Beyond the microcirculation: sequestration of infected red blood cells and reduced flow in large draining veins in experimental cerebral malaria.

Sequestration of infected red blood cells (iRBCs) in the microcirculation is a hallmark of cerebral malaria (CM) in post-mortem human brains. It remains controversial how this might be linked to the different disease manifestations, in particular brain swelling leading to brain herniation and death. The main hypotheses focus on iRBC-triggered inflammation and mechanical obstruction of blood flow. Here, we test these hypotheses using murine models of experimental CM (ECM), SPECT-imaging of radiolabeled iRBCs and cerebral perfusion, MR-angiography, q-PCR, and immunohistochemistry. We show that iRBC accumulation and reduced flow precede inflammation. Unexpectedly, we find that iRBCs accumulate not only in the microcirculation but also in large draining veins and sinuses, particularly at the rostral confluence. We identify two parallel venous streams from the superior sagittal sinus that open into the rostral rhinal veins and are partially connected to infected skull bone marrow. The flow in these vessels is reduced early, and the spatial patterns of pathology correspond to venous drainage territories. Our data suggest that venous efflux reductions downstream of the microcirculation are causally linked to ECM pathology, and that the different spatiotemporal patterns of edema development in mice and humans could be related to anatomical differences in venous anatomy.

Red Blood Cell Contribution to Thrombosis in Polycythemia Vera and Essential Thrombocythemia.

Polycythemia vera (PV) and essential thrombocythemia (ET) are myeloproliferative neoplasms (MPN) characterized by clonal erythrocytosis and thrombocytosis, respectively. The main goal of therapy in PV and ET is to prevent thrombohemorrhagic complications. Despite a debated notion that red blood cells (RBCs) play a passive and minor role in thrombosis, there has been increasing evidence over the past decades that RBCs may play a biological and clinical role in PV and ET pathophysiology. This review summarizes the main mechanisms that suggest the involvement of PV and ET RBCs in thrombosis, including quantitative and qualitative RBC abnormalities reported in these pathologies. Among these abnormalities, we discuss increased RBC counts and hematocrit, that modulate blood rheology by increasing viscosity, as well as qualitative changes, such as deformability, aggregation, expression of adhesion proteins and phosphatidylserine and release of extracellular microvesicles. While the direct relationship between a high red cell count and thrombosis is well-known, the intrinsic defects of RBCs from PV and ET patients are new contributors that need to be investigated in depth in order to elucidate their role and pave the way for new therapeutical strategies.

Hypoxically stored RBC resuscitation in a rat model of traumatic brain injury and severe hemorrhagic shock.

This study aims to investigate the effects of hypoxically stored Red Blood Cells (RBCs) in a rat model of traumatic brain injury followed by severe hemorrhagic shock (HS) and resuscitation. RBCs were made hypoxic using an O2 depletion system (Hemanext Inc. Lexington, MA) and stored for 3 weeks. Experimental animals underwent craniotomy and blunt brain injury followed by severe HS. Rats were resuscitated with either fresh RBCs (FRBCs), 3-week-old hypoxically stored RBCs (HRBCs), or 3-week-old conventionally stored RBCs (CRBCs). Resuscitation was provided via RBCs transfusion equivalent to 70 % of the shed blood and animals were followed for 2 h. The control group was comprised of healthy animals that were not instrumented or injured. Post-resuscitation hemodynamics and lactate levels were improved with FRBCs and HRBCs, and markers of organ injury in the liver (Aspartate aminotransferase [AST]), lung (chemokine ligand 1 [CXCL-1] and Leukocytes count), and heart (cardiac troponin, Interleukin- 6 [IL-6] and Tumor Necrosis Factor Alpha[TNF-α]) were lower with FRBCs and HRBCs resuscitation compared to CRBCs. Following reperfusion, biomarkers for oxidative stress, lipid peroxidation, and RNA/DNA injury were assessed. Superoxide dismutase [SOD] levels in the HRBCs group were similar to the FRBCs group and levels in both groups were significantly higher than CRBCs. Catalase levels were not different than control values in the FRBCs and HRBCs groups but significantly lower with CRBCs. Thiobarbituric acid reactive substances [Tbars] levels were higher for both CRBCs and HRBCs. Hypoxically stored RBCs show few differences from fresh RBCs in resuscitation from TBI + HS and decreased organ injury and oxidative stress compared to conventionally stored RBCs.

In vivo measurement of RBC survival in patients with sickle cell disease before or after hematopoietic stem cell transplantation.

ABSTRACT: Stable, mixed-donor-recipient chimerism after allogeneic hematopoietic stem cell transplantation (HSCT) for patients with sickle cell disease (SCD) is sufficient for phenotypic disease reversal, and results from differences in donor/recipient-red blood cell (RBC) survival. Understanding variability and predictors of RBC survival among patients with SCD before and after HSCT is critical for gene therapy research which seeks to generate sufficient corrected hemoglobin to reduce polymerization thereby overcoming the red cell pathology of SCD. This study used biotin labeling of RBCs to determine the lifespan of RBCs in patients with SCD compared with patients who have successfully undergone curative HSCT, participants with sickle cell trait (HbAS), and healthy (HbAA) donors. Twenty participants were included in the analysis (SCD pre-HSCT: N = 6, SCD post-HSCT: N = 5, HbAS: N = 6, and HbAA: N = 3). The average RBC lifespan was significantly shorter for participants with SCD pre-HSCT (64.1 days; range, 35-91) compared with those with SCD post-HSCT (113.4 days; range, 105-119), HbAS (126.0 days; range, 119-147), and HbAA (123.7 days; range, 91-147) (P<.001). RBC lifespan correlated with various hematologic parameters and strongly correlated with the average final fraction of sickled RBCs after deoxygenation (P<.001). No adverse events were attributable to the use of biotin and related procedures. Biotin labeling of RBCs is a safe and feasible methodology to evaluate RBC survival in patients with SCD before and after HSCT. Understanding differences in RBC survival may ultimately guide gene therapy protocols to determine hemoglobin composition required to reverse the SCD phenotype as it relates directly to RBC survival. This trial was registered at www.clinicaltrials.gov as #NCT04476277.

Red blood cell distribution width is associated with sarcopenia risk in early-stage non-small cell lung cancer.

BACKGROUND: Sarcopenia has received increasing attention in non-small cell lung cancer (NSCLC). Red blood cell distribution width (RDW) is a significant component of the complete blood count and indicates the heterogeneity of erythrocyte volume. Little information is known about RDW in relation to sarcopenia in early-stage (IA-IIIA) NSCLC. The purpose of the present study was to investigate the association between RDW and sarcopenia risk in early-stage NSCLC patients. METHODS: This study included 378 patients with pathologically confirmed stage IA-IIIA NSCLC. Sarcopenia was defined by measuring the skeletal muscle index (SMI) at the eleventh thoracic vertebra level. The maximum Youden index on the receiver operating characteristic (ROC) curve was used to estimate the cutoff value for RDW to predict sarcopenia. Logistic regression analyses were carried out to assess the independent risk factors for sarcopenia in NSCLC. RESULTS: The ROC curve indicated that the best cutoff point for RDW to predict sarcopenia was 12.9 (sensitivity of 43.80% and specificity of 76.76%, respectively). Moreover, there were significant differences in hemoglobin (p < 0.001), comorbidities (p = 0.001), histological type (p = 0.002), and cancer stage (p = 0.032) between the high RDW and low RDW groups. Logistic regression analyses revealed that high RDW is an independent risk factor for sarcopenia in early-stage NSCLC. CONCLUSION: RDW is associated with sarcopenia risk in early-stage NSCLC.

Prognostic value of red blood cell distribution width and D-Dimer in diffuse large B-cell lymphoma: Systematic review and meta-analysis.

BACKGROUND: The significant role of red blood cell distribution width (RDW) and D-Dimer as prognostic factors in patients with some blood malignancies has been reported recently. AIM: We designed and performed a meta-analysis to investigate the prognostic roles of RDW and D-Dimer in subjects with diffuse large B-cell lymphoma (DLBCL). MATERIALS AND METHODS: We systematically reviewed PubMed-Medline, SCOPUS, EMBASE, Web of Science Core Collection, and Google Scholar up to the present to look for publications on prognostic effects of RDW and D-Dimer in DLBCL patients. For investigation of the associations between RDW and D-Dimer with the overall survival (OS) and progression-free survival (PFS) of the DLBCL cases, hazard ratio (HR) with 95% confidence intervals (CIs) was used. RESULTS: We included 13 eligible studies in the present meta-analysis. The results of pooled analysis showed that increased levels of RDW was related to poor OS (HR = 2.01, 95% CI: 1.62-2.48, p value <.01, I2 = 0%) and poor PFS (HR = 1.52, 95% CI: 1.24-1.85, p value <.01, I2 = 16%) among the DLBCL patients. Similarly, a significant relationship was found between increased D-Dimer and poor OS (HR = 2.30, 95% CI: 1.03-5.14, p value <.05, I2 = 95%) of the DLBCL patients as well. In addition, there was no significant heterogeneity in OS (p value H = 0.65) and PFS (p value H = 0.31) related to RDW among studies included in the meta-analysis. CONCLUSION: Our finding clearly confirmed that elevated RDW levels and D-Dimer were associated with adverse OS and PFS in DLBCL.

Effect of octreotide on oxidative stress in the erythrocyte and kidney tissue in adriamycin-induced experimental nephrotic syndrome model.

INTRODUCTION: Nephrotic syndrome (NS) is one of the reasons of end-stage kidney disease, and elucidating the pathogenesis and offer new treatment options is important. Oxidative stress might trigger pathogenesis systemically or isolated in the kidneys. Octreotide (OCT) has beneficial antioxidant effects. We aimed to investigate the source of oxidative stress and the effect of OCT on experimental NS model. METHODS: Twenty-four non-uremic Wistar albino rats were divided into 3 groups. Control group, 2 mL saline intramuscular (im); NS group, adriamycin 5 mg/kg intravenous (iv); NS treatment group, adriamycin 5 mg/kg (iv) and OCT 200 mcg/kg (im) were administered at baseline (Day 0). At the end of 21 days, creatinine and protein levels were measured in 24-hour urine samples. Erythrocyte and renal catalase (CAT) and thiobarbituric acid reactive substance (TBARS) were measured. Renal histology was also evaluated. RESULTS: There was no significant difference among the 3 groups in terms of CAT and TBARS in erythrocytes. Renal CAT level was lowest in NS group, and significantly lower than the control group. In treatment group, CAT level significantly increased compared with NS group. In terms of renal histology, tubular and interstitial evaluations were similar in all groups. Glomerular score was significantly higher in NS group compared with control group and it was significantly decreased in treatment group compared to NS group. CONCLUSIONS: Oxidative stress in NS might be due to the decrease in antioxidant protection mechanism in kidney. Octreotide improves antioxidant levels and histology in renal tissue and might be a treatment option.

Erythrocytic α-Synuclein and the Gut Microbiome: Kindling of the Gut-Brain Axis in Parkinson's Disease.

BACKGROUND: Progressive spreading of α-synuclein via gut-brain axis has been hypothesized in the pathogenesis of Parkinson's disease (PD). However, the source of seeding-capable α-synuclein in the gastrointestinal tract (GIT) has not been fully investigated. Additionally, the mechanism by which the GIT microbiome contributes to PD pathogenesis remains to be characterized. OBJECTIVES: We aimed to investigate whether blood-derived α-synuclein might contribute to PD pathology via a gut-driven pathway and involve GIT microbiota. METHODS: The GIT expression of α-synuclein and the transmission of extracellular vesicles (EVs) derived from erythrocytes/red blood cells (RBCs), with their cargo α-synuclein, to the GIT were explored with various methods, including radioactive labeling of RBC-EVs and direct analysis of the transfer of α-synuclein protein. The potential role of microbiota on the EVs transmission was further investigated by administering butyrate, the short-chain fatty acids produced by gut microbiota and studying mice with different α-synuclein genotypes. RESULTS: This study demonstrated that RBC-EVs can effectively transport α-synuclein to the GIT in a region-dependent manner, along with variations closely associated with regional differences in the expression of gut-vascular barrier markers. The investigation further revealed that the infiltration of α-synuclein into the GIT was influenced significantly by butyrate and α-synuclein genotypes, which may also affect the GIT microbiome directly. CONCLUSION: By demonstrating the transportation of α-synuclein through RBC-EVs to the GIT, and its potential association with gut-vascular barrier markers and gut microbiome, this work highlights a potential mechanism by which RBC α-synuclein may impact PD initiation and/or progression. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.

Clonal hematopoiesis and acquired genetic abnormalities of the red cell: An historical review.

Several syndromes affecting the red cell that mimic those induced by germline mutations may result from a somatic mutation that accompanies a myeloid malignancy. These syndromes are most notable in cases of myelodysplastic syndrome, but they are not limited to any one category of myeloid neoplasm. Their occurrence in males exceed the male predominance that is evident in myeloid neoplasms. The syndromes include disorders of globin chain synthesis (α- and ß-thalassemia), heme synthesis (erythropoietic porphyria and erythropoietic uroporphyria), red cell membrane structure (elliptocytosis and spherocytosis), red cell enzyme activity (pyruvate kinase deficiency, glucose-6-phosphate dehydrogenase deficiency) and lowered expression of red cell ABO blood group antigens. This historical review describes the path to uncovering these acquired syndromes and their causal somatic mutations, where known. These syndromes often go unrecognized because of the dominant concern of the primary neoplasm. They may add to the healthcare needs of the patient.

The ratio of lymphocyte/red blood cells and platelets/lymphocytes are predictive biomarkers for lymph node metastasis in patients with breast cancer.

BACKGROUND: Axillary lymph node metastasis (LNM) affects the progression of breast cancer. However, it is difficult to preoperatively diagnose axillary lymph node status with high sensitivity. Therefore, we hypothesized that platelets/lymphocytes ratio (PLR) and lymphocytes/ red blood cells ratio (LRR) might help in the prognosis of lymph node metastasis in T1-T2 breast cancer. METHODS: 166 patients (Chang Ning Maternity & Infant Health Institute) were included in our study, and the associations of PLR and LPR with lymph node metastasis were investigated. Peripheral blood was collected one week before the surgery, and the patients were divided into different categories based on their PLR and LRR. RESULTS: The incidence of LNM was significantly increased in the high PLR group (p= 0.002) compared with the low PLR group; LNM was also significantly increased in the low LRR group (p= 0.036) compared with the high LPR group. Further, our study revealed that high PLR (p< 0.001, OR = 4.397, 95% CI = 2.005-9.645), low LRR (p= 0.017, OR = 0.336, 95%CI = 0.136-0.825) and high clinical T stage (p< 0.001, OR = 3.929, 95%CI = 1.913-8.071) are independent predictors of LNM. CONCLUSIONS: PLR and LRR could be identified as predictors of LNM in patients with T1/T2 breast cancer.

The association between human blood clot analogue computed tomography imaging, composition, contraction, and mechanical characteristics.

BACKGROUND: Clot composition, contraction, and mechanical properties are likely determinants of endovascular thrombectomy success. A pre-interventional estimation of these properties is hypothesized to aid in selecting the most suitable treatment for different types of thrombi. Here we determined the association between the aforementioned properties and computed tomography (CT) characteristics using human blood clot analogues. METHODS: Clot analogues were prepared from the blood of 4 healthy human donors with 5 red blood cell (RBC) volume suspensions: 0%, 20%, 40%, 60% and 80% RBCs. Contraction was measured as the weight of the contracted clots as a percentage of the original suspension. The clots were imaged using CT with and without contrast to quantify clot density and density increase. Unconfined compression was performed to determine the high strain compressive stiffness. The RBC content was analysed using H&E staining. RESULTS: The 5 RBC suspensions formed only two groups of clots, fibrin-rich (0% RBCs) and RBC-rich (>90% RBCs), as determined by histology. The density of the fibrin-rich clots was significantly lower (31-38HU) compared to the RBC-rich clots (72-89HU), and the density increase of the fibrin-rich clots was significantly higher (82-127HU) compared to the RBC-rich clots (3-17HU). The compressive stiffness of the fibrin-rich clots was higher (178-1624 kPa) than the stiffness of the RBC-rich clots (6-526 kPa). Additionally, the degree of clot contraction was higher for the fibrin-rich clots (89-96%) compared to the RBC-rich clots (11-77%). CONCLUSIONS: CT imaging clearly reflects clot RBC content and seems to be related to the clot contraction and stiffness. CT imaging might be a useful tool in predicting the thrombus characteristics. However, future studies should confirm these findings by analysing clots with intermediate RBC and platelet content.

Possible detection of physiological state disorders with the help of medicinal leeches Hirudo verbana Carena, 1820.

In the experiment, 160 medicinal leeches of the species Hirudo verbana Carena, 1820 were studied. Medicinal leeches were fed on the blood of animals and people (conditionally healthy and diseased). Four leeches were taken from each animal/person. The animals were studied for 3 weeks. Mortality was mostly observed in the first days after feeding on the blood of the host. We noted mortality, the appearance of constrictions on the leeches' body, the intensity of the host blood spitting from their body. The host's blood was taken from their stomach on the first day after feeding. Hematological and immunological indicators of blood were determined in the taken blood of the host. As a result of the study of the blood of the sick, significant changes were found, compared to conditionally healthy ones. It was manifested by an increase in erythrocytes and leukocytes. The leukocyte formula looked like in most pathological conditions of the inflammatory process. The obtained indicators of the experiment make it possible to quickly assess the presence of physiological disorders in the early stages of the disease.

New erythrocyte parameters derived from the Coulter principle relate with red blood cell properties-A pilot study in diabetes mellitus.

In routine hematological instruments, blood cells are counted and sized by monitoring the impedance signals induced during their passage through a Coulter orifice. However, only signals associated with centered paths in the aperture are considered for analysis, while the rejected measurements, caused by near-wall trajectories, can provide additional information on red blood cells (RBC), as recent publications suggest. To assess usefulness of two new parameters in describing alterations in RBC properties, we performed a pilot study to compare blood samples from patients with diabetes mellitus (DM), frequent pathological condition associated with impairment in RBC deformability, versus controls. A total of 345 blood samples were analyzed: 225 in the DM group and 120 in the control group. A diagram of [Formula: see text] and [Formula: see text], the two new parameters derived from the analysis of impedancemetry pulses, was used to compare distribution of RBC subpopulations between groups. To discriminate RBC from DM and control individuals, based on our multiparametric analysis, we built a score from variables derived from [Formula: see text] matrix which showed good performances: area under the receiving operating characteristic curve 0.948 (0.920-0.970), p<0.0001; best discriminating value: negative predictive value 94.7%, positive predictive value was 78.4%. These results seem promising to approach RBC alterations in routine laboratory practice. The related potential clinically relevant outcomes remain to be investigated.

Red blood cell distribution width has a prognostic value for gastric cancer patients after gastrectomy: A pooling-up analysis.

Our study aims to investigate whether preoperative red blood cell distribution width (RDW) has a prognostic value for patients after gastric cancer (GC) surgery. We searched articles in 3 databases including PubMed, Embase, and the Cochrane Library on May 16th, 2022. The prognostic indicators included overall survival (OS) and disease-free survival (DFS). RevMan 5.3 (The Cochrane Collaboration, London, United Kingdom) and Stata V16.0 were used for statistical analysis. The Risk Of Bias In Non-randomized Studies-of Interventions tool was used to assess risk of bias of the included studies. Ten articles involving 2740 patients were included. RDW was a prognostic factor for OS (hazard ratio = 1.81, 95% confidence interval [CI] = 1.38-2.37, P < .01) and DFS (hazard ratio = 1.99, I2 = 26%, 95% CI = 1.53-2.58, P < .01) for GC patients. Meanwhile, there were some differences between the high RDW group and the low RDW group. We found more patients older than 60 years old (OR = 2.58, 95% CI = 1.08-6.13, P = .03), larger tumor diameter (OR = 1.95, 95% CI = 1.33-2.85, P < .01) and later T stage (OR = 1.91, 95% CI = 1.07-3.42, P = .03) in the high RDW group than the low RDW group. No statistic difference was found in gender, N stage, tumor node metastasis stage, vascular invasion, differentiation, and adjuvant therapy between the 2 groups (P > .05). RDW was an independent prognostic factor for both OS and DFS of GC patients. High RDW level were strongly associated with poor survival.

Altered granulocyte count and erythrocyte measures in middle-aged, healthy carriers of APOE and PICALM risk genes for Alzheimer's disease.

APOE­Îµ4 genotype (apolipoprotein E, epsilon 4) is the strongest genetic risk factor for Alzheimer's disease (AD). Despite years of research, it is still not known how it contributes to dementia development. APOE has been implicated in many AD pathology mechanisms, like Aß clearance, brain metabolism, changes within microglia and other glial functions and inflammatory processes. In fact, immunological/inflammatory processes are recently discussed as an important factor in Alzheimer's development and granulocyte profiles changes are reported in patients. However, the exact link between the immune system and risk­genes is unknown. In particular, it is not known whether and how they interact throughout the lifetime, before the disease onset. The aim of the study was to investigate the relationship between granulocyte count and the APOE/PICALM genes in healthy individuals with an increased genetic risk of AD. An exploratory analysis regarding other blood cells was also conducted. Blood samples were collected from 77 healthy middle­aged (50-63 years old) participants, who were also asked to complete a health and life­style questionnaires. Groups with different AD risk­genes were compared. Differences in granulocyte profiles were found in healthy carriers of AD risk­genes who had slightly elevated eosinophil levels as compared to non-risk carriers. An exploratory analysis showed some alteration in mean corpuscular hemoglobin content and concentration (MCH/MCHC) levels between risk­carriers subgroups and non-risk carriers. No other differences in blood count or lipoprotein profile were found between healthy APOE/PICALM risk­carriers and non-risk carriers. Longitudinal studies will reveal if and how those changes contribute to the development of AD pathology.

Inflammatory Bowel Disease-Associated Colorectal Cancer: Translational and Transformational Risks Posed by Exogenous Free Hemoglobin Alpha Chain, A By-Product of Extravasated Erythrocyte Macrophage Erythrophagocytosis.

Colonic inflammatory bowel disease (IBD) encompasses ulcerative colitis (UC) and Crohn's colitis (CC). Patients with IBD are at increased risk for colitis-associated colorectal cancer (CACRC) compared to the general population. CACRC is preceded by IBD, characterized by highly heterogenous, pharmacologically incurable, pertinacious, worsening, and immune-mediated inflammatory pathologies of the colon and rectum. The molecular and immunological basis of CACRC is highly correlated with the duration and severity of inflammation, which is influenced by the exogenous free hemoglobin alpha chain (HbαC), a byproduct of infiltrating immune cells; extravasated erythrocytes; and macrophage erythrophagocytosis. The exogenous free HbαC prompts oxygen free radical-arbitrated DNA damage (DNAD) through increased cellular reactive oxygen species (ROS), which is exacerbated by decreased tissue antioxidant defenses. Mitigation of the Fenton Reaction via pharmaceutical therapy would attenuate ROS, promote apoptosis and DNAD repair, and subsequently prevent the incidence of CACRC. Three pharmaceutical options that attenuate hemoglobin toxicity include haptoglobin, deferoxamine, and flavonoids (vitamins C/E). Haptoglobin's clearance rate from plasma is inversely correlated with its size; the smaller the size, the faster the clearance. Thus, the administration of Hp1-1 may prove to be beneficial. Further, deferoxamine's hydrophilic structure limits its ability to cross cell membranes. Finally, the effectiveness of flavonoids, natural herb antioxidants, is associated with the high reactivity of hydroxyl substituents. Multiple analyses are currently underway to assess the clinical context of CACRC and outline the molecular basis of HbαC-induced ROS pathogenesis by exposing colonocytes and/or colonoids to HbαC. The molecular immunopathogenesis pathways of CACRC herein reviewed are broadly still not well understood. Therefore, this timely review outlines the molecular and immunological basis of disease pathogenesis and pharmaceutical intervention as a protective measure for CACRC.

Correlation between peripheral blood hemoglobin/erythrocyte distribution width ratio and prognosis of patients with primary colorectal cancer.

The prognostic value of peripheral blood hemoglobin/red blood cell distribution width ratio (HRR) in colorectal cancer (CRC) is unclear. The aim of this study was to analyze the correlation between peripheral blood HRR and the prognosis of CRC. A retrospective study analyzed the medical records of 284 CRC patients who attended Linyi People Hospital between June 1, 2017 and June 1, 2021. The optimal diagnostic cutoff value for hemoglobin (Hb)/erythrocyte distribution width was determined by ROC curve as 3.098, and patients were divided into high- and low-level groups for comparative analysis of clinical data. Kaplan-Meier method was used for survival analysis, and logrank test was used to assess survival differences. In univariate and multifactorial analyses, Cox proportional risk regression models were used to assess independent risk factors for overall survival (OS) and progression-free survival (PFS). All statistical tests were bilateral probability tests with α = 0.05, and P < .05 was considered statistically significant. There were 284 patients who were finally included in the statistical analysis. Gender, tumor stage, Hb, platelets, and CEA were associated with PFS and OS. Tumor stage, Hb, and HRR (P < .05) were independent risk factors for PFS and OS. Low-level HRR was associated with poor patient prognosis. Low-level HRR is associated with poor patient prognosis and is a potential tumor prognostic marker.

Summary of validation considerations with real-life examples using both qualitative and semiquantitative flow cytometry assays.

In the clinical laboratory, flow cytometry assays are critical to providing diagnostic and prognostic information to the treating clinicians. A validation or verification provides confidence that the assay will yield reliable results that can be trusted to make critical medical decisions. The following performance specifications should be included in a validation for laboratory developed tests as needed: accuracy (or trueness), precision (reproducibility and repeatability), detection capability, selectivity, reference range, and sample and reagent stability. We define these terms and present our approach to validation of several common flow cytometry assays, including examples of a leukemia/lymphoma assay and a paroxysmal nocturnal hemoglobinuria (PNH) assay.

Three-dimensional lab-on-a-foil device for dielectrophoretic separation of cancer cells.

A simple, low-cost, three-dimensional (3D) lab-on-a-foil microfluidic device for dielectrophoretic separation of circulating tumor cells (CTCs) is designed and constructed. Disposable thin films are cut by xurography and microelectrode array are made with rapid inkjet printing. The multilayer device design allows the studying of spatial movements of CTCs and red blood cells (RBCs) under dielectrophoresis (DEP). A numerical simulation was performed to find the optimum driving frequency of RBCs and the crossover frequency for CTCs. At the optimum frequency, RBCs were lifted 120 µm in z-axis direction by DEP force, and CTCs were not affected due to negligible DEP force. By utilizing the displacement difference, the separation of CTCs (modeled with A549 lung carcinoma cells) from RBCs in z-axis direction was achieved. With the nonuniform electric field at optimized driving frequency, the RBCs were trapped in the cavities above the microchannel, whereas the A549 cells were separated with a high capture rate of 86.3% ± 0.2%. The device opens not only the possibility for 3D high-throughput cell separation but also for future developments in 3D cell manipulation through rapid and low-cost fabrication.

Mathematical Analysis for the Flow of Sickle Red Blood Cells in Microvessels for Bio Medical Application.

Sickle cell disease (SCD) is an inherited monogenic disease which is characterized by distorted red blood cells (RBCs) that cause vaso-occlusion and vasculopathy. In the pathogenesis of SCD, polymerized hemoglobin turn RBCs into fragile, less deformable cells, and are subsequently more susceptible to endothelial adhesion after deoxygenation. Presently, electrophoresis and genotyping are used as routine tests for diagnosis of SCD. These techniques are expensive and require specialized laboratories. Lab-on-a-chip technology is a low-cost microfluidics-based diagnostic tool which holds significant promise for rapid screening of RBC deformability. To explore the sickle RBC mechanics for screening purposes, we present a mathematical model for the flow of single RBC with altered rheological properties and slip effect on capillary wall in microcirculation. We consider single-file flow of cells through the axis symmetrical cylindrical duct, applying lubrication theory as plasma trapped between successive red blood cells. The rheological parameters used from published literature for normal RBC and corresponding variation has been taken for the purpose of this simulation to present the condition of the disease. An analytical solution has been found for realistic boundary conditions and results are simulated using MATLAB. We found that the height of plasma film in the capillary increases with increase in cell deformability and compliance which affects the forward flow velocity in the capillary. Rigid RBCs with increased adhesion between cell and capillary wall shows reduction in velocity and occurrence of vaso-occlusion events in extreme conditions. These rheological properties of the cells coupled with microfluidics mechanics can mimic the physiological condition and provides unique insights with novel possibilities for the design of microfluidics base diagnostic kit towards effective therapeutic intervention of SCD.