Leukocyte Filtration Research Articles

Abstract 477: Platelet microRNAs Crosstalk with Vascular Endothelial Cells in Diabetes

Platelets are major source of extracellular miRNAs (miRs) constituting the plasma miR-pool. Here we generated miR-profile of leukocyte depleted platelets (LDP) isolated from type-2 diabetic (T2DM) or healthy subjects and investigated mechanism of miRNA mediated vascular crosstalk. LDPs were obtained by leukocyte filtration of platelet rich plasma isolated from blood withdrawn from T2DM (6 males, age: 54.8±7.6 yr; HbA1c: 8.9±0.6%; BMI: 30.9±2.8kg/m2) and healthy subjects (10 males, age: 49.2±6.8yr; HbA1c: 5.3±0.2%; BMI: 26.6±3.8kg/m2). MiR-profile was generated using total RNA isolated from T2DM or healthy LDPs. Students t-test applied to compare groups and p≤0.05 considered significant. Among all detected 735 miRs (cutoff<29 Ct), 19.5% miRs were depleted, whereas 7.5% showed >2 fold increase in T2DM-LDPs. Compared to healthy LDPs (n=10), depletion of 14 platelet-enriched miR candidates, including miR-21 and -22, were validated by RTPCR in individual T2DM-LDPs (n=11) irrespective of aspirin intake or presence of coronary heart disease.Ex-vivo thrombin stimulation of washed healthy platelets increased the release of miRNA into cellular-supernatants and confocal microscopy confirmed the cellular uptake of platelet released miRNA-carrying microparticles into human coronary artery endothelial cells (ECs) within 2-24h. In addition, EC overexpression of miR-21 or -22 decreased cell proliferation, whereas miR-22 also attenuated their tube forming capacity. MiR-21 or -22 modulations inversely regulated gene expression of their common predicted target, the cell-cycle regulator gene- CDK-6. MiR concentrations were also measured in particulate debris and plasma derived from coronary aspirates retrieved during stent implantation from 3 T2DM or 4 non-diabetic patients. In line, higher miR-21 or -22 ratios in plasma to particulate debris in T2DM vs non-diabetic aspirates indicated miR release from particulate debris into coronary plasma. This in turn provides relatively richer miR-21 or -22 microenvironment to the vicinal vascular ECs. T2DM platelets show distinct miR-profile compared to healthy platelets and depletion of miRNAs may represent their active secretion from diabetic platelets. Altered coronary microenvironment may lead to altered cellular responses to injury in diabetic vessels.

Inflammation, Atrial Fibrillation and Cardiac Surgery: Current Medical and Invasive Approaches for the Treatment of Atrial Fibrillation.

Atrial fibrillation (AF) is a cardiac dysrhythmia commonly seen in clinical practice especially after cardiac surgery. It is associated with increased morbidity and mortality for the patients. The pathogenesis of AF is not exactly understood yet, but there is growing data about the relationship between AF and inflammation. Cardiac surgery itself is a big source for inflammation. It causes major surgical trauma, ischemia/reperfusion injury, hypothermia, low arterial pressure, and the equipment of cardiopulmonary bypass makes a large foreign surface thus it activates inflammatory response. There is a large number of data about the treatment options of AF and there are also strategies, which are related to reduction of inflammatory activation during cardiopulmonary bypass. In order to review the relationship between cardiac surgery, inflammation, AF and treatment strategies in patients with AF, we conducted a search through Pubmed for articles in English using the keywords: "atrial fibrillation, cardiac surgery, inflammation, medical therapy, surgical therapy, ablation therapy" from January 2012 to present. We also searched separately for each alternative treatment modality on Pubmed. To identify further articles, we also looked into related citations in review articles and commentaries. We searched thoroughly the guidelines published by the European Society of Cardiology (2016), and the American Heart Association/ American College of Cardiology/ Heart Rhythm Surgery (2014). Many studies concluded that inflammation contributes in the occurrence of AF. Inflammatory markers, such as CRP, interleukins and complements have high sensitivity and specificity for prediction of AF whether the patient having cardiac surgery or not. Betablockers, diltiazem and amiodarone are the most commonly used drugs for rate control in AF following surgery. Although there are some new therapeutic approaches to reduce postoperative inflammatory activation, such as the use of vitamins, fatty acids, statins, or technical improvements to cardiopulmonary bypass unit like miniaturized bypass circuits, heparin coating of the circuits, leukocyte filters, or various surgical approaches like off-pump coronary bypass surgery, we still need more effective strategies to reduce both postoperative inflammation and postoperative AF risk after cardiac surgery. Today we use more advanced invasive and surgical treatment strategies for AF although we need far more advanced technics to reduce perioperative inflammatory activation, which actually causes AF.

A Leukocyte Filter Does Not Provide Further Benefit During Ex Vivo Lung Perfusion.

Normothermic ex vivo lung perfusion (EVLP) allows for assessment and reconditioning of donor lungs. Although a leukocyte filter (LF) is routinely incorporated into the EVLP circuit; its efficacy remains to be determined. Twelve pig lungs were perfused and ventilated ex vivo in a normothermic state for 12 hours. Lungs (n = 3) were allocated to four groups according to perfusate composition and the presence or absence of a LF in the circuit (acellular ± LF, cellular ± LF). Acceptable physiologic lung parameters were achieved during EVLP; however, increased amounts of pro-inflammatory cytokines (TNF-α and IL-6) and leukocytes in the perfusate were observed despite the presence or absence of a LF. Analysis of cells washed off the LF demonstrates that it trapped leukocytes although being ineffective throughout perfusion as it became saturated over 12 hours of EVLP. We conclude that there is no objective evidence to support the routine incorporation of a LF during EVLP as it does not provide further benefit and its removal does not appear to cause harm. The lack of hypothesized benefit to a LF may be because of the saturation of the LF with donor leukocytes, leading to similar amounts of circulating leukocytes still present in the perfusate with and without a LF.

Leukocyte filtration lesion impairs functional coagulation in banked whole blood.

Whole blood (WB) transfusion is a promising alternative to component therapy in hemostatic resuscitation. Use of banked WB requires filtration of white blood cells (leukoreduction) and an established shelf life during which WB retains coagulant capacities. The goal of this study was to define the time course of coagulation stability in leukoreduced compared to unfiltered WB under standard refrigeration conditions. Twelve WB units were donated by healthy volunteers after routine screening. Five units underwent standard leukocyte filtration and five did not. Two units were aliquoted into filtered and unfiltered samples, with platelets added to each sample on day 14. Units were stored at 4°C and sampled on days 0, 1, 2, 3, 4, 5, 6, 7, 10, 14, 21, 28, and 35 for immediate thromboelastography (TEG) analysis, and centrifuged and stored at -80°C for later calibrated automated thrombogram and coagulation factor assays. K-dependent factors and fibrinogen were low normal, decreased slightly over 35 days and were similar between unfiltered and filtered units. Labile factors were better preserved in filtered units, although unfiltered units did not show impaired coagulation over 35 days. Filtered blood had delayed clot initiation on days 0, 1, and 2 as measured by TEG R (p < 0.021); slower clot progression (TEG α-angle) on days 0, 1, 2, 3, 4, 5, and 6 (p < 0.023); weaker final clot (TEG MA) on all days (p < 0.0001). Thrombin generation was delayed on day 28 (p = 0.046) and decreased on days 10, 21, 28, and 35 (p < 0.034). Addition of platelets to filtered WB rescued TEG MA. Filtered WB had decreased functional clotting capacity and thrombin generation and may not be suitable for hemostatic resuscitation as the sole blood product. Therapeutic, level IV.

Autologous Blood Transfusion for Postpartum Hemorrhage.

Postpartum hemorrhage (PPH) is a leading contributor to maternal morbidity and mortality in the United States and globally. Although the rate of PPH is generally decreasing nationally, severity of PPH appears to be increasing, potentially related to the various comorbidities associated with women of childbearing age. There is increasing evidence of risks associated with allogeneic blood transfusion, which has historically been the classic therapeutic approach for treatment to PPH. Pregnant women are particularly susceptible to the implications of sensitization to red cell antigens, a common sequela to allogenic blood transfusion. Autologous blood transfusion eliminates the potential of communicable disease transmission as well as the conceivable threat of a blood transfusion reaction. Recent technological advances allow cell salvage coupled with the use of a leukocyte filter to be used as an alternative approach for improving the outcome for women experiencing a PPH. Modest changes in standard operating procedure and continued training in use and application of cell salvaged blood may assist in minimizing negative outcomes from PPH. Salvaged blood has been demonstrated to be at least equal and often superior to banked blood. We discuss nursing implications for application of this technology for women with PPH. Continued research is warranted to evaluate the impact that application of cell salvage with filtration has on the patient experiencing a PPH.

Leukocyte filtration of the cardiotomy suction. Does it affect systemic leukocyte activation or pulmonary function?

Cardiopulmonary bypass is thought to propagate a global systemic response through contact with the non-physiological surfaces of the extracorporeal circuit, leading to the stimulation of leukocytes, their adherence to endothelial cells and the release of cytotoxic molecules. This, in turn, has been shown to accelerate pulmonary injury. This study tested a new leukocyte-filtration system (RemoweLL) against a conventional system with no leukocyte-depleting capacity to determine the efficacy of the filtration system and its effects on pulmonary function. Thirty patients underwent coronary artery bypass graft surgery using either the RemoweLL filtration system (15 patients) or a conventional cardiopulmonary bypass circuit (15 patients). Data were collected on the total number of leukocytes, their differentiation and activation, using the leukocyte adhesion integrin CD11b as a surrogate marker. Pulmonary function was assessed using the Alveolar-arterial Oxygenation Index (AaOI) and patients were categorized using the Berlin definition of acute respiratory distress syndrome (ARDS). Both groups showed significant increases in leukocyte numbers during CPB (p<0.001), with no differences noted between the groups. CD11b showed a significant increase in both groups, with peak activation occurring at the end of CPB, but no difference between the groups (p=0.8). There was a trend towards lower AaOI increases in the filtration group, but this did not reach significance (p=0.075) and there was no difference in ARDS definitions (p=0.33). Leukocyte filtration of cardiotomy suction did not influence total leukocyte counts or activation as measured by CD11b upregulation. Furthermore, no evidence could be found to suggest improved pulmonary function.

Pathogen Inactivated Platelet Components Used without Leukofiltration, or Gamma Irradiation Show Comparable Therapeutic Efficacy and Safety to Gamma Irradiated and Non-Leukofiltered Conventional Products for the Support of HSCT Patients in Hong Kong

▪Introduction: Depending on the conditioning intensity and graft source, mosthematopoietic stem cell transplant(HSCT) patients invariably experience a cytopenic phase of at least 2-3 weeks and require transfusion support until engraftment. All transfused blood components post-transplant need to be gamma irradiated to prevent transfusion associated graft versus host disease (TA-GVHD). Platelet component (PC) manufacturing methodologies differ around the globe. Currently, Hong Kong HSCT patients are supported with individual platelet rich plasma (PRP) PCs without leukocyte filtration (NLF), tested by short-term aerobic cultures for bacterial contamination (STABC), and gamma irradiated. When new PC manufacturing methodologies are considered for adoption, it is important to assess the immediate and longer term effects on transfused patients. Amotosalen+UVA light (A-UVA) pathogen reduction technology (INTERCEPT™ Blood System, Cerus Corporation, Concord, CA) inactivates bacteria, viruses, parasites and leukocytes by efficient covalent adduct modification of DNA and RNA. Treatment with A-UVA prevents TA- GVHD in an animal model, inhibits clonal T cell proliferation, prevents allogeneic antigen stimulation in mixed lymphocyte reactions, and inhibits transcription mediated cytokine production and early activation antigen expression (Corash et al, BMT 2004). The common strategy of prescribing gamma irradiated PC only for suspected "high risk" patients is sub-optimal, as failure to identify recipients at risk for TA-GVHD was responsible for 50% of reported cases (Kopolovic et al. Blood 2016). Treatment of PC with A-UVA addresses two key issues for HSCT patients: bacterial contamination, and universal prevention of TA-GVHD. Here, we compared the clinical support of HSCT patients with conventional PCs (C-PC) vs. an equivalent HSCT group supported with A-UVA PCs (I-PC) using a sequential patient cohort design within a single HSCT clinical center.Methods: I-PCs were prepared from 5 pooled ABO-matched, NLF buffy coat PCs in platelet additive solution and treated with A-UVA replacing gamma irradiation and bacterial detection. C-PCs were prepared from 5 ABO-matched, STABC-negative, NLF PRP PCs, and treated with gamma irradiation. Each patient could receive up to 5 PC transfusions. The primary efficacy endpoint was the 1-hour corrected count increment (CCI) and the primary safety endpoint was the proportion of patients (P) with acute transfusion reactions (ATR). Follow-up data on mortality, hematopoiesis engraftment, and immune status were collected up to 100 days post HSCT.Results: Patient demographics and type of HSCT were similar between patient cohorts. 33 patients received 76 A-UVA PCs and 31 patients received 89 C-PCs. The mean days of platelet support (p=0.618) and mean 1-hour CCIs per patient averaged for all transfusions were comparable (p=0.296) between the I- PC and conventional C-PC cohorts (Table). The proportion of patients with AEs was lower (p=0.021) for the I-PC group (Table), and no related SAEs were observed during the entirety of trial. Survival at 100 days post HSCT and rates of remission were similar between the cohorts (Table). The ATR rate trended lower, although not significantly different (p=0.296), in the I- PC group (Table). Follow-up data showed that the patients had comparable neutrophil and platelet engraftment (Table) with comparable immune system reconstitution by 100 days post HSCT. No cases of TA-GVHD were observed in either cohort.Conclusions: A-UVA-treated PCs prepared without LF, gamma irradiation, and bacterial detection can replace C-PCs for support of HSCT patients resulting in comparable post transfusion CCI responses and short and intermediate term clinical outcomes, while offering additional protection against transfusion transmitted bacteria and emerging or untested pathogens. [Display omitted] DisclosuresSim Pui Yin:Cerus Corporation: Other: Investigator sponsored trial. Leung Yuk Yan:Cerus Corporation: Other: Investigator sponsored trial. Lam Chun Kit:Cerus Corporation: Other: Investigator sponsored trial. Tsoi Wai:Cerus Corporation: Other: Investigator sponsored trial. Lee Cheuk:Cerus Corporation: Other: Investigator sponsored trial. Lie Kwok Wai:Cerus Corporation: Other: Investigator sponsored trial. Huang:Cerus Corporation: Employment. Rico:Cerus Corporation: Employment. Lin:Cerus corp: Employment. Corash:Cerus Corporation: Employment. Stassinopoulos:Cerus Corporation: Employment.

Leukocyte filtration and postoperative infections

BackgroundLeukocyte filtration has been hypothesized to reduce the risk of postoperative infections by alleviating the immunosuppressive effect of whole blood. However, the literature regarding the clinical efficacy of leukocyte filtration remains conflicted. This meta-analysis investigates the impact of allogeneic and autologous leukocyte-filtered blood transfusions on the incidence of postoperative infections in adult surgical patients. MethodsA comprehensive literature search of PubMed, Google Scholar, and Cochrane Central Registry of Controlled trials (1966-2016) was completed for all published randomized controlled trials. Postoperative infections under “as-per-protocol” (APP) and “intention-to-treat” (ITT), length of stay, and mortality were analyzed. ResultsSixteen randomized controlled trials involving 6586 randomized (ITT) patients (4615 APP patients) in various clinical settings were evaluated. The leukocyte-filtered blood group demonstrated an overall 26% risk reduction in postoperative infections when analyzed by APP (relative risk [RR] = 0.74; 95% confidence interval [CI, 0.60-0.92]; P = 0.007) and a 22% risk reduction when analyzed by ITT (RR = 0.78; 95% CI [0.65-0.94]; P = 0.009). Leukocyte-filtered blood was also associated with a significant reduction in length of stay (standardized difference of mean [SDM] = −0.74; 95% CI [−1.32 to −0.15]; P = 0.014) and all-cause mortality (RR = 0.74; 95% CI [0.57-0.95]; P = 0.018). ConclusionsLeukocyte-filtered blood transfusions are associated with significantly lower postoperative infection rates in both the APP and ITT populations. Leukocyte filtration also shortens length of stay and decreases all-cause mortality in surgical patients and should be considered in all surgical patients.

Avoiding allogenic blood transfusions in endoscopic angiofibroma surgery.

BackgroundSurgical approaches for many tumours are often limited by blood loss, exposure and risk to vital anatomical structures. Therefore, the standard of care for certain skull base tumours has become endoscopic transnasal resection. Other surgical disciplines often use cell salvage techniques, but review of the otolaryngology literature revealed very few case reports. This study investigated the value and safety of salvage-type autologous blood transfusion during the endoscopic resection of juvenile nasopharyngeal angiofibromas (JNA).MethodsJNA is a rare vascular nasal tumour and the study extended over a 3-year period to obtain adequate patient numbers. All patients undergoing endoscopic resection during this period were included in the population sample. Ten patients with JNA were identified and underwent embolization prior to the endoscopic resection. In all cases the intraoperative blood salvage apparatus was used. Close post-operative monitoring was performed.ResultsHomologous blood transfusion could be avoided in all cases. Postoperative monitoring revealed transient bacteraemia in two cases where the leukocyte filter was not used, but no evidence of septicaemia.ConclusionsPerioperative cell saver and autologous blood transfusion in endonasal JNA surgery is safe. Homologous blood transfusion can be avoided by using this technique. The use of cell salvage allows for single stage surgery without the need to abandon surgery due to excessive blood loss and its future use is promising.

Abstract 16458: Paracrine Crosstalk Between Paltelets And Vascular Endothelial Cell is Mediated via MicroRNAs

Majority of extracellular miRNAs constituting plasma miRNA pool are released by platelets. This study investigated the miRNA signature of leukocyte depleted platelets (LDP) isolated from T2DM or healthy subjects and explored the mechanism of miRNA mediated vascular crosstalk. LDPs, obtained by leukocyte filtration of platelet rich plasma (PRP), were isolated from blood withdrawn from 6 T2DM patients (male, age: 54.8±7.6 yrs; HbA1c: 8.9±0.6%; BMI: 30.9±2.8kg/m2) and 10 healthy volunteers (male, age: 49.2±6.8yrs; HbA1c: 5.3±0.2%; BMI: 26.6±3.8kg/m2). Total RNA isolated from diabetic or healthy LDPs was subjected to PCR based miRNA profiling. Matrigel capillary formation and genomic BrdU incorporation were measured as readouts of angiogenesis and cell proliferation respectively. Cellular metabolic activities were measured by WST-1 assay. Student t-test was applied to compare groups and p&lt;0.05 considered significant. Total 735 (97%) miRNAs were detected (threshold&lt; 29 Ct) in all LDPs. Levels of 19.5% miRNAs were decreased in T2DM platelets, whereas 7.5% miRNAs showed increase at 2-50 fold cutoff value. Compared to healthy LDPs (n=10), decreased levels of 14 highly platelet enriched miRNAs candidates including miR-21 and miR-22 were confirmed by RTPCR validation of individual T2DM LDPs irrespective of aspirin intake or presence of coronary heart disease (n=11). Thrombin stimulation of washed healthy platelets increased miRNA release into cell supernatant and confocal microscopy confirmed the cellular uptake of platelet released micro-particles into human coronary artery endothelial cells within 2-24h. In addition, endothelial overexpression of miR-21 or miR-22 decreased cell proliferation and metabolic activity, whereas miR-22 also attenuated capillary formation capacity. MiR-21 and miR-22 modulation inversely regulated expression of cell-cycle regulator gene CDK-6 at mRNA and protein levels. Diabetic platelets show distinct miRNA signature compared to healthy platelets and lower levels of miRNAs in diabetic platelets may represent their active secretion during diabetes. Released miRNAs may mediate crosstalk with vascular cells by influencing cellular gene expression and function upon their uptake.

Acute limb ischemia: contemporary approach.

Acute limb ischemia is a critical condition with high mortality and morbidity even after surgical or endovascular intervention. Early recognition is important, but a delayed presentation is not uncommon. Viability of the limb is assessed by motor and sensory function and with interrogating Doppler flow signals in pedal arteries and popliteal veins as categorized by Rutherford. Category IIa indicates mild-to-moderate threat to limb salvage over a time frame without revascularization. Limb ischemia is critical without prompt revascularization in category IIb. Because the risk of reperfusion injury is high in this group of patients, perioperative management is important. In category III, reperfusion is not indicated except for embolism within several hours of onset. Intimal injury should be avoided by careful tactile control of a balloon with a smaller size catheter and under radiographic monitoring. Adjunctive treatment with catheter-directed thrombolysis or bypass surgery is sometimes necessary. Endovascular treatment is a promising option for thrombotic occlusion of an atherosclerotic artery. Ischemia-reperfusion injury is a serious problem. Controlled reperfusion with low-pressure perfusion at a reduced temperature and use of a leukocyte filter should be considered. The initial reperfusate is hyperosmolar, hypocalcemic, slightly alkaline, and contains free radical scavengers such as allopurinol. Immediate hemodialysis is necessary for acute renal injury caused by myoglobinemia. Compartment syndrome should be managed with assessment of intra-compartment pressure and fasciotomy.

Haematologic effects of leukoreduction on canine whole blood post-filtration and post-storage

Leukoreduction is the removal of leukocytes from a blood component to avoid the adverse reactions resulting from the exposure of leukocytes from a donor to the recipient during transfusion. A leukoreduced blood component has been defined as achieving a concentration of less than 5 × 106 residual donor leukocytes per final blood product while maintaining 85 % of the original red blood cell count. In the present study, we evaluated the efficiency of leukocyte filtration using a BioR 01-plus blood filter (Fresenius Hemocare, Germany) on samples of whole blood in citrate-phosphate-dextrose-adenine (CPDA-1) anticoagulant from 21 dogs. The blood cell count, red blood cells (RBCs), white blood cells (WBCs) and platelets; haemoglobin concentration; packed cell volume (PCV); mean corpuscular volume (MCV); RBC distribution width (RDW); morphologic index (MI) and RBC morphology were evaluated pre-filtration, post-filtration and after storage for 28 days at 4 °C. After filtration, a significant reduction in leukocyte (98 %) and platelet (95.1 %) counts were observed, while the RBCs, haemoglobin concentration and PCV remained within the recommended levels; storage did not change these parameters.

Challenge study of the pathogen reduction capacity of the THERAFLEX MB-Plasma technology.

Although most pathogen reduction systems for plasma primarily target viruses, bacterial contamination may also occur. This study aimed to investigate the bacterial reduction capacity of a methylene blue (MB) treatment process and its virus inactivation capacity in lipaemic plasma. Bacterial concentrations in plasma units spiked with different bacterial strains were measured before and after the following steps of the THERAFLEX MB-Plasma procedure: leucocyte filtration, MB/light treatment and MB filtration. Virus inactivation was investigated for three virus types in non-lipaemic, borderline lipaemic and highly lipaemic plasma. Leucocyte filtration alone efficiently eliminated most of the tested bacteria by more than 4 logs (Staphylococcus epidermidis and Staphylococcus aureus) or to the limit of detection (LOD) (≥ 4.8 logs; Escherichia coli, Bacillus cereus and Klebsiella pneumoniae). MB/light and MB filtration further reduced Staphylococcus epidermidis and Staphylococcus aureus to below the LOD. The small bacterium Brevundimonas diminuta was reduced by 1.7 logs by leucocyte filtration alone, and to below the LOD by additional MB/light treatment and MB filtration (≥ 3.7 logs). Suid herpesvirus 1, bovine viral diarrhoea virus and human immunodeficiency virus 1 were efficiently inactivated by THERAFLEX MB-Plasma, independent of the degree of lipaemia. THERAFLEX MB-Plasma efficiently reduces bacteria, mainly via the integrated filtration system. Its virus inactivation capacity is sufficient to compensate for reduced light transparency due to lipaemia.

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.

Effect of leukocyte filtration on the P-selectin expression of apheresis platelets.

The aim of this study was to investigate the effect of leukocyte filtration on the P-selectin (CD62P) surface expression of apheresis platelets during the retention period. Ten bags of apheresis platelets stored for 1 day (0-24 h) and 10 bags of apheresis platelets stored for 2 days (24-48 h) were used for leukocyte filtration (experimental group). Ten bags of apheresis platelets with the corresponding retention periods but without filtration were used as a negative control (control group). Thereafter, 100 μL of platelet suspensions from apheresis platelets with or without leukocyte filtration were sampled before and after leukocyte filtration for the detection of CD62P surface expression by flow cytometry. No statistical difference in the CD62P surface expression of apheresis platelets was observed before and after leukocyte filtration (P > 0.05), neither did the CD62P surface expression exhibit any change among the different retention periods. Leukocyte filtration does not affect the CD62P surface expression of apheresis platelets stored for up to 2 days, which indicates that leukocyte filtration does not damage the activation of apheresis platelets within the retention period.

Structural Characteristics of Blood Leukocyte-Filtration Material Produced by Electrospinning

Polymer—solvent systems for producing fibrous material for blood leukocyte filtration were selected. Electrospinning of the material was investigated. Its structure was characterized.

Impacts of leukocyte filtration and irradiation on coagulation factors in fresh frozen plasma.

This study aimed to compare and analyze the changes in the coagulation factors in fresh frozen plasma (FFP) prior to and following leukocyte filtration and irradiation. In total, 30 bags of FFP from healthy donors were processed: One-third of the FFP of each bag was left within the original bag (the A group), the other two-thirds of the FFP of each bag were passed through a disposable leukocyte filter, then divided equally into two parts. One of these was designated as the B group, and the other was designated the C group (subjected to 30 Gy irradiation). All samples were analyzed to evaluate 16 coagulation indicators. Analysis of variance revealed that there were statistically significant differences in the levels of fibrinogen (FbgC) and coagulation factor VIII (FVIII:C) among the groups (P=0.044 and P=0.015, respectively); the Dunnett’s t-test revealed that there was a statistically significant difference in the level of FbgC between the A and B groups (P=0.025), and there was a statistically significant difference in the level of FVIII:C between the A and C groups (P=0.009); while the remaining 14 coagulation parameters were not significantly different among the groups. Although the levels of FbgC and FVIII:C in the FFP were reduced following treatment, this would not affect the clinical effect of the FFP.

Leukodepletion as a point-of-care method for monitoring HIV-1 viral load in whole blood.

In order to limit the interference of HIV-1 cellular nucleic acids in estimating viral load (VL), the feasibility of leukodepletion of a small whole-blood (WB) volume to eliminate only leukocyte cell content was investigated, using a selection of filters. The efficacy of leukocyte filtration was evaluated by counting, CD45 quantitative PCR, and HIV-1 DNA quantification. Plasma HIV-1 was tested by real-time reverse transcription (RT)-PCR. A specific, miniaturized filter was developed and tested for leukocyte and plasma virus retention, WB sample dilution, and filtration parameters in HIV-1-spiked WB samples. This device proved effective to retain >99.9% of white blood cells in 100 μl of WB without affecting plasma VL. The Samba sample preparation chemistry was adapted to use a leukodepleted WB sample for VL monitoring using the point-of-care Samba-1 semiautomated system. The clinical performance of the assay was evaluated by testing 207 consecutive venous EDTA WB samples from HIV-1-infected patients attending a CD4 testing clinic. Most patients were on antiretroviral treatment (ART), but their VL status was unknown. Compared to the Roche Cobas AmpliPrep/Cobas TaqMan HIV-1 test, the new Samba assay had a concordance of 96.5%. The use of the Samba system with a VL test for WB might contribute to HIV-1 ART management and reduce loss-to-follow-up rates in resource-limited settings.

Attenuating the Systemic Inflammatory Response to Adult Cardiopulmonary Bypass: A Critical Review of the Evidence Base

A wide range of pharmacological, surgical, and mechanical pump approaches have been studied to attenuate the systemic inflammatory response to cardiopulmonary bypass, yet no systematically based review exists to cover the scope of anti-inflammatory interventions deployed. We therefore conducted an evidence-based review to capture “self-identified” anti-inflammatory interventions among adult cardiopulmonary bypass procedures. To be included, trials had to measure at least one inflammatory mediator and one clinical outcome, specified in the “Outcomes 2010” consensus statement. Ninety-eight papers satisfied inclusion criteria and formed the basis of the review. The review identified 33 different interventions and approaches to attenuate the systemic inflammatory response. However, only a minority of papers (35 of 98 [35.7%]) demonstrated any clinical improvement to one or more of the predefined outcome measures (most frequently myocardial protection or length of intensive care unit stay). No single intervention was supported by strong level A evidence (multiple randomized controlled trials [RCTs] or meta-analysis) for clinical benefit. Interventions at level A evidence included off-pump surgery, minimized circuits, biocompatible circuit coatings, leukocyte filtration, complement C5 inhibition, preoperative aspirin, and corticosteroid prophylaxis. Interventions at level B evidence (single RCT) for minimizing inflammation included nitric oxide donors, C1 esterase inhibition, neutrophil elastase inhibition, propofol, propionyl-L-carnitine, and intensive insulin therapy. A secondary analysis revealed that suppression of at least one inflammatory marker was necessary but not sufficient to confer clinical benefit. The most effective interventions were those that targeted multiple inflammatory pathways. These observations are consistent with a “multiple hit” hypothesis, whereby clinically effective suppression of the systemic inflammatory response requires hitting multiple inflammatory targets simultaneously. Further research is warranted to evaluate if combinations of interventions that target multiple inflammatory pathways are capable of synergistically reducing inflammation and improving outcomes after cardiopulmonary bypass.

Radiation Enhances Innate Immune Recognition of Cancer Stem Cells in Solid Tumors

Cancer stem cells (CSCs) are controversial and reported to be resistant to cytotoxic therapies such as radiation therapy (RT) and chemotherapy and responsible for cancer recurrences. They are defined by their ability to initiate tumors in vivo. A recent series of high impact studies have validated the critical importance of these cells. Previous studies demonstrate that natural killer (NK) cells are able to detect and reject allogeneic hematopoietic stem cells. We hypothesized that CSCs should be susceptible to NK immune attack and this susceptibility would be enhanced by the immunomodulatory effects of RT. All studies were approved by the UC Davis IRB and IACUC. Human cancer cells were xenografted into NOD-SCID IL2 receptor gamma chain null (NSG) mice. Primary human tumor tissue was obtained fresh from the UC Davis Cancer Center Biorepository. Human NK cells were isolated from leukocyte filters obtained from healthy donors. Localized RT was delivered using a clinical grade linear accelerator. Expression of CSC markers including aldehyde dehydrogenase (ALDH), CD133, CD24, and CD44 were analyzed by FACS using a BD Fortessa flow cytometer. Data were analyzed with FlowJo software version 7.2. Parametric and non-parametric statistical tests were performed as appropriate. We demonstrated that sorted CSCs were able to form tumors in vivo whereas non CSCs were very poorly tumorigenic. Coincubation with activated NK cells significantly reduced CSCs in cell lines (sarcoma 5±1% to 0.3±0.1%, breast 26±1.5 to 4±0.5%, pancreas 20±2% to 11±1.5%, P<0.05) and primary human tumors (liposarcoma 39% to 24%, pancreatic cancer 58% to 35%, P 120 days. This radio-immunotherapy regimen resulted in a synergistic decrease in CSC populations and enhanced survival in xenogeneic tumor models. These data suggest that NK cells can be used clinically to treat solid tumors and may provide the greatest benefit when used in combination with RT by targeting the population of CSCs that remain after RT has debulked the tumor of non-CSCs and increased the immunogenicity of the remaining CSC population. These finding could have profound consequences for recurrences after RT and provide additional evidence that combining RT and immunotherapy can lead to synergistic anti-tumor effects.