammatory chemokine interleukin-8 were elevated (P < .05) and the activity of superoxide dismutase, a marker of oxidative stress, was increased (P = .002). Hyperoxia did not augment these responses, with no significant differences detected between mild and severe hyperoxia. Our ex vivo model of ECMO revealed that the circuit itself triggers a pro-inflammatory and oxidative stress response, however, exposure to supra-physiologic oxygen does not amplify that response. Extended-duration studies and inclusion of an endothelial component could be beneficial in characterizing longer term changes.

Administration of mesenchymal stem cells during ECMO results in a rapid decline in oxygenator performance.

Mesenchymal stem cells (MSCs) have attracted attention as a potential therapy for Acute Respiratory Distress Syndrome (ARDS). At the same time, the use of extracorporeal membrane oxygenation (ECMO) has increased among patients with severe ARDS. To date, early clinical trials of MSCs in ARDS have excluded patients supported by ECMO. Here we provide evidence from an ex-vivo model of ECMO to suggest that the intravascular administration of MSCs during ECMO may adversely impact the function of a membrane oxygenator. The addition of clinical grade MSCs resulted in a reduction of flow through the circuit in comparison to controls (0.6 ±0.35 L min-1vs 4.12 ± 0.03 L min-1, at 240 minutes) and an increase in the transoygenator pressure gradient (101±9 mmHg vs 21±4 mmHg, at 240 minutes). Subsequent immunohistochemistry analysis demonstrated quantities of MSCs highly adherent to membrane oxygenator fibres. This study highlights the potential harm associated with MSC therapy during ECMO and suggests further areas of research required to advance the translation of cell therapy in this population.

Effect of ex vivo extracorporeal membrane oxygenation flow dynamics on immune response.

BACKGROUND: Extracorporeal membrane oxygenation is a life-saving support for heart and/or lung failure patients. Despite technological advancement, abnormal physiology persists and has been associated with subsequent adverse events. These include thrombosis, bleeding, systemic inflammatory response syndrome and infection. However, the underlying mechanisms are yet to be elucidated. We aimed to investigate whether the different flow dynamics of extracorporeal membrane oxygenation would alter immune responses, specifically the overall inflammatory response, leukocyte numbers and activation/adhesion surface antigen expression. METHODS: An ex vivo model was used with human whole blood circulating at 37°C for 6 hours at high (4 L/minute) or low (1.5 L/minute) flow dynamics, with serial blood samples taken for analysis. RESULTS: During high flow, production of interleukin-1ß (p < 0.0001), interleukin-6 (p = 0.0075), tumour necrosis factor-α (p = 0.0013), myeloperoxidase (p < 0.0001) and neutrophil elastase (p < 0.0001) were significantly elevated over time compared to low flow, in particular at 6 hours. While the remaining assessments exhibited minute changes between flow dynamics, a consistent trend of modulation in leukocyte subset numbers and phenotype was observed at 6 hours. CONCLUSION: We conclude that prolonged circulation at high flow triggers a prominent pro-inflammatory cytokine response and activates neutrophil granule release, but further research is needed to better characterize the effect of flow during extracorporeal membrane oxygenation.

Mesenchymal stem cells may ameliorate inflammation in an ex vivo model of extracorporeal membrane oxygenation.

INTRODUCTION: Mesenchymal stem cells exhibit immunomodulatory properties which are currently being investigated as a novel treatment option for Acute Respiratory Distress Syndrome. However, the feasibility and efficacy of mesenchymal stem cell therapy in the setting of extracorporeal membrane oxygenation is poorly understood. This study aimed to characterise markers of innate immune activation in response to mesenchymal stem cells during an ex vivo simulation of extracorporeal membrane oxygenation. METHODS: Ex vivo extracorporeal membrane oxygenation simulations (n = 10) were conducted using a commercial extracorporeal circuit with a CO2-enhanced fresh gas supply and donor human whole blood. Heparinised circuits (n = 4) were injected with 40 × 106-induced pluripotent stem cell-derived human mesenchymal stem cells, while the remainder (n = 6) acted as controls. Simulations were maintained, under physiological conditions, for 240 minutes. Circuits were sampled at 15, 30, 60, 120 and 240 minutes and assessed for levels of interleukin-1ß, interleukin-6, interleukin-8, interleukin-10, tumour necrosis factor-α, transforming growth factor-ß1, myeloperoxidase and α-Defensin-1. In addition, haemoglobin, platelet and leukocyte counts were performed. RESULTS: There was a trend towards reduced levels of pro-inflammatory cytokines in mesenchymal stem cell-treated circuits and a significant increase in transforming growth factor-ß1. Blood cells and markers of neutrophil activation were reduced in mesenchymal stem cell circuits during the length of the simulation. As previously reported, the addition of mesenchymal stem cells resulted in a reduction of flow and increased trans-oxygenator pressures in comparison to controls. CONCLUSIONS: The addition of mesenchymal stem cells during extracorporeal membrane oxygenation may cause an increase in transforming growth factor-ß1. This is despite their ability to adhere to the membrane oxygenator. Further studies are required to confirm these findings.

Platelet concentrates modulate myeloid dendritic cell immune responses.

Platelet transfusion has been reported to modulate the recipients' immune system. To date, the precise mechanism(s) driving poor patient outcomes (e.g., increased rate of mortality, morbidity, infectious complications and prolonged hospital stays) following platelet transfusion are largely undefined. To determine the potential for platelet concentrates (PC) to modulate responses of crucial immune regulatory cells, a human in vitro whole blood model of transfusion was established. Maturation and activation of human myeloid dendritic cells (mDC) and the specialized subset blood DC antigen (BDCA)3+ DC were assessed following exposure to buffy-coat derived PC at day (D)2 (fresh) and D5 (date-of-expiry). In parallel, to model recipients with underlying viral or bacterial infection, polyinosinic:polycytidylic acid or lipopolysaccharide was added. Exposure to PC had less of an impact on mDC responses than BDCA3+ DC responses. PC alone downregulated BDCA3+ DC expression of co-stimulatory molecules CD40 and CD80. In the model of viral infection, PC downregulated expression of CD83, and in the bacterial model of infection, PC downregulated CD80, CD83, and CD86. PC alone suppressed mDC production of interleukin (IL)-8, IL-12 and tumor necrosis factor (TNF)-α and BDCA3+ DC production of IL-8, IL-12, and IL-6. In the model of viral infection, production of IL-12 and interferon-gamma inducible protein (IP)-10 was reduced in both DC subsets, and IL-8 was reduced in BDCA3+ DC following PC exposure. When modeling bacterial infection, PC suppressed mDC and BDCA3+ DC production of IL-6 and IL-10 with a reduction in TNF-α evident in mDC. This study assessed the impact of PC "transfusion" on DC surface antigen expression and inflammatory mediator production and provided the first evidence that PC transfusion modulates blood mDC and BDCA3+ DC maturation and activation, particularly in the models of infection. Results of this study suggest that patients who receive PC, particularly those with underlying infectious complications, may fail to establish an appropriate immune response precipitating poor patient outcomes.

Immunomodulatory effect of cryopreserved platelets: altered BDCA3+ dendritic cell maturation and activation in vitro.

BACKGROUND: Cryopreservation of platelets (PLTs) is useful in remote areas to overcome logistic problems associated with supply and can extend the shelf life to 2 years. During cryopreservation, properties of PLTs are modified. Whether changes in the cryopreserved PLT (CPP) product are associated with modulation of recipients' immune function is unknown. We aimed to characterize the immune profile of myeloid dendritic cells (mDCs) and the specialized blood DC antigen (BDCA)3+ subset after exposure to CPPs. STUDY DESIGN AND METHODS: Using an in vitro whole blood model of transfusion, the effect of CPPs on mDC and BDCA3+ DC surface antigen expression and inflammatory mediator production was examined using flow cytometry. In parallel, polyinosinic:polycytidylic acid (poly(I:C)) or lipopolysaccharide (LPS) was utilized to model processes activated in viral or bacterial infection, respectively. RESULTS: Cryopreserved PLTs had minimal impact on mDC responses but significantly modulated BDCA3+ DC responses in vitro. Exposure to CPPs alone up regulated BDCA3+ DC CD86 expression and suppressed interleukin (IL)-8, tumor necrosis factor (TNF)-α, and interferon-γ inducible protein (IP)-10 production. In both models of infection-related processes, exposure to CPPs down regulated BDCA3+ DC expression of CD40, CD80, and CD83 and suppressed BDCA3+ DC production of IL-8, IL-12, and TNF-α. CPPs suppressed CD86 expression in the presence of LPS and IP-10 and IL-6 production with poly(I:C). CONCLUSION: Cryopreserved PLTs may be immunosuppressive, and this effect is more evident when processes associated with infection are concurrently activated, especially for BDCA3+ DCs. This suggests that transfusion of CPPs in patients with infection may result in impaired BDCA3+ DC responses.

Discrete responses of erythrocytes, platelets, and von Willebrand factor to shear.

Despite decades of technological advancements in blood-contacting medical devices, complications related to shear flow-induced blood trauma are still frequently observed in clinic. Blood trauma includes haemolysis, platelet activation, and degradation of High Molecular Weight von Willebrand Factor (HMW vWF) multimers, all of which are dependent on the exposure time and magnitude of shear stress. Specifically, accumulating evidence supports that when blood is exposed to shear stresses above a certain threshold, blood trauma ensues; however, it remains unclear how various constituents of blood are affected by discrete shears experimentally. The aim of this study was to expose blood to discrete shear stresses and evaluate blood trauma indices that reflect red cell, platelet, and vWF structure. Citrated human whole blood (n = 6) was collected and its haematocrit was adjusted to 30 ± 2% by adding either phosphate buffered saline (PBS) or polyvinylpyrrolidone (PVP). Viscosity of whole blood was adjusted to 3.0, 12.5, 22.5 and 37.5 mPa·s to yield stresses of 3, 6, 9, 12, 50, 90 and 150 Pa in a custom-developed shearing system. Blood samples were exposed to shear for 0, 300, 600 and 900 s. Haemolysis was measured using spectrophotometry, platelet activation using flow cytometry, and HMW vWF multimer degradation was quantified with gel electrophoresis and immunoblotting. For tolerance to 300, 600 and 900 s of exposure time, the critical threshold of haemolysis was reached after blood was exposed to 90 Pa for 600 s (P < 0.05), platelet activation and HMW vWF multimer degradation were 50 Pa for 600 s and 12 Pa for 300 s respectively (P < 0.05). Our experimental results provide simultaneous comparison of blood trauma indices and thus also the relation between shear duration and magnitude required to induce damage to red cells, platelets, and vWF. Our results also demonstrate that near-physiological shear stress (<12 Pa) is needed in order to completely avoid any form of blood trauma. Therefore, there is an urgent need to design low shear-flow medical devices in order to avoid blood trauma in this blood-contacting medical device field.

The "Intermediate" CD14 + CD16 + monocyte subpopulation plays a role in IVIG responsiveness of children with Kawasaki disease.

BACKGROUND: Kawasaki disease (KD) is an acute, self-limited febrile illness of unknown cause. Intravenous immunoglobulin (IVIG)-resistance are related to greater risk for permanent cardiac complications. We aimed to determine the correlation between monocytes and the phenotype of KD in relation to IVIG responsiveness in children. MATERIALS AND METHODS: The study cohort included 62 patients who were diagnosed with KD, 20 non febrile healthy controls (NFC), and 15 other febrile controls (OFC). In all enrolled patients, blood was taken at least 4 times and laboratory tests were performed. In addition, subtypes of monocytes were characterized via flow cytometry. RESULTS: The numbers of intermediate monocytes were significantly lower in IVIG-resistant group compared to IVIG-responsive group before IVIG infusion (p < 0.0001). After infusion, intermediate monocytes decreased in the responsive group, while a trend of increase was observed in the resistant group. Only intermediate monocytes were significant in logistic regression with adjusted OR of 0.001 and p value of 0.03. CONCLUSIONS: CD14 + CD16 + intermediate monocyte may play an important role in IVIG responsiveness among KD children. Low starting levels of intermediate monocytes, followed by a dramatic increase post-IVIG infusion during acute phase of KD are associated with IVIG-resistance. Functional studies on intermediate monocyte may help to reveal the pathophysiology.

Coagulation Dysfunction in Acute Respiratory Distress Syndrome and Its Potential Impact in Inflammatory Subphenotypes.

The Acute Respiratory Distress Syndrome (ARDS) has caused innumerable deaths worldwide since its initial description over five decades ago. Population-based estimates of ARDS vary from 1 to 86 cases per 100,000, with the highest rates reported in Australia and the United States. This syndrome is characterised by a breakdown of the pulmonary alveolo-epithelial barrier with subsequent severe hypoxaemia and disturbances in pulmonary mechanics. The underlying pathophysiology of this syndrome is a severe inflammatory reaction and associated local and systemic coagulation dysfunction that leads to pulmonary and systemic damage, ultimately causing death in up to 40% of patients. Since inflammation and coagulation are inextricably linked throughout evolution, it is biological folly to assess the two systems in isolation when investigating the underlying molecular mechanisms of coagulation dysfunction in ARDS. Although the body possesses potent endogenous systems to regulate coagulation, these become dysregulated and no longer optimally functional during the acute phase of ARDS, further perpetuating coagulation, inflammation and cell damage. The inflammatory ARDS subphenotypes address inflammatory differences but neglect the equally important coagulation pathway. A holistic understanding of this syndrome and its subphenotypes will improve our understanding of underlying mechanisms that then drive translation into diagnostic testing, treatments, and improve patient outcomes.

Anti-thrombogenic Surface Coatings for Extracorporeal Membrane Oxygenation: A Narrative Review.

Extracorporeal membrane oxygenation (ECMO) is used in critical care to manage patients with severe respiratory and cardiac failure. ECMO brings blood from a critically ill patient into contact with a non-endothelialized circuit which can cause clotting and bleeding simultaneously in this population. Continuous systemic anticoagulation is needed during ECMO. The membrane oxygenator, which is a critical component of the extracorporeal circuit, is prone to significant thrombus formation due to its large surface area and areas of low, turbulent, and stagnant flow. Various surface coatings, including but not limited to heparin, albumin, poly(ethylene glycol), phosphorylcholine, and poly(2-methoxyethyl acrylate), have been developed to reduce thrombus formation during ECMO. The present work provides an up-to-date overview of anti-thrombogenic surface coatings for ECMO, including both commercial coatings and those under development. The focus is placed on the coatings being developed for oxygenators. Overall, zwitterionic polymer coatings, nitric oxide (NO)-releasing coatings, and lubricant-infused coatings have attracted more attention than other coatings and showed some improvement in in vitro and in vivo anti-thrombogenic effects. However, most studies lacked standard hemocompatibility assessment and comparison studies with current clinically used coatings, either heparin coatings or nonheparin coatings. Moreover, this review identifies that further investigation on the thrombo-resistance, stability and durability of coatings under rated flow conditions and the effects of coatings on the function of oxygenators (pressure drop and gas transfer) are needed. Therefore, extensive further development is required before these new coatings can be used in the clinic.

In vitro Hemocompatibility Evaluation of the HeartWare Ventricular Assist Device Under Systemic, Pediatric and Pulmonary Support Conditions.

The development of adult use right ventricular assist devices (RVADs) and pediatric left ventricular assist devices (pediatric LVADs) have significantly lagged behind compared to adult use left ventricular assist devices (LVADs). The HeartWare ventricular assist device (HVAD) intended to be used for adult's systemic support, is increasingly used off-label for adult pulmonary and pediatric systemic support. Due to different hemodynamics and physiology, however, the HVAD's hemocompatibility profiles can be drastically different when used in adult pulmonary circulation or in children, compared to its intended usage state, which could have a direct clinical and developmental relevance. Taking these considerations in mind, we sought to conduct in vitro hemocompatibility testing of HVAD in adult systemic, pediatric systemic and adult pulmonary support conditions. Two HVADs coupled to custom-built blood circulation loops were tested for 6 hours using bovine blood at 37°C under adult systemic, pediatric systemic, and adult pulmonary flow conditions (flow rate = 5.0, 2.5, and 4.5 L/min; differential pressure = 100, 69, and 20 mm Hg, respectively). Normalized index of hemolysis for adult systemic, pediatric systemic, and adult pulmonary conditions were 0.0083, 0.0039, and 0.0017 g/100 L, respectively. No significant difference was seen in platelet activation for these given conditions. High molecular weight von Willebrand factor multimer degradation was evident in all conditions (p < 0.05). In conclusion, alterations in the usage mode produce substantial differences in hemocompatibility of the HVAD. These findings would not only have clinical relevance but will also facilitate future adult use RVAD and pediatric LVAD development.

Extracorporeal Membrane Oxygenation-Induced Hemolysis: An In Vitro Study to Appraise Causative Factors.

In vitro hemolysis testing is commonly used to determine hemocompatibility of ExtraCorporeal Membrane Oxygenation (ECMO). However, poor reproducibility remains a challenging problem, due to several unidentified influencing factors. The present study investigated potential factors, such as flow rates, the use of anticoagulants, and gender of blood donors, which could play a role in hemolysis. Fresh human whole blood was anticoagulated with either citrate (n = 6) or heparin (n = 12; 6 female and 6 male blood donors). Blood was then circulated for 360 min at 4 L/min or 1.5 L/min. Regardless of flow rate conditions, hemolysis remained unchanged over time in citrated blood, but significantly increased after 240 min circulation in heparinized blood (p ≤ 0.01). The ratio of the normalized index of hemolysis (NIH) of heparinized blood to citrated blood was 11.7-fold higher at 4 L/min and 16.5-fold higher at 1.5 L/min. The difference in hemolysis between 1.5 L/min and 4 L/min concurred with findings of previous literature. In addition, the ratio of NIH of male heparinized blood to female was 1.7-fold higher at 4 L/min and 2.2-fold higher at 1.5 L/min. Our preliminary results suggested that the choice of anticoagulant and blood donor gender could be critical factors in hemolysis studies, and should be taken into account to improve testing reliability during ECMO.

Characterizing preclinical sub-phenotypic models of acute respiratory distress syndrome: An experimental ovine study.

The acute respiratory distress syndrome (ARDS) describes a heterogenous population of patients with acute severe respiratory failure. However, contemporary advances have begun to identify distinct sub-phenotypes that exist within its broader envelope. These sub-phenotypes have varied outcomes and respond differently to several previously studied interventions. A more precise understanding of their pathobiology and an ability to prospectively identify them, may allow for the development of precision therapies in ARDS. Historically, animal models have played a key role in translational research, although few studies have so far assessed either the ability of animal models to replicate these sub-phenotypes or investigated the presence of sub-phenotypes within animal models. Here, in three ovine models of ARDS, using combinations of oleic acid and intravenous, or intratracheal lipopolysaccharide, we investigated the presence of sub-phenotypes which qualitatively resemble those found in clinical cohorts. Principal Component Analysis and partitional clustering identified two clusters, differentiated by markers of shock, inflammation, and lung injury. This study provides a first exploration of ARDS phenotypes in preclinical models and suggests a methodology for investigating this phenomenon in future studies.

Current Understanding of Leukocyte Phenotypic and Functional Modulation During Extracorporeal Membrane Oxygenation: A Narrative Review.

A plethora of leukocyte modulations have been reported in critically ill patients. Critical illnesses such as acute respiratory distress syndrome and cardiogenic shock, which potentially require extracorporeal membrane oxygenation (ECMO) support, are associated with changes in leukocyte numbers, phenotype, and functions. The changes observed in these illnesses could be compounded by exposure of blood to the non-endothelialized surfaces and non-physiological conditions of ECMO. This can result in further leukocyte activation, increased platelet-leukocyte interplay, pro-inflammatory and pro-coagulant state, alongside features of immunosuppression. However, the effects of ECMO on leukocytes, in particular their phenotypic and functional signatures, remain largely overlooked, including whether these changes have attributable mortality and morbidity. The aim of our narrative review is to highlight the importance of studying leukocyte signatures to better understand the development of complications associated with ECMO. Increased knowledge and appreciation of their probable role in ECMO-related adverse events may assist in guiding the design and establishment of targeted preventative actions.

Viability of Mesenchymal Stem Cells in an Ex Vivo Circulation System.

Extracorporeal membrane oxygenation (ECMO) is a well-known therapy for refractory cardiac and respiratory failure. Stem cell therapy has been investigated as an adjunctive treatment for use during ECMO, but little is known about the viability of stem cells during ECMO support. We evaluated the viability and activity of mesenchymal stem cells (MSCs) in ex vivo circulation (EVC) conditions. The experimental groups were divided into two subgroups: EVC with oxygenator (OXY group) and EVC without oxygenator (Non-OXY group). Mesenchymal stem cells (1.0 × 10) were injected into the EVC system. Cell counting, a lactate dehydrogenase (LDH) cytotoxicity assay, and the mitochondrial functions of viable MSCs were analyzed. The post-EVC oxygen consumption rate (OCR) was significantly lower than the pre-EVC OCR, regardless of whether the oxygenator was used. The LDH levels were significantly higher in the OXY group than in the Non-OXY group. The cellular loss was mainly due to lysis of the cells whereas the loss of cellular activity was attributed to the nonphysiologic condition itself, as well as the oxygenator. We concluded that direct infusion of MSCs during ECMO support did not serve as adjunctive therapy. Further studies are needed to improve the viability in an ECMO setting.

Heart Transplantation From Brain Dead Donors: A Systematic Review of Animal Models.

Despite advances in mechanical circulatory devices and pharmacologic therapies, heart transplantation (HTx) is the definitive and most effective therapy for an important proportion of qualifying patients with end-stage heart failure. However, the demand for donor hearts significantly outweighs the supply. Hearts are sourced from donors following brain death, which exposes donor hearts to substantial pathophysiological perturbations that can influence heart transplant success and recipient survival. Although significant advances in recipient selection, donor and HTx recipient management, immunosuppression, and pretransplant mechanical circulatory support have been achieved, primary graft dysfunction after cardiac transplantation continues to be an important cause of morbidity and mortality. Animal models, when appropriate, can guide/inform medical practice, and fill gaps in knowledge that are unattainable in clinical settings. Consequently, we performed a systematic review of existing animal models that incorporate donor brain death and subsequent HTx and assessed studies for scientific rigor and clinical relevance. Following literature screening via the U.S National Library of Medicine bibliographic database (MEDLINE) and Embase, 29 studies were assessed. Analysis of included studies identified marked heterogeneity in animal models of donor brain death coupled to HTx, with few research groups worldwide identified as utilizing these models. General reporting of important determinants of heart transplant success was mixed, and assessment of posttransplant cardiac function was limited to an invasive technique (pressure-volume analysis), which is limitedly applied in clinical settings. This review highlights translational challenges between available animal models and clinical heart transplant settings that are potentially hindering advancement of this field of investigation.

Low flow rate alters haemostatic parameters in an ex-vivo extracorporeal membrane oxygenation circuit.

BACKGROUND: Extracorporeal membrane oxygenation (ECMO) is a life-saving modality used to manage cardiopulmonary failure refractory to conventional medical and surgical therapies. Despite advances in ECMO equipment, bleeding and thrombosis remain significant complications. While the flow rate for ECMO support is well recognized, less is known about the minimum-rate requirements and haemostasis. We investigated the relationship between different ECMO flow rates, and their effect on haemolysis and coagulation. METHODS: Ten ex-vivo ECMO circuits were tested using donated, < 24-h-old human whole blood, with two flow rates: high-flow at 4 L/min (normal adult cardiac output; n = 5) and low-flow at 1.5 L/min (weaning; n = 5). Serial blood samples were taken for analysis of haemolysis, von Willebrand factor (vWF) multimers by immunoblotting, rotational thromboelastometry, platelet aggregometry, flow cytometry and routine coagulation laboratory tests. RESULTS: Low-flow rates increased haemolysis after 2 h (p = 0.02), 4 h (p = 0.02) and 6 h (p = 0.02) and the loss of high-molecular-weight vWF multimers (p = 0.01), while reducing ristocetin-induced platelet aggregation (p = 0.0002). Additionally, clot formation times were prolonged (p = 0.006), with a corresponding decrease in maximum clot firmness (p = 0.006). CONCLUSIONS: In an ex-vivo model of ECMO, low-flow rate (1.5 L/min) altered haemostatic parameters compared to high-flow (4 L/min). Observed differences in haemolysis, ristocetin-induced platelet aggregation, high-molecular-weight vWF multimers and clot formation time suggest an increased risk of bleeding complications. Since patients are often on ECMO for protracted periods, extended-duration studies are required to characterise long-term ECMO-induced haemostatic changes.

Packed Red Blood Cell Transfusion Modulates Myeloid Dendritic Cell Activation and Inflammatory Response In Vitro.

Transfusion of packed red blood cells (PRBCs) modulates patients' immune responses and clinical outcomes; however, the underpinning mechanism(s) remain unknown. The potential for PRBC to modulate myeloid dendritic cells (mDC) and blood DC antigen 3 was assessed using an in vitro transfusion model. In parallel, to model processes activated by viral or bacterial infection, toll-like receptor agonists polyinosinic:polycytidylic acid or lipopolysaccharide were added. Exposure to PRBC upregulated expression of CD83 and downregulated CD40 and CD80 on both DC subsets, and it suppressed production of interleukin (IL)-6, IL-8, IL-12, tumor necrosis factor-α, and interferon-gamma-inducible protein-10 by these cells. Similar effects were observed when modeling processes activated by concurrent infection. Furthermore, exposure to PRBC at date of expiry was associated with more pronounced effects in all assays. Our study suggests PRBC have an impact on recipient DC function, which may result in failure to establish an appropriate immune response, particularly in patients with underlying infection.